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Synthesis of autophagy inhibiting virantmycin analogs Zhang, Lingzhi 2015

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SYNTHESIS OF AUTOPHAGY INHIBITING VIRANTMYCIN ANALOGS  by  LINGZHI ZHANG  B.Eng., Jiangsu Normal University, 2012  A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF  MASTER OF SCIENCE in THE FACULTY OF GRADUATE AND POSTDOCTORAL STUDIES (Chemistry)  THE UNIVERSITY OF BRITISH COLUMBIA (Vancouver)  September 2015  © Lingzhi Zhang, 2015   ii Abstract  (–)-Virantmycin (1.12), first isolated from Streptomyces nitrosporeus in 1981, was found to be a potent inhibitor of autophagy with an IC50 of 0.5 µM against rapamycin-induced autophagy in MCF-7 cells. Recent studies showed that autophagy inhibition considerably reduced the growth of pancreatic ductal adenocarcinoma (PDAC) in mouse models. Therefore, virantmycin’s sub- µM potency as an early stage autophagy inhibitor makes it an interesting “lead compound” for the development of a treatment for PDAC. Previous attempts failed to make the benzoic acid from aryl iodide, using Kogen’s method. The current method for synthetic access to virantmycin analogs employs microwave irradiation to generate aryl nitriles, such as 2.144, for further installation of the carboxyl group at the aryl ring. Analogs 2.108 and 2.152 show the most potent autophagy inhibiting activity among the synthetic analogues prepared to date. The construction of simplified pharmacophore analogs 2.108 and 2.152 allows for scalable synthesis to provide quantities for animal testing.    iii Preface  Chapter 2 and 3 are based on work conducted at UBC. Virantmycin was isolated and characterized by Dr. Roberto Forestieri in the Andersen lab. I was responsible for planning synthetic routes, making and characterizing all synthetic analogues. Dr. Raymond J. Andersen provided technical suggestions. The biological data of all the synthetic compounds was collected by Aruna Balgi under the leadership of Dr. Michel Roberge in the Department of Biochemistry and Molecular Biology at UBC.   iv Table of Contents  Abstract .......................................................................................................................................... ii	  Preface ........................................................................................................................................... iii	  Table of Contents ......................................................................................................................... iv	  List of Tables .............................................................................................................................. viii	  List of Figures ............................................................................................................................... ix	  List of Schemes ............................................................................................................................. ix	  List of Symbols and Abbreviations .......................................................................................... xvi	  Acknowledgements ................................................................................................................... xxii	  Dedication ................................................................................................................................. xxiii	  Chapter 1: Introduction ................................................................................................................1	  1.1	   Drugs from nature .............................................................................................................. 1	  1.2	   Isolation of virantmycin as an inhibitor of autophagy ....................................................... 6	  1.2.1	   Autophagy ................................................................................................................... 6	  1.2.2	   Autophagy and cancer ................................................................................................. 8	  1.2.3	   Autophagy inhibitors ................................................................................................ 10	  1.2.4	   Virantmycin .............................................................................................................. 11	  1.3	   Tetrahydroquinoline natural products .............................................................................. 16	  1.4	   Proposed biosynthesis of bezastatins ............................................................................... 20	  Chapter 2: Synthesis of analogs of virantmycin for SAR studies ............................................23	  2.1	   Isolation and characterization of (–)-virantmycin ............................................................ 23	  2.2	   Prior syntheses ................................................................................................................. 24	    v 2.2.1	   Racemic synthesis by Raphael .................................................................................. 24	  2.2.2	   Racemic synthesis by Shirahama .............................................................................. 25	  2.2.3	   Asymmetric synthesis of unnatural (+)-virantmycin by Shirahama ......................... 27	  2.2.4	   Racemic synthesis by Corey ..................................................................................... 29	  2.2.5	   Enantioselective synthesis of (–)-virantmycin by Kogen ......................................... 31	  2.2.6	   Enantioselective synthesis of (–)-virantmycin by Back ............................................ 34	  2.3	   General synthetic analyses for SAR studies on virantmycin ........................................... 36	  2.4	   Retrosynthetic analysis of analog 2.1 .............................................................................. 37	  2.5	   Initial synthetic trials ........................................................................................................ 38	  2.6	   Synthesis of aryl nitriles ................................................................................................... 47	  2.7	   Synthesis of analogs with the saturated side chain .......................................................... 55	  Chapter 3: Conclusion .................................................................................................................61	  3.1	   Biological results ............................................................................................................. 61	  3.2	   Conclusion ....................................................................................................................... 62	  Chapter 4: Experimental .............................................................................................................64	  4.1	   Preparation of methyl ester 2.64 ...................................................................................... 65	  4.2	   Preparation of methyl ester 2.65 ...................................................................................... 67	  4.3	   Preparation of methyl ester 2.89 ...................................................................................... 69	  4.4	   Preparation of methyl ester 2.90 ...................................................................................... 71	  4.5	   Preparation of Weinreb amide 2.66 ................................................................................. 73	  4.6	   Preparation of Weinreb amide 2.91 ................................................................................. 75	  4.7	   Preparation of Weinreb amide 2.92 ................................................................................. 77	  4.8	   Preparation of bromide 2.94 ............................................................................................ 79	    vi 4.9	   Preparation of Grignard reagent 2.95 ............................................................................... 79	  4.10	   Preparation of ketone 2.97 ............................................................................................. 80	  4.11	   Preparation of ketone 2.96 ............................................................................................. 82	  4.12	   Preparation of alcohol 2.98 ............................................................................................ 84	  4.13	   Preparation of alcohol 2.99 ............................................................................................ 86	  4.14	   Preparation of alcohol 2.100 .......................................................................................... 88	  4.15	   Preparation of alcohol 2.101 .......................................................................................... 90	  4.16	   Preparation of ketone 2.102 ........................................................................................... 92	  4.17	   Preparation of alcohol 2.104 .......................................................................................... 94	  4.18	   Preparation of alcohol 2.105 .......................................................................................... 96	  4.19	   Preparation of tetrahydroquinoline 2.107 ...................................................................... 98	  4.20	   Preparation of alcohol 2.110 ........................................................................................ 100	  4.21	   Preparation of alcohol 2.111 ........................................................................................ 102	  4.22	   Preparation of alcohol 2.112 ........................................................................................ 104	  4.23	   Preparation of alcohol 2.106 ........................................................................................ 106	  4.24	   Preparation of tetrahydroquinoline 2.113 .................................................................... 108	  4.25	   Preparation of tetrahydroquinoline 2.114 .................................................................... 110	  4.26	   Preparation of tetrahydroquinoline 2.115 .................................................................... 112	  4.27	   Preparation of tetrahydroquinoline 2.108 .................................................................... 114	  4.28	   Preparation of nitrile 2.129 .......................................................................................... 116	  4.29	   Preparation of nitrile 2.130 .......................................................................................... 118	  4.30	   Preparation of ketone 2.140 ......................................................................................... 120	  4.31	   Preparation of ketone 2.141 ......................................................................................... 122	    vii 4.32	   Preparation of alcohol 2.142 ........................................................................................ 124	  4.33	   Preparation of alcohol 2.143 ........................................................................................ 126	  4.34	   Preparation of nitrile 2.144 .......................................................................................... 128	  4.35	   Preparation of alcohol 2.145 ........................................................................................ 130	  4.36	   Preparation of tetrahydroquinoline 2.146 .................................................................... 132	  4.37	   Preparation of alcohol 2.147 ........................................................................................ 134	  4.38	   Preparation of alcohol 2.148 ........................................................................................ 136	  4.39	   Preparation of tetrahydroquinoline 2.151 .................................................................... 138	  4.40	   Preparation of tetrahydroquinoline 2.152 .................................................................... 140	  References ...................................................................................................................................142	     viii List of Tables  Table 2.1 Optimization of the Grignard addition reaction conditions .......................................... 43	  Table 2.2 Attempts to install saturated side chains at C-2 ............................................................ 55	     ix List of Figures  Figure 1.1 Structures of plant-derived drugs .................................................................................. 1	  Figure 1.2 Structures of Spongothymidine (1.5), Ara-A (1.6), Ara-C (1.7) and AZT (1.8) ........... 3	  Figure 1.3 Amino-acid sequence and 3-D structure of ziconotide (1.9) ......................................... 4	  Figure 1.4 Halichondrin B (1.10) and E7389 (Eribulin, 1.11) ........................................................ 5	  Figure 1.5 Mechanism of autophagy .............................................................................................. 7	  Figure 1.6 Structure of (–)-virantmycin ........................................................................................ 11	  Figure 1.7 Virantmycin inhibiting rapamycin-induced autophagy64 ............................................ 12	  Figure 1.8 Effects of virantmycin (1.12) on autophagosome accumulation64 .............................. 13	  Figure 1.9 Stages of autophagy and sites of action of autophagy inhibitors64 .............................. 15	  Figure 1.10 Structures of benzastatins .......................................................................................... 17	  Figure 1.11 Structures of tetrahydroquinoline-based Pf-PFT inhibitors ....................................... 18	  Figure 1.12 γ-Secretase inhibitors derived from tetrahydroquinolines ......................................... 19	  Figure 1.13 Renin inhibitors derived from tetrahydroquinolines ................................................. 19	  Figure 2.1 Major structural elements in (–)-virantmycin (1.12) ................................................... 36	  Figure 2.2 Analogs 2.85 and 2.86 ................................................................................................. 37	  Figure 3.1 Virantmycin analogs synthesized ................................................................................ 61	  Figure 3.2 Current understanding of virantmycin’s structural features ........................................ 63	  Figure 4.1 1H and 13C NMR spectra of 2.64 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. .................................................................................................................................. 66	  Figure 4.2 1H and 13C NMR spectra of 2.65 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. .................................................................................................................................. 68	    x Figure 4.3 1H and 13C NMR spectra of 2.89 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. .................................................................................................................................. 70	  Figure 4.4 1H and 13C NMR spectra of 2.90 recorded in CDCl3 at 300 MHz and 75 MHz, respectively. .................................................................................................................................. 72	  Figure 4.5 1H and 13C NMR spectra of 2.66 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. .................................................................................................................................. 74	  Figure 4.6 1H and 13C NMR spectra of 2.91 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. .................................................................................................................................. 76	  Figure 4.7 1H and 13C NMR spectra of 2.92 recorded in CDCl3 at 300 MHz and 75 MHz, respectively. .................................................................................................................................. 78	  Figure 4.8 1H and 13C NMR spectra of 2.97 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. .................................................................................................................................. 81	  Figure 4.9 1H and 13C NMR spectra of 2.96 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. .................................................................................................................................. 83	  Figure 4.10 1H and 13C NMR spectra of 2.98 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. .................................................................................................................................. 85	  Figure 4.11 1H and 13C NMR spectra of 2.99 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. .................................................................................................................................. 87	  Figure 4.12 1H and 13C NMR spectra of 2.100 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. .................................................................................................................................. 89	  Figure 4.13 1H and 13C NMR spectra of 2.101 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. .................................................................................................................................. 91	    xi Figure 4.14 1H and 13C NMR spectra of 2.102 recorded in CDCl3 at 300 MHz and 150 MHz, respectively. .................................................................................................................................. 93	  Figure 4.15 1H and 13C NMR spectra of 2.104 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. .................................................................................................................................. 95	  Figure 4.16 1H and 13C NMR spectra of 2.105 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. .................................................................................................................................. 97	  Figure 4.17 1H and 13C NMR spectra of 2.107 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. .................................................................................................................................. 99	  Figure 4.18 1H and 13C NMR spectra of 2.110 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. ................................................................................................................................ 101	  Figure 4.19 1H and 13C NMR spectra of 2.111 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. ................................................................................................................................ 103	  Figure 4.20 1H and 13C NMR spectra of 2.112 recorded in CDCl3 at 600 MHz and 100 MHz, respectively. ................................................................................................................................ 105	  Figure 4.21 1H and 13C NMR spectra of 2.106 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 107	  Figure 4.22 1H and 13C NMR spectra of 2.113 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. ................................................................................................................................ 109	  Figure 4.23 1H and 13C NMR spectra of 2.114 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. ................................................................................................................................ 111	  Figure 4.24 1H and 13C NMR spectra of 2.115 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 113	    xii Figure 4.25 1H and 13C NMR spectra of 2.108 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 115	  Figure 4.26 1H and 13C NMR spectra of 2.129 recorded in CDCl3 at 400 MHz and 150 MHz, respectively. ................................................................................................................................ 117	  Figure 4.27 1H and 13C NMR spectra of 2.130 recorded in CDCl3 at 400 MHz and 150 MHz, respectively. ................................................................................................................................ 119	  Figure 4.28 1H and 13C NMR spectra of 2.140 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. ................................................................................................................................ 121	  Figure 4.29 1H and 13C NMR spectra of 2.141 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 123	  Figure 4.30 1H and 13C NMR spectra of 2.142 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. ................................................................................................................................ 125	  Figure 4.31 1H and 13C NMR spectra of 2.143 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 127	  Figure 4.32 1H and 13C NMR spectra of 2.144 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 129	  Figure 4.33 1H and 13C NMR spectra of 2.145 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 131	  Figure 4.34 1H and 13C NMR spectra of 2.146 recorded in CDCl3 at 400 MHz and 150 MHz, respectively. ................................................................................................................................ 133	  Figure 4.35 1H and 13C NMR spectra of 2.147 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 135	    xiii Figure 4.36 1H and 13C NMR spectra of 2.148 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 137	  Figure 4.37 1H and 13C NMR spectra of 2.151 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 139	  Figure 4.38 1H and 13C NMR spectra of 2.152 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. ................................................................................................................................ 141	    xiv List of Schemes  Scheme 1.1 Proposed biosynthesis of benzastatin family by Yoo78 ............................................. 20	  Scheme 1.2 Model study of indoline skeleton formation by epoxide cyclization78 ..................... 21	  Scheme 1.3 Model study of inter-conversion between tetrahydroquinolines and indolines78 ...... 22	  Scheme 2.1 Raphael’s total synthesis of (±)-virantmycin 2.1389 .................................................. 25	  Scheme 2.2 Shirahama’s total synthesis of (±)-2.14 and (±)-2.1590 ............................................. 26	  Scheme 2.3 Shirahama’s asymmetric total synthesis of (+)-virantmycin80 .................................. 28	  Scheme 2.4 Synthesis of building blocks 1.44 and 1.45 in Corey’s method92 ............................. 29	  Scheme 2.5 The completion of Corey’s total synthesis of (±)-virantmycin 2.4292 ...................... 30	  Scheme 2.6 Mechanism for the rearrangement of indolines to tetrahydroquinolines94 ................ 32	  Scheme 2.7 Kogen’s total synthesis of (–)-virantmycin94 ............................................................ 33	  Scheme 2.8 Desymmetrization of diester 1.73 to generate chiral center C295 ............................. 34	  Scheme 2.9 The completion of Back’s total synthesis of (–)-virantmycin 1.1295 ........................ 35	  Scheme 2.10 Retrosynthetic analysis of 2.63 ............................................................................... 38	  Scheme 2.11 Failed esterification of 2.63 ..................................................................................... 38	  Scheme 2.12 Synthesis of esters 2.89 and 2.90 ............................................................................ 39	  Scheme 2.13 Synthesis of alcohols of 2.98, 2.99, 2.100 and 2.101 .............................................. 40	  Scheme 2.14 Preparation of bromide 2.94 .................................................................................... 40	  Scheme 2.15 Rationale for stereochemical outcome of the addition reaction of 2.97 .................. 41	  Scheme 2.16 Metal-halogen exchanges resulting in deiodinated products .................................. 42	  Scheme 2.17 Attempts to synthesize analog 2.86 ......................................................................... 44	  Scheme 2.18 Attempts at early stage carboxylation ..................................................................... 44	    xv Scheme 2.19 Synthesis of tetrahydroquinolines of 2.113, 2.114, 2.115 and 2.108 ...................... 45	  Scheme 2.20 Attempts to install the carbxyl group on 2.115 ....................................................... 46	  Scheme 2.21 Attempts to install the carbonyl group on 2.112 ..................................................... 47	  Scheme 2.22 Attempts to convert 2.115 into 2.121 ...................................................................... 48	  Scheme 2.23 Plausible mechanism for the formation of formate 2.122 ....................................... 49	  Scheme 2.24 Factors in Pd-catalyzed cyanation reactions ........................................................... 50	  Scheme 2.25 Cyanation of aryl bromides assisted by microwave irradiation .............................. 52	  Scheme 2.26 Cyanation of aryl bromides assisted by microwave ................................................ 53	  Scheme 2.27 Attempts to hydrolyze the nitrile 2.130 to the ester 2.131 ...................................... 54	  Scheme 2.28 Attempts to install saturated side chains at C-2 ...................................................... 56	  Scheme 2.29 Hydrogenation of 2.97 ............................................................................................. 56	  Scheme 2.30 Synthesis of tetrahydroquinoline 2.146 ................................................................... 57	  Scheme 2.31 Synthesis of tetrahydroquinolines 2.151 and 2.152 ................................................ 58	  Scheme 2.32 Attempts to hydrolyze 2.146 to 2.153 ..................................................................... 59	  Scheme 2.33 Attempts to synthesize 2.155 .................................................................................. 60	  Scheme 3.1 Carboxylic acid 2.156 to be synthesized ................................................................... 62	     xvi List of Symbols and Abbreviations (±) racemic °C degrees Celsius % percent δ chemical shift in parts per million AD Alzheimer’s disease Ac acetyl B.C. Before Christ BINAPFu 2,2’-bis(diphenylphosphino)-3.3’-binaphtho[2,1-b] furan Boc t-butoxycarbonyl br broad BRSM based on recovered starting material Bu butyl Bu4NF tetra-n-butylammonium fluoride calcd calculated CQ chloroquine d doublet dba dibenzylideneacetone DCM dichloromethane dd doublet of doublets DEAD diethyl azodicarboxylate DIBAL diisobutylaluminium hydride   xvii DMAP 4-dimethylaminopyridine DMF N,N-dimethylformamide DMSO dimethyl sulfoxide DNA deoxyribonucleic acid DPPA diphenylphosphoryl azide dppp 1,3-bis(diphenylphosphino)propane ED50 half maximal effective concentration EGFP-LC3 enhanced green fluorescent protein-light chain 3 EtOAc ethyl acetate ESI-MS electrospray ionization mass spectrometry Et ethyl Et2O diethyl ether equiv. equivalent(s) FDA Food and Drug Administration g gram(s) Glu glutamic acid h hour(s) HCO2Ac acetic formic anhydride HCQ hydroxychloroquine HIV human immunodeficiency virus HOAc acetic acid HPLC high-performance liquid chromatography   xviii HREIMS high resolution electron impact mass spectroscopy HRESIMS high resolution electrospray ionization mass spectroscopy HTS high-throughput screening IC50 half maximal inhibitory concentration imid. imidazole i-PrMgCl isopropylmagnesium chloride i-PrPPh3I isopropyltriphenylphosphonium iodide J coupling constant KHMDS Potassium bis(trimethylsilyl)amide LC-MS liquid chromatography–mass spectrometry LC3 light chain 3 LiHMDS lithium bis(trimethylsilyl)amide LTBA lithium tri-tert-butoxyaluminum hydride M mole m multiplet MCF-7 Michigan Cancer Foundation-7 mCPBA meta-Chloroperoxybenzoic acid Me methyl MeCN acetonitrile MeOH methanol mg milligram(s) µg/mL microgram(s) per milliliter   xix MHz megahertz min minute(s) mL milliliter(s) µm micrometer(s) µM micromole(s) mM millimole(s) per milliliter(s) mmol millimol(s) mmol/mL millimole(s) per milliliter(s) MsOH methanesulfonic acid mTORC1 rapamycin complex 1 NBS N-bromosuccinimide (n-Bu)3P tributylphosphine NF-κB nuclear factor kappa-B nM nanomole(s) NMP N-methyl-2-pyrrolidone NMR nuclear magnetic resonance NOE nuclear overhauser effect NTBP 2,6-di-tert-butyl-4-methylpyridine PAS pre-autophagosomal structures PCC pyridinium chlorochromate PDAC pancreatic ductal adenocarcinoma Pf-PFT farnesyltransferase   xx PLE porcine liver esterase PPh3 triphenylphosphine PPTS pyridinium p-toluenesulfonate psi pound per square inch q quartet RNA ribonucleic acid ROS reactive oxygen species r.t. room temperature s singlet SAR structure-activity relationship satd. saturated SCUBA self-contained underwater breathing apparatus shRNA short hairpin ribonucleic acid t triplet TBAB tetra-n-butylammonium bromide TBAF tetra-n-butylammonium fluoride TBHP tert-butyl hydroperoxide TBSCl tert-butyldimethylsilyl chloride t-BuLi tert-butyllithium TEA triethylamine TEAC tetraethylammonium chloride THF tetrahydrofuran   xxi TIPSCl triisopropylsilychlorid TLC thin layer chromatography TMS trimethylsilyl TPSCl chlorotriphenylsilane TsCl 4-toluenesulfonyl chloride UBC University of British Columbia UV ultraviolet VO(acac)2 vanadyl acetylacetonate 3-MA 3-methyladenine   xxii Acknowledgements  I would first like to thank my supervisor, Dr. Raymond J. Andersen, for taking me into his group and his support for me to explore the world of chemistry. His patience and encouragement not only enabled me to be passionate about science, but also taught me the right attitude towards life and career. I feel extremely fortunate to work under his supervision.  I would like to thank Dr. David E. Williams for his helpful advices in purification of compounds. I would also like to thank Mike LeBlanc for his help and guidance in the lab. A great deal of thanks goes to our collaborators, Dr. Michel Roberge and Aruna Balgi who have greatly helped me and provided invaluable insights. I would like to extend my gratitude towards my great lab mates. Dr. Ping Cheng, who offered me enormous consultation, shared her chemistry wisdom and chatted together with joy, Dr. Luping Yan, a great mentor for me, who guided me in life and work, Dr. Ryan Centko for giving me a great deal of suggestions about practical lab and thesis writing, Rosanne for her great patience in teaching me lab techniques and unforgettable chats, Meng for introducing me fun places and bearing with me next to my fumehood, Kalindi for her delicious treats and her lovely boy, Jack for borrowing me chemicals and opening countless tightly sealed flasks and gas caps and Andrew for random chatting filled with laughter. Thanks also go to everybody that gathered data for me in the NMR and MS labs, especially Maria Ezhova in the NMR lab.  At last, I would like to thank my family: my father who taught me to be persistent and curious and my mother who always listens, understands me and provides me enormous support. I wish you happy and healthy.   xxiii Dedication      To my beloved mother Shi Liang    1 Chapter 1: Introduction 1.1 Drugs from nature Humans have relied on nature as an essential source of medicine dating back to prehistory. The earliest evidence, dating from 2600 BC, records the use of 1000 plant-derived substances to treat human disease in Mesopotamia.1 Furthermore, the documentation of the Egyptian, Chinese, Indian, Greek and Roman medicinal herbs demonstrates the important role of plants in the development of drug discovery.   Figure 1.1 Structures of plant-derived drugs  The first plant-based drug in history was morphine (1.1, Figure 1.1), one of the most widely used painkillers. It was isolated by the German pharmacist Friedrich Sertürner from a plant source, Papverum somniferum in 1804.2 This discovery offered researchers the inspiration of exploring natural sources for bioactive substances. Another example of a plant-derived drug is OHOHOHN(1.1) MorphineNONHHOH(1.2) QuinineNHN(1.3) ChloroquineClNNHN(1.4) HydroxychloroquineClNOH  2 the antimalarial drug, quinine (1.2, Figure 1.1), isolated from the bark of Cinchona species by the French pharmacists Gaventou and Pelletier.3 The discovery of quinine (1.2) provided the basis for the synthesis of the widely used antimalarial drugs chloroquine (CQ, 1.3, Figure 1.1) and hydroxychloroquine (HCQ, 1.4, Figure 1.1).3 Although plants have been utilized to search for chemotherapeutic agents throughout history, chemists and biologists have started to collect and investigate marine organisms since the 1950’s owing to improvements in scuba diving technology.4 The marine environment has proven to be an enormous source of novel biologically active compounds because oceans cover more than 70% of the earth’s surface. Studies show that marine organisms such as sponges, corals and algae produce potent compounds exhibiting significant bioactivities for drug screening.5 A systematic investigation of marine environments began in the 1970s due to the advent of sensitive analytical tools such as NMR spectroscopy, mass spectrometry, and HPLC. These technologies offered significant advances in separation and structure determination.4 Advances in NMR spectroscopy such as two-dimensional NMR methods and high-resolution spectroscopy have facilitated structural analysis on sub-milligram or milligram amounts of material at rapid rates. The combination of HPLC and mass spectrometry leading to HPLC-mass spectrometer (LC-MS) allows for dereplication of known compounds by correlating the molecular mass and the UV absorption data, allowing for easy comparison with a natural-product database. Therefore, improvements of these technologies have provided a rapid process for the isolation and structural characterization of novel compounds in days or months rather than years.6 Furthermore, the emergence of high-throughput screening (HTS)7 in the early 1990s accelerated the investigation of drug candidates in the pharmaceutical industry.8 The most   3 efficient HTS systems currently carry out tests on >250,000 samples per day per assay, facilitating rapid identification on the targeted pharmacophores.1 Spongothymidine (1.5, Figure 1.2), the first biologically active marine natural product reported in the literature, was isolated from the Caribbean sponge, Tethya crypta by Werner Bergman in the 1950s.9 10 11 This discovery led to the development of the antiviral/anticancer drugs Ara-A (1.6, Figure 1.2) and Ara-C (1.7, Figure 1.2) based on synthetic analog studies.12 These two compounds have been widely used in the clinic for decades. Later, related antiviral agents such as AZT (1.8, Figure 1.2), the first U.S. government approved anti-HIV drug, were developed from these sponge-derived lead compounds. 12 13   Figure 1.2 Structures of Spongothymidine (1.5), Ara-A (1.6), Ara-C (1.7) and AZT (1.8)  OOHHOOHNOOHHOOHNNHOOOHHOOHNNONH2ONNNNH2(1.5) Spongothymidine (1.6) Ara-A(1.7) Ara-CON3OH OHNHNOO(1.8) AZT  4 A great example of a marine-derived drug can be seen in one of the conotoxins, ziconotide (1.9, Figure 1.3). It was approved by the FDA in 2004 under the trade name Prialt for the treatment of chronic pain in a spinal cord injury.8 Ziconotide (1.9) is the synthetic equivalent of ω-conotoxin MVIIA, which is isolated from the venom of the marine snail, Conus magus.14 Ziconotide (1.9) can block neuronal N-type voltage-sensitive calcium channels selectively with higher potency than morphine.15 Moreover, clinical studies show that no addiction to ziconotide is observed in patients in contrast to opiates.14 Although the emergence of ziconotide as a selective blocker of a specific ion channel has inspired the investigation of other conotoxins, they have several limitations associated with most peptide-based drugs16, including short circulating half-life, poor proteolytic stability, and low oral bioavailability.17 Ziconotide (1.9) is limited to intrathecal administration.1 17 There are several strategies to modify peptides, including N- and C-termini modifications, cyclization, alkylation of amide nitrogen, side chain modifications, chirality changes, and amide bond surrogates, to overcome the limitations for designing peptide-based drug leads.18 Recent studies by Australian researchers demonstrated that conotoxins could be readily cyclized to improve their resistance to proteolysis without loss of their desirable biological activities.1 19  Figure 1.3 Amino-acid sequence and 3-D structure of ziconotide (1.9)   CKGKGAKCSRLMYDCCTGSCRSGKC CONH2H2N  5 A further example of a marine-derived drug lead is drin B (1.10, Figure 1.4), which was isolated by Uemura et al. in 1985 from the Japanese sponge, Halichondria okadai.20 Halichondrin B (1.10), one of a series of compounds originally isolated from the same sponge, was found to display remarkable in vitro and in vivo anticancer activities against murine melanomas and leukemias.13 21 Subsequently, a new source of halichondrin B (1.10), discovered from Lissodendoryx sp. in New Zealand, provided useful quantities of this compound for preclinical studies.8 This discovery sparked researchers to analyze the collection sea area, leading to the promising conclusion that the aquaculture of the deep-water Lissodendoryx could be carried out in water as shallow as 10 m, with the ability of producing a similar level of halichondrin complex found in the wild collection.4   Figure 1.4 Halichondrin B (1.10) and E7389 (Eribulin, 1.11)  Although the scarcity of the natural product could be partially solved by aquaculture, the structural complexity of halichondrin B (1.10) limited the efforts to develop the natural product as a new anticancer drug. A significant breakthrough was achieved by Kishi’s group, in cooperation with the Eisai pharmaceutical company, who reported a total synthesis of halichondrin B (1.10), requiring approximately 90 steps from commercially available starting OOOOOOOHOOH OHHHHHHOO O OHOOHO OO HO O OHOOHO OO HOOH2N OH(1.10) Halichondrin B (1.11) E7389 (Eribulin)  6 material.22 Kishi’s synthetic route allowed for structure-activity relationship (SAR) studies to identify the structural components that are responsible for its biological activity, revealing that the right-hand half of halichondrin B (1.10) retained the sub-nM activity of the parent compound.1 23 A structurally simpler analog E7389 (Eribulin, 1.11, Figure 1.4) was approved by the FDA in November 2010 for the treatment of refractory metastatic breast cancer.24 As of June 2014, the global marine derived pharmaceutical pipeline consists of six FDA-approved drugs, one EU registered drug, and 25 natural products or derivatives in different phases of the clinical pipeline.25 The preclinical pipeline remains active, continuing to provide several hundred novel compounds, with varying biological activities every year, to enhance the marine pharmaceutical clinical pipelines.25 26 However, the pharmaceutical industry is facing a decline in productivity due to the incompatibility of traditional natural product extract libraries with HTS and the subsequent advent of combinatorial chemistry.6 27 Recent technological improvements including simplification of crude mixtures and chemical synthesis help to address this issue of utilizing the diversity of marine nature products as lead compounds.6 1.2 Isolation of virantmycin as an inhibitor of autophagy 1.2.1 Autophagy Autophagy is a cellular catabolic process by which superfluous or damaged cellular contents are delivered to the lysosome for degradation and recycling.28 This process serves as a protective mechanism to maintain cellular homeostasis and to protect against various cellular stresses.29 There are three major types of autophagy: macroautophagy, microautophagy and chaperone-mediated autophagy.30 Macroautophagy, hereafter referred to as autophagy,30 the most common subtype, is characterized by the formation of unique double-membrane organelles called autophagosomes.   7 The process of autophagy, as shown in Figure 1.5, begins with the formation of phagophores, also known as pre-autophagosomal structures (PAS), which elongate to engulf the cytoplasmic constituents.31 The portions of the cytoplasm sequestered in the double-membrane bound organelles are called autophagosomes. The autophagosomes then fuse with acidic lysosomes to form autolysosomes.31 The sequestered cytoplasmic contents are degraded by lysosomal hydrolases and the monomeric degradants are transported back to the cytosol through permease for recycling.32 33  Figure 1.5 Mechanism of autophagy  In the absence of environmental stresses, basal autophagy plays an important role in maintaining cellular homeostasis by removing damaged or potentially toxic components as well as the turnover of long-lived proteins.33 34 On the other hand, autophagy can be stimulated by a broad range of stresses including starvation, hypoxia, oxidative stress and pathogen infection in order to promote cell survival.35 It has been reported that autophagy plays critical roles in diseases such as cancer, Parkinson’s disease, Alzheimer’s disease (AD) and Crohn’s disease.36  Pre-autophagosomalstructures (PAS)AutophagosomeLysosomeAutolysosome  8 1.2.2 Autophagy and cancer Although autophagy is cytoprotective under some conditions, the relationship between autophagy and cancer differs at different stages of cancer development. At the beginning, autophagy probably serves as a tumor suppressor by eliminating damaged organelles. However, once a tumor develops, this pathway enables the cancer cells to survive under stresses by generating nutrients and energy.30 37 The role of autophagy in cancer is complicated and it differs depending on tumor type or context. A great example illustrating how autophagy is protective against cancer is Beclin 1, the mammalian orthologue of the yeast Apg6/Vps30 gene.30 It was found to promote autophagy induced by nutrient deprivation in human MCF7 breast carcinoma cells. Moreover, the autophagy gene Beclin 1 showed inhibition against tumorigenesis.38 Initial evidence for the role of the autophagy gene Beclin 1 in tumor suppression was based on the study of a targeted mutant mouse model. Heterozygous disruption of beclin1 was found to promote hepatocellular carcinomas and cellular proliferation in mice with monoallelic deletion of Beclin 1. Furthermore, the heterozygous disruption of beclin1 resulted in an autophagy decrease in vivo, demonstrating that beclin 1 serves as a tumor-suppressor gene. Given the common deletion of beclin 1 in human breast, ovarian, and prostate cancer,39 the autophagy gene, beclin 1 would contribute to the study of human tumorigenesis. Recent studies reported by Mathew et al.40 have shown that deficient autophagy in mice led to an accumulation of the autophagic adaptor p62, which results in a high level of abnormal mitochondria, reactive oxygen species (ROS), and genome instability, thereby, promoting tumorigenesis. A puzzling aspect of this study is the reduced activation of transcription factor NF-κB by persistent p62, resulting from defective autophagy.41 This proposal is contrary to   9 previous studies,42 43 indicating p6244 as an activator of NF-κB. Based on the comparison of different cell systems, it is reasonable to propose that whether p62 is an inhibitor or activator of NF-κB may depend on the type of the cell and the stress. The cell system reported by Mathew et al. is of great interest because it provided a potential strategy for cancer chemoprevention with the clearance of p62 by stimulating autophagy. Conversely, autophagy has been shown to contribute to the survival of tumor cells. The initial interest in autophagy inhibition as a cancer therapy comes from research indicating that autophagy localizes to those regions stressed by hypoxia, chemotherapy, and radiotherapy.44 An interesting study by Amaravadi et al.45 indicated that the cooperation of autophagy inhibition with chemotherapy enhances the efficacy to induce cell apoptosis. This study was carried out in a lymphoma model and showed that inhibition of autophagy with CQ (1.3) or ATG5 short hairpin RNA (shRNA) promoted either p53 activation or alkylation drug therapy to induce tumor cell death.  Furthermore, the correlation of autophagy inhibition and pancreatic cancer has attracted great attention. Pancreatic ductal adenocarcinoma (PDAC) is highly lethal, with >40,000 cases diagnosed each year.46 It has a dismal five-year survival rate of 3% – 5% and is highly resistant to chemotherapy.47 Yang et al.48 observed highly elevated basal autophagy in pancreatic malignancies, even in nutrient-rich conditions. They proposed that PDAC cell lines depend on autophagy for sustained growth based on the fact that autophagy inhibition induced either by CQ (1.3) or by RNAi considerably reduced the growth of PDAC tumors in tumor xenograft mouse models.49 Therefore, exploration of autophagy inhibitors provides a potential way to reveal the interaction between autophagy and cancer and thus promotes the development of cancer therapies.   10 1.2.3 Autophagy inhibitors CQ (1.3) and its derivative, HCQ (1.4) have been widely used in humans as anti-malarial drugs. These compounds, known as lysosomotropic and acidotropic agents, block autophagosome degradation by inhibiting lysosomal acidification, resulting in the accumulation of undigested autophagosomes.50 Other autophagy inhibitors include bafilomycins,51 clomipramine,52 lucanthone,53 and Lys05.54 These compounds, along with CQ (1.3) and HCQ (1.4), inhibit autophagy after the formation of autophagosomes, and they interfere with lysosomal functions. Therefore, they are identified as late-stage inhibitors of autophagy. However, recent studies indicate that prolonged accumulation of abnormal autophagosomes in the cytoplasm could stimulate autophagy and be toxic to cells.55 56 Furthermore, although CQ (1.3) and HCQ (1.4) are the first generation of autophagy inhibitors and are safe for human use, recent studies show that the mechanism of inducing the cell death is unclear.55 Another cause for concern is the difficulty of achieving the high micromolar concentrations of HCQ (1.4) required to inhibit autophagy.57 58 On the other hand, researchers have been investigating early autophagy inhibitors suppressing autophagosome formation and sequestration, including 3-methyladenine (3-MA), wortmannin, and LY293002. However, they all have pharmacologically undesirable properties.59 The well-established autophagy inhibitor 3-MA is known to have effective inhibition of autophagic protein degradation at high concentrations, requiring an mM concentration for inhibition in vitro.60 61 It may also interfere with a variety of cellular processes independent of autophagy.59 Wortmannin and LY293002 are more potent than 3-MA, but they show strong inhibition of a number of lipid and protein kinases.    11 A recent study identified verteporfin62, an FDA-approved benzoporphyrin derivative, as an early autophagy inhibitor. It inhibits autophagy prior to autophagic sequestration but following the lipidation and membrane recruitment of LC3, the protein stably linked to the autophagosome membrane. It exhibits potent inhibition in vitro with a lower IC50 of 1µM, indicating its pharmacological potential.57 However, an ongoing study has shown that verteporfin inhibition results in the accumulation of abnormal p62 at the autophagosome membrane, which blocks its complete assembly. Therefore there is a need for more potent early autophagy inhibitors that have specific inhibition and are safe for human use to apply autophagy inhibition to cancer therapy. 1.2.4 Virantmycin (–)-Virantmycin (1.12, Figure 1.6), isolated by Roberto Forestieri in the Andersen Lab, was found to be a potent inhibitor of autophagy with an IC50 of 0.5 µM by our collaborator, Dr. Michel Roberge, at UBC.   Figure 1.6 Structure of (–)-virantmycin  A novel cell-based assay developed by Donohue et al.58 62 has been used to identify autophagy inhibitors in the Roberge Lab at UBC. In this assay, the breast cancer cell lines MCF-7 expressing enhanced green fluorescent protein-light chain 3 (EGFP-LC3) are exposed to the indicated concentrations of virantmycin (1.12), shown in Figure 1.7, with or without rapamycin HNClHOOCOMe(1.12) (–)-virantmycin  12 for 4 hours. Rapamycin,63 a mammalian target of rapamycin complex 1 (mTORC1) inhibitor, has been widely used to stimulate autophagy since autophagy is negatively regulated by mTORC1. EGFP antibody (top) is used to examine EGFP- LC3 degradation, and tubulin (bottom) is used as a loading control. When the cells are not treated with rapamycin or virantmycin (1.12), EGFP-LC3 is intact because the cells possess a very low basal level of autophagy. Rapamycin-stimulated autophagy results in the recruitment of EGFP-LC3 to autophagosomes and subsequent EGFP-LC3 degradation after the fusion between autophagosomes and lysosomes. The LC3 portion of the EGFP-LC3 protein is rapidly degraded, whereas the EGPF portion is degraded slowly, which leads to the accumulation of EGFP. Therefore, increased autophagy can be demonstrated by the levels of the EGFP band. This experiment shows that exposure to rapamycin in the presence of virantmycin (1.12) considerably reduces the levels of the EGFP band, indicating that virantmycin (1.12) inhibits rapamycin-stimulated autophagy. The effect of the inhibition depends on the concentration of virantmycin (1.12), with essentially complete inhibition at 5 µg/ml.  Figure 1.7 Virantmycin inhibiting rapamycin-induced autophagy64  EGFP%LC3)EGFP)tubulin)Virantmycin)(μg/ml))Rapamycin)(30nM))))%)))))+)))))%)))))+))))))%))))))+)))))%))))))+))))))%))))))+))))))%)))))+)))%)))))%)))))5)))))5)))))1))))))1))))0.5))0.5))0.1))0.1)0.05)0.05)  13 To study the effect of virantmycin (1.12) on autophagsome accumulation, MCF-7 cells expressing EGFP-LC3 are exposed to the indicated concentrations of virantmycin (1.12) alone or in combination with 75 µM CQ for 4 hours. As shown in Figure 1.8, this experiment indicates that virantmycin (1.12) alone causes the accumulation of autophagosomes with a lower level of 10000 units compared with the accumulation of autophagosomes caused by CQ with a level of 16000 units. When the cells are exposed to virantmycin (1.12) and CQ, the formation of auophagosomes is also reduced by virantmycin (1.12) to the same level as when cells are exposed to virantmycin (1.12) alone.   Figure 1.8 Effects of virantmycin (1.12) on autophagosome accumulation64  The site of action of virantmycin (1.12) in autophagy inhibition is well characterized based on the data shown above. Data obtained in the Roberge lab indicate that virantmycin (1.12) does not inhibit LC3 lipidation, LC3 recruitment to autophagosome membranes, LC3 puncta,   14 sequestration of cytoplasmic constituents, lysosome acidification or autophagosome formation. Therefore, virantmycin (1.12) inhibits autophagy prior to the stage where CQ and bafilomycins inhibit autophagy but following the formation of autophagosomes, as shown in Figure 1.9. Virantmycin’s sub-µM potency as an early-stage autophagy inhibitor makes it an interesting “lead compound” for the development of an anticancer drug candidate for the treatment of PDAC.   15  Figure 1.9 Stages of autophagy and sites of action of autophagy inhibitors64  The site of action of virantmycin (1.12) in autophagy inhibition is well characterized based on the data shown above. Data obtained in the Roberge lab indicate that virantmycin (1.12) does not inhibit LC3 lipidation, LC3 recruitment to autophagosome membranes, LC3 puncta, sequestration of cytoplasmic constituents, lysosome acidification or autophagosome formation. 3"MA%LY294002%Wortmannin%Ini5a5on%%%Nuclea5on%%%%%%Elonga5on%%%%%%%%Sequestra5on%%%%%%%Matura5on%%%%%%%%%Degrada5on%Upstream%signals%Maturing%autophagosome%Endosomes% MVBs%Lysosomes%Verteporfin%Virantmycin%Bafilomycins%Chloroquine%LC3%lipida5on:%NO%LC3%recruitment%to%mb:%NO%LC3%puncta:%NO%Sequestra5on:%NO%Autophagosomes%in%EM:%NO%EGFP"LC3%cleaved:%NO%Lysosomes%acidified:%YES%LC3%lipida5on:%YES%LC3%recruitment%to%mb:%YES%LC3%puncta:%NO%Sequestra5on:%NO%Autophagosomes%in%EM:%NO%EGFP"LC3%cleaved:%NO%Lysosomes%acidified:%YES%LC3%lipida5on:%YES%LC3%recruitment%to%mb:%YES%LC3%puncta:%YES%Autophagosomes%in%EM:%YES%Sequestra5on:%YES%EGFP"LC3%cleaved:%NO%Lysosomes%acidified:%YES%LC3%lipida5on:%YES%LC3%recruitment%to%mb:%YES%LC3%puncta:%YES%Autophagosomes%in%EM:%YES%Sequestra5on:%YES%EGFP"LC3%cleaved:%NO%Lysosomes%acidified:%NO%  16 Therefore, virantmycin (1.12) inhibits autophagy prior to the stage where CQ and bafilomycins inhibit autophagy but following the formation of autophagosomes, as shown in Figure 1.9. Virantmycin’s sub-µM potency as an early-stage autophagy inhibitor makes it an interesting “lead compound” for the development of an anticancer drug candidate for the treatment of PDAC. 1.3 Tetrahydroquinoline natural products A broad variety of natural products bearing tetrahydroquinoline skeletons have been discovered to have interesting biological activities. In 1996, Korean researchers reported the isolation of two tetrahydroquinoline alkaloids, which were named benzastatins C (1.13, Figure 1.10) and D (1.14, Figure 1.10).65 66 These metabolites were recovered from the culture broth of S. nitrosporeus 30643 and are structurally related to virantmycin (1.12, Figure 1.10). Benzastatins C (1.13) and D (1.14) were evaluated in rat liver microsomes and N18-RE-105 cell assays for analysis of inhibitory activity against lipid peroxidation and Glu toxicity respectively.67 They were found to possess weaker inhibitory activity than vitamin E against lipid peroxidation and similar inhibitory activity as vitamin E against glutamate toxicity. It was reported that free radicals were involved in the ischemic brain injury, thus it was important to screen radical scavengers or inhibitors of glutamate toxicity.68    17  Figure 1.10 Structures of benzastatins  The benzastatin family also includes aminobenzamide alkaloids, benzastatins A (1.15, Figure 1.10) and B (1.16, Figure 1.10) and indoline alkaloids, benzastatins E (1.17, Figure 1.10), F (1.18, Figure 1.10), and G (1.19, Figure 1.10)67 ,isolated from the same culture broth by the same research group. In recent years, substituted tetrahydroquinoline derivatives have attracted widespread attention as antimalarial agents. Previous studies69 showed that the enzyme farnesyltransferase (Pf-PFT) is an ideal drug target for the development of antimalarial because Pf-PFT inhibitors are highly cytotoxic to parasites and well tolerated in humans. A series of tetrahydroquinoline-based Pf-PFT inhibitors were synthesized and several compounds were discovered to inhibit Pf-PFT and the parasite at low nanomolar concentrations. Voorhis et al.69 synthesized tetrahydroquinoline-based analogs in five steps from commercially available 3-aminoquinoline. NH2H2NOCRR = CH2OMe, (1.15) benzastatin AR = CH3, (1.16) benzastatin BHNH2NOCOMeRR = Cl, (1.13) benzastatin CR = OH, (1.14) benzastatin DHNOHR1R2H2NOCR1 = CH2OMe, R2 = Me, (1.17) benzastatin ER1 = Me, R2 = Me, (1.18) benzastatin FR1 = Me, R2 = H, (1.19) benzastatin GHNClHOOCOMe(1.12) (–)-virantmycin  18 Compound 1.20 (Figure 1.11) exhibited an ED50 of 5 nM against (Malaria parasite) and inhibited Pf-PFT with an IC50 of 0.6 nM. Later, these authors70 investigated the SAR of the Pf-PFT inhibitory pharmacophore by variation of groups attached to the tetrahydroquinoline core. Compound 1.21 (Figure 1.11) emerged as the best antimalarial lead compound with an ED50 of 35 nM and 97% and 49% inhibition of Pf-PFT at 5 and 0.5 nM respectively.   Figure 1.11 Structures of tetrahydroquinoline-based Pf-PFT inhibitors  These tetrahydroquinolines underwent rapid metabolic degradation resulting in a high rate of clearance in vivo.71 72 Based on the structure of the enzyme–inhibitor complex and the study of the pathway of liver metabolism, the authors designed a series of 2-oxo-tetrahydroquinolines in which lone pairs on N-1 were less susceptible to oxidation by cytochrome P450s, the catalysts for the degradation  of tetrahydroquinolines. Compound 1.22 (Figure 1.11) showed high potency with an ED50 of 30 nM and 97% inhibition of Pf-PFT at 5 nM.72 Recently, a number of compounds containing tetrahydroquinoline frameworks were designed, synthesized and evaluated in the hopes of finding potent and novel γ-secretase inhibitors for the potential treatment of AD. Previous studies73 74 showed that γ-secretase plays NNC N SOO NNNN1.20NNC N SOO NNNNN1.21OONNC N SOO NNNN1.22OtBuOO  19 an important role in the production of β-amyloid, a signal for the onset and progression of AD. Therefore, γ-secretase has become an important target for AD therapy. Compounds 1.23 (Figure 1.12) displayed potent inhibition against γ-secretase in the SAR studies.  Figure 1.12 γ-Secretase inhibitors derived from tetrahydroquinolines  During the past decades, renin, an aspartyl protease, was shown to be a prime target for the treatment of hypertension. Many pharmaceutical companies75 76 77 designed and synthesized a series of tetrahydroquinoline-based compounds, among which 1.24 (Figure 1.13) and 1.25 (Figure 1.13) displayed potent renin inhibition and good permeability, solubility and metabolic stability for future improvement of oral bioavailability.  Figure 1.13 Renin inhibitors derived from tetrahydroquinolines  NSO OO NR1R2OCln = 3n1.23 NR1R2 = piperidyl, piperazyl etcHNNOO NRO OOMe1.24 R = (CH2)2NHCOMe, (CH2)2OCOMe etcNNN NH2H2NMeR1.25 R = (CH2)2NHCOMe, (CH2)2OCOMe etc  20 1.4 Proposed biosynthesis of bezastatins  In 1999, Yoo et al.78 proposed a possible biosynthetic pathway to the benzastatin family based on the coexistence of indoline and tetrahydroquinoline alkaloids. The author speculated that one of the double bonds in benzastatin A (1.15) was oxidized to an epoxide, which underwent intramolecular epoxide opening to give benzastatin D (1.14) and E (1.17) with indoline and tetrahydroquinoline skeletons respectively, as shown in Scheme 1.1.   Scheme 1.1 Proposed biosynthesis of benzastatin family by Yoo78   NH2H2NO ONH2H2NO OOHNH2NOCOMeOHHNOHOMeH2NOC(1.15) benzastatin A(1.14) benzastatin D (1.17) benzastatin ENOMeH2NOCH  21 The cyclization of an epoxide to the indoline ring system was confirmed by the model study shown in Scheme 1.2. Cyclized product 1.29 was spontaneously obtained after hydrogenation of the nitro group in 1.28 by with 10% Pd/C at room temperature. Thus the author proposed that the formation of the indoline skeleton was achieved by an intramolecular epoxide ring opening. Compound 1.29 prepared through an independent route was found to have the same spectral data as the one obtained from Scheme 1.2, which confirmed the structure of 1.29 in the model study.  Scheme 1.2 Model study of indoline skeleton formation by epoxide cyclization78  Furthermore, the author proposed the inter-conversion between the indoline and the tetrahydroquinoline skeletons via an aziridine intermediate, as shown in Scheme 1.3. They rationalized the inter-conversion by the treatment of prepared aziridine 1.30 with anhydrous HCl giving a mixture of tetrahydroquinoline 1.31 and indoline 1.32, as shown in Scheme 1.3. This inter-conversion provided a possible method to construct the tetrahydroquinoline skeleton from the indoline structure for further synthetic research.79 80 81 NO2CNNO2OMeONH2OMeOHNOHOMe1.26 1.27 1.281.29  22  Scheme 1.3 Model study of inter-conversion between tetrahydroquinolines and indolines78 N H dry HCl, CH2Cl2r.t., 30minHNClOMe +HNClMeO1.30 1.31 1.32OMe  23 Chapter 2: Synthesis of analogs of virantmycin for SAR studies 2.1 Isolation and characterization of (–)-virantmycin (–)-Virantmycin (1.12), an unusual chlorinated tetrahydroquinoline alkaloid, was isolated as colorless needles from cultures of a soil isolate AM-2722, subsequently identified as Streptomyces nitrosporeus.82 83 It exhibited very potent inhibitory activity against both RNA and DNA viruses in a plaque reduction test and displayed weak antifungal activity.84  (1.12) (–)-Virantmycin  The initial structural elucidation based on chemical transformation and NMR studies on the natural product was reported in 1981.84 However, the relative and absolute configuration at the two chiral centers C-2 and C-3 could not be assigned due to the absence of suitable derivatives for X-ray diffraction analysis. In 1988, Morimoto et al. reported the absolute configuration of the natural product based on NOE studies of synthetic (+)-virantmycin.85 The original assigned absolute configuration of (–)-virantmycin (1.12) had 2S, 3R at the two chiral centers, but this assignment was revised to 2R, 3R in 1990 and later confirmed by synthesis.86 87   HNClHOOCOMe234  24 2.2 Prior syntheses 2.2.1 Racemic synthesis by Raphael The first total synthesis (Scheme 2.1) of racemic virantmycin was completed by Raphael and Hill in 1986.88 89 The synthesis began with a Sonogashira coupling of iodobenzoate 2.3 with acetylenic alcohol 2.2, produced by an addition reaction of lithium acetylide with methoxyketone 2.1, to give the coupled product 2.4. A subsequent rearrangement of 2.4, catalyzed by acid, presumably, first produced unsaturated ketone 2.5, which underwent a Michael addition to give bicyclic ketone 2.6. Reduction of 2.6 and further dehydration of the resulting diastereomeric alcohols afforded bicyclic diene 2.7. Diene 2.7 was found to be thermally unstable and was thus converted to the stable N-formyl derivative 2.8 by treatment with formic acetic anhydride. Epoxidation of diene 2.8 yielded a diastereomeric mixture of bis-epoxides 2.9. Selective hydrogenolysis cleaved the only C-O bond at C-4 to give 2.10. This was subjected to de-epoxidation to regenerate the side-chain double bond, followed by N-deprotection to yield 2.11. The authors reported the formation of a single racemic diastereomeric secondary alcohol indicated that the epoxidation of the ring double bond proceeded stereoselectively and the resulting stereochemistry at C-3 was retained during hydrogenolysis. The relative configuration between hydroxyl group and adjacent methoxymethyl group in the racemic mixture was proven by NOE studies. Compound 2.11 was then advanced to 2.12 with retention of configuration at C-3. This occurred by the initial formation of aziridine with inversion followed by the ring opening with a chloride ion. Finally, methyl ester 2.12 was hydrolyzed to give (±)-virantmycin 2.13.   25  Scheme 2.1 Raphael’s total synthesis of (±)-virantmycin 2.1389  2.2.2 Racemic synthesis by Shirahama Five years after Raphael’s effort, Shirahama and Morimoto established a stereospecific synthetic route to the diastereomers 2.14 and 2.15.79 90 The key formation of the piperidine ring was completed by an intramolecular nitrene addition reaction followed by an aziridine ring opening with chloride ion. Aldehyde 2.17 was prepared easily from the known compound 2.16 by diazotization and the subsequent Lemieux-Johnson oxidation. The Wittig reaction of 2.17 with the prepared phosphorane 2.18 gave the (E)-olefin 2.20 with an E/Z ratio of >50:1. On the OHO +MeO2CNH2IPd(PPh3)2Cl2, CuIEt2NH, r.t., 16hMeO2CNH2OHOMsOH,THF, H2O40ºC, 20hHNMeO2COOMe HNMeO2COMemCPBA, NaHCO3CH2Cl, r.t., 72hNMeO2COMeOH2 (1atm), Pd/CDioxane, r.t., 21hOHCO2Ac, HCO2Hr.t., 12hNMeO2COMeOOHNMeO2COMeOHOHOHHNMeO2COMeOHSOCl2, CH2Cl240ºC, 5.5hHNMeO2COMeClHNHOOCOMeClLiOH, MeCN, H2O70ºC, 4h(±)-2.13OO2.12.2 2.3 2.42.6LiCCH(CH2NH2)2, C2H2benzene/THF, 35 to 50ºC OHO2.2MeO2CO2.5NH2OMe2.72.8 2.92.10 2.11 2.12  26 other hand, the Horner-Emmons reaction of 2.17 with the prepared phosphonate 2.19 yielded (E)-olefin 2.21 with an Z/E ratio of >50:1. The intramolecular nitrene addition reaction by photolysis of 2.20 and 2.21 was carried out stereospecifically to provide aziridines 2.22 and 2.23, respectively.  Scheme 2.2 Shirahama’s total synthesis of (±)-2.14 and (±)-2.1590  The methyl ester in aziridine 2.22 was chemoselectively reduced using 4 equiv. of LTBA to give the compound 2.24. However, the methyl ester in aziridine 2.23 was not selectively reduced under the same reaction conditions due to steric hindrance resulting from the cis-relationship between the methyl ester and the aromatic ring. LiAlH4 was used for reduction of HNHOOCOMeCl(±)-2.14HNHOOCOMeCl(±)-2.15NH2EtO2C2.16N3CHOEtO2C2.17MeO2CPPh32.182.19N3EtO2CCO2MeN3EtO2CCO2Me2.202.21EtO2CNMeOOCHEtO2CNMeOOCHEtO2CNHOHMeO2CNHOH2.222.232.252.24EtO2CNHOMe2.26MeO2CNHOMe2.27HNHOOCOMeCl(±)-2.14HNHOOCOMeCl(±)-2.15CH2Cl2, r.t., 38hKHMDS, 18-crown-6THF, –78 to –40ºC, 2.5hhv, toluene, r.t., 3hhv, toluene, r.t., 3hLTBA, THFr.t., 14h1) NaH, TBAB,THF, 0ºC, 30min2) MeI, –15ºC, 1h 2) TEAC, CH2Cl2, TFA–15ºC, 20min1) NaOH, MeOH,reflux, 3d1) KH,THF, 0ºC, 30min2) MeI, –15ºC, 1h 2) TEAC, CH2Cl2, TFA–15ºC, 20min1) NaOH, MeOH,reflux, 3dCO2MeP(OCH2CF3)2O  27 both the methyl and the ethyl esters to afford two hydroxyl groups, of which and only the benzylic alcohol was oxidized to the corresponding aldehyde, which was further converted to 2.25 by Corey’s method.91 After methylation of alcohols 2.24 and 2.25 and hydrolysis of esters 2.26 and 2.27, the highly regio and stereoselective ring opening of the aziridine with chloride ion provided (±)-2.14 and (±)-2.15 respectively. 2.2.3 Asymmetric synthesis of unnatural (+)-virantmycin by Shirahama In 1996, Shirahama and Morimoto reported the asymmetric total synthesis of (+)-virantmycin 2.41 (Scheme 2.3).80 At an early stage, benzoate 2.16 was elaborated into 2.28 after N-protection with TsCl followed by reduction of the ester and subsequent protection of the formed alcohol with TPSCl. Compound 2.28 was then converted to hemiacetal 2.29 by Lemieux-Johnson oxidation. The Wittig reaction of 2.29 with prepared phosphorane 2.18 stereoselectively afforded (E)-olefin 2.30 with an E/Z ratio of 30:1. Reduction of ester 2.30 gave an allylic alcohol, which was advanced to compound 2.31 through the sequence of reactions: alcohol protection as the TMS ether, N-protection then deprotection of TMS ether to lower the nucleophilicity of the nitrogen.  Allylic alcohol 2.31 underwent an asymmetric epoxidation using L-(+)-diethyl tartrate to give the endo epoxide as the only product. Methylation of this intermediate afforded compound 2.32, which was then converted to allylic acetate 2.35 with sodium iodide and zinc through the plausible mechanism via intermediate 2.33 and 2.34. Compound 2.35 was reduced with DIBAL to alcohol 2.36, which was then subjected to Vanadium-catalyzed asymmetric epoxidation to give 2.37 as a single diastereomer. The tetrahydroquinoline skeleton could be achieved by the reaction of 2.37 in the presence of TFA to yield the desired product 2.38 following Baldwin’s rules predictions. After protection of the diol 2.38, the regenerated benzylic alcohol was oxidized   28 to the carboxylic acid, followed by the sequence of N-deprotection, esterification of the carboxylic acid, deprotection of the diol, and methylation of the primary hydroxyl group to yield 2.39. Aziridine 2.40 was obtained by a Mitsunobu reaction of 2.39. After hydrolysis of methyl ester 2.40, the highly regio and stereospecific ring opening of aziridine with a chloride ion provided (+)-virantmycin (2.41).  Scheme 2.3 Shirahama’s asymmetric total synthesis of (+)-virantmycin80 NTsOTPSIOOZnNOTPSTsOAc2.33 2.34NH2EtO2CNHOTPSTs TsNOTPSOHMeO2CPPh3NHOTPSTsCO2Me2.16 2.28 2.292.302.181) OsO4, THF/H2Or.t., 30min2) NaIO4, 2h CH2Cl2, r.t., 52hNAcOTPSTsOMsO2.32NAcOTPSTsOH2.31NaI, Zn, DMF100ºC, 15minH+NHOTPSTsOAcNHOTPSTsOH2.352.36DIBAL, toluene–15ºC, 30minNHOTPSTsOHOOTPSTsNOHOH2.37 2.38TBHP, VO(acac)2CH2Cl2, 0ºC, 2.5h r.t., 6hTFA, tolueneMeO2CHNOMeOH MeO2CNHOMeHNClHOOCOMe2.39 2.40 (+)-2.41THF, r.t., 3hDEAD, PPh3  29 2.2.4 Racemic synthesis by Corey In 1999, Corey and Steinhagen established an efficient route for the synthesis of (±)-virantmycin 2.42.92 The key aspect of Corey’s strategy was the construction of the hydroquinoline system by the generation and trapping of an o-azaxylylene intermediate. This work showcased the application of the method to generate o-azaxylylenes through a [4 + 2] cycloaddition from chloromethylaniline derivatives and dienophiles, as reported from the same lab.93  Scheme 2.4 Synthesis of building blocks 1.44 and 1.45 in Corey’s method92 OHNH2ICl, HOAc23ºC, 2hOHNH2I TBSCl, imid., DMAPDMF, 23ºC, 2hOTBSNH2Iphosgene, DCMsatd. NaHCO3, 23ºC, 1hOTBSNCOI74% 97%99%O+HC CCO2EtLiCl, 2 mol% Pd(OAc)2HOAc, r.t., 3hOCO2EtCl1. DIBAL, toluene, CH2Cl22. TIPSCl, imid., CH2Cl2ClOH OTIPS(COCl)2, DMSOCH2Cl2, TEAClOTIPSO1. i-PrPPh3I, KHMDS    toluene, 16h, 40ºC2. TBAF, THF    23ºC, 15minClOH85% 55%94%92%NHClHOOCOMeOTBSNCOIClOH2.42 2.43 2.442.432.472.462.452.44 2.522.482.492.50 2.51  30 The Corey approach rested on the assembly of two simple building blocks 2.43 and 2.44, which were prepared as outlined in Scheme 2.4. The synthesis of isocyanate 2.43 was achieved starting from 2.45 through the sequence of iodination with ICl, TBSCl protection of the primary alcohol, and isocyanate formation with phosgene.  The other building block, compound 2.44, was synthesized from 2.48 and 2.49. The reaction of 2.48 with 2.49 in the prescence LiCl and of Pd(OAc)2 gave (Z)-keto ester 2.50 with a Z/E ratio of 93:7. Reduction of 2.50 using DIBAL, followed by selective TIPSCl protection of the primary hydroxyl group gave 2.51, which underwent Swern oxidation to provide 2.52. This intermediate was converted into allylic alcohol 2.44 by a Wittig reaction and subsequent deprotection.  Scheme 2.5 The completion of Corey’s total synthesis of (±)-virantmycin 2.4292  OTBSNCOIClOH+DMAP, CH2Cl223ºC, 1.5hOTBSNHIO OCl1. TBAF, THF2. SOCl2, TEA, CH2ClNHIO OClClINO OClNClOOI1. DIBAL, n-BuLi, THF;    then H3O+2. KH, THF, CH3INHClIOMeCO (1atm), DMF, MeOHPd(OAc)2, dppp, TEANHClMeO2COMe2.542.532.442.43Cs2CO3CH2Cl22.552.562.57NHClHO2COMe(±)-2.422.58LiOHMeCN/H2O  31 The reaction of isocyanate 2.43 with allylic alcohol 2.44 in the presence of DMAP gave carbamate 2.53, which was converted to carbamate 2.54 by alcohol deprotection and subsequent chlorination with SOCl2. The product was advanced to the hydroquinoline derivative 2.56 stereospecifically by way of a suprafacial (cis) cycloaddition of the intermediate o-azaxylylene 2.55. This intramolecular reaction (Scheme 2.5) was carried out in the presence of Cs2CO3 at 23 °C for 23 h to stereoselctively construct the hyroquinoline system in 90% yield. The carbamate moiety of 2.56 was easily cleaved upon treatment with DIBAL and n-BuLi to give the corresponding amino alcohol, which was methylated to give 2.57. The reaction of 2.57 with 1 atm of CO in DMF/MeOH in the presence of Pd(OAc)2, dppp and Et3N afforded the methyl ester 2.58, which was finally hydrolyzed to give (±)-virantmycin (2.42). 2.2.5 Enantioselective synthesis of (–)-virantmycin by Kogen The first total synthesis of natural (–)-virantmycin (1.12) was reported by Kogen and co-workers in 2003.94 They described a novel rearrangement of indoline-2-methanol to tetrahydroquinoline (Scheme 2.6).81 In this step, the crucial formation of contiguous quaternary and tertiary stereogenic centers was completed successfully. They designed the rearrangement reaction based on the proposed biosynthesis involving with the formation of an aziridine intermediate, followed by ring opening with an chloride ion. A similar ring-opening process to construct the tetrahydroquinoline skeleton was previously reported by Shirahama.80 A wide variety of optically active alcohols 2.59 were treated with PPh3 and CCl4 under reflux to provide the corresponding tetrahydroquinolines 2.62 as single isomers in moderate to good yield.    32  Scheme 2.6 Mechanism for the rearrangement of indolines to tetrahydroquinolines94  Consequently, this rearrangement reaction was utilized effectively in the total synthesis of (–)-virantmycin (1.12).94 The synthesis began with commercially available (S)-(–)-indoline-2-carboxylic acid (2.63), which was elaborated to 2.65 through acid-catalyzed esterification, followed by Boc-protection. Compound 2.65 was then converted to the Weinreb amide 2.66 by treatment with N,O-dimethylhydroxylamine hydrochloride and i-propylmagnesium chloride. Reaction of Weinreb amide 2.66 with methoxymethyl lithium, derived from Sn–Li exchange of MeOCH2Sn(n-Bu)3, provided ketone 2.67. This intermediate was then converted into iodide 2.68 in the presence of ICl and trisubstituted pyridine. Subsequently, iodide 2.68 underwent a highly diastereoselective 1, 2-addition of the Grignard to afford the corresponding alcohol 2.69 as the major isomer. Boc-deprotection of 2.69 in the presence of formic acid gave the rearrangement precursor 2.70. Subsequently, compound 2.70 was subjected to treatment with tri-n-butylphosphine and CCl4 to yield tetrahydroquinoline 2.71 as a single isomer.  HNH OHHNPPh3, CCl4ClHNHR'OHHNCl-R'RRPPh3CCl3H R'RRR'2.592.60 2.612.62  33  Scheme 2.7 Kogen’s total synthesis of (–)-virantmycin94  The authors described the formation of undesired side products, a deiodinated and an indole derivative when they used PPh3 in this reaction. After screening a number of aromatic and aliphatic phosphines, they found that tri-n-butylphosphine could be used as an alternative to suppress the detected side reactions. Carbonylation of tetrahydroquinoline 2.71 was carried out with 1 atm of CO in H2O/MeOH in the presence of Pd(OAc)2 and K2CO3 to provide (–)-virantmycin (1.12) in 53% yield (80% based on recovered starting material). Satisfyingly, the HN COOHHH2SO4, MeOH80ºC, 6h94%HN COOMeHBoc2O, CH2Cl2r.t., 18h92%BocN COOMeHBocNHNOOi-PrMgCl, Me(MeONH)•HClTHF, −20ºC to −10ºC72%BocNHOOMeBocNHOOMeIBocNI H OHICl, NTBMPCH2Cl2, 0ºC to r.t., 1hHCOOH, CH2Cl2r.t., 18h59%(n-Bu)3P, CCl4, CH2Cl240ºC, 30min45%MeOCH2Sn(n-Bu)3, n-BuLiTHF, -78ºC, 15min56%91% OMeTHF, -78ºC, 1h77% (dr = 19:1)HNI H OHOMeHNClIOMePd(OAc)2, K2CO3, H2O/MeOHCO (1atm), r.t., 18h53% 80%, brsmHNClHOOCOMe(–)-1.122.63 2.64 2.652.66 2.672.68 2.692.70 2.71MgBr  34 total synthesis of natural (–)-virantmycin (1.12) was achieved in only nine steps from commercially available starting material. 2.2.6 Enantioselective synthesis of (–)-virantmycin by Back Shortly after Kogen’s report, Back and Wulff95 announced a second enantioselective total synthesis of natural (–)-virantmycin (1.12). A key strategic principle here was the use of a Buchward-Hartwig aryl amination to construct the tetrahydroquinoline system.  As shown in Scheme 2.8, the construction of chiral center C2 was achieved through the desymmetrization of diester 2.72 by partial hydrolysis with porcine liver esterase (PLE) to provide 2.73. This intermediate was then converted separately into 2.74 and 2.75, acting as precursors of (+)- 2.41 and (–)-1.12 respectively.  Scheme 2.8 Desymmetrization of diester 1.73 to generate chiral center C295  Acylation of 2.76 with 2.75 afforded triester 2.77, which was converted to 2.78 through selective Krapcho decarboxylation of the β-keto ester moiety (Scheme 2.9). The ketone moiety of 2.78 was reduced to the hydroxyl functionalities of 2.79 and 2.80. This reaction proceeded MeOOOMeOOMeOOOHOOMeOOFOOFOOOOTMS2.72 2.732.752.74  35 with poor stereoselctivity, giving a 4:3 mixture of desired epimer 2.79 and undesired epimer 2.80, respectively. The authors reported that these two epimers were separable and ketone 2.78 was recycled from epimer 2.80 upon treatment with PCC.  Scheme 2.9 The completion of Back’s total synthesis of (–)-virantmycin 1.1295  After acetylation and selective TMS deprotection, 2.79 was converted into 2.81, which underwent a Curtius rearrangement with DPPA to give 2.82. Intramolecular Buchwald-Hartwig aryl amination of 2.82 provided the tetrahydroquinoline skeleton of 2.83. The authors discovered that ligand BINAPFu in the presence of Pd2(dba)3 improved the efficacy of coupling an ortho-substituted aryl bromide with an α-quaternary amine derivative after they conducted model studies on aminations of highly hindered amines using various ligands. Deacetylation and subsequent deformylation of 2.83 afforded amino alcohol 2.84, which was advanced to (–)-virantmycin (1.12) through the same sequence of aziridine formation and ring opening reported CO2MeBrMeO2C Me2OCMeO2COBrOOOTMS MeO2COBrOOOTMSMeO2CBrOOOTMSOHMeO2CBrOOOTMSOH+4:3MeO2CBrOOHOOAcMeO2CBrOOAcNHCHONOAcOCHOMeO2CNHOHOMeO2CNHClOHO2C(1.12) (–)-virantmycin2.76 2.77 2.782.79 2.802.81 2.822.84 2.831) LiHMDS, Et2O0ºC, 10min2) 2.75, Et2O 1h at 0ºC to r.t., 20h 10% aq. NaCl, DMSO125ºC, 20hNaBH4, MeOH0ºC, 2.5hPCC, CH2Cl2r.t., 3d1) DPPA, DMAP, NEt3,toluene, reflux, 2.25h2) NaBH4, THF, r.t., 12.5hPd2(dba)3, BINAPFuCs2CO3, toluene90ºC, 6.5h0.13 M NaOH in MeOHr.t., 24h  36 by Shirahama.80 The unnatural antipode (+)-2.41 was synthesized from 2.74 through a similar route as shown in Scheme 2.9. 2.3 General synthetic analyses for SAR studies on virantmycin  Figure 2.1 Major structural elements in (–)-virantmycin (1.12)  From the efforts described in the preceding pages, Kogen’s method94 was the most efficacious in synthesizing (–)-virantmycin (1.12) in nine steps, whereas previously reported synthetic routes 88 79 80 were lengthy and they involved undesirable repetition of protection-deprotection sequences. This methodology also offered a concise way to achieve higher stereoselectivity in constructing two contiguous chiral centers, compared with poor stereoselctivity of ketone reduction in Back’s route95 (Scheme 2.9). Furthermore, Kogen’s method (Scheme 2.7) provides high flexibility to a broad variety of analogs for SAR studies because the major structural elements: amino, carboxyl, chloro, methoxymethyl groups and olefin chain can be altered easily. Therefore, Kogen’s method was used because of its advantages over other methods of synthesis of  (–)-virantmycin (1.12). Modifications of this methodology were required to probe the SAR for the virantmycin autophagy inhibiting pharmacophore. HNClHOOCOMecarboxyl groupamino groupchloro groupmethoxymethyl groupolefin chainRRconfigurationconfiguration  37 2.4 Retrosynthetic analysis of analog 2.1 Based on the analysis on the major structural elements in (–)-virantmycin (1.12), we were interested in how modifications to the amino, carboxyl, chloro, methoxymethyl groups, the olefin chain and the configuration at C-2 and C-3 would affect the autophagy inhibiting activity of these compounds. The goal of the SAR study was to construct readily synthesized analogs to gain insights into the role of these functional groups in the autophagy inhibiting activity and to provide larger amounts of more accessible analogs for biological evaluation in vitro and in vivo.    Figure 2.2 Analogs 2.85 and 2.86  We attempted to construct analogs 2.85 (Figure 2.2) and 2.86 (Figure 2.2) with methyl groups at C-2 instead of methoxymethyl as a synthetic simplification that was presumed to still yield autophagy-inhibiting analogs. This structural simplification allowed us to avoid the complicated preparation procedure and the use of highly toxic tin reagent for installing the methoxymethyl group at C-2, as outlined in Kogen’s synthetic route.81 A retrosynthetic analysis of compound 2.85 based on Kogen’s route is presented in Scheme 2.10. The synthetic route to compound 2.85 begins with the construction of the alcohol 2.87, which is accomplished by diastereoselective 1,2-addition of a Grignard to ketone 2.88. This intermediate can be synthesized from commercially available (S)-(–)-indoline-2-carboxylic acid (2.63). HNHOOC ClHNHOOC Cl2.85 2.86  38  Scheme 2.10 Retrosynthetic analysis of 2.63  2.5 Initial synthetic trials Following Kogen’s procedure, the commercially available amino acid 2.63 was dissolved in methanol by a careful dropwise addition of sulfuric acid at room temperature (Scheme 2.11). The reaction mixture was stirred at 80 °C for 6 hours. Unfortunately, only a trace amount of the desired methyl ester 2.64 was obtained. Given the difficulty of controlling the addition rate of the sulfuric acid, this method was abandoned.   Scheme 2.11 Failed esterification of 2.63  Using an alternative procedure,96 carboxylic acid 2.63 was converted to methyl ester 2.64 in a yield of 98% by treatment with thionyl chloride in methanol (Scheme 2.12). Boc-protection of the methyl ester 2.64 gave compound 2.65, which was then brominated with NBS to afford BocNH OHXBocNHXOHN COOHHSHNHOOC Cl2.85 2.87 2.882.63HN COOHH2.63H2SO4, MeOH80ºC, 6hHN COOMeH2.64trace amount  39 bromide 2.89 in excellent yield. Furthermore, iodination of compound 2.65 with ICl and a trisubstituted pyridine gave iodide 2.90 in a yield of 87%.   Scheme 2.12 Synthesis of esters 2.89 and 2.90  Following Kogen’s protocol, compounds 2.89, 2.90 and 2.65 (Scheme 2.13) were converted to the corresponding Weinreb amides 2.91, 2.92 and 2.66, respectively, in moderate yields. Reactions of Weinreb amides 2.91 and 2.66 with Grignard reagent 2.95 in THF provided ketones 2.96 and 2.97 in yields of 38% and 35%, respectively.  HN COOHHSOCl2, MeOH40ºC, 4h98%HN COOMeHBoc2O, CH2Cl2r.t., 18h97%BocN COOMeHNBS, DMF, 0ºC, 2h97%BocN COOMeHBrBocN COOMeHIICl, 2,6-di-tert-butyl-4-methylpyridineCH2Cl2, 0ºC to r.t., 1h87%2.63 2.642.892.902.65  40  Scheme 2.13 Synthesis of alcohols of 2.98, 2.99, 2.100 and 2.101  Grignard reagent 2.95 was initially prepared from commercially available bromide 2.94. Due to the prohibition cost and a large amount of bromide 2.94 needed for the preparation of the key intermediates 2.96 and 2.97, bromide 2.94 was prepared from 2.93, as shown in Scheme 2.14, following the literature procedure,97 and was further purified by distillation under reduced pressure.   Scheme 2.14 Preparation of bromide 2.94  BocN COOMeHRR = Br, 2.89R =  I, 2.90R = H, 2.65i-PrMgCl, Me(MeONH)•HClTHF, –20ºC to –10ºCBocNHRNOOMgBrTHF, 0ºC, 1hBocNHROR = Br, 2.91, 75%R =  I, 2.92, 58%R = H, 2.66, 73%R = Br, 2.96, 38%R = H, 2.97, 35%2.95CH3Li, THFBocNR H OH–78ºC, 20minR = Br, 2.98, 66%R = H, 2.99, 56%MeMgBr, Et2O–40ºC, 2hBocNR H OHR = Br, 2.100R = H, 2.10195%BrO 1) MeMgBr, THF, reflux, 20min2) H2SO4/H2O (1:2), <10ºC, 30min2.93 2.94  41 The 1,2-addition of MeLi to ketones 2.96 and 2.97 proceeded with high diastereoselectivity and afforded the corresponding alcohols 2.98 and 2.99 in yields of 66% and 56%, respectively (Scheme 2.13). On the other hand, the reaction of ketones 2.96 and 2.97 with MeMgBr in Et2O gave diastereomeric alcohols 2.100 and 2.101, respectively, in excellent yields. The reversal of diastereoselectivity can be rationalized by the Felkin–Anh model presented in Scheme 2.15. Divalent Mg2+ can coordinate with a lone pair of electrons on the nitrogen together with a lone pair of electrons on the oxygen. Due to the chelation effect, the carbonyl oxygen atom and the nitrogen atoms are locked in an eclipsed conformation. The nucleophile can then attack from the least sterically hindered side to provide alcohol 2.101 with high diastereoselectivity.   Scheme 2.15 Rationale for stereochemical outcome of the addition reaction of 2.97 HNBocOMeLiBocNH OH2.97 2.992.97NMg2+HMeMgBrBocNH OH2.101Boc O  42 The reaction of the Weinreb amide 2.92 (Scheme 2.16) with the Grignard reagent 2.95 in THF gave a 4:1 mixture of the ketone 2.102 and the deiodinated ketone 2.97, which were not separated by flash column chromatography. The deiodinated ketone 2.97 was also observed and likely resulted from a metal-halogen exchange. Subsequent attempts at addition of MeLi to the mixture of the ketones 2.102 and 2.97 to access the iodinated alcohol 2.103 were unsuccessful. Ketone 2.102 was completely converted to the deiodinated alcohol 2.99.  Scheme 2.16 Metal-halogen exchanges resulting in deiodinated products  To overcome the undesired deiodination, a range of addition reaction conditions using the Grignard reagent MeMgBr, a less reactive nucleophile than MeLi, was screened. The results of these optimization studies are presented in Table 2.1. The results clearly suggest that Et2O is the solvent of choice. Running the reaction at a temperature higher than –40°C resulted in a higher ratio of the undesired alcohol 2.101. It was found that the addition reaction of the ketone 2.102 with MeMgBr, carried out in Et2O at –40°C for 2 hours, afforded a 7:3 mixture of the desired BocNHIOMgBrTHF, 0ºC, 1h38%2.95BocNHINOO+BocNHO4:1BocNHIO CH3Li, THF BocNH OH–78ºC, 20min2.92 2.102 2.972.102I2.103CH3Li, THF BocNH OH–78ºC, 20min2.99  43 alcohol 2.104 and the deiodinated alcohol 2.101 in yield of 91% (entry 2). Therefore, these optimized reaction conditions were utilized to generate the alcohol 2.104.  Entry Solvent T (°C) Time (h) Ratio (2.104/2.101) Yield (%) 1 Et2O –78 to –40 15 7:3 85 2 Et2O –40 2 7:3 91 3 Et2O –40 to –20 2 1:1 86 4 Et2O –40 to 0 1 3:7 85 5 THF –40 12 7:3 82 Table 2.1 Optimization of the Grignard addition reaction conditions  The mixture of the alcohols 2.104 and 2.101 was subjected to N-Boc-deprotection conditions using formic acid in CH2Cl2 at room temperature to give the alcohols 2.105 and 2.106 in a yield of 38% (Scheme 2.17). The undesired deiodinated alcohol 2.106 could only be partially separated from the desired alcohol 2.105 using flash column chromatography, which resulted in a 3:1 mixture of the alcohols 2.105 and 2.106. The rearrangement of this mixture in the presence of PPh3 and CCl4 in CH2Cl2 afforded a 7:3 mixture of 2.107 and 2.108. The deiodination observed in this reaction was previously reported in the literature.94 Unfortunately, attempts to suppress the formation of the undesired product failed when using tri-n-butylphosphine, an alternative reported by the authors.  BocNHIO BocNH OH2.102I2.104MeMgBr BocNH OH2.101+  44  Scheme 2.17 Attempts to synthesize analog 2.86  Subsequently, the mixture of 2.107 and 2.108 was subjected to carboxylation conditions using 1 atm CO in H2O/MeOH in the presence of Pd(OAc)2 and K2CO3 to give only a trace amount of the desired product 2.86. Attempts at purifying the crude reaction mixture, using preparative silica gel TLC followed by reversed phase HPLC, failed to separate the desired product 2.86 from unreacted starting tetrahydroquinolines 2.107 and 2.108.   Scheme 2.18 Attempts at early stage carboxylation HCOOH, CH2Cl2r.t., 18hHNI H OHPPh3, CCl4, CH2Cl240ºC, 30minHNI ClHNCl+62%38%BocNHIO BocNH OHBocNH OH+91% IMeMgBr Et2O, –40ºC, 2h7:32.102 2.104 2.1012.105HNH OH2.106+3:12.107 2.1087:3n-Bu3P, CCl4CH2Cl2, 40ºC, 30minHNI Cl2.107Pd(OAc)2, K2CO3, CO (1atm)H2O/MeOH, r.t. , 18hHNHOOC Cl2.86trace amountBocNHIO 1) MeLi, THF, −78ºC, 20min2) CO2, –78ºC to 0ºC, 12hBocNH OHHOOC2.102 2.109  45 Attempts were then made to install the carboxyl group at an early stage by carrying out a 1,2-addition of MeLi, followed by quenching with CO2 (bubbled into reaction) (Scheme 2.18). However, this method failed to afford the desired product 2.109, resulting in the formation of deiodinated alcohol 2.99.  Scheme 2.19 Synthesis of tetrahydroquinolines of 2.113, 2.114, 2.115 and 2.108   BocNR H OHR = Br, 2.98R = H, 2.99BocNR H OHR = Br, 2.100R = H, 2.101HCOOH, CH2Cl2r.t., 18hHNR H OHR = Br, 2.110R = H, 2.111HNR H OHR = Br, 2.112R = H, 2.106PPh3, CCl4, CH2Cl240ºC, 30minHNR ClR = Br, 2.115, 62%R = H, 2.108, 55%HNR ClR = Br, 2.113, 54%R = H, 2.114, 47%40%  46 Given the difficulty of installing the carboxyl group from the iodo intermediates, the bromo group was considered as an alternative to access the carboxyl group. The aryl bromides were more readily synthesized than the aryl iodides because the less reactive aryl bromides couldnot undergo metal-halogen exchange with MeLi or Grignard reagents. Therefore, alcohols 2.98, 2.99, 2.100 and 2.101 were transformed to tetrahydroquinolines 2.113, 2.114, 2.115 and 2.108, respectively, through the sequence of N-Boc-deprotection followed by diastereoselective PPh3–CCl4-mediated rearrangement (Scheme 2.19).  Scheme 2.20 Attempts to install the carbxyl group on 2.115  Given the use of metal reagents as nucleophiles in the addition reactions, the installation of carboxyl group was carried out using tetrahydroquinoline 2.115 (Scheme 2.20). t-BuLi was slowly added to a solution of 2.115 in Et2O at –78°C to generate the phenyllithium, which was quenched with dry CO2 gas. Unfortunately, the desired product 2.86 was not obtained (Scheme 2.20). This was thought to be due to the formed phenyl lithium being quenched by a trace amount of water in the CO2 gas. Given the difficulty of removing the water from CO2 gas, 1) t-BuLi, Et2O, –78ºC, 15min2) CO2, –78ºC to r.t. , 5h1) t-BuLi, Et2O, –78ºC, 15min2) ClCO2Me, –78ºC to r.t. , 5hBoc2O, TEA, CH2Cl240ºC, 18hNBocBrHNBr ClHNHOOC ClHNBr ClHNMeOOC Cl2.862.1152.1162.115HNBr Cl2.115 2.117Cl  47 anhydrous methyl chloroformate was chosen as the quenching reagent instead. Attempts to construct the methyl benzoate 2.116 by quenching the phenyllithium with methyl chloroformate failed. The proton of the free amine in the system may quench the phenyllithium. Thus N-Boc-protection of 2.115 was attempted in the presence of TEA in CH2Cl2. However, no reaction was observed probably due to the steric effect of the quaternary C-2. Having failed to install a carboxyl group on 2.115, construction of the carboxyl group on alcohol 2.112 was then attempted. As shown in Scheme 2.21, alcohol 2.112 was converted to the corresponding acetonide 2.118 by the treatment of Me2C(OMe)2 and PPTS in CH2Cl2 in a yield of 63%. Attempts to construct the methyl benzoate 2.120 through the sequence of lithium-halogen exchange, quenching phenyl lithium using methyl chloroformate followed by the deprotection of the acetonide failed. The reaction of acetonide 2.120 with t-BuLi in Et2O followed by the addition of methyl chloroformate gave complex mixture of products. NMR analysis of each purified fraction showed that none of them were the desired product 2.119.  Scheme 2.21 Attempts to install the carbonyl group on 2.112  2.6 Synthesis of aryl nitriles Having failed to construct the carboxyl group directly from both aryl iodides and aryl bromides, attempts were made to convert the bromo group into the cyano group, which could HNBr H OH2.112NBr H2.118OMe2C(OMe)2, PPTSCH2Cl2, r.t. , 7h1) t-BuLi, Et2O, –78ºC, 15min2) ClCO2Me, –78ºC to r.t. , 2hNMeOOC H2.119O63%Me2C(OMe)2, PPTSCH2Cl2, r.t. , 7hHNMeOOC H OH2.120  48 then be hydrolyzed to the desired carboxyl functionality. Furthermore, the cyano group can undergo cycloaddition to give the tetrazole ring, which is a bioisostere of the carboxyl group.  Scheme 2.22 Attempts to convert 2.115 into 2.121  Our first attempt to convert 2.115 into 2.121, under Rosenmund-von Braun cyanation conditions, resulted in the decomposition of the starting material 2.115 and no desired product 2.121 (Scheme 2.22). The decomposition of the starting material was result of either the long reaction time or high temperature employed. A search of the literature revealed an efficient route to access aromatic nitriles catalyzed under mild conditions with Pd2(dba)5.98 Tetrahydroquinoline 2.115 was subjected to NaCN and Pd2(dba)5 and t-Bu3P in MeCN/THF (Scheme 2.22). However, no reaction was observed, and the starting material 2.115 was recycled. The preparation of the Pd catalyst involved	  complicated procedures under rigorously oxygen and moisture-free conditions. Suspecting that the major difficulty was the preparation of the Pd catalyst, we sought a cyanation reaction catalyzed by Pd(PPh3)4, which is a commercially available catalyst. Pd(PPh3)4 was CuCN, DMA150–155ºC, 24hMeCN/THF, 70ºC, 2hNaCN (1.06 equiv), 1% Pd/t-Bu3P (1:1)HNBr Cl2.115HNNC Cl2.121HNBr Cl2.115HNNC Cl2.121Zn(CN)2, Pd(PPh3)4DMF, 90ºC, 4hHNBr Cl2.122HNBr OO H2.115  49 added to the solution of 2.115 with Zn(CN)2 in DMF in a glove box (Scheme 2.22). The reaction mixture was stirred at 90°C for 4 hours. Unfortunately, the desired product 2.121 was not obtained. NMR analysis of the crude reaction mixture showed formate 2.122 was the major side product.  Scheme 2.23 Plausible mechanism for the formation of formate 2.122  A possible mechanism for the formation of the formate 2.122 is shown in Scheme 2.23. The reaction begins with the attack of 2.115 at C-3 with DMF, followed by an attack of the imido ester 2.123 by oxygen lone pair of H2O to give 2.124. The C=O bond is regenerated after a proton transfer, followed by cleavage of the C-N bond of 2.125, resulting in 2.126 and dimethyl HNH NOHClHNBr ONClOH HHNBr OO NHHproton transferHNBr OO NH HHNBr OO HHCNHNHNBr OO H+    HCNBr2.115 2.1232.1242.1252.1262.122  50 amine as the leaving group. The deprotonation of the oxonium ion formed with CN- reveals the C=O bond to afford formate 2.124 and generates HCN in the process.   Scheme 2.24 Factors in Pd-catalyzed cyanation reactions  Trace amounts of water in the system may lead to the formation of HCN. HCN could deactivate Pd catalyst due to its higher reactivity toward Pd(0) than that of any haloarene substrate (Scheme 2.24, Step 1). Recent studies98 indicate that the reaction of HCN with Pd(0) to generate [Pd(CN)4]2- and/or [(CN)3Pd(H)]2- is a fast and irreversible process, resulting in the catalyst being poisoned. These authors showed that cyanide concentration is strictly comparable to the Pd catalyst concentration and hence reaction rates. Low cyanide concentration results in LnPdLnPd XArLnPd CNArAr CN[(CN)3PdAr]2-Ar CN [(CN)4Pd]2- + H2and/or[(CN)3PdH]2-excess CN-, H2O-L, -OH-CN-X-excess CN--Lexcess CN--L, -X-Step 2Step 1Step 3  51 slower reaction rates, whereas high cyanide concentration results in catalyst deactivation through irreversible displacement of the ligands on Pd by cyanide (Scheme 2.24, Step 2 and 3). Given the difficulty in controlling the rigorous conditions required for cyanation reactions catalyzed by transition metals, we attempted to construct the aryl nitrile via microwave-assisted cyanation reaction instead. A search of the literature revealed an efficient synthetic methodology to produce aryl nitriles from aryl bromides under irradiation with microwave.99 N-methyl-2-pyrrolidinone (NMP) was the solvent of choice because of its high boiling point and excellent solubility of CuCN, the cyanide source. Initial attempts to convert bromide 2.115 into the corresponding nitrile 2.121 at 200°C under microwave irradiation for 30 minutes resulted in a trace amount of the desired product 2.121 and a large amount of the side product 2.127 (Scheme 2.25). The higher reactivity of the chloro group toward CuCN than that of the bromo group was not expected. In order to avoid the formation of the side product, ketone 2.96 was chosen as the substrate for the cyanation reaction. Unexpectedly, the reaction of the ketone 2.96 with CuCN in NMP under microwave irradiation for 30 minutes, gave indole 2.129 in a yield of 54% (Scheme 2.25). The mechanisms for the release of the Boc group and the elimination occurring at the chiral center remain unclear.  Although the desired product 2.128 was not obtained, the successful conversion of the bromo group into the cyano group proved the efficiency of this methodology. Therefore, attention was turned to utilizing alcohol 2.112, the only intermediate without a halogen functionality or boc-protection. Irradiation of alcohol 2.112 in the presence of CuCN in NMP for 25 minutes afforded the desired product 2.130 in a yield of 38% (50% based on recovered starting material 2.112).   52 	  Scheme 2.25 Cyanation of aryl bromides assisted by microwave irradiation  Increasing the reaction time to 35 minutes resulted in the decomposition of both starting material 2.112 and desired product 2.130. Only trace amount of the desired product 2.130 was obtained. Furthermore, the concentration of CuCN reported in the literature resulted in no reaction. When the concentration of CuCN was reduced to 0.055 mmol/ml in NMP, the desired product 2.130 was obtained smoothly. When the reaction was scaled up to 200 mg, no reaction was observed. It was found that this microwave-assisted reaction was only optimal when carried HNNC ClHNBr ClCuCN, NMP200ºC, 30mintrace amount+2.115 2.121BocNBrO CuCN, NMP200ºC, 30minH2.96 2.128CuCN, NMP200ºC, 25min38%50%, brsmHNBr H OH2.112HNNC H OH2.130CuCN, NMP200ºC, 35minHNBr H OH2.112HNNC H OH2.130trace amountBocNHBrO BocNBr H OHHNBr H OH2.112HNBr Cl2.1152.96 2.100HNBr CNmajor2.127HNNCOCuCN, NMP200ºC, 30min54%2.129BocNNCOH  53 out on scales up to 30 mg of alcohol 2.112 with CuCN (0.055 mmol/mL in NMP, 1.3 equiv)(Scheme 2.25).  Scheme 2.26 Cyanation of aryl bromides assisted by microwave  Next, the nitrile 2.130 was subjected to Kogen’s PPh3–CCl4-mediated rearrangement conditions (Scheme 2.26). After stirring the reaction mixture at 40°C for 30 minutes, TLC monitoring indicated the presence of only starting material. Increasing the reaction time to 18 hours yielded only trace amount of the desired product 2.121. Attempts to optimize the rearrangement reaction by changing the solvent CH2Cl2 to CCl4 resulted in possible decomposition of the starting material 2.130 after the reaction mixture was stirred at 40°C in CCl4 for 3 hours. Thus, the reaction temperature was not increased and remained at 40°C for 15 hours to give a trace amount of the product 2.121. HNNC H OH2.130PPh3, CCl4, CH2Cl240ºC, 18hHNNC Cl2.121trace amountHNNC H OH2.130PPh3, CCl440ºC, 18hHNNC Cl2.121trace amountHNNC H OH2.130PPh3, CCl4, CH2Cl240ºC, 30minno reaction  54 Kogen et al. investigated the scope of the PPh3–CCl4-mediated rearrangement using various chiral alcohols. They found that the rearrangement proceeded efficiently with substrates when the aryl ring had bromo or ester groups, however no reaction occurred with substrates with carboxyl aryl substituents. Furthermore, treatment of the aryl amide under the same condition resulted in the reduction of the amide group to cyano group, and no further rearrangement of the aryl nitrile was reported. Based on their investigation and the failed attempts to optimize the rearrangement of nitrile 2.130, we decided to hydrolyze 2.130 to the corresponding ester 2.131 (Scheme 2.27).   Scheme 2.27 Attempts to hydrolyze the nitrile 2.130 to the ester 2.131  Considering the olefin chain of nitrile 2.130, hydrolysis of 2.130 under basic conditions was attempted. Hydrolysis of 2.130 by the treatment of NaOH in EtOH/H2O provided the corresponding carboxylic acid and subsequent methylation of the carboxylic acid with TMSCHN2 in methanol gave a trace amount of the desired methyl ester 2.131 (Scheme 2.27). Given the low yield of methyl ester 2.131 generated by hydrolyzing 2.130 under basic conditions, hydrolysis of 2.130 was attempted under acidic conditions. Due to the presence of the double bond, strong acids such as HCl were not utilized. Alternatively, the nitrile 2.130 was subjected to HNNC H OH2.130HNMeOOC H OH2.1311) NaOH, EtOH/H2O, reflux, 20h2) TMSCHN2, MeOH, r.t., 18htrace amountTMSCl, MeOH50ºC, 4hHNNC H OH2.130HNMeOOC H OH2.132Cl+HNMeOOC H OH2.131trace amountmajor  55 treatment with TMSCl in methanol. The reaction mixture was stirred at 50°C for 4 hours to give a trace amount of the desired methyl ester 2.131 and a large amount of the side product 2.132, which was generated by the addition of HCl across the double bond. 2.7 Synthesis of analogs with the saturated side chain Considering the elimination side product of the TMSCl hydrolysis reaction, construction of analogs with saturated side chains at C-2, instead of the analogs with olefin chains at C-2 was then attempted.   Entry Metal reagent Desired product Yield 1 n-BuLi  trace amount 2 i-PentylMgBr  trace amount 3 i-PrMgCl  trace amount Table 2.2 Attempts to install saturated side chains at C-2  BocNHBrNOOBocNHBrORTHFCH3LiTHFR-metalBocNBr HROH2.91 2.133 2.134BocNHBrO2.135BocNHBrO2.136BocNHBrO2.137  56 Several organometallic reagents such as n-BuLi, i-PentylMgBr, i-PrMgCl were screened, to install the saturated side chains. However, none were successful in yielding a desirable amount of the desired products. All reactions of Weinreb amide 2.91 with the organometallic reagents gave only trace amount of the desired products (Table 2.2). Furthermore, attempts to convert Weinreb amide 2.91 into ketone 2.138 and to further install the saturated side chain to afford alcohol 2.139 were unsuccessful (Scheme 2.28).  Scheme 2.28 Attempts to install saturated side chains at C-2  With the failed attempts of directly installing saturated side chains, we decided to reduce the olefin to the alkane using hydrogen. Substrates with no halogen substitution on the aryl ring were used due to the cleavage of C-X bond, which occurs under these reduction conditions.   Scheme 2.29 Hydrogenation of 2.97 BocNHBrNOOCH3LiTHFBocNHBrOTHFR-metal BocNBr HROH2.91 2.138 2.139H2 (1 atm), Pd/Cr.t., 18hBocNHOH2 (50 psi), Pd/Cr.t., 1hBocNHO BocNHO91%no reaction2.972.97BocNHO BocNH OHHNH OH2.1062.97 2.1012.140  57 Given the steric effect of alcohols 2.101 and 2.106, ketone 2.97 was chosen as the substrate for the hydrogenation reaction. Hydrogenation of 2.97 with 10% Pd/C under hydrogen (1 atm) for 18 hours gave no products. Elevating the pressure to 50 psi and stirring for 1 hour led to complete consumption of starting material 2.97, and formation of the desired product 2.140 in 91% yield (Scheme 2.29).  Scheme 2.30 Synthesis of tetrahydroquinoline 2.146  BocNHOMeMgBr, Et2O92%BocNH OHHCOOH, CH2Cl2r.t., 18h58%HNPPh3, CCl4, CH2Cl240ºC, 30min ClHN COOHHSOCl2,MeOH40ºC, 4h98%HN COOMeHBoc2O, CH2Cl2r.t., 18h97%BocN COOMeHi-PrMgCl, Me(MeONH)•HClTHF, −20ºC to −10ºC73%BocNHNOOMgBr , THF0ºC, 1h35%H2 (50 psi), Pd/Cr.t., 1hBocNHO91%0ºC, 2h96%NBS, DMF BocNHOBrBrHNH OHBr–40ºC, 2hCuCN, NMP200ºC, 25minTMSCl, MeOH50ºC, 4hMeO2C56% brsm40%58% brsm57%53%HNNC H OH2.1442.972.140HNMeO2C HOH2.1452.63 2.64 2.652.662.1412.142 2.1432.146  58 Next, bromination of ketone 2.140 using NBS gave bromide 2.141 in a yield of 96% (Scheme 2.30). Subsequent diastereoselective 1,2-addition of MeMgBr to 2.141 afforded a single diastereomer 2.142 in a high yield. Further deprotection of 2.142 with formic acid in CH2Cl2 yielded alcohols 2.143, which was subsequently converted to nitrile 2.144 using CuCN in NMP under microwave irradiation in a yield of 40%. Nitrile 2.144 was hydrolyzed to methyl ester 2.145 with TMSCl in methanol. Diastereoselective PPh3–CCl4-mediated rearrangement of methyl ester 2.145 provided tetrahydroquinoline 2.146 in a yield of 53%.  As shown in Scheme 2.31, ketone 2.140 was elaborated to alcohol 2.148 through the sequence of diastereoselective 1, 2-addition of the Grignard and N-Boc deprotection. Tetrahydroquinolines 2.151 and 2.152 were synthesized from alcohols 2.149 and 2.150 via PPh3–CCl4-mediated rearrangement.  Scheme 2.31 Synthesis of tetrahydroquinolines 2.151 and 2.152  As shown in Scheme 2.32, ester 2.146 was subjected to treatment with LiOH (2 equiv.) in MeCN/H2O by the same method used previously by Hill. It was found that this reaction resulted in trace amounts of the desired product 2.153 and the undesired product 2.154. The mechanism of formation of 2.154 probably involved the initial formation of an aziridine BocNHO2.140MeMgBr, Et2O100%BocNH OHHCOOH, CH2Cl2r.t. 18h86%HNH OH-40ºC, 2hHNH OHRPPh3, CCl4, CH2Cl240ºC, 30minHNClRR = Br, 2.149R = H, 2.150R = Br, 2.151, 53%R = H, 2.152, 69%2.147 2.148  59 intermediate followed by a ring opening with a hydroxide. Attempts to convert unreacted starting material 2.146 to 2.153 by increasing the amount of LiOH to 10 equiv. resulted in only trace amount of the desired product 2.153 and major amount of the undesired product 2.154 (Scheme 2.32). It was also found that the desired product 2.153 was not obtained when the reaction was carried out at room temperature for 5 days (Scheme 2.32). Attempts to hydrolyze ester 2.146 to carboxylic acid 2.153 under acidic conditions were unsuccessful (Scheme 2.32).  Scheme 2.32 Attempts to hydrolyze 2.146 to 2.153  Attempts were also made to install bromo group instead of chloro group at C-3 by carrying out the rearrangement using CBr4 instead of CCl4. The reaction of alcohol 2.145 in the presence of PPh3 and CBr4 at 40°C in CH2Cl2 for 6 hours gave tetrahydroquinoline 2.155 in 40% yield. Flash column chromatography purification failed to give clean NMR data of 2.155 so the LiOH (10 equiv.), MeCN/H2O70ºC, 4hHNClMeO2C2.146HNClHO2C2.153trace amountHNOHHO2C2.154major+HNClMeO2C2.146HCl (conc.),  THF/H2Or.t., 24hHNClHO2C2.153LiOH, MeCN/H2Or.t., 5dHNClMeO2C2.146HNClHO2C2.153HNClMeO2C2.146HCl (conc.), THF/H2Oreflux, 18hHNClHO2C2.153LiOH (2 equiv.), MeCN/H2O70ºC, 4hHNClMeO2C2.146HNClHO2C2.153trace amountHNOHHO2C2.154trace amount+  60 mixture was further purified by reversed phase HPLC. However, HPLC purification resulted in decomposition of 2.155.  Scheme 2.33 Attempts to synthesize 2.155HNMeO2C HOH2.145HNPPh3, CBr4, CH2Cl240ºC, 6h BrMeO2C40%2.155  61 Chapter 3: Conclusion 3.1 Biological results The biological activity of analogs 2.113, 2.115, 2.108, 2.130, 2.144, 2.145, 2.146, 2.151 and 2.152 was investigated by our collaborator Dr. Michel Roberge at UBC. An automated punctate EGFP-LC3 formation cell-based assay was used to measure autophagy-inhibiting activity of these analogs. In this assay, the cell line MCF-7 expressing EGFP-LC3 was exposed to these analogs alone and in the presence of 75 µM CQ.   Figure 3.1 Virantmycin analogs synthesized  Analog 2.108 was found to be the most potent inhibitor (5 µg/mL), inducing puncta on its own at 10 µg/mL and inhibiting puncta formation at 5 µg/mL in the presence of CQ. Analog 2.152 was found to inhibit autophagy at 5 µg/mL, without inducing puncta on its own. This indicates that the olefin moiety plays a role in the activity of inducing the formation of small autophagosomes (LC-GFP puncta). This unusual effect is the same as that of virantmycin on HNClMeO2C2.146HNNC H OH2.144HNMeO2C HOH2.145HNBr Cl2.113HNBr Cl2.115HNNC H OH2.130HNClBrHNCl2.151 2.152HNCl2.108  62 autophagy. Analog 2.115 with a bromine substitution on the aryl ring has essentially lost all activity and analog 2.151 shows faint autophagy inhibiting activity at 10 µg/mL. This indicates that the halogen substitution is detrimental to the autophagy-inhibiting activity. Analogs 2.130, 2.144, 2.145 and 2.146 were found to have no autophagy inhibiting activity. This indicates that the indoline skeleton does not possess autophagy-inhibiting activity and the tetrahydroquinoline skeleton is necessary for the activity in the assay. Analog 2.113 was found to have faint autophagy inhibiting activity at 10 µg/mL, while 2.115 was inactive. This suggests that the cis configuration between the olefin side chain and the chloro group might be better than the trans configuration for autophagy inhibition.  3.2 Conclusion All attempts to install carbonyl group on the aryl ring from the aryl iodide, using Pd(OAc)2 as a catalyst, under a CO atmosphere failed. In order to circumvent this problem, methodology has been developed in this thesis research to use microwave irradiation to generate aryl nitriles, such as 2.144, which provided the access to carbonyl group and the flexibility for preparation of other analogs. This methodology also provided the access to the epimeric configuration at C-2, using different organometallic reagents in a highly diastereoselective 1, 2-addition to ketone 2.17.  Scheme 3.1 Carboxylic acid 2.156 to be synthesized  HNClHO2C2.156HNNC H OH2.144  63 Nine virantmycin analogs 2.113, 2.115, 2.108, 2.130, 2.144, 2.145, 2.146, 2.151 and 2.152 were synthesized. Analogs 2.108 and 2.152 were found to be potent autophagy inhibitors. The methoxy group and methyl group at C-3’ were found to not be necessary for autophagy inhibiting activity. The olefin moiety was found to be responsible for inducing the formation of small autophagosomes. The presence of bromine and methyl ester substituents on the aryl ring had negative effects on autophagy-inhibiting activity. The presence of the tetrahydroquinoline skeleton seems to be a required structural feature necessary for autophagy inhibition. The importance of the carboxylic acid group is yet to be conclusively determined. Modifications of the current methodology are ongoing to yield carboxylic acid containing analogs. The construction of simplified pharmacophore analogs 2.108 and 2.152 allows for scalable synthesis to provide quantities for animal testing.  Figure 3.2 Current understanding of virantmycin’s structural features HNClHOOCOMethe role of carboxyl groupis to be determinedmethoxy group is not requiredolefin moiety is responsible for inducing the formation of small autophagosomes2R3Rcis configuration between olefin side chain and chloro group might be better than trans  configurationmethyl group is not requiredtetrahydroquinoline structure is required3'  64 Chapter 4: Experimental General Methods: All moisture and oxygen sensitive reactions were carried out in flame-dried glassware and under an Ar atmosphere unless otherwise noted. Air and moisture sensitive liquid reagents were manipulated via a dry syringe. Anhydrous tetrahydrofuran (THF) and diethylether (Et2O) were obtained from distillation over sodium. All other solvents and reagents were used as obtained from commercial sources without further purification. 1H and 13C spectra were obtained on Bruker Avance 400 direct, 400 inverse, 300 direct or Bruker Avance 600 CryoProbe spectrometers at room temperature. All microwave-heating reactions were carried out in a single-mode microwave instrument, Biotage Initiator 2.5. Flash column chromatography was performed using Silicycle Ultra-Pure sulica gel (230-400 mesh). Analytical thin-layer chromatography (TLC) plates were aluminum-backed ultrapure silica gel 250 µm. Electrospray ionization mass spectrometry (ESI-MS) spectra were recorded on a Micromass LCT instrument. All biological assays were carried out by researchers in laboratory of Dr. Michel Roberge at UBC.            65 4.1 Preparation of methyl ester 2.64  (S)-indoline-2 carboxylic acid (2.63) (10.0 g, 61.3 mmol) was dissolved in dry methanol (100 ml) and cooled to 0 °C. Thionyl chloride (4.92 ml, 67.5 mmol) was added dropwise to the solution via syringe. The mixture was heated at 40 °C for 4 h, after which the reaction was cooled to room temperature then neutralized with saturated NaHCO3 solution and extracted with EtOAc (250×3 ml). The organic extracts were combined, dried with MgSO4, filtered and then concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 4:1) to give the product 2.64 (10.6 g, 98%) as a white solid.  1H NMR (400 MHz, CDCl3) δ 3.33 (dd, J = 16.0, 5.6 Hz, 1H), 3.41 (dd, J = 16.0, 10.1 Hz, 1H), 3.76 (s, 3H), 4.37–4.41 (m, 1H), 4.51 (br, 1H), 6.72 (d, J = 7.8 Hz, 1H), 6.76 (t, J = 7.4 Hz, 1H), 7.05–7.11 (m, 2H); 13C NMR (100 MHz, CDCl3) δ 33.9, 52.6, 60.0, 110.2, 119.6, 124.6, 126.8, 127.8, 150.3, 174.8; HRESIMS [M+H]+ calcd for C10H12NO2 178.0868, found 178.0869. HN COOMeH2.64  66   Figure 4.1 1H and 13C NMR spectra of 2.64 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 7  6  5  4 7.11217.09307.06837.04926.78296.76446.74596.73046.71094.50794.41194.39784.38674.37263.75813.43863.41323.39853.37323.35683.34283.31673.30271.98280.97160.99090.93151.02093.07791.01711.0609"LZ 02-12-pdt"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain"[ppm] 160  140  120  100  80  60  40 174.7846150.2633127.8449126.8028124.6206119.6102110.198759.999752.577133.8710"LZ 02-12-pdt"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain"  67 4.2 Preparation of methyl ester 2.65  A solution of Boc2O (21.1 g, 96.9 mmol) in dry CH2Cl2 (25 ml) was added to a solution of methyl ester 2.64 (10.6 g, 60.0 mmol) in dry CH2Cl2 (25 ml) at room temperature. After stirring at room temperature for 18 h, the reaction mixture was concentrated. The crude product was purified by column chromatography (hexane–EtOAc = 100% to 4:1) to afford 2.65 (16.1 g, 97%) as colorless oil. 1H NMR (400 MHz, CDCl3) δ 1.50 (br s, 9H), 3.11 (dd, J = 16.5, 4.3 Hz, 1H), 3.50 (dd, J = 16.1, 11.5 Hz, 1H), 3.74 (s, 3H), 4.87 (br s, 1H), 6.94 (t, J = 7.4 Hz, 1H), 7.10 (d, J = 7.4 Hz, 1H), 7.19 (t, J = 7.6 Hz, 1H), 7.49 (br s, 0.3H), 7.89 (br s, 0.7H); 13C NMR (100 MHz, CDCl3) δ 28.4 (×3), 32.1*, 32.8, 52.4, 60.5, 81.4, 82.5*, 114.8, 122.7, 124.5, 124.8, 128.0, 142.6, 151.7, 172.6; HRESIMS [M+Na]+ calcd for C15H19NO4Na 300.1212, found 300.1211.     BocN COOMeH2.65  68   Figure 4.2 1H and 13C NMR spectra of 2.65 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 8  7  6  5  4  3  2 7.89007.49277.20857.18977.17077.10907.09066.95806.93956.92104.86793.74393.53363.50423.49273.46463.13213.12133.09083.07991.59611.49670.67000.27851.03051.04091.04941.03053.16681.11951.06099.6028"LZ 02-18-pdt"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain"[ppm] 150  100  50 172.6030151.7159142.6434128.0348M 124.8182124.4733122.6934114.7716M 82.527981.423960.499352.434232.808132.067528.3935"LZ 02-18-pdt"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain"  69 4.3 Preparation of methyl ester 2.89  NBS (5.40 g, 30.3 mmol) was added to a solution of 2.65 (8.40 g. 30.3 mmol) in DMF (25 ml) at 0 °C. The mixture was stirred for 2 h at 0 °C. After addition of water (35 ml), the aqueous solution was extracted with EtOAc (35×3 ml). The combined organic extracts were dried over MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 100% to 7:3) to afford the product 2.89 (10.5 g, 97%) as a yellow oil. 1H NMR (400 MHz, CDCl3) δ 1.48 (br s, 9H), 3.10 (d, J = 16.4 Hz, 1H), 3.45–3.50 (m, 1H), 3.75 (s, 3H), 4.84 (br s, 1H), 7.21(s, 1H), 7.29–7.88 (m, 2H); 13C NMR (100 MHz, CDCl3) δ 27.8 (×3), 31.2*, 31.9, 52.0, 59.7*, 60.1, 81.3, 82.4*, 114.5, 115.6, 126.9, 127.3*, 129.7, 130.3, 130.7*, 130.4*, 141.4, 151.0, 151.9*, 171.6; HRESIMS [M+Na]+ calcd for C15H18NO4BrNa 378.0317, found 378.0324. BocN COOMeHBr2.89  70  Figure 4.3 1H and 13C NMR spectra of 2.89 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 8  7  6  5  4  3  2 7.88257.77087.75897.30077.28977.20974.92084.85154.83913.74603.50053.47443.45483.09373.06641.58721.47930.69861.04400.99071.00003.04771.05441.03669.0550"LZ 02-28-pdt"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"[ppm] 160  140  120  100  80  60  40 171.6496151.8761150.9721141.3610140.4341130.7373130.3363129.7244127.3293126.9498115.5591114.467082.373681.282960.052359.732551.988031.853431.151427.7517"LZ 02-28-pdt"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"  71 4.4 Preparation of methyl ester 2.90  2,6-di-tert-butyl-4-methylpyridine (5.00 g, 24.4 mmol) was added to a solution of 2.65 (1.69 g, 24.4 mmol) in dry CH2Cl2 (15 ml) at room temperature after which ICl in CH2Cl2 (25.0 ml, 1.0M) was added to the mixture at 0 °C. The reaction mixture was stirred at room temperature for 1 h. After 1 h, the reaction mixture was diluted with EtOAc (20 ml). Saturated Na2S2O3 solution (40 ml) was poured into the mixture. The organic layer was then separated and the aqueous layer was extracted with EtOAc (40×3 ml). The combined extracts were washed with 1N HCl (30 ml), followed by saturated NaHCO3 solution (30 ml), dried with MgSO4 and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 9:1 to 7:3) to yield the product 2.90 (2.13 g, 87%) as a yellow oil. 1H NMR (300 MHz, CDCl3) δ 1.48 (br s, 9H), 3.06 (dd, J = 16.7, 4.1 Hz, 1H), 3.46 (dd, J = 16.6, 11.5 Hz, 1H), 3.73 (s, 3H), 4.84 (br s, 1H), 7.26 (br s, 0.3H), 7.38 (s, 1H), 7.47 (d, J = 8.3 Hz, 1H), 7.64 (br s, 0.6H); 13C NMR (75 MHz, CDCl3) δ 28.4 (×3), 32.3, 52.6, 60.5, 81.9, 85.2, 116.7, 130.7, 133.3, 136.9, 142.6, 151.6, 172.2; HRESIMS [M+Na]+ calcd for C15H18NO4NaI 426.0178, found 426.0174. BocN COOMeHI2.90  72  Figure 4.4 1H and 13C NMR spectra of 2.90 recorded in CDCl3 at 300 MHz and 75 MHz, respectively.  [ppm] 8  7  6  5  4  3  2 7.63777.48297.45517.38477.25974.84183.73473.50403.46553.44863.41023.09593.08233.04013.02651.47590.62670.94440.90770.31490.96812.95441.03470.99649.3113"LZ 01-174-51-70"  1  1  /Users/lingzhizhang/Desktop/Iodo[ppm] 160  140  120  100  80  60  40 172.1692M 151.5891142.6476136.9137133.3461130.7243116.743485.154381.898460.497852.582732.293128.3708"LZ 01-174-51-70-2"  2  1  /Users/lingzhizhang/Desktop/Iodo  73 4.5 Preparation of Weinreb amide 2.66  i-PrMgCl in THF (8.9 ml, 2M) was added to a solution of Me(MeO)NH•HCl (1.30 g, 13.3 mmol) and 2.65 (2.46 g, 8.88 mmol) in dry THF (20 ml) dropwise at –20 °C. Then the reaction was warmed up to –10 °C. After stirring at –10 °C for 20 mins, the reaction mixture was added Me(MeO)NH•HCl (1.30 g, 13.3 mmol) and i-PrMgCl in THF (8.9 ml, 2M) at –20 °C. The reaction was stirred at –10 °C for an additional 20 mins after which Me(MeO)NH•HCl (1.30 g, 13.3 mmol) and i-PrMgCl in THF (8.9 ml, 2M) were added to the reaction at –20 °C. After stirring for 10 mins at –10 °C, the reaction was quenched with saturated NH4Cl solution (60 ml) and extracted with EtOAc (60×3 ml). The organic extracts were combined, dried with MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 4:1 to 1:1) to yield the product 2.66 (1.97g, 73%) as a white solid. 1H NMR (600 MHz, CDCl3) δ 1.49 (br s, 6H), 1.60 (br s, 3H), 3.00 (t, J = 16.5, 1H), 3.23 (s, 3H), 3.50 (dd, J = 13.3 Hz, 1H), 3.75 (br s, 2H), 3.83 (br s, 1H), 5.18 (br s, 0.6H), 5.26 (br s, 0.4H), 6.91 (t, J = 6.8, 1H), 7.07 (d, J = 6.7, 1H), 7.17 (br s, 1H), 7.51 (br s, 0.4H), 7.93 (br s, 0.6H); 13C NMR (150 MHz, CDCl3) δ 27.9 (×3), 28.0*, 31.5, 32.2, 57.9*, 58.4, 60.9, 80.5, 81.5*, 114.2, 121.9, 123.8, 124.2, 127.2, 127.3*, 127.7*, 141.8*, 142.8 151.3, 152.3*, 171.5*, 172.1; HRESIMS [M+Na]+ calcd for C16H22N2O4Na 329.1477, found 329.1475. BocNHNOO2.66  74  Figure 4.5 1H and 13C NMR spectra of 2.66 recorded in CDCl3 at 600 MHz and 150 MHz, respectively.  [ppm] 8  7  6  5  4  3  2 [rel] 0  5  10  15  20  25 7.92707.51277.17397.07607.06496.91826.90666.89545.25855.17723.82703.75213.51993.49533.47573.22613.02783.00022.97281.60311.49260.57710.40001.09271.02861.04530.42690.60661.25852.11321.10273.19731.08773.42636.1430"Non-halogenlated Weinred Amide"  4  1  "/Users/lingzhizhang/Desktop/NMR/AV 600/nmr"[ppm] 160  140  120  100  80  60  40 [rel] 0  5  10  15  20  25 172.1285171.4574152.2672151.3008142.7641141.9094127.6939127.3462127.1919124.2376123.7707121.8536114.230681.535480.529860.913758.375757.880632.157331.509328.027427.8869"Non-halogenlated Weinred Amide"  5  1  "/Users/lingzhizhang/Desktop/NMR/AV 600/nmr"  75 4.6 Preparation of Weinreb amide 2.91  The procedure for the synthesis of Weinreb amide 2.91 from methyl ester 2.89 was the same as the synthesis of Weinreb amide 2.66 from methyl ester 2.65 and afforded a pale yellow soild (75%). 1H NMR (400 MHz, CDCl3) δ 1.48 (br s, 6H), 1.59 (br s, 3H), 2.97 (d, J = 16.1 Hz, 1H), 3.22 (s, 3H), 3.46 (dd, J = 16.2, 11.8 Hz, 1H), 3.74 (br s, 2H), 3.81 (br s, 1H), 5.15 (br, 1H), 7.17 (s, 1H), 7.27 (br s, 1H), 7.35 (br s, 0.3H), 7.80 (d, J = 7.9 Hz, 0.6H); 13C NMR (100 MHz, CDCl3) δ 28.4 (×3), 31.8*, 32.3, 32.8, 58.6*, 59.1, 61.5, 81.5, 82.5*, 114.7, 116.1, 127.4, 127.8, 130.7, 131.7*, 141.8*, 142.7, 151.7, 152.5*, 171.6*, 172.2; HRESIMS [M+Na]+ calcd for C16H21N2O4NaBr 407.0582, found 407.0584.  BocNHNOO2.91Br  76  Figure 4.6 1H and 13C NMR spectra of 2.91 recorded in CDCl3 at 400 MHz and 100 MHz, respectively.  [ppm] 8  7  6  5  4  3  2 7.80617.78637.34577.26707.17175.23965.17265.15403.81203.74473.49903.46903.45803.42893.21612.99082.95061.58741.48160.59380.31191.08870.98550.98611.00351.96971.01153.00001.03203.19426.0059"Bromo Weinred Amide"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"[ppm] 160  140  120  100  80  60  40 172.1783171.5652152.4785151.7038142.6713141.7777131.6960130.6773127.7884127.3939116.1266114.664582.518681.490161.514359.093058.648532.761732.329931.750128.4061"Bromo Weinred Amide"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"  77 4.7 Preparation of Weinreb amide 2.92  The procedure for the synthesis of Weinreb amide 2.92 from methyl ester 2.90 was the same as the synthesis of Weinreb amide 2.66 from methyl ester 2.65 and afforded a pale yellow oil (58%). 1H NMR (300 MHz, CDCl3) δ 1.45 (br s, 6H), 1.56 (br s, 3H), 2.92 (d, J = 16.2 Hz, 1H), 3.18 (s, 3H), 3.42 (dd, J = 16.8, 11.5 Hz, 1H), 3.71 (br s, 2H), 3.77 (br s, 1H), 5.13 (br m, 1H), 7.23 (br s, 0.3H), 7.32 (s, 1H), 7.42 (d, J = 8.6, 1H), 7.66 (d, J = 8.2, 0.6H); 13C NMR (75 MHz, CDCl3) δ 28.4 (×3), 31.6*, 32.2, 32.8, 58.5*, 59.0, 61.6, 81.5, 82.5*, 84.8, 116.7, 131.2, 132.1*, 133.2, 133.6*, 136.6, 142.5*, 143.5, 151.7, 152.4*, 171.6*, 172.1; HRESIMS [M+Na]+ calcd for C16H21N2O4NaI 455.0444, found 455.0447. BocNHNOO2.92I  78   Figure 4.7 1H and 13C NMR spectra of 2.92 recorded in CDCl3 at 300 MHz and 75 MHz, respectively. [ppm] 8  7  6  5  4  3  2 7.67437.64697.43447.40597.32177.23605.13183.77333.71263.46773.42943.41173.37373.18062.94702.89311.55601.45240.60260.98220.94040.31071.00000.94162.06081.05283.02031.02873.23785.9966"LZ 01-192-63-65"  1  1  /Users/lingzhizhang/Desktop/Iodo[ppm] 160  140  120  100  80  60  40 172.1243171.5904152.4430151.6867143.4568142.5456136.6486133.5726133.1963132.1495131.1600116.714384.771282.522681.500761.574558.959658.549332.771732.213331.643128.4367"LZ 01-192-63-65"  2  1  /Users/lingzhizhang/Desktop/Iodo  79 4.8 Preparation of bromide 2.94  A solution of cyclopropyl methyl ketone (15.0 ml, 160 mmol) in dry THF (22.4 ml) was added dropwise to a solution of MeMgBr (65.0 ml, 192 mmol) in dry Et2O under an argon atmosphere. The mixture was heated to reflux for 20 mins. When the reaction mixture was cooled to 0 °C, a cooled solution of concentrated sulphuric acid in water (1:2) was added to the reaction at a rate that the temperature does not raise above 10 °C. White solid and yellow solution were observed. The reaction was allowed to warm to room temperature and to stir for 30 mins after addition. The organic layer was then separated and the aqueous layer was extracted with Et2O. The combined organic extracts were washed with saturated NaHCO3 solution, dried with MgSO4, filtered and concentrated. The crude was purified by distillation under reduced pressure (b.p. 80 °C, 60 mmHg) to yield the product 2.94 (20 g, 77%) as a colorless oil. 4.9 Preparation of Grignard reagent 2.95  To a flam dried flask containing magnesium turnings (2.57 g, 105 mmol), previously washed by diluted HCl, water and acetone and iodine (2 small crstals) was added a solution of 2.5 (11.8 ml, 88.0 mmol) in dry THF (60 ml) over a period of 3 h. The dark violet solution was stirred for 1 min until it became colorless. The solution became dark grey and the resulting solution was heated to reflux for 1 h and then cooled to room temperature before it was ready to be used.  Br2.94MgBr2.95  80 4.10 Preparation of ketone 2.97   A solution of 5-bromo-2-methyl-2-pentene magnesium bromide 2.95 in THF was added dropwise to a solution of Weinreb Amide 2.66 (2.70 g, 8.83 mmol) in dry THF (15 ml) at 0 °C. The mixture was stirred at 0 °C for 1 h. The reaction was quenched with saturated NH4Cl solution (40 ml) and extracted with EtOAc (40×3 ml). The combined extracts were dried with MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 100% to 1:1) to afford the product 2.97 (1.02 g, 35%) as pale yellow oil. Based on recovered starting material 2.66 (1.00 g, 3.27 mmol), the yield of 2.97 was 56%. 1H NMR (400 MHz, CDCl3) δ 1.49 (br s, 6H), 1.59 (br s, 6H), 1.65 (s, 3H), 2.26 (q, J = 7.0 Hz, 2H), 2.43–2.57 (m, 2H), 2.93 (dd, J = 16.6, 5.0 Hz), 3.46 (br t, J = 13.0 Hz, 1H), 4.80 (br s, 1H), 5.04 (t, J = 6.3 Hz, 1H), 6.95 (t, J = 7.2 Hz, 1H), 7.10 (d, J= 7.1 Hz, 1H), 7.20 (br t, J = 7.3 Hz, 1H), 7.49 (br s, 0.3H), 7.90 (br s, 0.6H); 13C NMR (100 MHz, CDCl3) δ 17.8, 22.0, 25.8, 28.4 (×3), 31.9, 37.5, 66.7, 81.7, 115.0, 122.8 (×2), 124.6, 128.1 (×2), 133.1, 142.9, 151.8, 208.0; HRESIMS [M+Na]+ calcd for C20H27NO3Na 352.1889, found 352.1894. 2.97BocNHO  81  Figure 4.8 1H and 13C NMR spectra of 2.97 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 8  7  6  5  4  3  2 7.90257.4876M 7.22017.2005M 7.18387.11057.09276.96816.95006.93195.05165.0370M 5.01994.8047M 3.49573.45933.43092.96072.94802.91892.90662.57232.55482.53002.51162.49342.46932.44932.43312.28712.27012.25252.23501.65031.60511.58011.48920.59340.26731.05281.02931.03350.92250.98511.00001.06332.09061.94983.21716.30036.3844"LZ 02-34-pdt"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"[ppm] 200  150  100  50 208.0073M 151.7715M 142.8513133.0864128.1387M 128.0991124.6488M 122.8384122.8066114.968681.715966.656637.538731.853528.385725.777321.989217.7628"LZ 02-34-pdt"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"  82 4.11 Preparation of ketone 2.96  The procedure for the synthesis of ketone 2.96 from Weinreb amide 2.91 was the same as the synthesis of ketone 2.97 from Weinreb amide 2.66 and afforded a pale yellow oil (38%, 58% based on recovered 2.66). 1H NMR (600 MHz, CDCl3) δ 1.49 (br s, 6H), 1.61 (br s, 6H), 1.68 (br, s 3H), 2.28 (q, J = 7.1 Hz, 2H), 2.39–2.55 (m, 2H), 2.93 (dd, J = 16.8, 4.5 Hz, 1H), 3.47 (br t, J = 13.8 Hz, 1H), 4.81 (br, 1H), 5.05 (br t, J = 7.1 Hz, 1H), 7.23 (s, 1H), 7.28 (s, 0.3H), 7.33 (d, J = 6.2 Hz, 1H), 7.80 (d, J = 7.0 Hz, 0.6H); 13C NMR (150 MHz, CDCl3) δ 17.2, 21.4, 25.2, 27.7 (×3), 30.8, 37.2, 66.4, 81.5, 114.6, 115.7, 122.0, 124.0, 127.1, 130.5, 132.7, 141.6, 151.1, 206.8; HRESIMS [M+Na]+ calcd for C20H26NO3NaBr 430.0994, found 430.0992. 2.96BocNHOBr  83  Figure 4.9 1H and 13C NMR spectra of 2.96 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.80757.79587.33897.32857.28147.23295.05374.93394.91844.82554.81353.48883.46393.44292.94772.94022.91972.91222.55042.53472.52322.49082.47732.46462.42662.39772.38612.28902.27721.67591.60631.59440.63121.00780.30700.90421.00881.08121.07781.00002.11242.02943.18816.02796.4314"LZ 02-112-17-39"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"[ppm] 200  150  100  50 206.7520M 151.1189M 141.5882M 132.7455130.4660127.1075124.0181122.0327115.7155114.611281.530666.041137.207130.836427.732825.231321.362417.2183"LZ 02-112-17-39"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"  84 4.12 Preparation of alcohol 2.98  A solution of MeLi (3.5 ml, 1.6M) in diethyl ether was added dropwise to a solution of 2.96 (425 mg, 1.04 mml) in dry THF (5 ml) at –78 °C. The mixture was stirred for 20 min at –78 °C. The reaction was quenched with saturated NH4Cl solution at –78 °C and warmed up to room temperature. The aqueous solution was extracted with EtOAc (15×3 ml). The combined organic extracts were dried with MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 100% to 7:3) to yield the product 2.98 (291 mg, 66%) as a colorless oil. 1H NMR (600 MHz, CDCl3) δ 0.89 (s, 3H), 1.47–1.54 (m, 2H), 1.58 (s, 9H), 1.63 (s 3H), 1.69 (s 3H), 2.14–2.17 (m, 2H), 2.76 (d, J = 16.6 Hz, 1H), 3.30 (dd, J = 16.7, 10.4 Hz, 1H), 4.60 (d, J = 9.7 Hz, 1H), 5.12 (t, J = 7.0 Hz, 1H), 7.23–7.31 (m, 3H); 13C NMR (150 MHz, CDCl3) δ 17.3, 20.6, 21.2, 25.3, 27.9 (×3), 30.3, 39.0, 66.2, 75.7, 82.6, 115.1, 117.3, 123.9, 126.9, 129.7, 131.4, 133.3, 141.5, 154.9; HRESIMS [M+Na]+ calcd for C21H30NO3NaBr 446.1307, found 446.1302. 2.98BocNBr H OH  85  Figure 4.10 1H and 13C NMR spectra of 2.98 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.30207.27107.23485.13305.12125.10964.60664.59053.32603.30873.29823.28092.77672.74902.17322.16282.15682.14672.13561.69001.63111.57601.54371.53641.52661.51911.50821.49681.48961.48431.47941.47331.46750.88713.38630.96971.01620.99940.98492.01812.93402.98069.13252.34413.0682"LZ 02-50-pdt"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Bromo"[ppm] 160  140  120  100  80  60  40  20 154.8935141.4630133.3200131.4212129.7219126.9127123.9016117.2899115.148582.561375.672766.170238.998130.282927.886225.293821.197420.574017.2660"LZ 02-50-pdt"  3  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Bromo"  86 4.13 Preparation of alcohol 2.99  The procedure for the synthesis of alcohol 2.99 from ketone 2.97 was the same as the synthesis of alcohol 2.98 from ketone 2.96 and afforded a colorless oil (56%). 1H NMR (600 MHz, CDCl3) δ 0.88 (s, 3H), 1.50–1.53 (m, 2H), 1.59 (s, 9H), 1.64 (s 3H), 1.69 (s 3H), 2.17–2.20 (m, 2H), 2.77 (d, J = 16.4 Hz, 1H), 3.33 (dd, J = 16.5, 10.4 Hz, 1H), 4.62 (d, J = 9.8 Hz, 1H), 5.14 (t, J = 7.1 Hz, 1H), 6.98 (t, J = 7.4 Hz, 1H), 7.12 (d, J = 7.3 Hz, 1H), 7.16 (t, J = 7.7 Hz, 1H), 7.45 (br s, 1H); 13C NMR (150 MHz, CDCl3) δ 17.3, 20.5, 21.2, 25.3, 27.9 (×3), 30.5, 39.2, 65.9, 75.8, 82.2, 116.0, 122.7, 123.9, 124.1, 126.8, 130.9, 131.3, 142.2, 155.5; HRESIMS [M+Na]+ calcd for C21H31NO3Na 368.2202, found 368.2209. 2.99BocNH OH  87  Figure 4.11 1H and 13C NMR spectra of 2.99 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.44817.17297.16017.14727.12637.11416.97506.96276.95045.14755.13575.12404.62534.60903.35073.33333.32323.30592.78702.75962.19522.18352.17912.16791.69441.63941.59011.55271.54251.53721.52531.52041.51511.50951.50340.87890.90690.90301.06220.99750.92471.13831.02621.01711.96683.10872.97279.08932.42523.2714"LZ 02-36-pdt"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Non halogenated"[ppm] 160  140  120  100  80  60  40 155.4834142.1680131.2682130.9220126.7954124.0760123.8923122.6555115.980482.210075.758965.929439.207930.547427.934025.304421.228520.535217.2668"LZ 02-36-pdt"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Non halogenated"  88 4.14 Preparation of alcohol 2.100  To a solution of 2.96 (292 mg, 0.71 mmol) in dry Et2O (10 ml) was added dropwise a solution of MeMgBr (1.2 ml, 3M) in dry Et2O at –40 °C. After stirring at –40 °C for 2 h, the reaction was quenched with saturated NH4Cl solution (15 ml) and extracted with EtOAc (15×3 ml). The combined organic extracts were dried with MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 100% to 7:3) to afford the product 2.100 (288 mg, 95%) as colorless oil. 1H NMR (600 MHz, CDCl3) δ 1.19 (s, 3H), 1.40–1.44 (m, 1H), 1.58 (s, 12H), 1.66 (s 3H), 1.71–1.77 (m, 1H), 2.01–2.05 (m, 1H), 2.10–2.14 (m, 1H), 2.94 (d, J = 16.3 Hz, 1H), 3.30 (dd, J = 16.6, 10.3 Hz, 1H), 4.54 (d, J = 9.1 Hz, 1H), 5.03 (t, J = 6.7 Hz, 1H), 7.25–7.36 (m, 3H); 13C NMR (150 MHz, CDCl3) δ 17.2, 21.5, 23.4, 25.2, 27.9 (×3), 30.1, 35.8, 67.5, 75.6, 82.2, 115.2, 117.4, 124.1, 126.8, 129.5, 131.2, 133.7, 141.7, 154.3; HRESIMS [M+Na]+ calcd for C21H30NO3NaBr 446.1307, found 446.1302. BocNBr H OH2.100  89  Figure 4.12 1H and 13C NMR spectra of 2.100 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.36317.27837.25145.3100M 5.04055.0288M 5.01834.54614.53103.32713.30993.29943.28222.95392.92682.14392.12242.11332.10342.04842.03682.02742.01772.00551.77321.75311.73931.70841.65871.58541.41821.40431.18863.10610.94671.01411.11551.00001.02351.01710.93263.169812.22291.28943.0671"LZ 02-148-pdt"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Bromo"[ppm] 160  140  120  100  80  60  40  20 M 154.3471141.6830133.6909131.2135129.5235126.7619124.1210117.4193115.177382.169875.565967.542235.755630.067027.873725.248223.351221.521817.2100"LZ 02-148-pdt"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Bromo"  90 4.15 Preparation of alcohol 2.101  The procedure for the synthesis of alcohol 2.101 from ketone 2.97 was the same as the synthesis of alcohol 2.100 from ketone 2.96 and afforded colorless oil (95%). 1H NMR (600 MHz, CDCl3) δ 1.19 (s, 3H), 1.23–1.25 (m, 1H), 1.38–1.43 (m, 1H), 1.54 (s, 3H), 1.58 (s 9H), 1.62 (s 3H), 1.97–2.04 (m, 1H), 2.07–2.13 (m, 1H), 2.92 (d, J = 16.4 Hz, 1H), 3.32 (dd, J = 16.5, 10.5 Hz, 1H), 4.54 (d, J = 9.7 Hz, 1H), 4.99 (t, J = 6.0 Hz, 1H), 6.96 (t, J = 7.4 Hz, 1H), 7.12 (d, J = 7.3 Hz, 1H), 7.15 (t, J = 7.8 Hz, 1H), 7.48 (br s, 1H); 13C NMR (150 MHz, CDCl3) δ 17.2, 21.5, 23.6, 25.2, 27.9 (×3), 30.4, 35.5, 67.6, 75.7, 81.9, 116.1, 122.7, 123.8, 124.3, 126.7, 131.0, 131.2, 142.4, 154.7; HRESIMS [M+Na]+ calcd for C21H31NO3Na 368.2202, found 368.2209. BocNH OH2.101  91  Figure 4.13 1H and 13C NMR spectra of 2.101 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 [rel] 0  5  10  15  20  25  30 7.47727.16597.15317.14007.12847.11626.97576.96346.95104.99814.98814.97804.55044.53433.34373.32623.31623.29882.93082.90342.13452.12522.11232.10292.09312.08292.07332.03702.02712.01682.00711.99661.98451.97441.62161.58161.53901.43181.42071.40591.40111.38690.88190.98561.00331.02050.95451.03911.04821.00001.00901.02793.11649.02433.06951.16711.08763.1756"LZ 03-196-28-36"  3  1  /Users/lingzhizhang/Desktop[ppm] 7  6  5  4  3  2 [rel] 0  5  10  15  20  25  30 7.47727.16597.15317.14007.12847.11626.97576.96346.95104.99814.98814.97804.55044.53433.34373.32623.31623.29882.93082.90342.13452.12522.11232.10292.09312.08292.07332.03702.02712.01682.00711.99661.98451.97441.62161.58161.53901.43181.42071.40591.40111.38690.88190.98561.00331.02050.95451.03911.04821.00001.00901.02793.11649.02433.06951.16711.08763.1756"LZ 03-196-28-36"  3  1  /Users/lingzhizhang/Desktop  92 4.16 Preparation of ketone 2.102  The procedure for the synthesis of ketone 2.102 from Weinreb amide 2.92 was the same as the synthesis of ketone 2.97 from Weinreb amide 2.66 and afforded a yellow oil (35%, 55% based on recovered 2.37).  1H NMR (300 MHz, CDCl3) δ 1.51 (br s, 6H), 1.61 (br s, 6H), 1.68 (br s, 3H), 2.29 (q, J = 7.1 Hz, 2H), 2.45–2.58 (m, 2H), 2.93 (dd, J = 16.8, 4.8 Hz, 1H), 3.47 (br t, J = 13.8 Hz, 1H), 4.83 (br s, 1H), 5.06 (br t, J = 6.7 Hz, 1H), 7.43 (s, 1H), 7.52 (d, J = 8.6 Hz, 1H), 7.69 (br s, 0.6H); 13C NMR (150 MHz, CDCl3) δ 17.2, 21.4, 25.2, 27.7 (×3), 30.7, 36.9, 65.9, 81.6, 84.7, 114.4, 116.3, 122.0, 127.6, 130.2, 132.7, 136.5, 142.3, 151.1, 207.5; HRESIMS [M+Na]+ calcd for C20H26NO3NaI 478.0855, found 478.0847. BocNHIO2.102  93  Figure 4.14 1H and 13C NMR spectra of 2.102 recorded in CDCl3 at 300 MHz and 150 MHz, respectively. [ppm] 8  7  6  5  4  3  2 7.68967.53637.50767.42697.28837.14277.11827.00186.97736.95355.08305.06055.03824.83213.50063.44843.40862.96212.94582.90602.89012.57752.54512.52062.49672.45022.32292.29952.27592.25241.68311.61291.50550.56650.90940.81740.31560.19100.17070.91780.97190.98631.00001.96631.88523.37026.25855.9376"LZ 02 -64-pdt"  1  1  "/Users/lingzhizhang/Desktop/NMR/AV 300/nmr"[ppm] 200  150  100  50 207.4534206.6992151.0531142.3228136.4652132.8973132.7306132.5550130.1611127.6005124.0866124.0259122.2964122.2052122.0448116.2999114.386984.732381.555281.159366.103165.922637.201536.949531.275830.690927.731825.232921.364617.2198"LZ 01-194-47-51"  2  1  "/Users/lingzhizhang/Desktop/NMR/AV 600/nmr"  94 4.17 Preparation of alcohol 2.104  The procedure for the synthesis of alcohol 2.104 from ketone 2.102 was the same as the synthesis of alcohol 2.100 from ketone 2.96 and afforded yellow oil (91%). 1H NMR (400 MHz, CDCl3) δ 1.18 (s, 3H), 1.22–1.29 (m, 1H), 1.37–1.44 (m, 1H), 1.57 (s, 12H), 1.66 (s 3H), 1.96–2.06 (m, 1H), 2.08–2.16 (m, 1H), 2.92 (d, J = 16.8 Hz, 1H), 3.28 (dd, J = 16.8, 10.4 Hz, 1H), 4.51 (d, J = 10.2 Hz, 1H), 5.02 (t, J = 7.1 Hz, 1H), 7.24 (br s, 1H), 7.43–7.47 (m, 2H); 13C NMR (100 MHz, CDCl3) δ 17.8, 22.1, 21.5, 24.0, 25.8, 28.4 (×3), 30.5, 36.3, 68.0, 76.2, 82.8, 86.1, 123.2, 124.7, 131.8, 133.2, 134.6, 136.1, 143.0, 155.1; HRESIMS [M+Na]+ calcd for C21H30NO3NaI 494.1168, found 494.1174. BocNH OHI2.104  95  Figure 4.15 1H and 13C NMR spectra of 2.104 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 7  6  5  4  3  2 7.46577.44447.43277.2672M 7.23865.03565.01815.00034.51964.49413.31883.29293.27683.25092.93952.89762.15832.14332.10882.09482.07972.06202.04392.02892.01572.00001.98051.96491.65061.57111.44451.43891.43171.41051.40322.05390.89491.17181.13471.35661.29391.33521.28073.025512.62891.39061.23393.0000"LZ 02-66-pdt"  1  1  "/Users/lingzhizhang/Desktop/NMR/AV 400 DIR/nmr"[ppm] 160  140  120  100  80  60  40  20 [rel]- 0  1  2  3  4  5  6 M 155.0635143.0246136.1364134.6191133.1693131.8155124.6906123.240586.119282.784676.150968.011736.324130.501928.443325.806023.998122.101717.7716"LZ 02-66-pdt"  2  1  "/Users/lingzhizhang/Desktop/NMR/AV 400 DIR/nmr"  96 4.18 Preparation of alcohol 2.105  The procedure for the synthesis of alcohol 2.105 from alcohol 2.104 was the same as the synthesis of alcohol 2.143 from alcohol 2.142 and afforded a colorless solid (38%). 1H NMR (400 MHz, CDCl3) δ 1.23 (s, 3H), 1.38–1.44 (m, 1H), 1.55–1.60 (m, 1H), 1.64 (s 3H), 1.70 (s 3H), 2.02–2.14 (m, 2H), 2.91–3.07 (m, 2H), 3.87 (t, J = 9.7 Hz, 1H), 5.13 (t, J = 6.3 Hz, 1H), 6.42 (d, J = 8.2 Hz, 1H), 7.27–7.29 (m, 2H), 7.34 (s, 1H); 13C NMR (100 MHz, CDCl3) δ 17.8, 22.4, 24.9, 25.9, 30.4, 37.5, 68.2, 72.7, 80.1, 111.8, 124.4, 132.1, 132.4, 133.5, 136.0, 150.6; HRESIMS [M+H]+ calcd for C16H23NOI 372.0824, found 372.0826.  HNI H OH2.105  97  Figure 4.16 1H and 13C NMR spectra of 2.105 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 7  6  5  4  3  2 [rel] 0  5  10  15 7.33977.29377.26796.43346.41305.14505.13085.11333.89903.87493.85063.06633.05203.04153.02633.00112.97222.94892.93212.90862.14112.12552.11352.09942.08392.07062.05572.05182.04092.02281.70201.63561.59551.58711.58021.56031.55181.44051.42661.42031.41260.93891.13090.94331.10851.30152.35222.41093.32923.14931.11371.33322.5600"LZ 02-176-46-60"  1  1  "/Users/lingzhizhang/Desktop/NMR/AV 400 DIR/nmr"[ppm] 140  120  100  80  60  40  20 [rel] 0  5  10  15  20 150.5567135.9943133.4762132.4223132.1179124.3679111.759780.141372.676668.232137.531930.384825.852424.934622.415217.8395"LZ 02-176-46-60"  2  1  "/Users/lingzhizhang/Desktop/NMR/AV 400 DIR/nmr"  98 4.19 Preparation of tetrahydroquinoline 2.107  The procedure for the synthesis of tetrahydroquinoline 2.107 from alcohol 2.105 was the same as the synthesis of tetrahydroquinoline 2.146 from alcohol 2.145 and afforded a pale yellow oil (62%).  1H NMR (600 MHz, CDCl3) δ 1.23 (s, 3H), 1.45–1.50 (m, 1H), 1.62 (s, 3H), 1.69 (s, 3H), 1.71–1.74 (m, 1H), 2.05–2.13 (m, 2H), 3.04 (dd, J = 16.9, 8.3 Hz, 1H), 3.22 (dd, J = 17.0, 5.4 Hz, 1H), 3.79 (br s, 1H), 4.14 (dd, J = 8.3, 5.4 Hz, 1H), 5.09 (t, J = 6.4, 1H), 6.29 (d, J = 9.0 Hz, 1H), 7.25 (br m, 2H); 13C NMR (150 MHz, CDCl3) δ 17.3, 20.9, 21.1, 25.3, 34.1, 39.4, 55.0, 60.0, 77.6, 123.3, 127.0, 128.7, 131.8, 135.5, 137.1, 142.3; HREIMS [M+H]+ calcd for C16H22NClI 390.0486, found 390.0484. HNI Cl2.107  99  Figure 4.17 1H and 13C NMR spectra of 2.107 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 [rel] 0  5  10  15  20  25  30 7.24916.29386.27885.10115.09925.08975.07804.15414.14514.14034.13133.78643.23653.22733.20803.19913.06413.05023.03593.02202.13222.12002.10772.09742.08862.07982.07022.06072.04781.73951.73181.72801.72291.71601.71111.70531.68771.61241.50171.49241.48401.47791.47461.70910.99211.26131.25530.91111.14421.20962.34041.30903.50553.11702.7020"LZ 02-182"  1  1  "/Users/lingzhizhang/Desktop/NMR/AV 600/nmr"[ppm] 140  120  100  80  60  40  20 [rel] 0  2  4  6 142.2678137.1229135.5299131.7951128.7498126.9619123.330277.611459.952655.025839.382734.110425.269321.140620.941517.2706"LZ 02-92-pdt"  2  1  "/Users/lingzhizhang/Desktop/NMR/AV 600/nmr"  100 4.20 Preparation of alcohol 2.110  The procedure for the synthesis of alcohol 2.110 from alcohol 2.98 was the same as the synthesis of alcohol 2.143 from alcohol 2.142 and afforded a colorless solid (40%). 1H NMR (400 MHz, CDCl3) δ 1.14 (s, 3H), 1.44–1.59 (m, 2H), 1.63 (s 3H), 1.70 (s 3H), 2.02–2.20 (m, 2H), 2.93–3.07 (m, 2H), 3.90 (t, J = 9.6 Hz, 1H), 5.13 (t, J = 6.3 Hz, 1H), 6.52 (d, J = 8.2 Hz, 1H), 7.09–7.11 (m, 1H), 7.16 (s, 1H); 13C NMR (100 MHz, CDCl3) δ 17.9, 21.4, 22.5, 25.8, 30.9, 40.8, 67.8, 72.9, 111.3 (×2), 124.2, 127.8, 130.1, 131.9, 132.3, 149.5; HRESIMS [M+H]+ calcd for C16H23NOBr 324.0963, found 324.0966. HNBr H OH2.110  101  Figure 4.18 1H and 13C NMR spectra of 2.110 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 7  6  5  4  3  2 7.16407.11337.09717.09276.53196.51145.15025.13275.11873.92783.90383.87973.06513.04043.02502.99522.97092.95492.93082.20462.18922.12792.11212.09542.06982.05352.04412.03562.01891.69821.63381.58771.57871.57411.55341.54751.53951.52721.51751.51331.50261.49081.48371.47581.46851.45670.95520.95350.96320.98291.09812.08782.31912.97923.00002.36532.9885"LZ 01-166-32-52"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Bromo"[ppm] 140  120  100  80  60  40  20 149.4861132.2782131.9322130.0749127.7766124.2399111.264572.866967.802240.827530.896125.837522.522821.387117.8602"LZ 01-166-32-52"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Bromo"  102 4.21 Preparation of alcohol 2.111  The procedure for the synthesis of alcohol 2.111 from alcohol 2.23 was the same as the synthesis of alcohol 2.143 from alcohol 2.142 and afforded a colorless solid (40%). 1H NMR (400 MHz, CDCl3) δ 1.17 (s, 3H), 1.45–1.61 (m, 2H), 1.65 (s 3H), 1.71 (s 3H), 2.05–2.11 (m, 1H), 2.13–2.17 (m, 1H), 2.98 (dd, J = 15.7, 9.4 Hz, 1H), 3.06 (dd, J = 15.6, 10.2 Hz, 1H), 3.89 (t, J = 9.7 Hz, 1H), 5.15 (t, J = 6.8 Hz, 1H), 6.65 (d, J = 7.2 Hz, 1H), 6.73 (t, J = 7.4 Hz, 1H), 7.02 (t, J = 7.5 Hz, 1H), 7.09 (d, J = 7.2 Hz, 1H); 13C NMR (100 MHz, CDCl3) δ 17.9, 21.5, 22.6, 25.9, 31.0, 41.1, 67.5, 72.7, 109.8, 119.4, 124.4, 124.8, 127.4, 129.4, 132.1, 150.9; HRESIMS [M+H]+ calcd for C16H24NO 246.1858, found 246.1866. HNH OH2.111  103  Figure 4.19 1H and 13C NMR spectra of 2.111 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 7  6  5  4  3  2 7.09547.07747.04297.02407.00526.74886.73046.71206.65716.63785.16895.15185.13493.91723.89303.86863.08773.06193.04843.02313.00802.98462.96892.94532.17242.16072.14522.12622.10672.09072.08042.06332.05121.71141.64671.60731.59351.57291.55971.54621.52931.51291.49991.48561.46640.96891.03440.97140.98350.99741.20670.98361.06541.44971.22073.04893.04422.08913.1159"LZ 02-14-18-28"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Non halogenated"[ppm] 140  120  100  80  60  40  20 150.8560132.0949129.4204127.3600124.8251124.4381119.3743109.800972.698967.528041.116130.974025.851522.621421.457517.8540"LZ 02-14-18-28"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Non halogenated"  104 4.22 Preparation of alcohol 2.112  The procedure for the synthesis of alcohol 2.112 from alcohol 2.100 was the same as the synthesis of alcohol 2.143 from alcohol 2.142 and afforded a colorless solid (40%). 1H NMR (600 MHz, CDCl3) δ 1.23 (s, 3H), 1.40–1.44 (m, 1H), 1.55–1.61 (m, 1H), 1.63 (s 3H), 1.69 (s 3H), 2.02–2.08 (m, 1H), 2.09–2.15 (m, 1H), 2.95 (dd, J = 16.1, 9.3 Hz, 1H), 3.04 (dd, J = 16.0, 10.1 Hz, 1H), 3.89 (t, J = 9.7 Hz, 1H), 5.12 (t, J = 7.0 Hz, 1H), 6.51 (d, J = 8.2 Hz, 1H), 7.09–7.11 (m, 1H), 7.16 (s, 1H); 13C NMR (100 MHz, CDCl3) δ 17.3, 21.8, 24.4, 25.3, 30.0, 37.0, 67.8, 72.0, 110.4, 111.5, 123.8, 127.2, 129.4, 131.4, 131.6, 149.2; HRESIMS [M+H]+ calcd for C16H23NOBr 324.0963, found 324.0966. HNBr H OH2.112  105  Figure 4.20 1H and 13C NMR spectra of 2.112 recorded in CDCl3 at 600 MHz and 100 MHz, respectively. [ppm] 7  6  5  4  3  2 7.10707.10527.09407.09337.09156.51576.50205.13495.12325.11153.90153.88523.86923.06123.04443.03453.01772.96862.95312.94192.92632.15292.14812.14402.13152.12082.11132.10132.09082.08172.07102.06102.05192.04112.02912.01861.69291.62741.60531.59621.58641.58241.57491.56361.55451.43990.88490.94100.93590.90821.13120.98410.98891.52041.06792.92382.80061.27481.19683.1314"LZ 03-08-41-60"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Bromo"[ppm] 140  120  100  80  60  40  20 149.2058131.5819131.3627129.4302127.2032123.7627110.5229110.431872.045967.825136.981129.947325.284124.417921.844417.2685"LZ 03-08-41-60"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Bromo"  106 4.23 Preparation of alcohol 2.106  The procedure for the synthesis of alcohol 2.106 from alcohol 2.101 was the same as the synthesis of alcohol 2.143 from alcohol 2.142 and afforded a colorless solid (40%). 1H NMR (600 MHz, CDCl3) δ 1.28 (s, 3H), 1.42–1.47 (m, 1H), 1.56–1.61 (m, 1H), 1.63 (s 3H), 1.69 (s 3H), 2.03–2.09 (m, 1H), 2.10–2.16 (m, 1H), 2.39 (br s, 1H), 2.98 (dd, J = 15.7, 9.1 Hz, 1H), 3.09 (dd, J = 15.6, 10.4 Hz, 1H), 3.91 (t, J = 9.7 Hz, 1H), 5.13 (t, J = 6.8 Hz, 1H), 6.73 (d, J = 7.7 Hz, 1H), 6.77 (t, J = 7.4 Hz, 1H), 7.04 (t, J = 7.6 Hz, 1H), 7.10 (d, J = 7.3 Hz, 1H); 13C NMR (150 MHz, CDCl3) δ 17.3, 21.9, 24.6, 25.3, 30.0, 37.3, 67.7, 71.8, 110.0, 119.6, 123.8, 124.4, 126.9, 129.3, 131.5, 149.3; HRESIMS [M+H]+ calcd for C16H24NO 246.1858, found 246.1866. HNH OH2.106  107  Figure 4.21 1H and 13C NMR spectra of 2.106 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.10457.09247.05367.04107.02846.78506.77286.76056.73206.71925.14325.13195.12063.92523.90903.89283.10683.08933.08073.06342.99692.98172.97072.95552.39062.16432.15412.14192.13222.12272.11262.10202.09302.08202.07232.06322.05252.04042.03061.69441.63131.61341.60811.59411.58541.56150.95560.96840.92940.87231.02180.94620.98151.06870.80721.08161.22463.09723.16611.24501.17873.3586"LZ 03-184-34-40"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Non halogenated"[ppm] 140  120  100  80  60  40  20 M 149.2644131.4757129.3471126.8507124.3539123.8440119.6055110.019371.845267.724037.270730.007525.286324.639921.908517.2700"LZ 03-184-34-40"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Non halogenated"  108 4.24 Preparation of tetrahydroquinoline 2.113  The procedure for the synthesis of tetrahydroquinoline 2.113 from alcohol 2.110 was the same as the synthesis of tetrahydroquinoline 2.146 from alcohol 2.145 and afforded a yellow oil (54%). 1H NMR (400 MHz, CDCl3) δ 1.26 (s, 3H), 1.47–1.56 (m, 1H), 1.61 (s, 3H), 1.69 (s, 3H), 1.73–1.80 (m, 1H), 1.99–2.11 (m, 2H), 3.05 (dd, J = 17.3, 7.4 Hz, 1H), 3.24 (dd, J = 17.2, 5.3 Hz, 1H), 3.84 (br s, 1H), 4.12 (dd, J = 7.3, 5.4 Hz, 1H), 5.10 (t, J = 7.0, 1H), 6.39 (d, J = 9.1 Hz, 1H), 7.09–7.11 (m, 2H); 13C NMR (100 MHz, CDCl3) δ 17.8, 22.0, 24.9, 25.8, 34.1, 34.9, 55.4, 61.4, 109.3, 116.4, 120.1, 123.9, 130.3, 131.9, 132.3, 141.4; HREIMS [M]+ calcd for C16H21N35Cl79Br 341.05459, found 341.05438.  HNClBr2.113  109   Figure 4.22 1H and 13C NMR spectra of 2.113 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 7  6  5  4  3  2 7.10727.09166.39746.37465.11825.10065.08314.13494.12134.11664.10313.83853.26443.25103.22133.20803.08493.06643.04173.02332.11142.09322.07662.06702.05032.03472.00801.99021.80271.78771.77731.76911.76261.75491.74341.72911.69131.60781.55541.54451.52931.51781.86600.92181.02601.03260.94110.99331.01092.19351.08153.25612.91731.37293.0244"LZ 02-74-pdt"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Bromo"[ppm] 140  120  100  80  60  40 141.4385132.3069131.9152130.3020123.8892120.1334116.3900109.334361.416455.351734.901934.120725.845324.876022.044917.7913"LZ 02-74-pdt"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Bromo"  110 4.25 Preparation of tetrahydroquinoline 2.114  The procedure for the synthesis of tetrahydroquinoline 2.114 from alcohol 2.111was the same as the synthesis of tetrahydroquinoline 2.146 from alcohol 2.145 and afforded pale yellow oil (47%). 1H NMR (400 MHz, CDCl3) δ 1.28 (s, 3H), 1.50–1.58 (m, 1H), 1.60 (s, 3H), 1.69 (s, 3H), 1.73–1.80 (m, 1H), 2.00–2.10 (m, 2H), 3.10 (dd, J = 17.0, 7.9 Hz, 1H), 3.26 (dd, J = 17.0, 5.4 Hz, 1H), 3.82 (br s, 1H), 4.16 (dd, J = 7.9, 5.5 Hz, 1H), 5.11 (t, J = 7.0, 1H), 6.50 (d, J = 8.0 Hz, 1H), 6.67 (t, J = 7.4 Hz, 1H), 6.98 (d, J = 7.5 Hz, 1H), 7.02 (t, J = 7.7 Hz, 1H); 13C NMR (100 MHz, CDCl3) δ 17.8, 22.1, 25.0, 25.8, 34.4, 34.5, 55.3, 62.5, 114.9, 117.8, 118.3, 124.1, 127.5, 129.4, 132.1, 142.4; HRESIMS [M+H]+ calcd for C16H23NCl 264.1519, found 264.1515. HNCl2.114  111  Figure 4.23 1H and 13C NMR spectra of 2.114 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 7  6  5  4  3  2 7.03667.01826.99836.98536.96666.69276.67426.65596.51376.49385.12395.10645.08914.17624.16254.15654.14283.82133.29003.27643.24743.23393.12773.10803.08523.06542.09762.08102.05732.03992.02071.99911.80181.78611.77741.76831.76181.75371.74291.72821.68621.60201.57951.56331.55351.54001.06000.99701.00220.98511.17691.08100.96421.00001.04982.42661.24243.68573.28351.59263.7359"LZ 02-20-30-32"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Non halogenated"[ppm] 140  120  100  80  60  40 142.4015132.0959129.4055127.5146124.1265118.2689117.8460114.857062.455855.285034.462234.411425.831925.017022.050217.7646"LZ 02-20-30-32"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeLi/Non halogenated"  112 4.26 Preparation of tetrahydroquinoline 2.115  The procedure for the synthesis of tetrahydroquinoline 2.115 from alcohol 2.112 was the same as the synthesis of tetrahydroquinoline 2.146 from alcohol 2.145 and afforded a pale yellow oil (62%). 1H NMR (600 MHz, CDCl3) δ 1.24 (s, 3H), 1.46–1.55 (m, 1H), 1.62 (s, 3H), 1.69 (s, 3H), 1.71–1.73 (m, 1H), 2.05–2.12 (m, 2H), 3.06 (dd, J = 16.9, 8.3 Hz, 1H), 3.23 (dd, J = 16.9, 5.3 Hz, 1H), 3.77 (br s, 1H), 4.15 (dd, J = 8.2, 5.4 Hz, 1H), 5.10 (t, J = 6.4, 1H), 6.39 (d, J = 9.1 Hz, 1H), 7.08–7.10 (m, 2H); 13C NMR (150 MHz, CDCl3) δ 17.3, 21.1, 21.2, 25.3, 33.7, 39.1, 55.1, 59.3, 108.6, 115.7, 119.2, 123.1, 129.7, 131.2, 131.9, 141.3; HREIMS [M]+ calcd for C16H21N35Cl79Br 341.05459, found 341.05438. HNBr Cl2.115  113  Figure 4.24 1H and 13C NMR spectra of 2.115 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.09557.08496.39326.37815.10735.09635.08604.16504.15604.15134.14233.77423.25273.24393.22453.21573.07863.06483.05043.03662.11592.10492.09602.08772.05111.73121.72671.71941.71501.70861.69301.61811.55371.52431.50801.49861.49021.48151.92691.01311.00010.89910.89821.00641.01382.02401.02493.27442.96121.26013.1572"LZ 02-158-14-30"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Bromo"[ppm] 140  120  100  80  60  40 141.2655131.9413131.2358129.7213123.1333119.2076115.7327108.584059.314555.085839.143633.714925.271021.208221.101817.2759"LZ 02-158-14-30"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Bromo"  114 4.27 Preparation of tetrahydroquinoline 2.108  The procedure for the synthesis of tetrahydroquinoline 2.108 from alcohol 2.106 was the same as the synthesis of tetrahydroquinoline 2.146 from alcohol 2.145 and afforded a pale yellow oil (55%). 1H NMR (600 MHz, CDCl3) δ 1.28 (s, 3H), 1.50–1.55 (m, 1H), 1.62 (s, 3H), 1.69 (s, 3H), 1.77–1.82 (m, 1H), 2.10–2.16 (m, 2H), 3.11 (dd, J = 16.7, 8.7 Hz, 1H), 3.27 (dd, J = 16.8, 5.4 Hz, 1H), 4.22 (dd, J = 8.6, 5.5 Hz, 1H), 5.11 (t, J = 7.0, 1H), 6.61 (br s, 1H), 6.71 (t, J = 7.0 Hz, 1H), 6.99 (d, J = 7.4 Hz, 1H), 7.04 (t, J = 7.5 Hz, 1H); 13C NMR (150 MHz, CDCl3) δ 17.3, 20.9, 21.1, 25.3, 34.1, 39.3, 55.1, 59.9, 114.3, 117.2, 117.4, 123.3, 127.0, 128.8, 131.8, 142.0; HRESIMS [M+H]+ calcd for C16H23NCl 264.1519, found 264.1515. HNCl2.108  115   Figure 4.25 1H and 13C NMR spectra of 2.108 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.04837.03587.02326.99486.98246.72376.71206.70046.60545.12045.10885.09714.23544.22634.22114.21203.28953.28053.26153.25263.13153.11703.10363.08912.15722.13012.11762.10421.82231.80831.79901.79411.78801.78241.77071.68991.62001.55361.54361.53611.52910.93480.88340.84090.84460.95900.91470.98980.98491.96681.16063.04403.26831.66513.3569"LZ 03-186-46-65"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Non halogenated"[ppm] 140  120  100  80  60  40 M 141.9832131.8097128.7507126.9680123.3026117.4354117.2126114.270959.922355.088639.320734.091125.262121.143220.920517.2677"LZ 03-186-46-65"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain/MeMgBr/Non halogenated"  116 4.28 Preparation of nitrile 2.129  The procedure for the synthesis of nitrile 2.129 from ketone 2.96 was the same as the synthesis of nitrile 2.144 from alcohol 2.143 and afforded a white solid (54%). 1H NMR (400 MHz, CDCl3) δ 1.65 (br s, 3H), 1.69 (s, 3H), 2.48 (q, J = 7.4 Hz, 2H), 3.01 (t, J = 7.5 Hz, 2H), 5.17 (t, J = 7.1 Hz, 1H), 7.26 (s, 1H), 7.54 (d, J = 7.7 Hz, 2H), 8.10 (s, 1H), 9.67 (br s, 1H); 13C NMR (150 MHz, CDCl3) δ 17.3, 22.9, 25.3, 38.2, 104.0, 108.7, 112.9, 119.5, 121.8, 126.8, 127.8, 128.6, 133.0, 136.4, 138.0 192.7; HRESIMS [M+Na]+ calcd for C16H16N2ONa 275.1160, found 275.1163. HNNCO2.129  117  Figure 4.26 1H and 13C NMR spectra of 2.129 recorded in CDCl3 at 400 MHz and 150 MHz, respectively. [ppm] 8  6  4  2 9.67058.10007.56237.54807.53817.52377.25995.18445.17275.16093.02273.01042.99782.50132.48922.47692.46491.69090.99260.99222.07081.70030.98552.11392.03533.15843.8004"LZ 02-190-25-41"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"[ppm] 150  100  50 192.7286137.9617136.4496133.0130128.5845127.8360126.8065121.8452119.4945112.9186108.6973103.952838.221825.263222.850817.3107"LZ 02-190-25-41"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"  118 4.29 Preparation of nitrile 2.130  The procedure for the synthesis of nitrile 2.130 from alcohol 2.112 was the same as the synthesis of nitrile 2.144 from alcohol 2.143 and afforded a yellow oil (38%, 50% based on recovered 2.112). 1H NMR (400 MHz, CDCl3) δ 1.24 (s, 3H), 1.39–1.44 (m, 1H), 1.54–1.59 (m, 1H), 1.62 (s 3H), 1.69 (s 3H), 2.04–2.09 (m, 1H), 2.10–2.16 (m, 1H), 3.02 (d, J = 9.3 Hz, 2H), 3.96 (t, J = 9.2 Hz, 1H), 4.47 (br s, 1H), 5.11 (t, J = 6.4 Hz, 1H), 6.56 (d, J = 8.1 Hz, 1H), 7.26 (s, 1H), 7.29 (d, J = 8.0 Hz, 1H); 13C NMR (150 MHz, CDCl3) δ 17.3, 21.7, 24.0, 25.3, 29.5, 36.3, 67.5, 72.8, 99.9, 108.0, 120.1, 123.6, 127.6, 129.1, 131.9, 132.6,154.2; HRESIMS [M+Na]+ calcd for C17H22N2ONa 293.1630, found 293.1635. HNNC H OH2.130  119  Figure 4.27 1H and 13C NMR spectra of 2.130 recorded in CDCl3 at 400 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.30097.28757.25586.56496.55145.12545.11505.10414.47183.97693.96143.94613.03073.01522.16102.14912.12682.12042.11022.09952.08902.07772.06042.05042.03741.68751.62421.59441.58081.57711.57161.56271.55311.54181.43611.42721.41801.01061.29280.98090.95010.78470.99142.08480.97671.16853.08283.07991.29861.24843.1827"LZ 03-72-91-115"  1  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"[ppm] 140  120  100  80  60  40  20 154.1725132.6005131.9100129.0941127.6281123.5677120.1384107.962799.885872.763967.493836.292729.451325.283323.969621.713917.2835"LZ 03-72-91-115"  2  1  "/Users/lingzhizhang/Desktop/Unsaturated Sidechain"  120 4.30 Preparation of ketone 2.140  Pd/C (10% wt, 13 mg, 0.058 mmol) was added to a solution of 2.97 (192 mg, 0.58 mmol) in methanol (3 ml). The reaction mixture was stirred under H2 (50 psi) at room temperature for 1 h, after which the solid catalyst filtered of and washed with EtOAc. The organic phase was washed with H2O (10×3 ml), dried with MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 100% to 4:1) to yield the product 2.140 (175 mg, 91%) as colorless oil. 1H NMR (400 MHz, CDCl3) δ 0.86 (d, J = 6.6 Hz, 6H), 1.11–1.17 (m, 2H), 1.49–1.62 (m, 12H), 2.39–2.51 (m, 2H), 2.93 (dd, J = 16.6, 5.0 Hz, 1H), 3.47 (br t, J = 13.1 Hz, 1H), 4.81 (br, 1H), 6.95 (t, J = 7.4 Hz, 1H), 7.11 (d, J = 7.3 Hz), 7.20 (br t, J = 7.0 Hz), 7.49 (br s, 0.3H), 7.90 (br s, 0.6H); 13C NMR (100 MHz, CDCl3) δ 21.1, 22.6 (×2), 28.0, 28.4 (×3), 32.0, 37.7, 38.6, 66.7, 81.7, 115.0, 122.8, 124.7, 128.1, 129.1, 142.9, 151.9, 208.4; HRESIMS [M+Na]+ calcd for C20H29NO3Na 354.2045, found 354.2041. BocNHO2.140  121  Figure 4.28 1H and 13C NMR spectra of 2.140 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 8  6  4  2 7.90447.4937M 7.22227.20377.18717.11557.09726.96986.95136.93284.87684.81073.50663.47013.44092.96082.94842.91932.90692.51252.48812.46942.45192.38621.61951.60281.59411.58591.58121.56241.54721.54461.53151.51471.49781.49101.16721.14991.12731.11010.57990.29241.01931.00761.00080.99141.01361.00802.014012.43902.13276.3547"LZ 03-98-25-27"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain"[ppm] 200  150  100  50 208.3984151.8545142.8941129.0857128.1341124.6552122.8335114.973381.714666.666638.618437.663631.962328.425727.988422.598222.577221.1428"LZ 03-98-25-27"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain"  122 4.31 Preparation of ketone 2.141  NBS (103 mg, 0.58 mmol) was added to a solution of 2.140 (191 mg, 0.58 mmol ) in DMF (3ml) at 0 °C. The mixture was stirred for 2 h at 0 °C. After addition of water (10 ml), the aqueous solution was extracted with EtOAc (10×3 ml). The combined organic extracts were dried over MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 9:1 to 7:3) to give the product 2.141 (228 mg, 96%) as a yellow oil. 1H NMR (600 MHz, CDCl3) δ 0.86 (d, J = 6.7 Hz, 6H), 1.12–1.16 (m, 2H), 1.47 (br s, 6H), 1.50–1.59 (m, 6H), 2.33–2.47 (m, 2H), 2.91 (dd, J = 16.8, 4.7 Hz, 1H), 3.46 (br t, J = 14.0 Hz, 1H), 4.80 (br s, 1H), 7.21 (s, 1H), 7.31 (d, J = 6.24 Hz, 1H), 7.78 (d, J = 7.38 Hz, 1H); 13C NMR (150 MHz, CDCl3) δ 20.6, 22.0 (×2), 27.4, 27.8 (×3), 30.9, 37.3, 38.0, 66.0, 81.5, 114.6, 115.7, 127.1, 127.4,130.5, 141.6, 151.1, 207.1; HRESIMS [M+Na]+ calcd for C20H28NO3NaBr 432.1150, found 432.1141. BocNHOBr2.141  123  Figure 4.29 1H and 13C NMR spectra of 2.141 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 6  4  2 7.78957.77727.31827.30787.21494.89134.87634.81234.79963.48063.45493.43402.92652.91862.89852.89062.46912.45782.36042.33261.59241.58431.58091.56791.56401.51821.50701.49511.47451.16001.14801.13381.12210.86540.85430.66911.20160.95340.98991.02861.00002.02806.41616.04622.13666.1603"LZ 03-104-41-44"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"[ppm] 200  150  100  50 207.1033151.0991141.6151130.4705127.4475127.1145115.7242114.606381.502266.017138.037837.342530.948527.784727.422422.022720.5591"LZ 03-104-41-44"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"  124 4.32 Preparation of alcohol 2.142  To a solution of 2.41 (292 mg, 0.71 mmol) in dry Et2O (10 ml) was added dropwise a solution of MeMgBr (1.2 ml, 3M) in dry Et2O at –40 °C. After stirring at –40 °C for 2 h, the reaction was quenched with saturated NH4Cl solution (15 ml) and extracted with EtOAc (15×3 ml). The combined organic extracts were dried with MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 100% to 7:3) to afford the product 2.142 (279 mg, 92%) as a colorless oil. 1H NMR (400 MHz, CDCl3) δ 0.83 (d, J = 6.6 Hz, 6H), 1.03–1.10 (m, 2H), 1.16  (s, 3H), 1.21–1.51 (m, 5H), 1.57 (s, 9H), 2.91 (d, J = 16.6 Hz, 1H), 3.28 (dd, J = 16.7, 10.3 Hz, 1H), 4.51 (d, J = 8.7 Hz, 1H), 7.23–7.33 (m, 3H); 13C NMR (100 MHz, CDCl3) 21.2, 22.7, 22.8, 24.0, 28.2, 28.5 (×3), 30.7, 36.6, 39.8, 68.1, 76.3, 82.7, 115.7, 117.9, 127.3, 130.1, 134.3, 142.2, 154.8; HRESIMS [M+Na]+ calcd for C21H32NO3NaBr 448.1463, found 448.1469. BocNH OHBr2.142  125  Figure 4.30 1H and 13C NMR spectra of 2.142 recorded in CDCl3 at 400 MHz and 100 MHz, respectively. [ppm] 7  6  5  4  3  2  1 7.33537.25987.23977.23274.52464.50283.31543.28973.27363.24792.93342.89181.56601.51391.49711.48051.46391.40581.39131.32341.31671.28931.27331.26181.25281.23551.21041.16021.09591.08851.07360.83090.87201.32340.97161.04931.08428.84375.06682.96421.99192.90523.0790"LZ 03-106-87-95"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"[ppm] 140  120  100  80  60  40 M 154.7623M 142.2246134.3398130.0854127.2939117.9216115.739382.683976.273768.116239.7979M 36.642630.654928.459228.227323.986722.760222.696221.2235"LZ 03-106-87-95"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"  126 4.33 Preparation of alcohol 2.143  To a solution of 2.142 (233 mg, 0.57 mmol) in dry CH2Cl2 (5 ml) was added formic acid (10 ml) at room temperature. After stirring for 18 h at room temperature, the reaction mixture was neutralized with saturated NaHCO3 solution and extracted with EtOAc (10×3 ml). The combined organic extracts were dried with MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 9:1 to 3:2) to afford the product 2.143 (107 mg, 58%) as a white solid.  1H NMR (600 MHz, CDCl3) δ 0.89 (d, J = 6.8 Hz, 6H), 1.18–1.20 (m, 2H), 1.22 (s, 3H), 1.29–1.45 (m, 3H), 1.48–1.59 (m, 2H), 2.19 (br, 1H), 2.92 (dd, J = 16.0, 9.2 Hz, 1H), 3.05 (dd, J = 15.9, 10.4 Hz, 1H), 3.88 (t, J = 9.8 Hz, 1H), 6.52 (d, J = 8.3 Hz, 1H), 7.10 (d, J = 8.2 Hz, 1H), 7.16 (s, 1H); 13C NMR (150 MHz, CDCl3) δ 20.9, 22.1, 22.3, 24.6, 27.5, 29.9, 37.5, 39.2, 67.9, 71.9, 110.7 (×2), 127.2, 129.4, 131.5, 149.0; HRESIMS [M+Na]+ calcd for C16H24NONa79Br 348.0939, found 348.0937. HNH OHBr2.143  127  Figure 4.31 1H and 13C NMR spectra of 2.143 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.16227.10807.09436.52786.51403.89773.88123.86523.06943.05213.04293.02562.94712.93172.92052.90512.19281.59241.58151.57051.55941.54831.53721.52501.50891.50391.48781.48361.45431.44091.43191.42241.41581.40681.39861.38491.36921.36331.34391.32601.31931.31591.30050.92890.94810.96791.15091.05041.01440.79182.13593.27013.23121.87706.0787"LZ 03-114-65-70"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"[ppm] 140  120  100  80  60  40  20 148.9972131.5357129.4306127.2174110.7335110.660971.905167.896639.178937.509329.857227.504624.626922.256422.128520.9298"LZ 03-114-65-70"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"  128 4.34 Preparation of nitrile 2.144  CuCN (9.0 mg, 0.099 mmol) and NMP (1.8 ml) was added to 2.143 (25 mg, 0.076 mmol) and the mixture was stirring at 200 °C in microwave reactor for 25 min. When the reaction mixture was cooled to room temperature, iced water was added to the mixture and white precipitate was observed. The ammonia solution was used to rinse the mixture to give blue cuprous complex aqueous solution, which was extracted with EtOAc (10 ×4 ml). The combined organic extracts were dried with MgSO4, filtered and concentrated. The residual NMP in crude mixture was removed by vacuum pump for 1 day. The crude mixture was purified by column chromatography (hexane–EtOAc = 9:1 to 1:1) to give the product 2.144 (8.2 mg, 40%) as a yellow oil. Based on recovered starting material 2.143 (8 mg, 0.025 mmol), the yield of 2.144 was 58%.   1H NMR (600 MHz, CDCl3) δ 0.88 (d, J = 6.7 Hz, 6H), 1.16–1.20 (m, 2H), 1.24 (s, 3H), 1.29–1.43 (m, 3H), 1.48–1.58 (m, 2H), 3.01 (dd, J = 16.3, 9.7 Hz, 1H), 3.06 (dd, J = 16.3, 9.4 Hz, 1H), 3.98 (t, J = 9.5 Hz, 1H), 6.61 (d, J = 8.1 Hz, 1H), 7.27 (s, 1H), 7.31 (d, J = 8.16 Hz, 1H); 13C NMR (150 MHz, CDCl3) δ 20.8, 22.1, 22.2, 24.2, 27.5, 29.4, 37.0, 39.1, 67.6, 72.5, 100.5, 108.5, 120.0, 127.7, 129.5, 132.6, 153.7; HRESIMS [M+Na]+ calcd for C17H24N2ONa 295.1786, found 295.1784. HNNC H OH2.144  129  Figure 4.32 1H and 13C NMR spectra of 2.144 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2  1 7.31577.30217.27426.61506.60153.99103.97523.95943.07883.06303.05163.03603.01993.00902.99271.58341.57231.56131.55021.53911.52761.50611.50101.48501.47551.42721.40991.40111.39231.38051.36521.35921.34191.33321.32661.32211.30711.29491.23861.20411.19921.18861.18361.17860.92010.86860.96770.99030.94431.02502.09043.38903.24582.00786.2588"LZ 03-128-110-125"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"[ppm] 160  140  120  100  80  60  40  20 153.7150132.5517129.4700127.6854120.0116108.5099100.484772.465567.615739.094737.004329.381327.500724.219322.218722.098320.7774"LZ 03-128-110-125"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"  130 4.35 Preparation of alcohol 2.145  TMSCl (0.8 ml, 62.3 mmol) was added to a solution of the nitrile 2.144 (35 mg, 0.13 mmol) in dry methanol (0.75 ml) under an argon atmosphere at room temperature. The reaction mixture was heated at 40 °C for 5 h. When the reaction was cooled to room temperature, water (0.2 ml) was added to the mixture followed by the addition of NaHCO3 until the aqueous layer was neutral and the mixture stopped bubbling. Then the reaction mixture was dried with MgSO4, filtered and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 4:1 to 1:1) to give the product 2.145 (22 mg, 57%) as a yellow oil. 1H NMR (600 MHz, CDCl3) δ 0.88 (d, J = 6.7 Hz, 6H), 1.16–1.21 (m, 2H), 1.26 (s, 3H), 1.30–1.44 (m, 3H), 1.50–1.58 (m, 2H), 3.01 (dd, J = 14.9, 9.5 Hz, 1H), 3.08 (dd, J = 16.0, 9.7 Hz, 1H), 3.85 (s, 3H), 3.98 (t, J = 9.5 Hz, 1H), 6.66 (d, J = 8.2 Hz, 1H), 7.74 (s, 1H), 7.78 (d, J = 8.2 Hz, 1H); 13C NMR (150 MHz, CDCl3) δ 20.8, 22.1, 22.2, 24.4, 27.5, 29.4, 37.2, 39.1, 51.3, 68.0, 72.3, 108.5, 120.9, 125.8, 128.9, 130.1, 153.75, 166.8; HRESIMS [M+Na]+ calcd for C18H27NO3Na 328.1889, found 328.1894. HNMeO2C HOH2.145  131  Figure 4.33 1H and 13C NMR spectra of 2.145 recorded in CDCl3 at 600 MHz and 150 MHz, respectively.  [ppm] 7  6  5  4  3  2  1 7.78267.76897.74586.66666.65303.99553.97973.96383.85203.10003.08383.07333.05733.03203.01633.00562.98971.57701.56591.55491.54361.52401.51931.50301.49931.43671.39911.39371.35941.33191.32771.31401.29971.26111.20971.19531.19211.18431.01750.91130.97731.05132.99632.07122.13382.15281.29883.33622.17976.0628"LZ 03-134-50-72"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"[ppm] 160  140  120  100  80  60  40 166.8120153.4939130.0848128.9455125.8263120.8833108.460572.304468.011451.269839.116737.221129.405927.503024.384322.231022.100320.8378"LZ 03-134-50-72"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"  132 4.36 Preparation of tetrahydroquinoline 2.146  CCl4 (0.11 ml, 1.18 mmol) was added to a solution of 2.145 (36 mg, 0.12 mmol) in dry CH2Cl2 (1 ml). The reaction was heated to 40 °C followed by the addition of PPh3 (93 mg, 0.35 mmol). The reaction was stirred under reflux conditions for 30 min, then cooled to room temperature and concentrated. The crude reaction mixture was purified by column chromatography (hexane–EtOAc = 100% to 4:1) to give the product 2.146 (20 mg, 53%) as a pale yellow oil.  1H NMR (400 MHz, CDCl3) δ 0.88 (d, J = 6.5 Hz, 6H), 1.15–1.21 (m, 2H), 1.29 (s, 3H), 1.35–1.43 (m, 2H), 1.45–1.50 (m, 1H), 1.52–1.58 (m, 1H), 1.68–1.75 (m, 1H), 3.11 (dd, J = 16.8, 8.3 Hz, 1H), 3.29 (dd, J = 16.8, 5.1 Hz, 1H), 3.85 (s, 3H), 4.20 (dd, J = 8.2, 5.2 Hz, 1H), 6.54 (d, J = 7.8 Hz, 1H), 7.70–7.72 (m, 2H); 13C NMR (150 MHz, CDCl3) δ 20.1, 21.7, 22.1, 22.2, 27.4, 33.7, 38.7, 39.9, 51.2, 55.5, 59.1, 113.0, 116.1, 118.3, 129.1, 131.0, 146.3, 166.8; HRESIMS [M+H]+ calcd for C18H27NO2Cl 324.1730, found 324.1733. HNClMeO2C2.146  133  Figure 4.34 1H and 13C NMR spectra of 2.146 recorded in CDCl3 at 400 MHz and 150 MHz, respectively. [ppm] 8  6  4  2 7.71917.70116.54506.52564.21414.20104.19354.18053.84663.31263.29973.27063.25783.14383.12293.10183.08101.75161.73731.71751.71011.69661.67601.58171.56711.55051.53381.51831.50071.49451.47891.45621.44541.43101.40721.38821.36871.35091.28531.21181.20391.19521.86220.89980.98803.05511.01081.03950.98781.34081.06342.13172.92792.21206.5267"LZ 03-148-38-41"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"[ppm] 160  140  120  100  80  60  40 166.7597146.2698131.0143129.1151118.2710116.1229113.040759.093155.524251.177639.921138.739033.652027.430322.205922.060921.657820.0899"LZ 03-148-38-41-600"  4  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Bromo"  134 4.37 Preparation of alcohol 2.147  The procedure for the synthesis of alcohol 2.147 from ketone 2.140 was the same as the synthesis of alcohol 2.142 from ketone 2.141 and afforded a white solid (100%). 1H NMR (600 MHz, CDCl3) δ 0.81–0.82 (m, 6H), 1.03–1.08 (m, 2H), 1.17  (s, 3H), 1.21–1.34 (m, 3H), 1.38–1.50 (m, 2H), 1.58 (s, 9H), 2.91 (d, J = 16.4 Hz, 1H), 3.31 (dd, J = 16.5, 10.4 Hz, 1H), 4.53 (d, J = 9.8 Hz, 1H), 6.96 (t, J = 7.4 Hz, 1H), 7.12 (d, J = 7.4 Hz, 1H), 7.15 (t, J = 7.8 Hz, 1H), 7.47 (br s, 1H); 13C NMR (150 MHz, CDCl3) δ 20.7, 22.1, 22.2, 23.6, 27.7, 27.9 (×3), 30.4, 35.9, 39.3, 67.5, 75.8, 81.8, 116.0, 122.7, 123.7, 126.6, 131.3, 142.4, 154.8; HRESIMS [M+Na]+ calcd for C21H33NO3Na 370.2358, found 370.2360. BocNH OH2.147  135  Figure 4.35 1H and 13C NMR spectra of 2.147 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2  1 7.47367.16257.14967.13657.12387.11156.97076.95846.94594.54134.52503.33313.31583.30563.28842.92512.89781.58321.49641.48571.47451.46381.45271.44171.42761.41951.41081.40121.39471.38891.38291.37511.34071.32021.29611.28101.27121.26171.25391.24491.21790.89391.02980.98331.02331.00481.05690.99789.14382.07483.00083.05221.95046.0327"LZ 02-188"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Non halogenated"[ppm] 140  120  100  80  60  40 154.7843142.3856131.2721126.6303123.7105122.6631116.036081.806275.794767.549739.270935.899430.416227.938127.672823.579422.218822.146720.6567"LZ 02-188"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Non halogenated"  136 4.38 Preparation of alcohol 2.148  The procedure for the synthesis of alcohol 2.148 from 2.147 was the same as the synthesis of alcohol 2.143 from alcohol 2.142 and afforded a white solid (86%). 1H NMR (600 MHz, CDCl3) δ 0.90 (d, J = 6.8 Hz, 6H), 1.16–1.21 (m, 2H), 1.24 (s, 3H), 1.31–1.47 (m, 3H), 1.52–1.61 (m, 2H), 2.95 (dd, J = 15.7, 9.2 Hz, 1H), 3.06 (dd, J = 15.6, 10.5 Hz, 1H), 3.88 (t, J = 9.8 Hz, 1H), 6.67 (d, J = 7.7 Hz, 1H), 6.74 (t, J = 7.4 Hz, 1H), 7.03 (t, J = 7.6 Hz, 1H), 7.09 (d, J = 7.3 Hz, 1H); 13C NMR (150 MHz, CDCl3) δ 21.0, 22.2, 22.3, 24.8, 27.5, 29.9, 37.7, 39.2, 67.6, 71.8, 109.4, 118.9, 124.3, 126.7, 129.1, 150.1; HRESIMS [M+H]+ calcd for C16H26NO 248.2014, found 248.2016. HNH OH2.148  137  Figure 4.36 1H and 13C NMR spectra of 2.148 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2  1 7.09597.08377.03947.02677.01406.75086.73856.72626.67176.65883.89473.87793.86203.08643.06883.06033.04282.96852.95322.94232.92712.30631.60771.59661.57451.56331.55231.54461.54011.52391.51941.46971.46121.45151.44491.44221.43511.42921.42041.40871.40541.39981.38011.35921.35271.34791.34221.33341.32631.32311.32081.31040.97500.94990.95830.94941.26051.00141.03280.79992.11313.12393.16942.01516.0407"LZ 02-198-35-55"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Non halogenated"[ppm] 140  120  100  80  60  40 150.1410129.0568126.7483124.2864118.9367109.383971.796967.609439.244537.683329.926427.517624.828022.275122.150521.0034"LZ 02-198-35-55"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Non halogenated"  138 4.39 Preparation of tetrahydroquinoline 2.151  The procedure for the synthesis of tetrahydroquinoline 2.151 from alcohol 2.149 was the same as the synthesis of tetrahydroquinoline 2.146 from alcohol 2.145 and afforded a pale yellow oil (53%). 1H NMR (600 MHz, CDCl3) δ 0.87 (d, J = 6.6 Hz, 6H), 1.12–1.21 (m, 2H), 1.25 (s, 3H), 1.34–1.41 (m, 2H), 1.43–1.48 (m, 1H), 1.51–1.57 (m, 1H), 1.64–1.70 (m, 1H), 3.06 (dd, J = 17.0, 8.2 Hz, 1H), 3.23 (dd, J = 17.0, 5.3 Hz, 1H), 4.17 (dd, J = 7.9, 5.5 Hz, 1H), 6.47 (br, 1H), 7.10–7.11 (m, 2H); 13C NMR (150 MHz, CDCl3) δ 20.1, 21.1, 22.1, 22.2, 27.4, 33.7, 38.8, 39.4, 55.6, 59.2, 109.4, 116.3, 119.8, 129.8, 131.3, 143.0; HREIMS [M]+ calcd for C16H23N35Cl79Br 343.07024, found 343.07026. HNClBr2.151  139  Figure 4.37 1H and 13C NMR spectra of 2.151 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2 7.11487.10166.47464.17834.16914.16514.15593.24963.24083.22133.21253.08073.06713.05253.03881.70341.69531.68081.67571.66361.65331.63751.57411.56761.55631.54521.53411.52311.51191.47841.46981.45851.45061.43151.40921.39751.38761.37691.36471.35641.34431.24961.20771.19601.83440.88201.07411.00111.06481.21061.49341.36542.30423.58292.35166.5238"LZ 03-178-1-5"  1  1  "/Users/lingzhizhang/Desktop/NMR/AV 600/nmr"[ppm] 140  120  100  80  60  40  20 M 142.9680131.3067129.8077M 119.8153116.2789M 109.396659.1550M 55.573439.364938.769933.669827.434922.210222.076621.080620.0850"LZ 03-178-1-5"  2  1  "/Users/lingzhizhang/Desktop/NMR/AV 600/nmr"  140 4.40 Preparation of tetrahydroquinoline 2.152  The procedure for the synthesis of tetrahydroquinoline 2.152 from alcohol 2.150 was the same as the synthesis of tetrahydroquinoline 2.146 from alcohol 2.145 and afforded a pale yellow oil (69%). 1H NMR (600 MHz, CDCl3) δ 0.88 (d, J = 6.5 Hz, 6H), 1.14–1.20 (m, 2H), 1.23 (s, 3H), 1.32–1.48 (m, 3H), 1.54–1.58 (m, 1H), 1.67–1.71 (m, 1H), 3.10 (dd, J = 16.7, 9.1 Hz, 1H), 3.25 (dd, J = 16.6, 5.3 Hz, 1H), 3.73 (br s, 1H), 4.20 (dd, J = 8.9, 5.5 Hz, 1H), 6.52 (d, J = 8.0 Hz, 1H), 6.66 (t, J = 7.3 Hz, 1H), 6.97 (d, J = 7.5 Hz, 1H), 7.01 (t, J = 7.5 Hz, 1H); 13C NMR (150 MHz, CDCl3) δ 20.0, 21.0, 22.1, 22.3, 27.5, 34.1, 38.9, 39.9, 55.0, 60.0, 114.1, 117.1, 117.4, 127.0, 128.8, 142.3; HREIMS [M]+ calcd for C16H24N35Cl 265.15973, found 265.15955. HNCl2.152  141  Figure 4.38 1H and 13C NMR spectra of 2.152 recorded in CDCl3 at 600 MHz and 150 MHz, respectively. [ppm] 7  6  5  4  3  2  1 7.02597.01347.00086.97786.96536.67416.66196.64966.52706.51374.21114.20184.19614.18703.72723.26483.25593.23723.22833.11863.10363.09093.07571.71711.70961.69541.68861.67511.66681.57851.56751.55651.54541.53541.48031.47391.45931.45301.43331.42281.41201.40241.39331.38611.38041.37571.36491.35601.34321.33991.32871.31741.22851.19701.19260.99521.00000.91760.93830.91790.84561.03251.03971.17251.68323.62483.22042.56876.9970"LZ 03-176-64-80"  1  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Non halogenated"[ppm] 140  120  100  80  60  40 142.3350128.7563126.9550117.3818117.0602114.136060.049655.003139.937238.859234.144127.482722.273322.098121.020820.0017"LZ 02-200"  2  1  "/Users/lingzhizhang/Desktop/Saturated Chain/Non halogenated"  142 References 1.	   Cragg,	   G.	   M.;	   Newman,	   D.	   J.,	   Natural	   products:	   a	   continuing	   source	   of	   novel	   drug	  leads.	  Biochimica	  et	  biophysica	  acta	  2013,	  1830	  (6),	  3670–3695.	  2.	   Schmitz,	  R.,	  Friedrich	  Wilhelm	  Serturner	  and	  the	  discovery	  of	  morphine.	  Pharmacy	  in	  history	  1985,	  27	  (2),	  61–74.	  3.	   Cechinel-­‐Filho,	   V.,	   Plant	   bioactives	   and	   drug	   discovery:	   principles,	   practice,	   and	  perspectives.	  2012,	  2–4.	  4.	   Newman,	   D.	   J.;	   Cragg,	   G.	   M.,	   Marine	   natural	   products	   and	   related	   compounds	   in	  clinical	   and	   advanced	   preclinical	   trials.	   Journal	   of	   natural	   products	  2004,	   67	   (8),	   1216–1238.	  5.	   Imhoff,	   J.	   F.;	   Labes,	   A.;	  Wiese,	   J.,	   Bio-­‐mining	   the	  microbial	   treasures	   of	   the	   ocean:	  new	  natural	  products.	  Biotechnology	  advances	  2011,	  29	  (5),	  468–482.	  6.	   Koehn,	  F.	  E.;	  Carter,	  G.	  T.,	  The	  evolving	  role	  of	  natural	  products	   in	  drug	  discovery.	  Nature	  reviews.	  Drug	  discovery	  2005,	  4	  (3),	  206–220.	  7.	   Butler,	   M.	   S.,	   The	   role	   of	   natural	   product	   chemistry	   in	   drug	   discovery.	   Journal	   of	  natural	  products	  2004,	  67	  (12),	  2141–2153.	  8.	   Molinski,	   T.	   F.;	   Dalisay,	   D.	   S.;	   Lievens,	   S.	   L.;	   Saludes,	   J.	   P.,	   Drug	   development	   from	  marine	  natural	  products.	  Nature	  reviews.	  Drug	  discovery	  2009,	  8	  (1),	  69–85.	  9.	   Bergmann,	  W.;	   Feeney,	  R.	   J.,	   Contributions	   to	   the	   study	  of	  marine	  products.	  XXXII.	  The	  nucleosides	  of	  sponges.	  I.	  Journal	  of	  organic	  chemistry	  1951,	  16,	  981–987.	  10.	   Bergmann,	  W.;	  Burke,	  D.	  C.,	  Contributions	  to	  the	  study	  of	  marine	  products.	  XL.	  The	  nucleosides	  of	  sponges.	  IV.	  Spongosine.	  Journal	  of	  organic	  chemistry	  1956,	  22,	  226–228.	    143 11.	   Bergmann,	  W.;	  stempien,	  M.	  F.,	  Contributions	  to	  the	  study	  of	  marine	  products.	  XLIII.	  The	   nucleosides	   of	   sponges.	   V.	   The	   synthesis	   of	   spongosine.	   Journal	  of	  organic	   chemistry	  1957,	  2,	  1557–1575.	  12.	   Zhang,	  L.;	  An,	  R.;	  Wang,	  J.;	  Sun,	  N.;	  Zhang,	  S.;	  Hu,	  J.;	  Kuai,	  J.,	  Exploring	  novel	  bioactive	  compounds	  from	  marine	  microbes.	  Current	  opinion	  in	  microbiology	  2005,	  8	  (3),	  276–281.	  13.	   Newman,	   D.	   J.;	   Cragg,	   G.	   M.,	   Advanced	   preclinical	   and	   clinical	   trials	   of	   natural	  products	  and	  related	  compounds	  from	  marine	  sources.	  Current	  medicinal	  chemistry	  2004,	  11	  (13),	  1693–1713.	  14.	   Miljanich,	   G.	   P.,	   Ziconotide:	   neuronal	   calcium	   channel	   blocker	   for	   treating	   severe	  chronic	  pain.	  Current	  medicinal	  chemistry	  2004,	  11	  (23),	  3029–3040.	  15.	   Bowersox,	   S.	   S.;	   Luther,	   R.,	   Pharmacotherapeutic	   potential	   of	   omega-­‐conotoxin	  MVIIA	   (SNX-­‐111),	   an	   N-­‐type	   neuronal	   calcium	   channel	   blocker	   found	   in	   the	   venom	   of	  Conus	  magus.	  Toxicon	  :	  official	  journal	  of	  the	  International	  Society	  on	  Toxinology	  1998,	  36	  (11),	  1651–1658.	  16.	   Hamman,	  J.	  H.;	  Enslin,	  G.	  M.;	  Kotze,	  A.	  F.,	  Oral	  delivery	  of	  peptide	  drugs:	  barriers	  and	  developments.	   BioDrugs	   :	   clinical	   immunotherapeutics,	   biopharmaceuticals	   and	   gene	  therapy	  2005,	  19	  (3),	  165–177.	  17.	   Brady,	  R.	  M.;	  Baell,	  J.	  B.;	  Norton,	  R.	  S.,	  Strategies	  for	  the	  development	  of	  conotoxins	  as	  new	  therapeutic	  leads.	  Marine	  drugs	  2013,	  11	  (7),	  2293–2313.	  18.	   Adessi,	  C.;	  Soto,	  C.,	  Converting	  a	  peptide	  into	  a	  drug:	  strategies	  to	  improve	  stability	  and	  bioavailability.	  Current	  medicinal	  chemistry	  2002,	  9	  (9),	  963–978.	    144 19.	   Clark,	  R.	   J.;	  Akcan,	  M.;	  Kaas,	  Q.;	  Daly,	  N.	  L.;	  Craik,	  D.	   J.,	  Cyclization	  of	  conotoxins	   to	  improve	   their	   biopharmaceutical	   properties.	  Toxicon	   :	   official	   journal	  of	   the	   International	  Society	  on	  Toxinology	  2012,	  59	  (4),	  446–455.	  20.	   Hirata,	   Y.;	   Uemura,	   D.,	   Halichondrins	   -­‐	   Antitumor	   Polyether	   Macrolides	   from	   a	  Marine	  Sponge.	  Pure	  applied	  chemistry	  1986,	  58	  (5),	  701–710.	  21.	   Uemura,	   D.;	   Takahashi,	   K.;	   Yamamoto,	   T.;	   Katayama,	   C.;	   Tanaka,	   J.;	   Okumura,	   Y.;	  Hirata,	   Y.,	   Norhalichondrin-­‐a	   -­‐	   an	  Antitumor	   Polyether	  Macrolide	   from	   a	  Marine	   Sponge.	  Journal	  of	  the	  American	  chemical	  society	  1985,	  107	  (16),	  4796–4798.	  22.	   Aicher,	  T.	  D.;	  Buszek,	  K.	  R.;	  Fang,	  F.	  G.;	  Forsyth,	  C.	  J.;	  Jung,	  S.	  H.;	  Kishi,	  Y.;	  Matelich,	  M.	  C.;	   Scola,	   P.	   M.;	   Spero,	   D.	   M.;	   Yoon,	   S.	   K.,	   Total	   Synthesis	   of	   Halichondrin-­‐B	   and	  Norhalichondrin-­‐B.	  Journal	  of	  the	  American	  chemical	  society	  1992,	  114	  (8),	  3162–3164.	  23.	   Towle,	  M.	  J.;	  Salvato,	  K.	  A.;	  Budrow,	  J.;	  Wels,	  B.	  F.;	  Kuznetsov,	  G.;	  Aalfs,	  K.	  K.;	  Welsh,	  S.;	   Zheng,	  W.	   J.;	   Seletsky,	   B.	  M.;	   Palme,	  M.	  H.;	  Habgood,	   G.	   J.;	   Singer,	   L.	   A.;	   DiPietro,	   L.	   V.;	  Wang,	  Y.;	  Chen,	  J.	  J.;	  Quincy,	  D.	  A.;	  Davis,	  A.;	  Yoshimatsu,	  K.;	  Kishi,	  Y.;	  Yu,	  M.	  J.;	  Littlefield,	  B.	  A.,	   In	   vitro	   and	   in	   vivo	   anticancer	   activities	  of	   synthetic	  macrocyclic	   ketone	  analogues	  of	  halichondrin	  B.	  Cancer	  Research	  2001,	  61	  (3),	  1013–1021.	  24.	   Cortes,	   J.;	   O'Shaughnessy,	   J.;	   Loesch,	   D.;	   Blum,	   J.	   L.;	   Vahdat,	   L.	   T.;	   Petrakova,	   K.;	  Chollet,	   P.;	   Manikas,	   A.;	   Dieras,	   V.;	   Delozier,	   T.;	   Vladimirov,	   V.;	   Cardoso,	   F.;	   Koh,	   H.;	  Bougnoux,	   P.;	   Dutcus,	   C.	   E.;	   Seegobin,	   S.;	   Mir,	   D.;	   Meneses,	   N.;	   Wanders,	   J.;	   Twelves,	   C.;	  Breast,	   E.	   E.	  M.,	   Eribulin	  monotherapy	   versus	   treatment	   of	   physician's	   choice	   in	   patients	  with	  metastatic	  breast	  cancer	  (EMBRACE):	  a	  phase	  3	  open-­‐label	  randomised	  study.	  Lancet	  2011,	  377	  (9769),	  914–923.	    145 25.	   Mayer,	  A.	  M.	  S.;	  Glaser,	  K.	  B.;	  Cuevas,	  C.;	  Jacobs,	  R.	  S.;	  Kem,	  W.;	  Little,	  R.	  D.;	  McIntosh,	  J.	  M.;	  Newman,	  D.	   J.;	  Potts,	  B.	  C.;	  Shuster,	  D.	  E.,	  The	  odyssey	  of	  marine	  pharmaceuticals:	  a	  current	  pipeline	  perspective.	  Trends	  pharmacol	  sciences	  2010,	  31	  (6),	  255–265.	  26.	   Mayer,	   A.	   M.;	   Rodriguez,	   A.	   D.;	   Taglialatela-­‐Scafati,	   O.;	   Fusetani,	   N.,	   Marine	  pharmacology	   in	   2009-­‐2011:	   marine	   compounds	   with	   antibacterial,	   antidiabetic,	  antifungal,	   anti-­‐inflammatory,	   antiprotozoal,	   antituberculosis,	   and	   antiviral	   activities;	  affecting	  the	  immune	  and	  nervous	  systems,	  and	  other	  miscellaneous	  mechanisms	  of	  action.	  Marine	  drugs	  2013,	  11	  (7),	  2510–2573.	  27.	   Bajorath,	   J.,	   Integration	   of	   virtual	   and	   high-­‐throughput	   screening.	  Nature	   reviews.	  Drug	  discovery	  2002,	  1	  (11),	  882–894.	  28.	   Thorburn,	   A.;	   Thamm,	   D.	   H.;	   Gustafson,	   D.	   L.,	   Autophagy	   and	   cancer	   therapy.	  Molecular	  pharmacology	  2014,	  85	  (6),	  830–838.	  29.	   Donohue,	   E.;	   Balgi,	   A.	   D.;	   Komatsu,	   M.;	   Roberge,	   M.,	   Induction	   of	   Covalently	  Crosslinked	   p62	   Oligomers	   with	   Reduced	   Binding	   to	   Polyubiquitinated	   Proteins	   by	   the	  Autophagy	  Inhibitor	  Verteporfin.	  Plos	  one	  2014,	  9	  (12),	  e114964.	  30.	   Jiang,	  P.;	  Mizushima,	  N.,	  Autophagy	  and	  human	  diseases.	  Cell	  research	  2014,	  24	  (1),	  69–79.	  31.	   Ravikumar,	   B.;	   Sarkar,	   S.;	   Davies,	   J.	   E.;	   Futter,	   M.;	   Garcia-­‐Arencibia,	   M.;	   Green-­‐Thompson,	  Z.	  W.;	  Jimenez-­‐Sanchez,	  M.;	  Korolchuk,	  V.	  I.;	  Lichtenberg,	  M.;	  Luo,	  S.;	  Massey,	  D.	  C.;	  Menzies,	   F.	  M.;	  Moreau,	  K.;	  Narayanan,	  U.;	  Renna,	  M.;	   Siddiqi,	   F.	  H.;	  Underwood,	  B.	  R.;	  Winslow,	  A.	  R.;	  Rubinsztein,	  D.	  C.,	  Regulation	  of	  mammalian	  autophagy	   in	  physiology	  and	  pathophysiology.	  Physiological	  reviews	  2010,	  90	  (4),	  1383–1435.	    146 32.	   Kondo,	   Y.;	   Kanzawa,	   T.;	   Sawaya,	   R.;	   Kondo,	   S.,	   The	   role	   of	   autophagy	   in	   cancer	  development	  and	  response	  to	  therapy.	  Nature	  reviews	  cancer	  2005,	  5	  (9),	  726–734.	  33.	   Yang,	   Z.;	   Klionsky,	   D.	   J.,	   Mammalian	   autophagy:	   core	   molecular	   machinery	   and	  signaling	  regulation.	  Current	  opinion	  in	  	  cell	  biology	  2010,	  22	  (2),	  124–131.	  34.	   Rabinowitz,	   J.	  D.;	  White,	  E.,	  Autophagy	  and	  metabolism.	  Science	  2010,	  330	   (6009),	  1344–1348.	  35.	   Russell,	  R.	  C.;	  Yuan,	  H.	  X.;	  Guan,	  K.	  L.,	  Autophagy	  regulation	  by	  nutrient	  signaling.	  Cell	  research	  2014,	  24	  (1),	  42–57.	  36.	   Jones,	  S.	  A.;	  Mills,	  K.	  H.;	  Harris,	  J.,	  Autophagy	  and	  inflammatory	  diseases.	  Immunology	  and	  cell	  biology	  2013,	  91	  (3),	  250–258.	  37.	   Carew,	   J.	   S.;	  Kelly,	  K.	  R.;	  Nawrocki,	  S.	  T.,	  Autophagy	  as	  a	   target	   for	  cancer	   therapy:	  new	  developments.	  Cancer	  management	  and	  research	  2012,	  4,	  357–365.	  38.	   Liang,	  X.	  H.;	  Jackson,	  S.;	  Seaman,	  M.;	  Brown,	  K.;	  Kempkes,	  B.;	  Hibshoosh,	  H.;	  Levine,	  B.,	   Induction	  of	  autophagy	  and	   inhibition	  of	   tumorigenesis	  by	  beclin	  1.	  Nature	  1999,	  402	  (6762),	  672–676.	  39.	   Aita,	  V.	  M.;	  Liang,	  X.	  H.;	  Murty,	  V.	  V.;	  Pincus,	  D.	  L.;	  Yu,	  W.;	  Cayanis,	  E.;	  Kalachikov,	  S.;	  Gilliam,	  T.	  C.;	  Levine,	  B.,	  Cloning	  and	  genomic	  organization	  of	  beclin	  1,	  a	  candidate	  tumor	  suppressor	  gene	  on	  chromosome	  17q21.	  Genomics	  1999,	  59	  (1),	  59–65.	  40.	   Mathew,	   R.;	   Karp,	   C.	   M.;	   Beaudoin,	   B.;	   Vuong,	   N.;	   Chen,	   G.;	   Chen,	   H.	   Y.;	   Bray,	   K.;	  Reddy,	   A.;	   Bhanot,	   G.;	   Gelinas,	   C.;	   Dipaola,	   R.	   S.;	   Karantza-­‐Wadsworth,	   V.;	   White,	   E.,	  Autophagy	  suppresses	  tumorigenesis	  through	  elimination	  of	  p62.	  Cell	  2009,	  137	  (6),	  1062–1075.	    147 41.	   Moscat,	   J.;	   Diaz-­‐Meco,	   M.	   T.,	   p62	   at	   the	   crossroads	   of	   autophagy,	   apoptosis,	   and	  cancer.	  Cell	  2009,	  137	  (6),	  1001–1004.	  42.	   Sanz,	   L.;	   Diaz-­‐Meco,	   M.	   T.;	   Nakano,	   H.;	   Moscat,	   J.,	   The	   atypical	   PKC-­‐interacting	  protein	  p62	  channels	  NF-­‐kappaB	  activation	  by	  the	  IL-­‐1-­‐TRAF6	  pathway.	  The	  EMBO	  journal	  2000,	  19	  (7),	  1576–1586.	  43.	   Komatsu,	  M.;	  Waguri,	  S.;	  Koike,	  M.;	  Sou,	  Y.	  S.;	  Ueno,	  T.;	  Hara,	  T.;	  Mizushima,	  N.;	  Iwata,	  J.;	   Ezaki,	   J.;	   Murata,	   S.;	   Hamazaki,	   J.;	   Nishito,	   Y.;	   Iemura,	   S.;	   Natsume,	   T.;	   Yanagawa,	   T.;	  Uwayama,	   J.;	   Warabi,	   E.;	   Yoshida,	   H.;	   Ishii,	   T.;	   Kobayashi,	   A.;	   Yamamoto,	   M.;	   Yue,	   Z.;	  Uchiyama,	   Y.;	   Kominami,	   E.;	   Tanaka,	   K.,	   Homeostatic	   levels	   of	   p62	   control	   cytoplasmic	  inclusion	  body	  formation	  in	  autophagy-­‐deficient	  mice.	  Cell	  2007,	  131	  (6),	  1149–1163.	  44.	   White,	   E.;	   DiPaola,	   R.	   S.,	   The	   double-­‐edged	   sword	   of	   autophagy	   modulation	   in	  cancer.	  Clinical	  cancer	  research	   :	  an	  official	   journal	  of	   the	  American	  Association	   for	  Cancer	  Research	  2009,	  15	  (17),	  5308–5316.	  45.	   Amaravadi,	  R.	  K.;	  Yu,	  D.;	  Lum,	  J.	  J.;	  Bui,	  T.;	  Christophorou,	  M.	  A.;	  Evan,	  G.	  I.;	  Thomas-­‐Tikhonenko,	   A.;	   Thompson,	   C.	   B.,	   Autophagy	   inhibition	   enhances	   therapy-­‐induced	  apoptosis	  in	  a	  Myc-­‐induced	  model	  of	  lymphoma.	  The	  Journal	  of	  clinical	  investigation	  2007,	  117	  (2),	  326–336.	  46.	   Jemal,	  A.;	  Siegel,	  R.;	  Xu,	   J.;	  Ward,	  E.,	  Cancer	  statistics,	  2010.	  CA:	  a	  cancer	  journal	  for	  clinicians	  2010,	  60	  (5),	  277–300.	  47.	   Hezel,	  A.	  F.;	  Kimmelman,	  A.	  C.;	  Stanger,	  B.	  Z.;	  Bardeesy,	  N.;	  Depinho,	  R.	  A.,	  Genetics	  and	   biology	   of	   pancreatic	   ductal	   adenocarcinoma.	   Genes	   &	   development	   2006,	   20	   (10),	  1218–1249.	    148 48.	   Yang,	  S.;	  Wang,	  X.;	  Contino,	  G.;	  Liesa,	  M.;	  Sahin,	  E.;	  Ying,	  H.;	  Bause,	  A.;	  Li,	  Y.;	  Stommel,	  J.	  M.;	  Dell'antonio,	  G.;	  Mautner,	  J.;	  Tonon,	  G.;	  Haigis,	  M.;	  Shirihai,	  O.	  S.;	  Doglioni,	  C.;	  Bardeesy,	  N.;	   Kimmelman,	   A.	   C.,	   Pancreatic	   cancers	   require	   autophagy	   for	   tumor	   growth.	   Genes	   &	  development	  2011,	  25	  (7),	  717–729.	  49.	   Yang,	   S.;	   Kimmelman,	   A.	   C.,	   A	   critical	   role	   for	   autophagy	   in	   pancreatic	   cancer.	  Autophagy	  2011,	  7	  (8),	  912–913.	  50.	   Schneider,	   P.;	   Korolenko,	   T.	   A.;	   Busch,	   U.,	   A	   review	   of	   drug-­‐induced	   lysosomal	  disorders	   of	   the	   liver	   in	  man	   and	   laboratory	   animals.	  Microscopy	   research	  and	   technique	  1997,	  36	  (4),	  253–275.	  51.	   Carr,	  G.;	  Williams,	  D.	  E.;	  Diaz-­‐Marrero,	  A.	  R.;	  Patrick,	  B.	  O.;	  Bottriell,	  H.;	  Balgi,	  A.	  D.;	  Donohue,	   E.;	   Roberge,	   M.;	   Andersen,	   R.	   J.,	   Bafilomycins	   produced	   in	   culture	   by	  Streptomyces	   spp.	   isolated	   from	   marine	   habitats	   are	   potent	   inhibitors	   of	   autophagy.	  Journal	  of	  natural	  products	  2010,	  73	  (3),	  422–427.	  52.	   Rossi,	  M.;	  Munarriz,	  E.	  R.;	  Bartesaghi,	  S.;	  Milanese,	  M.;	  Dinsdale,	  D.;	  Guerra-­‐Martin,	  M.	   A.;	   Bampton,	   E.	   T.;	   Glynn,	   P.;	   Bonanno,	   G.;	   Knight,	   R.	   A.;	   Nicotera,	   P.;	   Melino,	   G.,	  Desmethylclomipramine	   induces	   the	   accumulation	   of	   autophagy	   markers	   by	   blocking	  autophagic	  flux.	  Journal	  of	  cell	  science	  2009,	  122	  (Pt	  18),	  3330–3339.	  53.	   Carew,	  J.	  S.;	  Espitia,	  C.	  M.;	  Esquivel,	  J.	  A.,	  2nd;	  Mahalingam,	  D.;	  Kelly,	  K.	  R.;	  Reddy,	  G.;	  Giles,	   F.	   J.;	   Nawrocki,	   S.	   T.,	   Lucanthone	   is	   a	   novel	   inhibitor	   of	   autophagy	   that	   induces	  cathepsin	  D-­‐mediated	  apoptosis.	  The	  Journal	  of	  biological	  chemistry	  2011,	  286	   (8),	  6602–6613.	    149 54.	   McAfee,	  Q.;	  Zhang,	  Z.;	  Samanta,	  A.;	  Levi,	  S.	  M.;	  Ma,	  X.	  H.;	  Piao,	  S.;	  Lynch,	  J.	  P.;	  Uehara,	  T.;	  Sepulveda,	  A.	  R.;	  Davis,	  L.	  E.;	  Winkler,	  J.	  D.;	  Amaravadi,	  R.	  K.,	  Autophagy	  inhibitor	  Lys05	  has	  single-­‐agent	  antitumor	  activity	  and	  reproduces	  the	  phenotype	  of	  a	  genetic	  autophagy	  deficiency.	  Proceedings	  of	   the	  aational	  academy	  of	  sciences	  of	   the	  United	  States	  of	  America	  2012,	  109	  (21),	  8253–8258.	  55.	   Bristol,	  M.	  L.;	  Emery,	  S.	  M.;	  Maycotte,	  P.;	  Thorburn,	  A.;	  Chakradeo,	  S.;	  Gewirtz,	  D.	  A.,	  Autophagy	   inhibition	   for	   chemosensitization	   and	   radiosensitization	   in	   cancer:	   do	   the	  preclinical	   data	   support	   this	   therapeutic	   strategy?	   The	   Journal	   of	   pharmacology	   and	  experimental	  therapeutics	  2013,	  344	  (3),	  544–552.	  56.	   Juhasz,	   G.,	   Interpretation	   of	   bafilomycin,	   pH	   neutralizing	   or	   protease	   inhibitor	  treatments	  in	  autophagic	  flux	  experiments:	  novel	  considerations.	  Autophagy	  2012,	  8	  (12),	  1875–1876.	  57.	   Donohue,	  E.;	  Thomas,	  A.;	  Maurer,	  N.;	  Manisali,	   I.;	  Zeisser-­‐Labouebe,	  M.;	  Zisman,	  N.;	  Anderson,	   H.	   J.;	   Ng,	   S.	   S.;	   Webb,	   M.;	   Bally,	   M.;	   Roberge,	   M.,	   The	   autophagy	   inhibitor	  verteporfin	   moderately	   enhances	   the	   antitumor	   activity	   of	   gemcitabine	   in	   a	   pancreatic	  ductal	  adenocarcinoma	  model.	  Journal	  of	  cancer	  2013,	  4	  (7),	  585–596.	  58.	   Donohue,	   E.,	   Characterization	   of	   verteporfin	   as	   an	   inhibitor	   of	   autophagosome	  formation	  and	  its	  therapeutic	  potential	  in	  cancer.	  Ph.D	  thesis,	  University	  of	  British	  Columbia	  2013.	  59.	   Klionsky,	  D.	  J.;	  Cuervo,	  A.	  M.;	  Seglen,	  P.	  O.,	  Methods	  for	  monitoring	  autophagy	  from	  yeast	  to	  human.	  Autophagy	  2007,	  3	  (3),	  181–206.	    150 60.	   Bischoff,	  P.;	  Josset,	  E.;	  Dumont,	  F.	  J.,	  Novel	  pharmacological	  modulators	  of	  autophagy	  and	  therapeutic	  prospects.	  Expert	  opinion	  on	  therapeutic	  patents	  2012,	  22	  (9),	  1053–1079.	  61.	   Fuertes,	  G.;	  Martin	  De	  Llano,	  J.	  J.;	  Villarroya,	  A.;	  Rivett,	  A.	  J.;	  Knecht,	  E.,	  Changes	  in	  the	  proteolytic	   activities	   of	   proteasomes	   and	   lysosomes	   in	   human	   fibroblasts	   produced	   by	  serum	   withdrawal,	   amino-­‐acid	   deprivation	   and	   confluent	   conditions.	   The	   Biochemical	  journal	  2003,	  375	  (Pt	  1),	  75–86.	  62.	   Donohue,	  E.;	  Tovey,	  A.;	  Vogl,	  A.	  W.;	  Arns,	  S.;	  Sternberg,	  E.;	  Young,	  R.	  N.;	  Roberge,	  M.,	  Inhibition	  of	  autophagosome	  formation	  by	  the	  benzoporphyrin	  derivative	  verteporfin.	  The	  Journal	  of	  biological	  chemistry	  2011,	  286	  (9),	  7290–7300.	  63.	   Balgi,	  A.	  D.;	  Fonseca,	  B.	  D.;	  Donohue,	  E.;	  Tsang,	  T.	  C.;	  Lajoie,	  P.;	  Proud,	  C.	  G.;	  Nabi,	  I.	  R.;	  Roberge,	   M.,	   Screen	   for	   chemical	   modulators	   of	   autophagy	   reveals	   novel	   therapeutic	  inhibitors	  of	  mTORC1	  signaling.	  Plos	  one	  2009,	  4	  (9),	  e7124.	  64.	   Personal	  communicaion	  from	  Dr.	  Michel	  Roberge,	  Department	  of	  Biochemistry	  and	  Molecular	  Biology,	  UBC.	  65.	   Kim,	  W.	  G.;	  Kim,	  J.	  P.;	  Kim,	  C.	  J.;	  Lee,	  K.	  H.;	  Yoo,	  I.	  D.,	  Benzastatins	  A,	  B,	  C,	  and	  D:	  new	  free	  radical	  scavengers	   from	  Streptomyces	  nitrosporeus	  30643	  I.	  Taxonomy,	   fermentation,	  isolation,	  physico-­‐chemical	  properties	  and	  biological	  activities.	  Journal	  of	  antibiotics	  1996,	  49,	  20–25.	  66.	   Kim,	  W.	  G.;	  Kim,	  J.	  P.;	  Kim,	  C.	  J.;	  Lee,	  K.	  H.;	  Yoo,	  I.	  D.,	  Benzastatins	  A,	  B,	  C,	  and	  D:	  new	  free	  radical	  scavengers	  from	  Streptomyces	  nitrosporeus	  30643	  II.	  Structure	  determination.	  Journal	  of	  antibiotics	  1996,	  49,	  26–30.	    151 67.	   Kim,	  W.	  G.;	  Kim,	  J.	  P.;	  Koshino,	  H.;	  Shin-­‐Ya,	  K.;	  Seto,	  H.;	  Yoo,	  I.	  D.,	  Benzastatins	  E,	  F,	  G:	  new	   alkaloids	   with	   neuronal	   cell	   protecting	   activity	   from	   Streptomyces	   nitrosporeus.	  Tetrahedron	  1997,	  53	  (12),	  4309–4316.	  68.	   Kinouchi,	  H.;	  Epstein,	  C.	  J.;	  Mizui,	  T.;	  Carlson,	  E.;	  Chen,	  S.	  F.;	  Chan,	  P.	  H.,	  Attenuation	  of	   focal	   cerebral	   ischemic	   injury	   in	   transgenic	   mice	   overexpressing	   CuZn	   superoxide	  dismutase.	  Proceedings	  of	  the	  national	  academy	  of	  sciences	  of	  the	  United	  States	  of	  America	  1991,	  88,	  11158–11162.	  69.	   Nallan,	  L.;	  Bauer,	  K.	  D.;	  Bendale,	  P.;	  Rivas,	  K.;	  Yokoyama,	  K.;	  Hornéy,	  C.	  P.;	  Pendyala,	  P.	   R.;	   Floyd,	   D.;	   Lombardo,	   L.	   J.;	   Williams,	   D.	   K.;	   Hamilton,	   A.;	   Sebti,	   S.;	   Windsor,	   W.	   T.;	  Weber,	   P.	   C.;	   Buckner,	   F.	   S.;	   Chakrabarti,	   D.;	   Gelb,	   M.	   H.;	   Van	   Voorhis,	   W.	   C.,	   Protein	  farnesyltransferase	   inhibitors	   exhibit	   potent	   antimalarial	   activity.	   Journal	   of	   medicinal	  chemistry	  2005,	  48	  (11),	  3704–3713.	  70.	   Bendale,	  P.;	  Olepu,	  S.;	  Suryadevara,	  P.	  K.;	  Bulbule,	  V.;	  Rivas,	  K.;	  Nallan,	  L.;	  Smart,	  B.;	  Yokoyama,	  K.;	  Ankala,	  S.;	  Pendyala,	  P.	  R.;	  Floyd,	  D.;	  Lombardo,	  L.	  J.;	  Williams,	  D.	  K.;	  Buckner,	  F.	   S.;	   Chakrabarti,	   D.;	   Verlinde,	   C.	   L.;	   Van	   Voorhis,	  W.	   C.;	   Gelb,	   M.	   H.,	   Second	   generation	  tetrahydroquinoline-­‐based	  protein	   farnesyltransferase	   inhibitors	  as	  antimalarials.	   Journal	  of	  medicinal	  chemistry	  2007,	  50	  (19),	  4585–4605.	  71.	   Bulbule,	   V.	   J.;	   Rivas,	   K.;	   Verlinde,	   C.	   L.;	   Van	   Voorhis,	   W.	   C.;	   Gelb,	   M.	   H.,	   2-­‐Oxotetrahydroquinoline-­‐based	   antimalarials	   with	   high	   potency	   and	   metabolic	   stability.	  Journal	  of	  medicinal	  chemistry	  2008,	  51	  (3),	  384–387.	    152 72.	   Kaur,	   K.;	   Jain,	   M.;	   Reddy,	   R.	   P.;	   Jain,	   R.,	   Quinolines	   and	   structurally	   related	  heterocycles	  as	  antimalarials.	  European	  journal	  of	  medicinal	  chemistry	  2010,	  45	  (8),	  3245–3264.	  73.	   Guo,	   T.;	   Gu,	  H.;	   Hobbs,	   D.	  W.;	   Rokosz,	   L.	   L.;	   Stauffer,	   T.	  M.;	   Jacob,	   B.;	   Clader,	   J.	  W.,	  Design,	   synthesis,	   and	   evaluation	   of	   tetrahydroquinoline	   and	   pyrrolidine	   sulfonamide	  carbamates	   as	   γ-­‐secretase	   inhibitors.	   Bioorganic	   &	   medicinal	   chemistry	   letters	   2007,	   17	  (11),	  3010–3013.	  74.	   Asberom,	  T.;	  Bara,	  T.	  A.;	  Clader,	  J.	  W.;	  Greenlee,	  W.	  J.;	  Guzik,	  H.	  S.;	  Josien,	  H.	  B.;	  Li,	  W.;	  Parker,	   E.	   M.;	   Pissarnitski,	   D.	   A.;	   Song,	   L.;	   Zhang,	   L.;	   Zhao,	   Z.,	   Tetrahydroquinoline	  sulfonamides	   as	   γ-­‐secretase	   inhibitors.	  Bioorganic	  &	  medicinal	  chemistry	   letters	  2007,	  17	  (1),	  205–207.	  75.	   Holsworth,	  D.	  D.;	  Cai,	  C.;	  Cheng,	  X.	  M.;	  Cody,	  W.	  L.;	  Downing,	  D.	  M.;	  Erasga,	  N.;	  Lee,	  C.;	  Powell,	  N.	  A.;	  Edmunds,	  J.	  J.;	  Stier,	  M.;	  Jalaie,	  M.;	  Zhang,	  E.;	  McConnell,	  P.;	  Ryan,	  M.	  J.;	  Bryant,	  J.;	   Li,	   T.;	   Kasani,	   A.;	   Hall,	   E.;	   Subedi,	   R.;	   Rahim,	   M.;	   Maiti,	   S.,	   Ketopiperazine-­‐based	   renin	  inhibitors:	  optimization	  of	   the	   "C"	   ring.	  Bioorganic	  &	  medicinal	  chemistry	  letters	  2006,	  16	  (9),	  2500–2504.	  76.	   Powell,	  N.	  A.;	  Ciske,	  F.	  L.;	  Cai,	  C.;	  Holsworth,	  D.	  D.;	  Mennen,	  K.;	  Van	  Huis,	  C.	  A.;	  Jalaie,	  M.;	  Day,	   J.;	  Mastronardi,	  M.;	  McConnell,	   P.;	  Mochalkin,	   I.;	   Zhang,	  E.;	  Ryan,	  M.	   J.;	  Bryant,	   J.;	  Collard,	   W.;	   Ferreira,	   S.;	   Gu,	   C.;	   Collins,	   R.;	   Edmunds,	   J.	   J.,	   Rational	   design	   of	   6-­‐(2,4-­‐diaminopyrimidinyl)-­‐1,4-­‐benzoxazin-­‐3-­‐ones	  as	  small	  molecule	  renin	  inhibitors.	  Bioorganic	  &	  medicinal	  chemistry	  2007,	  15	  (17),	  5912–5949.	    153 77.	   Holsworth,	  D.	  D.;	  Jalaie,	  M.;	  Belliotti,	  T.;	  Cai,	  C.;	  Collard,	  W.;	  Ferreira,	  S.;	  Powell,	  N.	  A.;	  Stier,	   M.;	   Zhang,	   E.;	   McConnell,	   P.;	   Mochalkin,	   I.;	   Ryan,	  M.	   J.;	   Bryant,	   J.;	   Li,	   T.;	   Kasani,	   A.;	  Subedi,	   R.;	   Maiti,	   S.	   N.;	   Edmunds,	   J.	   J.,	   Discovery	   of	   6-­‐ethyl-­‐2,4-­‐diaminopyrimidine-­‐based	  small	   molecule	   renin	   inhibitors.	   Bioorganic	   &	  medicinal	   chemistry	   letters	  2007,	   17	   (13),	  3575–3580.	  78.	   Yoo,	  S.	  E.;	  Kim,	  J.	  H.;	  Yi,	  K.	  Y.,	  Model	  study	  for	  the	  biogenesis	  of	  benzastatins.	  Bulletin	  of	  the	  Korean	  Chemical	  Society	  1999,	  20	  (2),	  139–140.	  79.	   Morimoto,	   Y.;	   Matsuda,	   F.;	   Shirahama,	   H.,	   Total	   synthesis	   of	   (±)-­‐virantmycin	   and	  determination	  of	  its	  stereochemistry.	  Synlett	  1991,	  202–203.	  80.	   Morimoto,	  Y.;	  Shirahama,	  H.,	  Synthetic	  studies	  on	  virantmycin.	  2.	   total	  synthesis	  of	  unnatural	  (+)-­‐virantmycin	  and	  determination	  of	  its	  absolute	  stereochemistry.	  Tetrahedron	  1996,	  52	  (32),	  10631–10652.	  81.	   Ori,	   M.;	   Toda,	   N.;	   Takami,	   K.;	   Tago,	   K.;	   Kogen,	   H.,	   Stereospecific	   construction	   of	  contiguous	   quaternary	   and	   tertiary	   stereocenters	   by	   rearrangement	   from	   indoline-­‐2-­‐methanol	   to	   2,2,3-­‐trisubstituted	   tetrahydroquinoline:	   application	   to	   an	   efficient	   total	  synthesis	  of	  natural	   virantmycin.	  Angewandte	  chemie	  international	  edition	  2003,	  42	   (22),	  2540–2543.	  82.	   Omura,	   S.;	   Nakagawa,	   A.;	   Hashimoto,	   H.;	   Oiwa,	   R.;	   Iwai,	   Y.;	   A.,	   H.;	   Shibukawa,	   N.;	  Kojima,	  Y.,	  Virantmycin,	  a	  potent	  antiviral	  antibiotic	  produced	  by	  a	  strain	  of	  streptomyces.	  The	  Journal	  of	  antibiotics	  1980,	  33,	  1395.	    154 83.	   Nakagawa,	  A.;	  Iwai,	  Y.;	  Hashimoto,	  H.;	  Miyazaki,	  N.;	  Oiwa,	  R.;	  Takahashi,	  Y.;	  Hirano,	  A.;	  Shibukawa,	  N.;	  Kojima,	  Y.;	  Omura,	  S.,	  Virantmycin,	  a	  new	  antiviral	  antibiotic	  produced	  by	  a	  strain	  of	  Streptomyces.	  The	  Journal	  of	  antibiotics	  1981,	  34,	  1480.	  84.	   Omura,	   S.;	   Nakagawa,	   A.,	   Structure	   of	   virantmycin,	   a	   novel	   antiviral	   antibiotic.	  Tetrahedron	  letters	  1981,	  22	  (23),	  2199–2202.	  85.	   Morimoto,	   Y.;	   ODA,	   K.;	   Shirahama,	   H.;	   Matsumoto,	   T.;	   Omura,	   S.,	   Assignment	   of	  absolution	   configuration	   fro	   virantmycin	   and	   synthesis	   of	   its	   antipode.	   Chemistry	   letters	  1988,	  909–912.	  86.	   Pearce,	  C.	  M.;	  Sanders,	  J.	  K.	  M.,	  Stereochemistry	  of	  (–)-­‐Virantmycin.	  Journal	  Chemistry	  Society	  Perkin	  Transactions	  1	  1990,	  409–411.	  87.	   Morimoto,	   Y.;	   Matsuda,	   F.;	   Shirahama,	   H.,	   Total	   synthesis	   of	   (±)-­‐virantmycin	   and	  determination	  of	  its	  stereochemsitry.	  Synlett	  1991,	  202–203.	  88.	   Hill,	  M.	  L.;	  Raphael,	  R.	  A.,	  Total	  synthesi	  of	  the	  antiviral	  (±)	  virantmycin.	  Tetrahedron	  letters	  1986,	  27	  (11),	  1293–1296.	  89.	   Hill,	   M.	   L.;	   Raphael,	   R.	   A.,	   Total	   synthesis	   of	   the	   antiviral	   (±)-­‐virantmycin.	  Tetrahedron	  1990,	  46,	  4587–4594.	  90.	   Morimoto,	  Y.;	  Matsuda,	  F.;	  Shirahama,	  H.,	  Synthetic	  studies	  on	  virantmycin.	  1.	   total	  synthesis	   of	   	   (±)-­‐virantmycin	   and	   determination	   of	   its	   relative	   stereochemistry.	  Tetrahedron	  1996,	  52	  (32),	  10609–10630.	  91.	   Corey,	  E.	  J.;	  Gilman,	  N.	  W.;	  Ganem,	  B.	  E.,	  New	  methods	  for	  the	  oxidation	  of	  aldehydes	  to	   carboxylic	   acids	   and	   esters.	   Journal	   of	   the	   American	   chemical	   Society	   1968,	   90	   (20),	  5616–5617.	    155 92.	   Steinhagen,	   H.;	   Corey,	   E.	   J.,	   A	   simple	   convergent	   approach	   to	   the	   synthesis	   of	   the	  antiviral	  agent	  virantmycin.	  Organic	  letters	  1999,	  1	  (5),	  823–824.	  93.	   Steinhagen,	  H.;	  Corey,	  E.	   J.,	  A	  convenient	  and	  versatile	  route	  to	  hydroquinolines	  by	  inter-­‐	   and	   intramolecular	   Aza-­‐Diels–Alder	   pathways.	   Angewandte	   chemie	   international	  edition	  1999,	  38,	  1928–1931.	  94.	   Ori,	  M.;	  Toda,	  N.;	  Takami,	  K.;	  Tago,	  K.;	  Kogen,	  H.,	   Stereospecific	   synthesis	   of	   2,2,3-­‐trisubstituted	  tetrahydroquinolines:	  application	  to	  the	  total	  syntheses	  of	  benzastatin	  E	  and	  natural	  virantmycin.	  Tetrahedron	  2005,	  61	  (8),	  2075-­‐2104.	  95.	   Back,	   T.	   G.;	  Wulff,	   J.	   E.,	   A	   stereodivergent	   synthesis	   of	   virantmycin	  by	   an	   enzyme-­‐mediated	   diester	   desymmetrization	   and	   a	   highly	   hindered	   aryl	   amination.	   Angewandte	  chemie	  international	  edition	  2004,	  43	  (47),	  6493–6469.	  96.	   Jones,	   G.	   B.;	   Heaton,	   S.	   B.;	   Chapman,	   B.	   J.;	   Guzel,	   M.,	   On	   the	   origins	   of	  enantioselectivity	   in	   oxazaborolidine	   mediated	   carbonyl	   reductions.	   Tetrahedron-­‐asymmetry	  1997,	  8	  (21),	  3625-­‐3636.	  97.	   Palais,	  L.;	  Alexakis,	  A.,	  Copper-­‐Catalyzed	  Asymmetric	  Conjugate	  Addition	  with	  Chiral	  SimplePhos	  Ligands.	  Chemistry-­‐	  a	  European	  journal	  2009,	  15	  (40),	  10473-­‐10485.	  98.	   Ushkov,	  A.	  V.;	  Grushin,	  V.	  V.,	  Rational	  Catalysis	  Design	  on	   the	  Basis	  of	  Mechanistic	  Understanding:	   Highly	   Efficient	   Pd-­‐Catalyzed	   Cyanation	   of	   Aryl	   Bromides	   with	   NaCN	   in	  Recyclable	   Solvents.	   Journal	   of	   the	   American	   chemical	   society	   2011,	   133	   (28),	   10999–11005.	  99.	   Cai,	  L.	  Z.;	  Liu,	  X.;	  Tao,	  X.	  C.;	  Shen,	  D.,	  Efficient	  microwave-­‐assisted	  cyanation	  of	  aryl	  bromide.	  Synthetic	  Communications	  2004,	  34	  (7),	  1215–1221.	  

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