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Clonal analysis of human mammary cell transformation and X-ray sensitivity

University of British Columbia

Intra-tumoural biological, transcriptional and genomic heterogeneity are hallmarks of human breast cancers. However, tumour propagating activity appears confined to subsets of cells within each tumour. This finding is thought to indicate a persistence of mechanisms that maintain a hierarchical growth and differentiation structure in the normal mammary gland. Because little is known about the responses of either primary normal or malignant human mammary cells to existing therapies, this thesis sought to examine the intrinsic sensitivity of different purified human mammary colony-forming cell (CFC) types and tumours derived from them to ionizing radiation. Luminal progenitor (LP) CFCs were found to be ~1.5-fold more radioresistant than basal cell (BC) CFCs and LPs also showed evidence of checkpoint adaptation, slower repair activity and greater predisposition of γH2AX foci accumulation. Two human breast cancer cell lines (MDA-MB231 and SUM149) and a non-tumorigenic human mammary cell line (MCF-10A) were all found to be more radioresistant than the normal LP-CFCs. CFCs isolated from 8-week tumours generated in mice transplanted with normal human BCs or LPs transduced with KRASG¹²D showed even greater radioresistance and this was further increased in serially passaged derivative lines with more aggressive growth properties. To examine the responsiveness of malignant cells with tumor-initiating cell (TIC) activity in vivo, a dose-response analysis was first undertaken of their frequencies in the MDA-MB231 and SUM149 cells. Limiting dilution analysis (LDA) and single-cell transplants showed the frequency of TICs in both to be very high (<10%), but with increasingly marked inhibition of their detection in tumours initiated from innocula containing 2x10⁴ cells or more. Analogous LDA measurements of the TICs in KRASG12D-transduced BCs and LPs, yielded frequencies of ~0.2% and ~0.07%, respectively, in contrast to the frequencies of 0.02-0.5% for BCs and 0.01-0.6% for LPs for tumours initiated from 3x10⁴-10⁶ cells. These results demonstrate the heterogeneity of treatment responses in normal human mammary cells with innate proliferative ability that can be heightened by transformation. They also reveal the complex clonal dynamics operative in the growth of TICs in vivo that may confound interpretation of treatment effects assessed only by measuring immediate changes in tumor size.

Text

2018-04-16

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/65394

Interdisciplinary Oncology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/