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A cross species approach to identify potential therapeutic targets through synthetic lethal interactions

University of British Columbia

Chromosome instability (CIN) is characterized by the loss or gain of large portions of DNA and is characteristic of ~85% of solid tumours. Sequencing of the human orthologues of ~200 genes that cause CIN in yeast identified mutations in approximately 25% of tumours tested. Mutations in cohesin genes and CDC4 represented the two major mutational categories identified. Large scale genetic interaction networks in model organisms can provide insight into the biology underlying tumour mutations and can identify potential therapeutic targets. This approach is based on the concept of synthetic lethality (SL); cell death resulting from the combination of two sub-lethal mutations. Therapies that take advantage of SL distinguish a cancer cell from a normal cell based on their genetic background. This thesis investigated genetic interactions of three cohesin genes, SMC1, SCC1, and SCC2, using high throughput synthetic genetic array (SGA) methods in S. cerevisiae. The overlay of these three genome wide SGA screens and validation using growth curve analysis found that sub-optimal cohesin requires the presence of proteins that mediate replication fork progression. The protruding vulva assay was developed to identify genetic interactions in the somatic cells of C. elegans. It was used to test whether the cohesin interactions were conserved in a multicellular animal. 80% of the validated interactions identified with cohesin in yeast are conserved in C. elegans. Additional fork mediators, namely the pme/PARP family of genes was found to interact with him-1/SMC1 in both C. elegans and human cells. Human cells depleted of SMC1 by siRNA are selectively sensitive to the PARP inhibitor olaparib, currently being evaluated in phase II clinical trials. Additional genetic interaction testing found that CDC4 has a distinct genetic interaction profile from that of cohesin, suggesting different mutational consequences. Work in C. elegans with the lin-23 mutant suggested LIN-23 is involved in controlling CYE-1/Cyclin E levels. lin-23 mutants and human CDC4-/- cells are unable to properly respond to alkylating DNA damage, suggesting CDC4 is important for the DNA damage response. Keywords: colon tumours, chromosome instability, cohesin, CDC4, genetic interactions, SGA, replication fork, PARP.

Text

2011-10-20

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/38145

Medical Genetics

University of British Columbia

UBCV

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