Specific synthetic lethal killing of RAD54B-deficient human colorectal cancer cells by FEN1 silencing

Abstract

Mutations that cause chromosome instability (CIN) in cancer cells produce ''sublethal'' deficiencies in an essential process (chromosome segregation) and, therefore, may represent a major untapped resource that could be exploited for therapeutic benefit in the treatment of cancer. If second-site unlinked genes can be identified, that when knocked down, cause a synthetic lethal (SL) phenotype in combination with a somatic mutation in a CIN gene, novel candidate therapeutic targets will be identified. To test this idea, we took a cross species SL candidate gene approach by recapitulating a SL interaction observed between rad54 and rad27 mutations in yeast, via knockdown of the highly sequence-and functionally-related proteins RAD54B and FEN1 in a cancer cell line. We show that knockdown of RAD54B, a gene known to be somatically mutated in cancer, causes CIN in mammalian cells. Using high-content microscopy techniques, we demonstrate that RAD54B-deficient human colorectal cancer cells are sensitive to SL killing by reduced FEN1 expression, while isogenic RAD54B proficient cells are not. This conserved SL interaction suggests that extrapolating SL interactions observed in model organisms for homologous genes mutated in human cancers will aid in the identification of novel therapeutic targets for specific killing of cancerous cells exhibiting CIN.cancer therapeutics ͉ chromosome instability ͉ synthetic lethality

DNA Base Excision Repair: Evolving Biomarkers for Personalized Therapies in Cancer

“…These results suggest that FEN-1 might be a potential marker for selecting patients at high risk and therapy [84]. Interestingly, synthetic lethality (SL) has been observed in RAD54B-deficient human colorectal cancer cell line by iatrogenic reduction of FEN1 expression thus demonstrating it to be a potential novel therapeutic biological target [85].…”

Synthetic Lethal Genetic Interactions That Decrease Somatic Cell Proliferation in Caenorhabditis elegans Identify the Alternative RFCCTF18as a Candidate Cancer Drug Target

“…On the other hand, work by Tarailo et al (2007) that focused on the spindle assembly checkpoint suggested that a much higher fraction of interactions, on the order of 50%, are conserved between S. cerevisiae and C. elegans. Recent work by McManus et al (2009) demonstrated the usefulness of a cross-species candidate approach for drug target identification. They showed that an interaction first identified in yeast between RAD27 and RAD54 (Symington, 1998) was conserved in HCT116 cells.…”

“…Second-site mutations that are nondetrimental to normal somatic cells, but that cause SL in combination with any of these known cancer mutations represent potential cancer therapeutic targets (Hartwell et al, 1997;Kaelin, 2005;McManus et al, 2009). Cross-species candidate genes for such second-site SL partners of primary CIN mutations can be derived from yeast SL genetic interaction networks and tested for conservation of SL in mammalian cell lines using a combination of siRNA and knockout mutations (McManus et al, 2009). This is a relatively expensive and time-consuming task in tissue culture that involves testing the SL gene pairs singly and in combination and then assessing cell proliferation.…”

Droplet microfluidic technology for single-cell high-throughput screening

“…First, it permitted analysis of the overall utility and biocompatibility of the droplet format, by allowing direct measurement of the viability of encapsulated cells. Second, it enabled cytotoxicity screening capabilities that are important in cancer research to either assess drug cytotoxicity or to screen cancer cells for therapeutic targets via synthetic lethality (19). This droplet workflow is generic and could be used with different types of libraries (e.g., DNA, siRNA, drug), fluorescent assay read-outs, or additional droplet manipulation modules.…”