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Approaches to Identifying Synthetic Lethal Interactions in Cancer YALE JOURNAL OF BIOLOGY AND MEDICINE 88 (2015), pp.145-155. REvIEw Approaches to Identifying Synthetic Lethal Interactions in Cancer Jordan M. Thompson, Quy H. Nguyen, Manpreet Singh, and Olga v. Razorenova* Molecular Biology and Biochemistry Department, University of California Irvine, Irvine, California Targeting synthetic lethal interactions is a promising new therapeutic approach to exploit specific changes that occur within cancer cells. Multiple approaches to investigate these interactions have been developed and successfully implemented, including chemical, siRNA, shRNA, and CRISPR library screens. Genome- wide computational approaches, such as DAISY, also have been successful in predicting synthetic lethal in - teractions from both cancer cell lines and patient samples. Each approach has its advantages and disadvantages that need to be considered depending on the cancer type and its molecular alterations. This review discusses these approaches and examines case studies that highlight their use. INTRODUCTION formed genome [1,2]. Indeed, most of the hallmarks of A major challenge in developing new anti-cancer cancer are associated with changes to the cancer cell’s therapies is to identify compounds that have sufficient genetic makeup in order to enable it to endlessly prolif - therapeutic windows comprised of the concentrations erate [3]. These changes can be exploited to specifically causing tumor regression with minimal normal tissue tox - target and kill cancer cells while sparing normal cells icity. Most cancer therapies suffer from narrow thera - [2,4]. Traditional chemotherapy, including irradiation or peutic windows causing numerous side effects that DNA damaging agents, relies on this principle. Most can - greatly reduce a patient’s quality of life. Further elucida - cer cells have defects in their cell cycle checkpoints tion of the unique differences between cancer and nor - where DNA would normally be repaired before replica - mal tissue allows for the development of targeted tion [5,6]. Thus, radiotherapy and DNA damaging agents therapies with reduced side effects. During malignant target DNA in the continuously dividing cancer cells, progression, cancer cells acquire multiple mutations, in - causing cell death. Normal cells, with their intact cell cluding the activation of proto-oncogenes, the inactiva - cycle checkpoints, repair the damage before dividing tion of tumor suppressors, and other additional genetic [5,6]. However, these therapies often have serious side or epigenetic alterations resulting in a drastically trans - effects and greatly reduce a patient’s quality of life [7]. *To whom all correspondence should be addressed: Olga V. Razorenova, University of California Irvine, 845 Health Sciences Road, Gross Hall Room 3010, Mail Code 3900, Irvine, CA 92697; Tele: 949-824-8156; Email: [email protected]. †Abbreviations: AML, acute myeloid luekemia; APC, adenomatous polyposis coli; ATM, ataxia telangiectasia mutated; ATR, ataxia telangiectasia and Rad3-related protein kinase; BID, BH3 interacting-domain death agonist; BRCA, breast cancer early onset; BRG1 , Brahma related gene 1; BRM, Brahma; CCNE1, cycline E1; CPT, camptothecin; DECODER, Deep Coverage Design shRNA Library; DISC, death-inducing signaling complex; FA, Fanconi anemia; FANCD2, Fanconi anemia group D2; gRNA, guide RNA; HR, homologous recombination; IR, ionizing radiation; KRAS, Kirsten rat sarcoma; mESC, mouse embryonic stem cells; MSH2, MutS protein homolog 2; NSAIDs, nonsteroidal anti-inflammatory drugs; PARP1, Poly [ADP-ribose] Polymerase 1; RCC, renal cell carci - nomas; RNAi, RNA interference; SCNA, somatic copy number alteration; SDL, synthetic dosage lethality; SoF, survival of the fittest; SWI/SNF, switch/sucrose NonFermentable; TOP, topoisomerase; TRAIL, tumor necrosis factor-related apoptosis-inducing ligand; VHL, Von Hippel Lindau. Keywords: synthetic lethality, synthetic lethal interactions, drug discovery, drug screening assays, antitumor, cancer, neoplasms, li - brary screens, siRNA screen, siRNA library, shRNA screen, shRNA library, CRISPR screen, CRISPR library, computational screen, DAISY Author contributions: JMT: Wrote the paper and compiled Figures 1, 2, and 3; QHN: wrote the CRISPR section and compiled Figure 4; MS: wrote the DAISY section and compiled Figure 5; OVR: Outlined the review content and helped to finalize the text and fig - ures. QHN and MS contributed equally to the work. All authors contributed to literature search and revising the article. This review was supported by ACS fund IRG 98-27907 to OVR. Copyright © 2015 145 146 Thompson et al.: Approaches to identifying synthetic lethal interactions at any level, and the loss of function does not have to occur in the same manner for both co-essential genes. For exam - ple, one gene can be lost due to deletion and the other in - hibited by a chemical compound. while synthetic lethality is best known in the context of loss-of-function mutants [9], other perturbations including gene overexpression [11,12], epigenetic changes [13], and cell extrinsic differ - ences [14,15] can cause these interactions to occur [10] (Figure 1). In identifying genes that are synthetically lethal within each cancer type, we gain an understanding of the molecular biology of those cancers and how it can be specifically exploited. There are multiple approaches that have been successfully used to identify synthetic lethal in - teractions in cancer. DIFFERENT APPROACHES TO IDENTIFY SYNTHETIC LETHAL INTERACTIONS Hypothesis-Driven Approach It is possible to identify synthetic lethal interactions using a candidate or hypothesis-driven approach if the al - terations to the molecular pathways in the cancer are well established. For instance, if a tumor suppressor is fre - quently lost and the resulting changes to the gene expres - Figure 1. Synthetic lethal interactions spare normal sion profile are known, then it may be possible to use RNA cells while selectively killing cancer cells. A) In the loss of function, phenotype cancer cells have lost the function interference (RNAi †) or chemical compounds to inhibit the of gene X due to genetic loss, epigenetic changes, cell ex - genes that are expected to be upregulated to compensate trinsic changes, and more. When cells that express gene for the loss. This may result in a synthetic lethal interaction. X are treated to inhibit gene X’s synthetic lethal partner Mutations in the breast cancer early onset ( BRCA ) 1 and gene Y, they remain viable, but cancer cells lacking gene BRCA2 genes are known to occur frequently in breast and X die. B) In the gain of function phenotype, also called syn - ovarian cancers. Knowing that BRCA1/2 are involved in thetic dosage lethal (SDL), cancer cells have an overex - homologous recombination (HR) and DNA double-strand pression or overactivation of gene X due to oncogenic break repair, both Byant et al. [16] and Farmer et al. [17] mutation, generation of fusion proteins, changes in up - stream regulators, epigenetic changes, cell extrinsic hypothesized that inhibition of Poly (ADP-Ribose) Poly - changes, and more. When cells with wild type (WT) gene merase (PARP), responsible for DNA single-strand break X are treated to inhibit gene X’s synthetic lethal partner repair, would be synthetically lethal with loss of BRCA1/2 . gene Y they survive, but cancer cells with the gene X gain This hypothesis is based on the knowledge that PARP de - of function die. ficiency results in spontaneous single-strand breaks at the DNA replication fork, which require HR for repair. Cells Synthetic lethality represents a more targeted approach deficient in BRCA1/2 are unable to provide HR for repair that exploits the specific changes within the cancer at a of DNA single- and double-strand breaks caused by chem - single gene level that separate it from healthy tissue. ical or genetic PARP inhibition, causing synthetic lethality. The synthetic lethal interaction was extended to in vivo studies, followed by clinical studies where treatment of SYNTHETIC LETHALITY BRCA1/2 -deficient tumors with a DNA damaging agent Synthetic lethality is defined as the interaction be - combined with a PARP inhibitor extended patient survival tween two co-essential genes such that inhibiting the func - over chemotherapy alone [18,19]. tion of either gene separately results in cell survival, but Synthetic lethal interactions also may explain why inhibiting the function of both genes results in cell death particular chemical compounds have increased efficacy in (Figure 1) [8-10]. Inhibition of a gene may be achieved by cancers characterized with specific genetic alterations. chemical or genetic means. For instance, the genetic inhi - while clinical trials for PARP inhibitors in ovarian cancer bition of a gene’s function may occur through RNAi, mu - were successful in increasing progression free survival by tation, deletion, epigenetic changes, or perturbations of 62 percent overall [20], in a follow-up analysis of the same upstream regulators. Chemical inhibition of the gene’s study, it found that the BRCA1/2 -deficient group had a function may be achieved by treatment with a chemical much higher rate of 82 percent [21]. Another study by Lei - compound. The inhibition of co-essential genes can occur bowitz et al. [22] hypothesized that non-steroidal anti-in - Thompson et al.: Approaches to identifying synthetic lethal interactions 147 Figure 2. Approaches to generating matched cell lines for synthetic lethality screens. There are multiple ap - proaches to generate
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