Non-oncogene addiction in cancer cells

Division Of Basic Sciences - Nci

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Abstract

BACKGROUND. We define non-oncogene addiction (NOA) as a cancer cell's reliance on stress-response genes for survival. The malignant state itself generates unique oncogenic stress phenotypes, such as DNA damage, metabolic stress, proteotoxic stress, an inflammatory microenvironment, and immune surveillance. This stress makes cancer cells highly dependent on, and thus sensitive to the disruption of, stress-response pathways. In contrast, normal cells don't experience such oncogenic stress, making them far less sensitive to these same perturbations. This fundamental difference between cancer and normal cells' dependency on stress-response pathways forms the theoretical basis of NOA. Distinct from oncogene addiction (OA), NOA involves stress-response genes rarely mutated in cancer. NOA represents a broad category of synthetic lethal and collateral dependency mechanisms, offering a unique, orthogonal approach to current targeted cancer therapies. OBJECTIVES. The objectives of this project are: 1) to identify NOA genes and molecular pathways in cancer cells using genetic screens and hypothesis-driven approaches; 2) to investigate the molecular mechanisms that underly NOA; and 3) to explore the therapeutic implication of NOA genes as potential cancer drug targets. We will carry out these objectives primarily using cancer cells harboring the KRAS oncogene as our model system because KRAS mutant tumors tend to have worse prognosis than KRAS wildtype tumors. MAJOR ACTIVITIES, SIGNIFICANT RESULTS AND KEY OUTCOMES. 1) NOA of KRAS mutant cells to the RNA splicing factor ERH. Through a synthetic lethal RNAi screen in KRAS mutant cells, we have identified the ERH gene as a synthetic lethal partner in KRAS mutant cells. The ERH gene encodes an evolutionarily conserved protein of the same name, but with poorly understood function. Using mass-spectrometry, we identified that ERH is associated with the spliceosome protein SNRPD3, and it is therefore a component of the RNA splicing machinery. We discovered that ERH is required for the proper splicing and expression of a subset of mitotic genes, including CENPE and ATR, that are critical for the maintenance of chromosomal stability in KRAS mutant cells. This work defined a new function for the ERH protein and uncovered a previously unknown form of NOA to the RNA splicing machinery in KRAS mutant cancer cells. This work suggests that perturbation of selected components of the RNA splicing machinery could be a potential approach to target KRAS mutant cancer cells. This work has been completed and published. 2) NOA of KRAS mutant cells to the SUMOylation pathway for oncogenic transformation. Through a synthetic lethal RNAi screen in KRAS mutant cells, we have identified the SUMOylation pathway including the SUMO E2 ligase, UBE2, to play an important role in the survival and anchorage-independent growth of KRAS mutant cancer cells. Using mass-spectrometry, we identified multiple proteins whose SUMOylation are disrupted in KRAS mutant cancer cells. Using genetic rescue approaches, we showed that several SUMO target proteins, including KAP1, CHD1 and EIF3L, are critical for the growth of KRAS mutant cells under anchorage-independent conditions. This work identifies a new role of the SUMO pathway in KRAS-driven oncogenesis and suggests the SUMO pathway as a potential drug target for KRAS mutant tumors. This work has been published. We are currently exploring the effect of small-molecule SUMO inhibitors in preclinical models of KRAS mutant cancer. 3) NOA of KRAS mutant cells to autophagy. Through a combinatorial RNAi analysis in KRAS mutant cells, we identified the kinase CRAF, among many other signaling effectors downstream of KRAS, as the major driver that mediates the oncogenic signaling of mutant KRAS. We discovered that the autophagy E1 ligase ATG7 as an important NOA mechanism that supports the survival of KRAS mutant cancer cells when CRAF is inhibited. This work identifies the autophagy pathway as a co-target of CRAF in KRAS mutant cancer cells. This work has been published. We are currently investigating the mechanism by which autophagy contributes to the growth and survival of KRAS mutant tumors, and we are evaluating the effect of pharmacological inhibitor targeting the autophagy E1 ligase ATG7 in combination with KRAS and MAPK inhibitors on the growth and survival of KRAS mutant cancer cells.