Smallcelllungcancer(SCLC)isthemostaggressiveformoflungcancer,withamediansurvivalof24 monthsandafive-yearoverallsurvivalof7%.Cisplatin-basedchemotherapyisthefirst-linetreatmentforSCLC, functioning by creating DNA crosslinks leading to cancer cell death. While initially effective, many patients ultimatelydevelopcisplatinresistanceandexperiencetumorrecurrence.TheDNAdamageresponse(DDR)is asignalingnetworkthatrecognizeschallengestogenomeintegrityandcoordinatesdiverseDNArepairandcell cycle checkpoint pathways. Targeting DDR proteins critical to the cellular response to cisplatin in SCLC may overcome acquired cisplatin resistance, but the cellular response to cisplatin remains unclear because many proteins that respond to cisplatin-induced DNA damage have yet to be identified. To address this issue, a syntheticlethalscreenusingacustomsiRNAlibraryof1008nuclearenzymeswasperformedinH128cisplatin- resistantSCLCcellstoidentifygenescriticalformediatingcisplatinresistance.EnhancerOfZeste2Polycomb Repressive Complex 2 Subunit (EZH2), a H3 K27 methyltransferase, was identified as one of the strongest syntheticlethalhitsinthescreen,whereknockdownofEZH2stronglysensitizedH128cellstocisplatin. EZH2isoverexpressedinSCLCaswellasothercancersandhasbeenimplicatedincancerprogression. Moreover,EZH2inhibitorshaveshownefficacyinseveralcancers,includingSCLC;?however,thethemolecular underpinnings of the efficacy of EZH2 depletion or inhibition in SCLC and the mechanistic role of EZH2 in mediatingcisplatinresistanceinSCLCareunclear.Inthisregard,IvalidatedthatEZH2depletionincisplatin- resistantH128andH146SCLCcelllinesresultsincisplatinhypersensitivity,andfurthermorefoundthatEZH2 localizestoDNAdamagesitesinducedbylasermicroirradiation,suggestingthatEZH2mayfunctiondirectlyin mediating DNA damage resistance in SCLC. In addition, using mass spectrometry analysis of purified EZH2 fromcells,IidentifiedanovelinteractionbetweenEZH2andDNADamage-BindingProtein1(DDB1),anE3 ubiquitinligasecomponentthatpromotesnucleotideexcisionrepair(NER)ofcisplatin-inducedDNAintrastrand crosslinks,whichIvalidatedbyco-immunoprecipitation.Assuch,IhypothesizethatEZH2playsacriticalrolein mediatingcisplatinresistanceinSCLCbypromotingDNAdamageresponse(DDR)activitiesandfurthermore thatEZH2inhibitionwillsensitizeresistantSCLCcellsandtumorswithdysregulatedDDRpathwaystocisplatin treatment.
Aim1 willdeterminethemechanismbywhichEZH2mediatescisplatinresistanceinSCLC.
Aim2 will determine if EZH2 inhibition sensitizes resistant SCLC cells and tumors with dysregulated DDR protein expressiontocisplatintreatmentinvitroandinvivo.CompletionoftheseaimswilldefineanovelroleforEZH2 in the DDR in mediating cisplatin resistance in SCLC and elucidate the effectiveness and specific biological context for which EZH2 can function as a novel therapeutic target for improving the efficacy of treatment for patientswithSCLCwhofailinitialtreatment.
Small cell lung cancer (SCLC) is associated with extremely poor outcomes because most patients develop resistance to initial chemotherapy treatment. Here, we have identified Enhancer Of Zeste 2 Polycomb Repressive Complex 2 Subunit (EZH2) as a mediator of chemotherapy resistance in SCLC and provide evidence for its role in the DNA damage response (DDR). Completion of these aims will define a novel role for EZH2 in the DDR in mediating cisplatin resistance in SCLC and elucidate the effectiveness and specific biological context for which EZH2 can function as a novel therapeutic target for improving the efficacy of treatment for patients with SCLC who fail initial treatment.
