Abstract

Amajorchallengeforthecancergenomicscommunityisdeterminingwhichsomaticmutationsarecontributing totumorigenesisandwhichare?passenger?(neutral)mutations.SomaticmutationsthatcausealteredmRNA splicinghaverecentlybeenappreciatedasoncogenicdriverevents,suchasthecasewithskippingofexon14 intheMETproto-oncogene.Additionally,lungcancerpatientswithskippingofMETexon14respondwellto targeteddruginhibitors.ThroughcomputationalanalysisofbothDNAandmRNAsequencingdataof495lung cancerdonorsamples,wehaveidentified635casesofexon-skippingassociatedwithsomaticmutation;? however,welackexperimentalevidencetoknowwhichoftheseexonsmightalsorepresentoncogenicdriver events.Thisprojectwilltestthehypothesisthatasubsetofexon-skippingeventsassociatedwith somaticmutationsarenoveloncogenicalterations.Totestthishypothesis,ourfirstaimwilldetermineif previouslyuncharacterizedexon-skippingevents,associatedwithsomaticmutationsinRASpathwaygenes, areoncogenic.RASpathwaygenesareknowntoberecurrentlymutatedinlungcancerandmanyare targetablealterations;?therefore,wewillfocusourinitialstudiesonexon-skippingeventswhicharemostlikely tobeoncogenicandmostlikelytoleadtoatherapeutictarget.
Our secondaim willdevelopanovelhigh- throughputCRISPR-Cas9screentargetingall635exonstotestifanyoftheseexonsareoncogenicwhenthey areskipped.
Our finalaim willusehomology-directedCRISPR-Cas9toknock-incandidatesplicemutations andvalidatethatthesemutationscauseexon-skipping,whichleadstoanoncogenicgenealteration. Completionofthisstudywillsettheframeworkformorebroadstudiesofsomaticmutationsthataffectgene functionthroughalternativesplicingandtoinvestigatenoveltargetedtherapies.

Public Health Relevance

Throughcomputationalanalysisoflungcancerdonorsamples,wefoundhundredsofgeneswithalteredgene processing,termedalternativemRNAsplicing,thatcanbeassociatedwithasomaticDNAmutation.Giventhat thecancerbiologyfieldknowsverylittleabouthowalternativesplicingaffectsgenefunction,itisunclearifthese splicingalterationscontributetocancer.WeproposetouseCRISPR-Cas9genomeeditingapproachestotest allsplicingchangesfortheirabilitytocausenormalcellstotransformintocancerouscells?pointingtoapotential genetictargetforfuturetherapies.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21CA238600-01
Application #
9727822
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Schwartz, Elena Ivan
Project Start
2019-02-07
Project End
2021-02-28
Budget Start
2019-02-07
Budget End
2020-02-29
Support Year
1
Fiscal Year
2019
Total Cost
Indirect Cost

  Institution

Name
University of California Santa Cruz
Department
Engineering (All Types)
Type
Biomed Engr/Col Engr/Engr Sta
DUNS #
125084723
City
Santa Cruz
State
CA
Country
United States
Zip Code
95064