title: Development of siRNA payloads to target KRAS-mutant cancer.
abstract: RNAi is a powerful tool for target identification and can lead to novel therapies for pharmacologically intractable targets such as KRAS. RNAi therapy must combine potent siRNA payloads with reliable in vivo delivery for efficient target inhibition. We used a functional "Sensor" assay to establish a library of potent siRNAs against RAS pathway genes and to show that they efficiently suppress their targets at low dose. This reduces off-target effects and enables combination gene knockdown. We administered Sensor siRNAs in vitro and in vivo and validated the delivery of KRAS siRNA alone and siRNA targeting the complete RAF effector node (A/B/CRAF) as promising strategies to treat KRAS-mutant colorectal cancer. We further demonstrate that improved therapeutic efficacy is achieved by formulating siRNA payloads that combine both single-gene siRNA and node-targeted siRNAs (KRAS + PIK3CA/B). The customizable nature of Sensor siRNA payloads offers a universal platform for the combination target identification and development of RNAi therapeutics.
  
  
  CONCLUSIONS
  To advance RNAi therapy for KRAS-mutant cancer, we developed a validated siRNA library against RAS pathway genes that enables combination gene silencing. Using an in vivo model for real-time siRNA delivery tracking, we show that siRNA-mediated inhibition of KRAS as well as RAF or PI3K combinations can impair KRAS-mutant colorectal cancer in xenograft models.
authors.affiliation: Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, CA 94158, USA.
authors.forename: Tina L
authors.initials: TL
authors.lastname: Yuan
authors.affiliation: Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.
authors.forename: Christof
authors.initials: C
authors.lastname: Fellmann
authors.affiliation: Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.
authors.forename: Chih-Shia
authors.initials: CS
authors.lastname: Lee
authors.affiliation: Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, CA 94158, USA.
authors.forename: Cayde D
authors.initials: CD
authors.lastname: Ritchie
authors.affiliation: Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.|Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
authors.forename: Vishal
authors.initials: V
authors.lastname: Thapar
authors.affiliation: Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.
authors.forename: Liam C
authors.initials: LC
authors.lastname: Lee
authors.affiliation: Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.
authors.forename: Dennis J
authors.initials: DJ
authors.lastname: Hsu
authors.affiliation: Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.|Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.
authors.forename: Danielle
authors.initials: D
authors.lastname: Grace
authors.affiliation: Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.
authors.forename: Joseph O
authors.initials: JO
authors.lastname: Carver
authors.affiliation: Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.|Research Institute of Molecular Pathology (IMP), Dr. Bohr-Gasse 7, 1030 Vienna, Austria.
authors.forename: Johannes
authors.initials: J
authors.lastname: Zuber
authors.affiliation: Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA.
authors.forename: Ji
authors.initials: J
authors.lastname: Luo
authors.affiliation: Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, CA 94158, USA.
authors.forename: Frank
authors.initials: F
authors.lastname: McCormick
authors.affiliation: Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.|Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA.|Howard Hughes Medical Institute, New York, NY 10065, USA.
authors.forename: Scott W
authors.initials: SW
authors.lastname: Lowe
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D016000:Cluster Analysis; D004195:Disease Models, Animal; D016503:Drug Delivery Systems; D020869:Gene Expression Profiling; D055785:Gene Knockdown Techniques; D015723:Gene Library; D018014:Gene Transfer Techniques; D011905:Genes, ras; D006801:Humans; D051379:Mice; D009154:Mutation; D053758:Nanoparticles; D009369:Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000008:administration & dosage / Q000235:genetics*; D015203:Reproducibility of Results; D015398:Signal Transduction; D047368:Tumor Burden / Q000235:genetics; D023041:Xenograft Model Antitumor Assays
chemical_list: D034741:RNA, Small Interfering; D019869:Phosphatidylinositol 3-Kinases
references.citation: Mol Pharm. 2009 May-Jun;6(3):659-68
references.pmid: 19267452
references.citation: Cancer Cell. 2011 May 17;19(5):652-63
references.pmid: 21514245
references.citation: Cell. 1992 Sep 18;70(6):923-35
references.pmid: 1356076
references.citation: Cell. 2007 Jun 1;129(5):957-68
references.pmid: 17540175
references.citation: Nat Genet. 2005 Nov;37(11):1289-95
references.pmid: 16200064
references.citation: Nature. 2002 Jul 11;418(6894):244-51
references.pmid: 12110901
references.citation: Clin Cancer Res. 2010 Apr 15;16(8):2450-7
references.pmid: 20332327
references.citation: Pharmaceuticals (Basel). 2013;6(1):85-107
references.pmid: 23667320
references.citation: Cell. 2011 Oct 14;147(2):344-57
references.pmid: 22000013
references.citation: Cancer Cell. 2009 Jun 2;15(6):489-500
references.pmid: 19477428
references.citation: Genes Cancer. 2011 Mar;2(3):359-72
references.pmid: 21779505
references.citation: Cancer Cell. 2013 Nov 11;24(5):617-30
references.pmid: 24229709
references.citation: Nat Rev Drug Discov. 2013 Jan;12(1):35-50
references.pmid: 23274470
references.citation: Nat Struct Mol Biol. 2004 Jul;11(7):599-606
references.pmid: 15170178
references.citation: BMC Bioinformatics. 2006 Nov 30;7:520
references.pmid: 17137497
references.citation: Clin Cancer Res. 2012 Apr 15;18(8):2316-25
references.pmid: 22261800
references.citation: Proc Natl Acad Sci U S A. 2007 Apr 3;104(14):5715-21
references.pmid: 17379663
references.citation: Nat Biotechnol. 2003 Jun;21(6):635-7
references.pmid: 12754523
references.citation: Nat Methods. 2006 Mar;3(3):199-204
references.pmid: 16489337
references.citation: Nat Biotechnol. 2009 Jun;27(6):549-55
references.pmid: 19465925
references.citation: Proc Natl Acad Sci U S A. 2012 Dec 26;109(52):E3659-67
references.pmid: 23236152
references.citation: Cancer Cell. 2013 May 13;23(5):594-602
references.pmid: 23680146
references.citation: Mol Cell. 2011 Mar 18;41(6):733-46
references.pmid: 21353615
references.citation: Nat Rev Mol Cell Biol. 2008 Jul;9(7):517-31
references.pmid: 18568040
references.citation: Proc Natl Acad Sci U S A. 2014 Aug 26;111(34):E3553-61
references.pmid: 25114235
references.citation: Cancer Res. 2005 Oct 1;65(19):8984-92
references.pmid: 16204072
references.citation: Nature. 2010 Mar 18;464(7287):427-30
references.pmid: 20179705
references.citation: Nature. 2010 Apr 15;464(7291):1067-70
references.pmid: 20305636
references.citation: J Thorac Oncol. 2010 Oct;5(10):1630-6
references.pmid: 20802351
references.citation: Cell. 2010 Jan 22;140(2):209-21
references.pmid: 20141835
references.citation: Nature. 2010 Mar 18;464(7287):431-5
references.pmid: 20130576
references.citation: Nat Rev Drug Discov. 2002 Sep;1(9):727-30
references.pmid: 12209152
references.citation: Cancer Discov. 2011 Jul;1(2):128-36
references.pmid: 22043453
references.citation: J Clin Oncol. 2004 Nov 15;22(22):4456-62
references.pmid: 15483017
references.citation: Cell. 2004 Jan 23;116(2):281-97
references.pmid: 14744438
references.citation: Mol Cell. 2013 Feb 21;49(4):751-8
references.pmid: 23352452
references.citation: Nat Cell Biol. 2014 Jan;16(1):10-8
references.pmid: 24366030
references.citation: EMBO J. 2010 Mar 17;29(6):1091-104
references.pmid: 20150892