Table S1: Time-Dependent Cancer Cell Proliferation Inhibition Potency by LY3295668

Table S1: Time-dependent cancer cell proliferation inhibition potency by LY3295668.

Cell proliferation inhibition IC50 (mM) Time (Hours) NCI-H446 Hela Calu6 4 hours 9.269 ± 3.59 1.262 ± 1.06 1.909 ± 0.507 8 hours 1.575 ± 0.306 0.961 ± 0.058 1.059 ± 0.078 16 hours 0.637 ± 0.414 0.487 ± 0.050 0.208 ± 0.030 24 hours 0.415 ± 0.200 0.331 ± 0.054 0.135 ± 0.107 48 hours 0.090 ± 0.021 0.108 ± 0.034 0.076 ± 0.033

Note: The Cell proliferation inhibition IC50 was measured using Cell Titer Glo assay. Table S2 Kinase profile in NCI-H446 cell

LY3295668 Kinase Sequence Labeling Site 1µM ABL, ARG LMTGDTYTAHAGAKFPIK Activation Loop 5.6 ABL, ARG YSLTVAVKTLKEDTMEVEEFLKLys1 -7.4 ACK TVSVAVKCLKPDVLSQPEAMDDFIRLys1 12.4 AGK ATVFLNPAACKGK ATP 17.2 AKT1 GTFGKVILVK ATP Loop -2.5 AKT1, AKT3 YYAMKILK Lys1 -8.7 AKT2, AKT3 GTFGKVILVR ATP Loop -7.1 AMPKa1, AMPKa2 DLKPENVLLDAHMNAK Lys2 -1 ARAF DLKSNNIFLHEGLTVK Lys2 -38.7 ATR FYIMMCKPK ATP 0.7 AurA FILALKVLFK Lys1 88.4 AurA DIKPENLLLGSAGELK Lys2 89.9 AurA, AurB, AurC GKFGNVYLAR ATP Loop -140.7 AurB SHFIVALKVLFK Lys1 -256.5 BRAF DLKSNNIFLHEDLTVK Lys2 -4.9 BRSK1, BRSK2 VAIKIVNR Lys1 11.5 CaMK1a LVAIKCIAK Lys1 7.1 CaMK1d LFAVKCIPK Lys1 -4.4 CaMK2d IPTGQEYAAKIINTKK Lys1 -13 CaMK2g TSTQEYAAKIINTK Lys1 -5.4 CaMK4 DLKPENLLYATPAPDAPLK Lys2 -0.1 CaMKK1 DIKPSNLLLGDDGHVK Lys2 5.9 CaMKK2 DIKPSNLLVGEDGHIK Lys2 3.1 CASK ETGQQFAVKIVDVAK Lys1 -29.4 CCRK DLKPANLLISASGQLK Lys2 18.4 CDC2 DLKPQNLLIDDKGTIK Protein Kinase Domain 12.2 CDK10 DLKVSNLLMTDK Lys2 -8.8 CDK11, CDK8 DLKPANILVMGEGPER Lys2 -5.4 CDK2 DLKPQNLLINTEGAIK Lys2 -0.3 CDK4 DLKPENILVTSGGTVK Lys2 -26.5 CDK5 DLKPQNLLINR Lys2 -22.2 CDK7 DLKPNNLLLDENGVLK Lys2 2.1 CDK9 DMKAANVLITR Lys2 1 CHK1 DIKPENLLLDER Lys2 -8.1 CHK2 DLKPENVLLSSQEEDCLIK Lys2 -17.4 CK1a DIKPDNFLMGIGR Lys2 -33.2 CK1d, CK1e DVKPDNFLMGLGKK Lys2 -12.4 CK1g2 DVKPENFLVGRPGTK Lys2 -8.7 CLK1 LTHTDLKPENILFVQSDYTEAYNPKLys2 20.5 CLK2 LTHTDLKPENILFVNSDYELTYNLEKLys2 31 CSK VSDFGLTKEASSTQDTGKLPVKActivation Loop 2.9 DGKA IDPVPNTHPLLVFVNPKSGGKATP -8.7 DGKH ATFSFCVSPLLVFVNSKSGDNQGVKATP -1.7 DGKQ GRLLTALVLPDLLHAKLPPDSCPLLVFVNPKSGGLKATP -8.5 DNAPK EHPFLVKGGEDLR ATP 7.9 DNAPK KGGSWIQEINVAEK ATP -25.3 eEF2K YIKYNSNSGFVR ATP 4 EGFR IPVAIKELR Lys1 -721.5 EGFR LLGAEEKEYHAEGGKVPIK Activation Loop -980.9 EphA2 VLEDDPEATYTTSGGKIPIR Activation Loop 92 EphA7 VIEDDPEAVYTTTGGKIPVR Activation Loop 19.3 EphB1 YLQDDTSDPTYTSSLGGKIPVRActivation Loop -8.8 EphB2 FLEDDTSDPTYTSALGGKIPIRActivation Loop 10.7 EphB4 FLEENSSDPTYTSSLGGKIPIRActivation Loop 12.6 Erk1 DLKPSNLLINTTCDLK Lys2 -11.6 Erk2 DLKPSNLLLNTTCDLK Lys2 -10.6 Erk3 DLKPANLFINTEDLVLK Lys2 -2.2 Erk5 DLKPSNLLVNENCELK Lys2 -0.2 FAK CIGEGQFGDVHQGIYMSPENPALAVAIKTCKLys1 -2.3 FER TSVAVKTCKEDLPQELK Lys1 -3.1 FGFR1,PDGFRb,FMS,TYRO3VAVKMLK Lys1 46.4 FRAP IQSIAPSLQVITSKQRPR ATP 0.4 FYN, SRC, YES QGAKFPIKWTAPEAALYGR Activation Loop -41.6 GCK DIKGANLLLTLQGDVK Lys2 5.3 GCN2 DLKPVNIFLDSDDHVK Lys2 -3.7 GSK3A DIKPQNLLVDPDTAVLK Lys2 3.5 GSK3B DIKPQNLLLDPDTAVLK Lys2 -1.9 HER3/ErbB3 GVWIPEGESIKIPVCIKVIEDKLys1 -6.5 HER4/ErbB4 GIWVPEGETVKIPVAIKILNETTGPKLys1 10 IKKa DLKPENIVLQDVGGK Lys2 15.4 IKKb DLKPENIVLQQGEQR Lys2 4.1 ILK WQGNDIVVKVLK Lys1 8.1 ILK ISMADVKFSFQCPGR Protein Kinase Domain -7.4 IRAK1 AIQFLHQDSPSLIHGDIKSSNVLLDERLys2 -6.5 IRAK4 DIKSANILLDEAFTAK Lys2 -4.9 IRE1 DLKPHNILISMPNAHGK Lys2 -5.8 JAK1 domain1 QLASALSYLEDKDLVHGNVCTKNLLLARProtein Kinase Domain 14.7 JAK1 domain2 IGDFGLTKAIETDKEYYTVK Activation Loop 22.1 JAK1 domain2 YDPEGDNTGEQVAVKSLKPESGGNHIADLKKLys1 10 JAK2 domain2 IGDFGLTKVLPQDKEYYK Activation Loop -45.2 JNK1, JNK2, JNK3 DLKPSNIVVK Lys2 2.3 JNK2 YQQLKPIGSGAQGIVCAAFDTVLGINVAVKKLys1 -23 KHS1 NVHTGELAAVKIIK Lys1 0.8 KHS2 NVNTGELAAIKVIK Lys1 -8 LATS1 ALYATKTLR Lys1 15.1 LATS2 DIKPDNILIDLDGHIK Lys2 -15.8 LKB1 DIKPGNLLLTTGGTLK Lys2 -3.4 LOK DLKAGNVLMTLEGDIR Lys2 -1.3 LYN VAVKTLKPGTMSVQAFLEEANLMKLys1 -0.8 MAP2K1 IMHRDVKPSNILVNSR Lys2 -28.3 MAP2K1, MAP2K2 KLIHLEIKPAIR Lys1 -2 MAP2K1, MAP2K2 DVKPSNILVNSR Lys2 -10.1 MAP2K3 DVKPSNVLINK Lys2 4.9 MAP2K4 DIKPSNILLDR Lys2 1.8 MAP2K5 DVKPSNMLVNTR Lys2 -4.2 MAP2K6 DVKPSNVLINALGQVK Lys2 6.2 MAP2K7 DVKPSNILLDER Lys2 2.5 MAP3K1 DVKGANLLIDSTGQR Lys2 12.8 MAP3K2 ELAVKQVQFDPDSPETSKEVNALECEIQLLKLys1 9.3 MAP3K2, MAP3K3 DIKGANILR Lys2 4.7 MAP3K3 ELASKQVQFDPDSPETSKEVSALECEIQLLKLys1 0.3 MAP3K4 DIKGANIFLTSSGLIK Lys2 6 MAP3K5 DIKGDNVLINTYSGVLK Lys2 13 MAP3K6 DIKGDNVLINTFSGLLK Lys2 29.9 MARK2, MARK3 DLKAENLLLDADMNIK Lys2 -8.8 MARK3 EVAIKIIDKTQLNPTSLQK Lys1 -11.9 MARK3, MARK4 EVAIKIIDK Lys1 5.4 MARK4 DLKAENLLLDAEANIK Lys2 9.4 MAST1, MAST2 DLKPDNLLITSMGHIK Lys2 8.3 MAST3 DLKPDNLLITSLGHIK Lys2 32.4 MASTL GAFGKVYLGQK ATP Loop -12.9 MASTL LYAVKVVK Lys1 -22.8 MELK DLKPENLLFDEYHK Lys2 -13.4 MER, TYRO3 KIYSGDYYR Activation Loop 23.4 MLK1 DLKSSNILILQK Lys2 12.5 MLK3 DLKSNNILLLQPIESDDMEHKLys2 -1.5 MLK4 DLKSSNILLLEK Lys2 3.6 MPSK1 DLKPTNILLGDEGQPVLMDLGSMNQACIHVEGSRLys2 -8.9 MSK1 domain1 DIKLENILLDSNGHVVLTDFGLSKLys2 5.8 MSK2 domain1 DLKLENVLLDSEGHIVLTDFGLSKLys2 10.7 MST1 ETGQIVAIKQVPVESDLQEIIKLys1 3.4 MST2 ESGQVVAIKQVPVESDLQEIIKLys1 0.6 MST3 DIKAANVLLSEHGEVK Lys2 5.6 MST4, YSK1 DIKAANVLLSEQGDVK Lys2 6.8 MYT1 DTLLALAHLHSQGLVHLDVKPANIFLGPRLys2 0.1 NDR1 DIKPDNLLLDSK Lys2 -3.8 NDR2 DIKPDNLLLDAK Lys2 16.9 NEK1 DIKSQNIFLTK Lys2 -5.3 NEK2 DLKPANVFLDGK Lys2 -12.6 NEK3 SKNIFLTQNGK Activation Loop -13.1 NEK4 DLKTQNVFLTR Lys2 7.8 NEK6 TVALKK Lys1 2.6 NEK6, NEK7 DIKPANVFITATGVVK Lys2 -10.1 NEK7 AACLLDGVPVALKK Lys1 -1.7 NEK8 DLKTQNILLDK Lys2 0.5 NEK9 DIKTLNIFLTK Lys2 11 NLK DIKPGNLLVNSNCVLK Lys2 -16.6 NuaK1 VVAIKSIR Lys1 -0.4 NuaK2 LVAIKSIR Lys1 -14.7 OSR1 DVKAGNILLGEDGSVQIADFGVSAFLATGGDITRLys2 11.2 p38a DLKPSNLAVNEDCELK Lys2 -3.8 p38a QELNKTIWEVPER Protein Kinase Domain -14.6 p38d, p38g DLKPGNLAVNEDCELK Lys2 -12.7 p70S6K DLKPENIMLNHQGHVK Lys2 -4.9 p70S6Kb DLKPENIMLSSQGHIK Lys2 -6 PAK2 IGQGASGTVFTATDVALGQEVAIKQINLQKLys1 -40.3 PAK2 KNPQAVLDVLKFYDSNTVK Protein Kinase Domain -6.4 PAN3 VMDPTKILITGK Protein Kinase Domain -4.8 PCTAIRE1 SKLTDNLVALKEIR Lys1 -14.7 PCTAIRE2, PCTAIRE3 SKLTENLVALKEIR Lys1 17.8 PEK DLKPSNIFFTMDDVVK Lys2 3.3 PFTAIRE1 LVALKVIR Lys1 3.2 PHKg1 DLKPENILLDDNMNIK Protein Kinase Domain 14.5 PI4K2B SEEPYGQLNPKWTK ATP 4 PI4KA, PI4KAP2 SGTPMQSAAKAPYLAK ATP 6.8 PI4KB VPHTQAVVLNSKDK ATP -8.9 PIK3C2B VIFKCGDDLRQDMLTLQMIRATP -2.3 PIK3C3 TEDGGKYPVIFKHGDDLR ATP 7.6 PIK3CA RPLWLNWENPDIMSELLFQNNEIIFKNGDDLRQDMLTLQIIRATP -40.9 PIK3CB VFGEDSVGVIFKNGDDLRQDMLTLQMLRATP -11.9 PIP4K2A AKELPTLKDNDFINEGQK ATP -8.8 PIP4K2B AKDLPTFKDNDFLNEGQK ATP 1.5 PIP4K2C TLVIKEVSSEDIADMHSNLSNYHQYIVKATP -16.5 PIP5K3 GGKSGAAFYATEDDRFILK ATP 4.2 PITSLRE DLKTSNLLLSHAGILK Lys2 8.9 PKCi IYAMKVVK Lys1 -6.9 PKD1 DVAIKIIDK Lys1 3 PKD2 DVAVKVIDK Lys1 -10.9 PKD3 DVAIKVIDK Lys1 -16.7 PKR DLKPSNIFLVDTK Lys2 0.3 PLK1 CFEISDADTKEVFAGKIVPK Lys1 -252.3 PRP4 CNILHADIKPDNILVNESK Lys2 -3.3 PRPK FLSGLELVKQGAEAR ATP Loop -9.9 RAF1 DMKSNNIFLHEGLTVK Lys2 -10.9 RIPK1 DLKPENILVDNDFHIK Lys2 15.8 ROCK1 KLQLELNQER Protein Kinase Domain 5.2 ROCK1, ROCK2 DVKPDNMLLDK Lys2 -4 RSK1 domain1 DLKPENILLDEEGHIKLTDFGLSKEAIDHEKLys2 -11.9 RSK1 domain1, RSK2 domain1,DLKPENILLDEEGHIK RSK3 domain1 Lys2 1.7 RSK1 domain2 DLKPSNILYVDESGNPECLR Lys2 1.4 RSK2 domain1 DLKPENILLDEEGHIKLTDFGLSKESIDHEKLys2 -19.5 RSK2 domain2 DLKPSNILYVDESGNPESIR Lys2 -4.7 RSK3 domain1 DLKPENILLDEEGHIKITDFGLSKLys2 -0.4 RSKL1 VLGVIDKVLLVMDTR ATP -3.6 SGK3 FYAVKVLQK Lys1 4.6 SLK DLKAGNILFTLDGDIK Lys2 7 SMG1 DTVTIHSVGGTITILPTKTKPKATP -7.2 SNRK DLKPENVVFFEK Lys2 2.3 SRC VAIKTLKPGTMSPEAFLQEAQVMKKLys1 -25.8 SRPK1 IIHTDIKPENILLSVNEQYIR Lys2 -4.9 SRPK1, SRPK2 FVAMKVVK Lys1 10.9 STK33 DLKLENIMVK Lys2 -1 STLK3 DLKAGNILLGEDGSVQIADFGVSAFLATGGDVTRLys2 15.2 STLK5 YSVKVLPWLSPEVLQQNLQGYDAKActivation Loop -37.7 TAK1 DLKPPNLLLVAGGTVLK Lys2 9.8 TAO1, TAO3 DIKAGNILLTEPGQVK Lys2 11.7 TAO2 DVKAGNILLSEPGLVK Lys2 19.7 TBK1 TGDLFAIKVFNNISFLRPVDVQMRLys1 -0.7 TLK1 YLNEIKPPIIHYDLKPGNILLVDGTACGEIKLys2 -9.2 TLK2 YLNEIKPPIIHYDLKPGNILLVNGTACGEIKLys2 -11.2 TNK1 SVPVAVKSLR Lys1 -15.8 TYK2 domain2 IGDFGLAKAVPEGHEYYR Activation Loop -3.1 ULK1 DLKPQNILLSNPAGR Lys2 22.1 ULK2 DLKPQNILLSYANR Lys2 17.7 ULK3 NISHLDLKPQNILLSSLEKPHLKLys2 -11.9 VACAMKL NYNQPSEVTDRYDLGQVIKTEEFCEIFRATP Loop -3 VRK2 MLDVLEYIHENEYVHGDIKAANLLLGYKLys2 5.3 Wee1 YIHSMSLVHMDIKPSNIFISR Lys2 -5 Wnk1, Wnk2 GSFKTVYK ATP Loop 11.3 Wnk1, Wnk2, Wnk3 DLKCDNIFITGPTGSVK Lys2 18.5 YANK3 DVKPDNILLDER Lys2 7.7 YSK1 EVVAIKIIDLEEAEDEIEDIQQEITVLSQCDSPYITRLys1 -16.5 ZAK WISQDKEVAVKK Lys1 -14.1 ZC1/HGK TGQLAAIKVMDVTEDEEEEIKLEINMLKKLys1 -20.4 ZC1/HGK, ZC2/TNIK, ZC3/MINKDIKGQNVLLTENAEVK Lys2 -4.8 ZC2/TNIK TGQLAAIKVMDVTGDEEEEIKQEINMLKKLys1 -28.4 Lys1 Conserved Lysine 1 Lys2 Conserved Lysine 2 ATP Loop ATP binding loop, Activation Loop Activation loop ATP ATP site in non-canonical kinase (e.g. lipid kinase) Protein Kinase DomainOther lysine within kinase domain, possibly not in ATP binding site Other Labeling of residue outside of the protein kinase domain, possibly not in ATP binding site

>90% Inhibition 75 - 90% Inhibition 50 - 75% Inhibition 35 - 50% Inhibition No change 50 -100% increase in MS signal > 100% increase in MS signal Not colored Variable or weak data not reaching significance despite >35% inhibition Table S3 Kinase profile in NCI-H446 xenograft LY3295668 50 LY3295668 Kinase mg/kg 3hour 50mg/kg 24hour ABL, ARG 41.35 -8.05 ACK 20.3 11.1 AGK -13.4 -23.15 AKT1 1.6 4.75 AKT1, AKT3 9.55 24.05 AKT2, AKT3 4.7 12.8 AMPKa1, AMPKa2 -27.5 -0.75 ARAF 22.85 -95.25 ATR -17.05 -6.45 AurA 191.2 157.6 AurA, AurB, AurC 62.8 -3.15 AurB 59.85 -80.75 BRAF -4.75 16.15 CaMK1a 13.6 6.25 CaMK1d 10.1 -5.6 CaMK2b 6.25 -8.7 CaMK2d 9.5 -30 CaMK2g 15.65 -9.45 CaMK4 36.15 22.65 CaMKK1 15.85 7.3 CaMKK2 -0.35 7.55 CASK -6.6 -7.95 CCRK 3.35 -4.8 CDC2 26.9 -12.85 CDK10 -4 3.95 CDK11, CDK8 -33 19.3 CDK2 -1.6 -18.75 CDK4 -0.55 21.5 CDK5 6.8 -2 CDK7 5.4 10.5 CDK9 -20.55 7.6 CHK1 22.25 40.9 CHK2 -4.35 -5.95 CK1a 0.4 9.85 CK1d, CK1e -7.8 -3.7 CRK7 -13.7 3.2 CSK -5.1 -7.75 DGKA -9.8 -19 DGKH -21.25 3.4 DLK 8.35 16.45 DNAPK 10.95 -7.05 eEF2K 15.5 19.4 EGFR -8.1 0.35 EphA7 5.85 10.95 EphB1 -5.35 -22.65 EphB2 -34.85 -22.95 EphB4 -6.1 5.8 Erk1 -9.15 -2.55 Erk2 -23.75 -7.8 Erk5 -12.5 6.35 FAK -33.95 -6.45 FER -2.15 -15.1 FRAP 1.95 -27.95 FYN, SRC, YES 1.75 3.65 GCK -26.65 18.75 GCN2 -14.25 -12.95 GSK3A 3.5 -4.8 GSK3B 12.65 -0.25 HER4/ErbB4 -11.4 -1.05 IKKa -48.05 -34.35 IKKb -20.25 -9.95 ILK -59.15 -51.1 IRAK1 -25.75 -0.3 IRAK4 -24.35 20.5 IRE1 1.65 -7.55 ITPK1 26.2 -24.7 JAK1 domain1 -17.6 -8.7 JAK1 domain2 3.8 -26.95 JNK1, JNK2, JNK3 11.1 12.45 KHS1 4.15 -9.05 KHS2 -10 -18.6 KSR1, KSR2 2.45 4.7 LATS1 -2.65 5.5 LATS2 5.2 2.5 LKB1 -4.3 0.4 LOK -9.3 5.15 LRRK2 20.8 -9.85 MAP2K1 20.9 -13.1 MAP2K1, MAP2K2 -21.35 -13.45 MAP2K3 2.45 6.3 MAP2K4 -30.25 -15.4 MAP2K5 -38.05 1.5 MAP2K6 -37.05 -6.1 MAP2K7 -0.5 -1.55 MAP3K1 -7.5 10.4 MAP3K2 5.65 11.85 MAP3K2, MAP3K3 -22.15 23.9 MAP3K3 9.9 2.2 MAP3K4 -176.1 -55.15 MAP3K5 5.7 12.25 MAP3K6 -2.8 16.15 MARK1 22.35 -5.5 MARK2, MARK3 15.85 17.3 MARK3 12.95 9.85 MARK3, MARK4 1.85 12.2 MARK4 11.1 5.2 MAST1, MAST2 -16.35 3.85 MAST3 7.9 25.85 MAST4 -6.2 -10.95 MASTL 30.05 -5.55 MELK 16.7 -0.05 MLK3 -14.05 10 MLK4 -30.65 8.4 MPSK1 7.35 20.3 MRCKb -6.05 -14.4 MSK1 domain1 10.8 2.4 MSK2 domain1 43.05 18.6 MST1 7.5 -2.5 MST2 8.45 8.15 MST3 -20.5 6.2 MST4, YSK1 -26.2 17.75 NDR1 -1.85 10.35 NDR2 -27.05 11.75 NEK1 -34.5 -11.9 NEK2 15.4 -13.85 NEK3 -2.3 -6.05 NEK4 11.95 -6.85 NEK6 12.1 25.1 NEK6, NEK7 -5.9 -7.65 NEK7 -7.35 -11.9 NEK8 -48.6 -15.15 NEK9 -7.8 -5.45 NLK 18.6 7.2 NuaK1 1.95 11.7 NuaK2 3.95 9.35 OSR1 -51.7 18.4 p38a -5.35 -4.45 p38d, p38g -25.45 -0.95 p70S6K -3.55 11.8 p70S6Kb 17.5 9.6 PAK2 -3.8 -16.95 PAN3 -10.55 13.9 PCTAIRE1 -1.5 -5.6 PCTAIRE2, PCTAIRE3 19 16.75 PDK1 2.4 -13.25 PEK -4.35 6.05 PFTAIRE1 -7.2 -13.45 PHKg2 -11.15 9.3 PI4KA, PI4KAP2 -7.85 7.6 PI4KB -11.3 -3.6 PIK3C2B -11.4 4.95 PIK3C3 9.05 -15.65 PIK3CB -44.75 -21.35 PIP4K2A 0.25 -33.55 PIP4K2B 2.85 -22.8 PIP4K2C 5.7 0.85 PIP5K3 6.05 -2.1 PITSLRE 20.35 7.2 PKCi 16.8 5.6 PKD1 -9.45 6.95 PKD2 -24.95 -3.75 PKR -4.6 8.35 PLK1 -5 -163.15 PRP4 -7.35 4.25 PRPK 12.85 -4.9 ROCK1 2.15 -14.7 ROCK1, ROCK2 9.85 3.55 RSK1 domain1 -11.4 -50.5 RSK1 domain1, RSK2 domain1,4.7 RSK3 domain1-11.75 RSK1 domain2 -35.2 -13.4 RSK2 domain1 -10.35 -36.45 RSK2 domain2 -13.95 -1.4 RSK4 domain1 22.6 -8.1 RSKL1 -21.55 -24.45 SGK3 23.35 15.6 SLK -22.15 8.1 SMG1 0.4 -30.7 smMLCK 6.95 12.85 SNRK -45.15 -6.15 SRPK1 14.7 -17 SRPK1, SRPK2 6.7 -13 STK33 -27.15 6.15 STLK3 -32.3 30.5 STLK5 -14.55 -3.7 SYK 8.7 14.25 TAK1 4.3 6.65 TAO1, TAO3 -2.3 21.35 TAO2 -19.7 11.95 TBK1 -54.55 -42.95 TLK1 -0.75 -7.5 TLK2 -8.05 -11.05 TYK2 domain2 6.3 -2.3 ULK1 -26.85 -0.2 ULK2 -9.05 -3.45 ULK3 -6.5 -1.45 VACAMKL 0.5 -7 Wee1 -1.3 6.9 Wnk1, Wnk2 -18.05 -6.25 Wnk1, Wnk2, Wnk3 -53.1 -20.45 YANK3 4.3 10.15 YSK1 19.4 -29.05 ZAK -18.6 -1.65 ZC1/HGK -8.8 -22.2 ZC1/HGK, ZC2/TNIK, ZC3/MINK-22.7 7.55 ZC2/TNIK -20.1 -8.65 Table S4 Kinase profile in mouse spleen LY3295668 50 LY3295668 50 Kinase mg/kg 3hr mg/kg 24hr AurA 99.9 93.5 CDC2 27.6 42.65 MASTL 36.3 38.65 PLK1 -16.55 34.02 CHK2 28.45 47.55 CDK2 24 45.5 CHK1 15.6 45.15 AurB -3.65 44.25 HRI 36.8 39.8 LATS1 32.7 37.2 PKR 14.9 35.55 FER 13.1 26 NDR2 14.45 25.9 TLK1 1.85 22.6 eEF2K 1.05 22.3 AMPKa1 9.5 20.3 CDK6 -0.3 18.9 FAK -19.85 18.05 MARK3 11.25 18 CDK4 3.75 15.9 NDR1 -1.7 15.45 AMPKa1, AMPKa2 4.5 15.05 CK1a -2.75 15.05 MAP2K4 -2.35 14.9 KHS1 -1.05 14.45 NEK6 5.05 13.7 ACK 15.5 13.3 LKB1 6.85 13.2 EGFR -0.65 11.75 ATR -5.35 11.75 ULK1 20.65 11.35 CHED 23.75 10.75 QSK 16.45 10.75 IRE1 -0.75 10.7 MAP3K4 10.75 10.55 RSK1 domain2 0.25 10.15 ROCK1 0.5 10.1 MER, TYRO3 -11.45 9.8 ROCK1, ROCK2 -6.3 9.3 CK2a1 7.8 9.25 FES -3.25 9.25 PCTAIRE2, PCTAIRE3 -0.4 8.85 MAP2K3 9.9 8.55 VRK2 21.35 8.5 CaMKK2 0.4 8.25 Wnk1, Wnk2, Wnk3 4.25 7.85 CDK7 9.45 7.85 MAPKAPK2 13.05 7.3 Erk2 6.85 7.25 MAP2K2 -1.45 7.25 PIK3CG -4.9 7.05 MAP3K2 5.2 6.85 TAK1 11.05 6.3 p70S6K 8.25 6.1 ZAK 6.1 6.05 GSK3A -4.05 6 MER -10.3 5.95 ZAP70 13.3 5.75 MARK3, MARK4 -0.25 5.6 MARK2 11.4 5.55 TLK1, TLK2 5.05 4.75 Wee1 -1.9 4.5 Wnk1, Wnk2 -5.05 4.4 Wnk1, Wnk2, Wnk4 4.2 4.35 smMLCK -14.7 4.3 MAP3K5, MAP3K6 3.35 4.2 MARK1, MARK2, MARK3 5.1 3.5 OSR1 9.55 3.35 TAO2 -15.55 2.65 Erk1 0.4 2.65 LATS2 3.1 2.5 SLK 11.7 2.35 IRAK1 14.55 2.25 PAK2 4.6 2.2 CASK -10.5 2.2 LYN -12.5 2.15 MAP3K3 4.7 2 PIP5K3 -1.6 1.05 NEK8 12.35 0.25 MAP2K1, MAP2K2 -2.45 0.25 MAP2K1 -10.55 0.05 MAP3K5 1.25 0 PRP4 -10.2 -0.05 CDK9 1.7 -0.25 NEK1 7.6 -1.2 MST2 2.95 -1.2 GSK3B -13 -1.6 Erk5 -6.85 -1.7 RSKL1 domain1 11.55 -1.75 RSK2 domain2 -8.95 -1.9 BRAF 16.05 -2 MLK3 -4.2 -2.1 RIPK3 13.7 -2.45 HCK -10.55 -2.75 STLK5 10.45 -2.95 NuaK1 -0.95 -3.1 NEK9 15.5 -3.2 PCTAIRE1, PCTAIRE2, PCTAIRE3-2.35 -3.8 MAP3K1 5.3 -4.05 CaMK1a -0.75 -4.15 IKKe 15.2 -4.25 MAP2K7 9.85 -4.65 NEK4 6.55 -4.95 PKD2 -9 -5.05 PITSLRE -22.45 -5.25 JAK1 domain2 -9.05 -5.3 YSK1 9.45 -5.55 PKD3 2.55 -5.6 FGR -12.85 -5.8 SMG1 -4.4 -5.85 CaMK1d -4.7 -6.1 RSK1 domain1, RSK2 domain1, -8.6RSK3 domain1 -6.1 RSK3 domain1 1.1 -6.2 MAP2K6 -10.2 -6.25 MLKL -9.6 -6.5 PYK2 22 -7.4 JAK1 domain1 4.65 -7.7 PIK3CD -12.55 -7.75 CDK11, CDK8 -5.85 -7.8 ANPb 9.35 -8 CCRK -5 -8.25 CaMK2g -20 -8.3 SGK3 2.1 -8.5 ABL, ARG 14.7 -8.7 MAP3K2, MAP3K3 3.4 -8.7 MAP2K5 -1.7 -8.9 PIP4K2A -14.5 -9.55 p38a -10.3 -10.25 PIP4K2B -3.3 -10.4 PIK3CB -13.05 -11 TAO1, TAO3 -14.75 -11.25 p70S6Kb 13.2 -11.7 p38d, p38g -17.3 -11.85 IRAK4 12.85 -12 IKKe, TBK1 -5.25 -13.4 PIK3C3 -11.5 -13.85 JNK1, JNK2, JNK3 -4.15 -14.1 BARK1 -11 -14.1 SRPK1, SRPK2 -30.2 -14.45 PIP4K2C -0.6 -14.5 KHS2 -12.9 -15.05 PI4KA -18.1 -15.15 ILK 15.55 -15.25 CaMK2a, CaMK2b, CaMK2d, CaMK2g-7.75 -15.5 SNRK -12.15 -15.65 LOK 1.55 -15.95 FYN, SRC, YES -2.7 -16 MST3 -25.2 -16.1 AXL -3.85 -16.25 MST1, MST2 -8.7 -16.35 MST4, YSK1 -3.3 -16.45 p38b -14.45 -17.15 CSK -14.65 -17.25 TBK1 -7.9 -17.35 PI4KB -2.85 -17.4 PIK3CA -16.1 -18.5 PFTAIRE1 -15.7 -18.55 CaMK2d -5.05 -18.6 GCK -3.5 -18.7 MST1 -3.05 -18.75 AKT1 -5.9 -18.8 MAST4 -11.3 -19.35 CDK5 1.45 -19.75 PKD1, PKD2 -16.95 -19.85 NEK7 -1.8 -20.05 HPK1 -1.4 -20.95 ZC1/HGK, ZC2/TNIK, ZC3/MINK-19.85 -21.95 KSR1, KSR2 4.55 -22.5 BTK -10.95 -22.75 MAST3 -23.85 -23.15 IRAK3 16.85 -23.6 CaMK4 -12.55 -23.6 FRAP -10.65 -24.25 SYK -13.9 -29.2 RSK1 domain1 -24.85 -29.75 ULK3 -4.3 -34.8 NuaK2 -9.15 -38.6 LRRK2 -16.25 -40.25 RSK2 domain1 -46 -45.35 GCN2 domain2 -60.4 -61.05 MSK2 domain1 -50.75 -77.15 ARAF 21.45 -326.3 Table S5 Biochimical and cell assay correlation Variable by Variable Count Spearman ρ Log (Aurora B biochemical assay IC50) Log (Aurora A biochemical assay IC50) 2940 0.5258 Log (Aurora A cell-based assay IC50) Log (Aurora A biochemical assay IC50) 496 0.7264 Log (Aurora A cell-based assay IC50) Log (Aurora B biochemical assay IC50) 477 0.5597 Log (Aurora B cell-based assay IC50) Log (Aurora A biochemical assay IC50) 452 0.3241 Log (Aurora B cell-based assay IC50) Log (Aurora B biochemical assay IC50) 728 0.7139 Log (Aurora B cell-based assay IC50) Log (Aurora A cell-based assay IC50) 365 0.6253 Log (Mitotic arrest EC50) Log (Aurora A biochemical assay IC50) 706 0.5999 Log (Mitotic arrest EC50) Log (Aurora B biochemical assay IC50) 663 0.5406 Log (Mitotic arrest EC50) Log (Aurora A cell-based assay IC50) 360 0.8901 Log (Mitotic arrest EC50) Log (Aurora B cell-based assay IC50) 365 0.6889