Anders Dohlman BD2K-LINCS DCIC Ma’ayan Lab 11/29/16 Predicting Adverse Cardiovascular Events for Tyrosine Kinase Inhibitors from Molecular Features Anders Dohlman BD2K-LINCS DCIC Ma’ayan Lab 11/29/16

Cardiac Effects of TK Inhibition Hypertension high blood pressure Hypertrophy thickened ventricular septum Cardiomyopathy decrease in contractility Ejection fraction decrease less blood leaving heart Pulmonary odema buildup of fluid in the lung

ACEs Bit Matrix + FAERS FDA Drug Labels EBGM score: how much more likely a drug-event pair are to occur together versus the two occurring independently a two sided 90% confidence interval is given, we used an EB05 score of 2 or greater to consider an association ie. “we are 95% sure than is adverse event occurs at least twice as often with the drug than they do independently” FDA Drug Labels

ELISA –– Kinase Inhibition No ACE Decreased Ejection Fraction Cardiac Failure Hypertension ELISA = enzyme immunoassay Measures kinase affinities for small molecules relative to an immobilized ligand Reports as % inhibition of kinase relative to immobilized ligand Data from Uitdehaag et al. (PLoS ONE, 2014)

Random Forest Classification Scheme Reported F1 score is essentially an average of precision & sensitivity precision =  # true positives / # test outcome positives sensitivity = # true positives / # condition positive Not Reported INPUT TKI signatures Molecular features OUTPUT ACE Predictions Feature importance

ELISA Canvas Analysis Any Cardiac Failure Decreased EF Hypertension Least important Most important Tiles are arranged to place co-inhibited kinases adjacently. Each canvas is a torus. Any Cardiac Failure Decreased EF Hypertension PDGFRA PDGFRA_V561D FLT4 KIT EGFR ABL_E255K ABL ARG CDK4/CycD3 LKB1 CDK9/CycT1 CDK3/CycE1 KDR KIT_T670I FLT1 FLT4 RET_M918T

Hypertension is predicted by VEGFRs, cKIT, RET ADR no ADR Hypertension is predicted by VEGFRs, cKIT, RET FLT1 and KDR form a heterodimer PI3K/AKT promotes neural growth PI3K/AKT pathway is involved in Long-term Potentiation (PI3K necessary & sufficient for LTP) % Inhibition FLT1 (VEGFR1) KDR (VEGFR2) KIT –– proto-oncogene implicated in GIST, AML RET –– cadherin family, involved in neuronal navigation / promote angiogenesis & vascular development

Cardiac Failure is predicted by ABLs, cKIT, PDGFR ADR no ADR Cardiac Failure is predicted by ABLs, cKIT, PDGFR EGFR regulates PI3-K-Akt Signaling, STAT, Ras-Raf-Mek-Erk pathways % Inhibition ABL (ABL1) ARG (ABL2) KIT –– proto-oncogene implicated in GIST, AML PDGFRA –– organ development, wound healing, proliferation / NRTKs, regulate cytoskeleton remodeling, motility

LINCS Transcripomics –– L1000 Based on FLEXMAP 3D System by Luminex Measures 1000 “landmark” genes per experiment, and uses these to infer expression of the full transcriptome Minimally redundant Widely expressed across cellular contexts Over 1.5 million gene expression profiles in total 20,413 chemical compounds 22,119 genetic perturbations 77 cell lines profiled 80%

LINCS Transcripomics –– DToxS Seeks to obtain cellular signatures related to adverse effects caused by FDA-approved drugs Four adult PromoCell cardiomyocyte cell lines 38 drugs 570 transcriptional experiments 114 proteomic experiments

Trace of Performance L1000 F1 DToxS F1 Genes

Trace of Performance L1000 F1 ~50 DToxS F1 ~12 Genes

Pathway Analysis Using Gene Set Enrichment ACEs M f Kinase Inhibitors Gene Expression Enrichment Features

Pathway Analysis Using Gene Set Enrichment 500 down 500 up Gene Signature ACEs M f Kinase Inhibitors Gene Expression Enrichment Features

Pathway Analysis Using Gene Set Enrichment 500 down 500 up Gene Signature ACEs M f Kinase Inhibitors Gene Expression Enrichment Features

Pathway Analysis Using Gene Set Enrichment 500 down 500 up Gene Signature Enrichment Vector ACEs M f Kinase Inhibitors Gene Expression Enrichment Features

Pathway Analysis Using Gene Set Enrichment Hypertension Cardiac Failure Decreased Ejection Fraction Indeed, a study of Imatinib suggests that calcium-induced loss of mitochrochondrial membrane potential and a reduction of cellular ATP leads to cell death Gini

Pathway Analysis Using Gene Set Enrichment Hypertension Cardiac Failure Decreased Ejection Fraction Indeed, a study of Imatinib suggests that calcium-induced loss of mitochrochondrial membrane potential and a reduction of cellular ATP leads to cell death Significant enrichment for transmembrane ion channels, particularly Ca2+ ...these channels play a fundamental role in neuromuscular signaling, muscle contraction

Conclusions Screening ELISA binding data to identify kinases predictive of adverse cardiac events Cardiac Failure: ABL1, ABL2 Decreased Ejection Fraction: cyclin-kinase complexes Hypertension: VEGFRs, c-Kit Binding of drug-resistant c-Kit mutants are stronger predictors of hypertension than the c-Kit wild-type ATP-binding site: V654A, T670I Screening of LINCS L1000, DToxS data for genetic biomarkers of adverse cardiac events Peak performance with 12 or 50 genes, respectively Enrichment analysis of predictive genes reveals potential mechanisms of hypertension & cardiac dysfunction Ca2+-induced mitochondrial membrane potential collapse Potentially due to deregulation of PKA in cardiomyocytes F1 score is essentially an average of precision & sensitivity precision =  # true positives / # test outcome positives sensitivity = # true positives / # condition positive

Acknowledgements Avi Ma’ayan Darrell Abernethy Shadia Zaman ... Qiaonan Duan Simon Koplev Zichen Wang The Ma’ayan Lab F1 score is essentially an average of precision & sensitivity precision =  # true positives / # test outcome positives sensitivity = # true positives / # condition positive