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Bardenhagen, Jennifer

Bradley, Michael

Bristow, Christopher A

Carroll, Christopher L

Carugo, Alessandro

Conner, Michael

Cross, Jason

Di Francesco, M. Emilia

Feng, Ningping

Fnu, Sonal

Gao, Guang

Geck Do, Mary

Han, Jing

Harris, Angela

He, Meng

Heffernan, Timothy P

Jiang, Yongying

Jones, Philip

Joseph, Sarah

Kang, Zhijun

Kovacs, Jeffrey

Le, Kang

Leonard, Paul

Linares, Jennifer

Lorca, Sabina A

Lucas, Reagan

Mahaeshwari, Sweta

Marszalek, Joe

Mullinax, Robert A

Nguyen, Phuong

Peoples, Michael

Perry, Teresa

Quill, Thomas M

Ramaswamy, Vijayan

Reiling, Jan

Robinson, Frederick

Rodriguez, Christian

Rogers, Norma

Schmidt, Stephanie T, PhD

Seth, Sahil

Shepard, Hannah E

Soth, Michael

Spencer, Nakia D

Suarez, Catalina

Uprety, Nadima

Wu, Qi

Xu, Alan

Yu, Simon S

English

OpenWorks @ MD Anderson

Conclusions• CDK11 was identified from an in vivo functional genomics screenas a novel therapeutic target in PCAD patients.• Pharmacological inhibition with potent and selective toolcompounds revealed a disconnect with genetic validation data.• Minimal tumor growth inhibition was observed in mouse efficacystudies as exposures were limited due to toxicity.• Taken together, our multidisciplinary approach to evaluate geneticand pharmacological inhibition of CDK11 suggests that while thereis a dependency of a subset of tumor cells on CDK11, systemicinhibition of CDK11 in a whole organism is not tolerated, therebylimiting the therapeutic potential.Multi-disciplinary efforts to evaluate the therapeutic potential of CDK11, a novel transcription associated cyclin dependent kinase.Christopher L. Carroll1, Jan Reiling2, Jeffrey J. Kovacs2, Michael Bradley1, Meng He2, Nadima Uprety2, Michael Peoples2, Frederick S. Robinson2, Alessandro Carugo2, Michael Conner1, Christian Rodriguez1, Jennifer Bardenhagen1, Vijayan Ramaswamy1, Paul G. Leonard1, Zhijun Kang1, Catalina Suarez1, Teresa L. Perry2, Jing Han2, Sonal Fnu2, Nakia D. Spencer2, Guang Gao2, Angela L. Harris2, Alan Xu1, Jennifer Linares1, Thomas M. Quill1, Phuong K. Nguyen1, Qi Wu1, Stephanie T. Schmidt2, Sahil Seth2, Christopher A. Bristow2, Norma E. Rogers2, Simon S. Yu1, Hannah E. Shepard1, Robert A. Mullinax2, Sabina A. Lorca2, Kang Le1, Reagan Lucas1, Sweta Mahaeshwari1, Sarah Joseph3, Ningping Feng2, Yongying Jiang1, Joe Marszalek2, Michael J. Soth1, Jason B. Cross1, Mary K. Geck Do1, M. Emilia Di Francesco1, Timothy P. Heffernan2, Philip Jones11IACS (Institute for Applied Cancer Science); 2TRACTION (Translational Research to AdvanCe Therapeutics and Innovation in Oncology), University of Texas MD Anderson Cancer Center, Houston, Texas 77054, United States;3Department of Genomic Medicine, University of Texas MD Anderson Cancer Center, Houston, Texas 77030, United StatesPILOT in vivo functional genomics screen0 3 6 9 1 2 1 5 1 8 2 1 2 4 2 7 3 0 3 3 3 6 3 9 4 201 0 02 0 03 0 04 0 05 0 06 0 07 0 08 0 09 0 0P A T C 1 2 4D a y s  o n  T re a tm e n tTumor volume (mm3)s h L u c  O ff  D O Xs h L u c  O n  D O Xs h 3  O ff  D O Xs h 3  O n  D O X0 3 6 9 12 15 18 21 24 27 30020040060080010001200140016001800PATC53Days on TreatmentTumor volume (mm3)shLUC off doxshLUC ON doxsh3 off doxsh3 on DOXProfiles of in vivo tool compoundsPotent and selective tool compounds with suitablepharmacokinetic properties for oral or intraperitoneal (IP)dosing were identified.Compounds are low nanomolar in biochemical andcellular target engagement assays and induce a potentanti-proliferative response in PATC53 cells. Bothcompounds exhibit selectivity over the CDK family andthe broader kinome (>100x over kinome panel).Series 2 compound, IACS-054647, exhibits an excellentin vitro and in vivo pharmacokinetic profile, suitable fororal dosing in mouse models.Series 3 compound, IACS-074527, suffers from poorpermeability and low solubility, limiting exposures withoral dosing. However, low metabolic clearance allows forexcellent exposure with IP dosing.Identification of small molecule inhibitorsGiven the biological findings, CDK11 represented anopportunity to develop a first-in-class small moleculeinhibitor.A screening funnel was established to enable thedevelopment of high quality tool compounds.Two chemically distinct leads were identified from ahigh-throughput screening campaign. Weekly profiling ofanalogs in activity assays and in vitro pharmacokineticassays allowed for rapid optimization.Select compounds were then profiled in the PATC53cellular phenotypic assay, kinase selectivity panels andmouse pharmacokinetic studies to identify in vivo toolcompounds.Differential regulation of cell cycle PATC53shLUC37.2 % G143.4 % S16.7 %G2PATC53shP234.8 % G136.4 % S25.7 %G2PATC53sh339.8 % G129.7 % S28.8 %G2PATC53shP2+rescue36.2 % G145.5 % S16.3 %G2PATC53sh3+rescue42.6 % G136.4 % S19.9 %G2PATC124shLUC41.2 % G145.9 % S11.0 %G2PATC124shP246.0 % G138.4 % S13.9 %G2PATC124sh347.3 % G136.0 % S14.0 %G2G1SG2G1SG2RNRSelective alterations of replication-dependent histone gene expressionNRRNo Dox Dox No Dox1 20.00.51.01.5Relative colony formationshlucshlucsh3sh31 20.00.51.01.5Relative colony formationshlucshlucsh3No Doxsh3Doxshlucshlucsh3No Doxsh3Doxshlucsh3shlucsh30.00.51.01.5PATC69Relative colony formationshlucshlucsh3sh3Doxshlucsh3shlucsh30.00.51.01.5Relative colony formationPATC124Pleiotropic functions of CDK11Mechanism of action studies uncovered pleiotropic functionsof CDK11, including time dependent impact on RNAmetabolism, altered cell cycle function, and selectivealteration of replication-dependent histone gene expression.Identification of CDK11 as a novel therapeutic targetWe previously employed our in vivo target discovery platform, PILOT(Patient-based In vivo Lethality to Optimize Treatment), to identifynew vulnerabilities that can be exploited therapeutically to optimizecancer treatment. Leveraging the PILOT platform, cyclin dependentkinase 11 (CDK11) was identified as a top-scoring hit from an in vivofunctional genomics screen across multiple patient-derived PDACxenograft models.Differential response across PDAC cell linesGenetic disruption of CDK11 expression induced a differentialresponse across a panel of PDAC cell lines in vitro. Knockdown ofCDK11 inhibited the growth established xenografts in vivo.Kinase activity is critical to rescue CDK11 knockdown in cellsOverexpression of CDK11 wild type (WT) but not kinase defective mutants (562 and 580) rescue the growth inhibitory effect of CDK11 knockdown in PDAC cell lines.CDK11 knockdown alters splicing patterns Significant increase of differential splicing events (q < 0.05) in PATC530.0001 0.01 1020406080100IACS-54647 [M]survival (AU) PATC53PATC153PATC124PATC1480.0001 0.01 1020406080100IACS-74418 [M]survival (AU) PATC53PATC153PATC124PATC148Disconnect with pharmacological inhibitionPharmacological inhibition of CDK11 induced a potent anti-proliferative response across a panel of PDAC cell lines, in contrast to the differential response observed with genetic disruption. (IACS-074418 is closely related to IACS-074527 with a comparable profile) 0 5 10 15 20-30-20-10010Treatment (Days)Body weight change (%)Mean +/- SEM0 5 10 15 20-30-20-10010Treatment (Days)Body weight change (%)Mean +/- SEMVehicle, IP, QDIACS-074527, 40 mpk IP, QDIACS-074527, 5 mpk IP, QDDrug discovery screening funnelIACS-054647(Series 2)IACS-074527(Series 3)CDK11A/CycD3 IC50 (M) 0.006 0.001CDK11A/CycL2 Cell IC50 (M) 0.013 0.018PATC53 Phenotypic EC50 (M) 0.303 0.320CDK9/T1 2 mM ATP IC50 (M) 3.6 2.5CLint mouse LM (mL/min/kg) 53 15MDCK Papp A-B (x10-6 cm/s) 19 <0.4MDR1 ER ((B-A)/(A-B)) 1.9 >90Solubility pH = 7.4 (M) 62 16CLp, IV 1 mg/kg (mL/min/kg) 26.6 22.2Vss, IV 1 mg/kg (L/kg) 8.3 13.2T1/2, IV 1 mg/kg (h) 5.8 10.3Cmax, 10 mg/kg (M) 2.81 ± 0.773† 4.07 ±1.21‡AUClast,10 mg/kg (h*M) 25.8† 8.13‡F, 10 mg/kg (%) 135† 60.5‡In VitroMouse PK†Oral dosing; ‡Intraperitoneal dosing*40 mpk dose not tolerated (required dosing holidays for days 8 – 15) *20 mpk dose not tolerated (4 mice required dosing holidays) 0 5 10 15 20050010001500Treatment (Days)Tumor volume (mm3)(mean +/- SEM)VehicleIACS-054647, 20 mpk PO, QD*IACS-054647, 10 mpk PO, QD0 5 10 15 20050010001500Treatment (days)Tumor volume (mm3 )Mean +/- SEMVehicle, IP QDIACS-074527, 40 mpk IP, QD*IACS-074527, 5 mpk IP, QDNo differential response observed in PDAC panelMinimal tumor growth inhibition observed at tolerated doses in PATC53 mouse efficacy modelsSelectivity AssayCDK9/CyclinT1 LANCECellular Target Engagement AssayCDK11A/CyclinL2 NanoBRET(HEK293)Cellular Phenotypic AssayCyQuant (PATC53)Biochemical AssayCDK11A/CyclinD3 RDAIC50 ≤0.1 MWeeklyIC50 ≤0.2 MIC50 ≤0.5 M, >100x selective%F >50In Vitro Pharmacokinetics LM’s, PPB, Sol.Mouse Pharmacokinetics Selectivity PanelsCDK family/KinomeIn Vivo tool compoundsKinase selectivity panels Compounds were screened in a select panel of kinases, including the entire CDK family, based on full kinome scans with representative compounds in each series.  Gene modulation in PATC53• 22 genes with 2 fold decrease  • 44 genes with 1.5 fold decreaseIACS-054647CDK9/CycT1 151xCDK7/CycH/MAT1 902xCDK13/CycK 1486xCDK12/CycK 1653xCDK10/CycQ 2338xAll other CDKs >5000HIPK4 317xMEK2 1319xIACS-074527CDK13/CycK 272xCDK9/CycT1 332xCDK10/CycQ 398xCDK12/CycK 414xCDK16/CycY 506xAll other CDKs >800xTAOK2 284xERK7 482xFold selectivity based on IC50 values Fold selectivity based on IC50 valuesKinase-deadKinase-deadIACS-054647 mouse efficacy/tolerability studyBody weight changeBody weight changeIACS-074527 mouse efficacy/tolerability study

Multi-disciplinary efforts to evaluate the therapeutic potential of CDK11, a novel transcription associated cyclin dependent kinase.

Christopher L. Carroll

Jan Reiling

Jeffrey Kovacs

Michael Bradley

Meng He

Nadima Uprety

Michael Peoples

Fredierick Robinson

Alessandro Carugo

Michael Conner

Christian Rodriguez

Jennifer Bardenhagen

Vijayan Ramaswamy

Paul Leonard

Zhijun Kang

Catalina Suarez

Teresa Perry

Jing Han

Sonal Fnu

Nakia D. Spencer

Guang Gao

Angela Harris

Alan Xu

Jennifer Linares

Thomas M. Quill

Phuong Nguyen

Qi Wu

Stephanie T. Schmidt

Sahil Seth

Christopher A. Bristow

Norma Rogers

Simon S. Yu

Hannah E. Shepard

Robert A. Mullinax

Sabina A. Lorca

Kang Le

Reagan Lucas

Sweta Mahaeshwari

Sarah Joseph

Ningping Feng

Yongying Jiang

Joe Marszalek

Michael Soth

Jason Cross

Mary Geck Do

M. Emilia Di Francesco

Timothy P. Heffernan

Philip Jones

Crossref

https://openworks.mdanderson.org/cgi/viewcontent.cgi?article=1043&context=leading-edge

Multi-disciplinary efforts to evaluate the therapeutic potential of CDK11, a novel transcription associated cyclin dependent kinase.

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