The Path from Chemical Tool to Approvable Drug
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Part 2 of Ohio State's Drug Development Bootcamp
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The Path from Chemical Tool to Approvable Drug
- 1. THE PATH FROM CHEMICAL TOOL TO APPROVABLE DRUG An Integrated Approach to Drug Discovery EVERY STEP OF THE WAY EVERY STEP OF THE WAY
- 2. The Path to a Clinical Candidate
- 3. ‘END TO END’ INTEGRATED DRUG DISCOVERY 3 EVERY STEP OF THE WAY B/D Molecular Biology Cell Line Generation FBDDStructural Biology CRISPR Adenoviral Platform Human 1o Cells Functional Genomics Library Design Analytical & Purification Process ChemistryCADD Synthetic Chemistry Formulation Pharmaceutics Chemo- genomics D/P S/I Safety Assessment Safety Pharmacology Non-GLP/GLP Toxicology Anatomic & Clinical Pathology Imaging Animal Model Development Large Animal Efficacy Models Discovery Pathology In Vivo Efficacy In Vivo Validation PK/PD Dose to Human Predictions ADME Bioanalysis Targets Clinical Candidate Pharmacology in vitro/in vivo Hit Finding: HTS, HCS IND Enabling Studies Medicinal Chemistry Biomarker Development Target Discovery & Validation DP DP Ch Discovery Pathway Chemistry Biology/ Discovery Technologies DMPK/Pharmacology/Safety/ In vivo models
- 4. 4 EVERY STEP OF THE WAY Are we there yet?
- 5. BEGIN WITH THE END IN MIND! Properties of Drug-Like Molecules 5 EVERY STEP OF THE WAY • Potent • Modulate the target in a predictable way • Selective • How selective is selective enough? • Formulatable • Reasonable synthetic route or method of production • Good safety profile • Understanding of toxicity • No such thing as a “magic” therapeutic index • Well-behaved pharmacokinetics • PK relates to the pharmacodynamics • Amenable to QD dosing (BiD acceptable in some cases) • Rapid, predictable onset of action • Consistent metabolism • Clearance is important • No accumulation • Understand drug-drug interactions • Available biomarker
- 6. THE FIRST PART OF THE JOURNEY 6 EVERY STEP OF THE WAY • A chemical tool is a fit for purpose compound. • “Good enough” with respect to potency, selectivity and PK • Will never become a drug • Should serve as a starting point for a synthetic chemistry strategy • Must be modifiable in a sensible manner • Shouldn’t have serious red flags (e.g. highly reactive intermediates) • Should be able to provide proof of concept for the project Identification of a Chemical Tool
- 7. Inhibition of Cyclin-Dependent Kinase-4: A Targeted Approach to Cancer Therapy A Case Study in Cancer Drug Discovery Peter Toogood, David Fry, W. R. Leopold CASE STUDY Road from Tool to Approved Drug
- 8. Cyclin D Cdk4/6 Cdk2 Cyclin E E2F/DP1 Rb The Eukaryotic Cell Cycle Growth Factors G0 P P G1 SG2 M Transcription stops, DNA replication starts E2F/DP1 Rb P P Rb P P E2F/DP1 Transcription E2F/DP1 P P P Rb P PCyclin A P P Rb P P P P Rb P P Cdk1 Cyclin B p16 p27 C. J. Sherr Science 1996, 274, 1672. S. A. Ezhevsky et al. Mol. Cell. Biol. 2001, 21, 4773. (Kip1) (INK4a) Cdk1 Cdk3 Cyclin A Cdk2 HDAC HDAC
- 9. RATIONALE FOR TARGETING CDK4/6 Beginning with the End in Mind • Most human tumors are deregulated in some part of the Cyclin D/CDK4/p16/Rb/E2F pathway: • e.g. p16 deletion, cyclin D over-expression, Rb phosphorylation, Rb deletion • Inhibition of CDK4/6 will restore cell cycle control at the G1 checkpoint • CDK4/6 inhibition anticipated to be cytostatic not cytotoxic • Rb-deleted tumors not anticipated to be sensitive
- 10. N N N OHN N N N OHN 2 6 2 6 8 PD 0168553 Cdk4/D IC50 > 10 µM Cdk1/B IC50 > 10 µM Cdk2/A IC50 > 10 µM Cdk2/E IC50 > 10 µM PDGFr IC50 = 0.184 µM FGFr IC50 = 1.479 µM cSrc IC50 = 1.754 µM PD 0166447 Cdk4/D IC50 = 0.620 µM Cdk1/B IC50 = 1.015 µM Cdk2/A IC50 = 0.129 µM Cdk2/E IC50 = 0.410 µM PDGFr IC50 = 1.785 µM FGFr IC50 = 3.295 µM cSrc IC50 = 21.50 µM S/T Kinases Tyr-Kinases IDENTIFICATION OF NOVEL INHIBITORS Transitioning From Tyr Kinase Inhibition to Ser/Thr Kinase Inhbition
- 11. Crystal Structure of PD 0172803 Bound to Cdk2 0.00 0.10 0.20 0.30 40 nM 20 nM 10 nM control 1 / [ATP](mM) 1 Rate of pRb phosphorylation Increasing [Inhibitor] 0.5 1.0 1.5 2.0 INHIBITORS ARE ATP-COMPETITIVE Understanding Mechanism
- 12. Electron donating, polar substituents provide optimal potency and physical properties Cycloalkyl groups preferred over alkyl- or aryl- substituents Are any substituents tolerated here? M. Barvian et al. J. Med. Chem. 2000, 43, 4606. N N NHN O N N H 5 6 2 8 PD 0205606 INITIAL OPTIMIZATION FOR POTENCY A Rational Synthetic Strategy
- 13. N N NHN O N N H 2 8 0205606 Cdk4/D: Cdk2/A: FGFr: HCT-116: IC50 (µM) 0.006 0.015 0.081 0.138 PD 0205606 is a potent Cdk inhibitor with reasonable solubility (0.12 mg/mL) But…. CL = 142 mL/min/Kg AUC (PO) = 145 ng/hr/mL t 1/2(PO) = 1.6 h Loss of selectivity! Poor Pharmacokinetics! THE FIRST CHEMICAL TOOL
- 14. PD 0202020 Cdk4/D IC50 = 0.720 µM Cdk1/B IC50 = 0.193 µM Cdk2/A IC50 = 0.012 µM Cdk2/E IC50 = 0.218 µM PDGFr IC50 = 10.82 µM FGFr IC50 = 3.900 µM cSrc IC50 = 7.150 µM A potent inducer of apoptosis in HT-29 colon carcinoma cells Metabolized at C5-C6 double bond N N NHN O N 5 6 O O O p450 Proposed biotransformation ENGINEERING OUT METABOLIC LIABILITY
- 15. N N NHN O N N H 5 6 N N NHN O N N H 0205606 Cdk4/D IC50 = 0.006 µM Cdk2/A IC50 = 0.024 µM FGFr IC50 = 0.081 µM PDGFr IC50 = 0.5 µM 0217048 (R = H) Cdk4/D IC50 = 0.014 µM Cdk2/A IC50 > 5 µM FGFr IC50 = 4.23 µM PDGFr IC50 = 1.81 µM 2 R • Substituents at C5 larger than methyl are not tolerated • Effect of the C5 methyl group is modified by the C6 substituent R Cdk4/D IC50(µM) Cdk1/B IC50(µM) Cdk2/A IC50(µM) Cdk2/E IC50(µM) Et 0.025 4.119 1.538 1.650 Cl 0.016 >5 1.625 1.500 Br 0.005 2.615 0.439 0.950 Ac 0.002 >5 0.230 1.15 CO2Et 0.006 >5 >5 >5 ATTACKING THE C5 – C6 POSITION C5-Methylation Affords Cdk4 Selectivity
- 16. N N NHN O N N H O PD 0326562 Enzyme IC50 (µM) Cdk4/D Cdk1/B Cdk2/A Cdk2/E FGFr PDGFr VEGFr Lck MDA-MB-435 0.002 >5 0.23 1.15 3.39 3.10 1.69 1.744 IC50 = 0.03 µM PD 0326562 is efficacious at 75 mg/Kg but has a narrow therapeutic index with significant toxicity at 100 mg/Kg HOW SELECTIVE IS SELECTIVE ENOUGH ?
- 17. 0174413 Cdk4/D: Cdk2/A: 0.210 µM 0.012 µM 0204661 Cdk4/D: Cdk2/A: 0.092 µM 0.002 µM 0205783 Cdk4/D: Cdk2/A: 0.145 µM 5.010 µM N N NHN O N N NHN O N N N NHN O N PD 0204661 is toxic….. but PD 205783 is somewhat selective for Cdk4 UNDERSTANDING TOXICITY A Re-examination of the Pharmacophore
- 18. N N NHN O N N H 2 8 6 Br N N NHN O N N N H Br Rat PK CL = 53 mL/min/Kg IV t1/2 = 2 h F = 10% Enzyme IC50 (µM) Cdk4/D Cdk1/B Cdk2/A Cdk2/E FGFr PDGFr 0.016 >5 >5 >5 1.16 4.36 0325056 causes a clean G1 arrest up to 10.0 µM 0221933 0325056 DISCOVERY OF EXQUISITE SELECTIVITY OF CDK4
- 19. PD Number 0221933 0325056 0324275 0332725 0326562 0332991 Cdk4/D 0.002 0.016 0.006 0.123 0.002 0.011 Cdk1/B 0.281 >5 3.530 - >5 >5 Cdk2/A 0.042 >5 0.556 >5 0.23 >5 FGFr 0.080 1.16 0.535 - 3.39 >5 Cdk2/E 0.172 >5 3.500 - 1.15 >5 PDGFr 0.056 4.36 0.300 - 3.10 > 5 Potency and selectivity for Cdk4: IC50 (µM) 1 2 3 Structure 1 1 2 2 3 3 A CH N CH N CH N N N NHN O A N N H Br N N NHN O A N N H O N N NHN O A N N H O THE SELECTIVITY EFFECT IS GENERAL
- 20. Protein Kinase IC50 (mM) Cdk4/D1 Cdk4/D3 Cdk6/D2 Cdk2/E2 Cdk2/A Cdk1/B Cdk5/p25 Gsk3β EGFr FGFr PDGFr VEGFr Lck Insulin Receptor AMPK Chk1 0.011 0.009 0.015 >10 >10 >10 >10 >10 >10 >10 >10 >10 >10 >10 >10 >10 Protein Kinase IC50 (mM) CK-1 SGK C-src JNK MAPKAP-K1a MKK p70S6K PDK1 PHK SAPK3 MAPK2 p70S6K PKA PKB PKC DYRK 1A >10 >10 >10 >10 8.0 >10 >10 >10 >10 >10 >10 >10 >10 >10 >10 2.0 PD0332991 IS SELECTIVE FOR CDK4/6 VS OTHER KINASES
- 21. Cell Line Cell Type Rb IC50 (µM) MDA-MB-435 Breast Carcinoma + 0.16 ZR-75-1 Breast Carcinoma + 0.17 T-47D Breast Carcinoma + 0.04 MCF-7 Breast Carcinoma + 0.10 H1299 Lung Carcinoma + 0.12 Colo205 Colon Carcinoma + 0.13 CRRF-CEM Acute lymphoblastic leukemia (ALL) + 0.25 K562 CML + 0.40 MDA-MB-468 Breast Carcinoma - >3 (20%) H2009 Lung Carcinoma - >3 (0%) IN VITRO ACTIVITY OF PD0332991 Dependence on Rb
- 22. Biological Effect Rb phos. (Ser780) Cellular Proliferation Initiate G1 arrest IC50 (µM) 0.066 0.032 0.040 Data obtained using MDA-MB-435 and MDA-MB-453 breast carcinoma cell lines RELATING TARGET MODULATION TO BIOLOGICAL EFFECTS Target Biomarker Correlates with Biological Effects
- 23. 1 10 100 1000 10000 0 5 10 15 20 25 30 Time (Hours) Subject Rat05 Subject Rat06 Subject Rat07 Subject Rat08 Mean Data Individual and Mean Plasma 0332991 Concentration-Time Profiles Following a Single 20 mg/kg PO Dose to Fasted Male Sprague-Dawley Rats Time (h) Concentration(ng/mL) t1/2 = 2.8 h F = 56% PD 0332991 IS ORALLY BIOAVAILABLE Well Behaved PK Profile
- 24. Days Post Tumor Implant 20 30 40 50 MedianTumorMass(mg) 150 200 250 300 400 500 600 700 800 900 1500 100 1000 control 36 mg/kg 58 mg/kg 93 mg/kg 150 mg/kg 240 mg/kg Rx days:12-39 Route: Oral Schedule: Q1D Duration of therapy PD0332991 INHIBITS TUMOR GROWTH IN VIVO MDA-MB-435 Breast Cancer Line
- 25. Days Post Tumor Implant 20 25 30 35 40 45 50 55 60 65 70 75 80 85 MedianTumorMass(mg) 60 70 80 90 150 200 250 300 400 500 600 700 800 900 100 1000 control 12.5 mg/kg 37.5 mg/kg 75 mg/kg 150 mg/kg Rx days:18-31 Route: Oral Schedule: Q1D Limit of palpation Duration of therapy PD0332991 INHIBITS TUMOR GROWTH IN VIVO Colo-205
- 26. MedianTumorMass(mg) Days Post Implant Daily PO Therapy MDA-MB-468 60 70 80 90 100 200 300 400 20 25 30 35 40 45 50 55 60 Control 240 mg/kg (3/10 NSD) 150 mg/kg 75 mg/kg mg/kg 37.5 mg/kg PD0332991 IS INACTIVE IN Rb NEGATIVE TUMORS Predicted by Hypothesis
- 27. DEVELOPMENT OF PD 0332991 FOR CLINICAL USE Observed Clinical Toxicity is Exaggerated Pharmacology PD 0332991 was nominated for development in 2002 An Investigational New Drug application was filed in 2003 Phase I clinical trials commenced in 2004 2007 ASCO Annual Meeting Abstract (J. Clin. Oncology 2007, 25, no. 18S) A Phase I Dose Escalation Trial of Daily Oral CDK4/6 Inhibitor PD 0332991 O’Dwyer et al - Principal and dose-limiting toxicity is myelosuppresion. - Of patients receiving PD 0332991 for 21 days in a 28 day cycle, MTD = 125 mg QD. - 6 patients achieved stable disease, 3 of them for at least 20 cycles (breast, colon, ovarian)
- 28. CLINICAL ACTIVITY AGAINST ER-POSITIVE BREAST CANCER Promising Early Results ASCO Annual Meeting, Abstract no. 3060 (J. Clin. Oncology 2010, 28, 15s) Phase I/II Study of PD 0332991 and Letrozole in ER-Positive Breast Cancer Slamon et al PD 0332291 (125 mg) QD for 21 days of a 28 day cycle with letrozole 2.5 mg QD • 12 Patients enrolled (post-menopausal, ER-positive, HER-2 negative) • 3 Patients discontinued due to disease progression • DLT = grade 4 neutropenia • 3 Patients with partial response • 9 Patients with stable disease
- 29. PHASE I CLINICAL DATA IN MULTIPLE MYELOMA Other Indications 52nd ASH Annual Meeting, Abstract 860. Lentzsch et al. • A Phase I Study of PD 0332991: Complete CDK4/6 Inhibition and Tumor Response in Sequential Combination with Bortezemib and Dexamethasone for Relapsed and Refractory Multiple Myeloma • Schedule B: Patients received PD 0332991 (100 or 125 mg) QD days 1-12 of a 21 day cycle, with (IV) bortezomib and (oral) dexamethasone administered on days 8, 11, 15 and 18. “Encouraging antitumor activity was observed in this heavily treated MM population” Of 12 patients receiving schedule B , there was one partial response , three patients with stable disease (≥ 3 months) and one very good partial response with a patient who had relapsed on lenalidomide, bortezomib, and carfilzomib therapies and a stem cell transplant
- 30. WE HAVE A DRUG!!! 30 Pfizer’s Palbociclib (PD-0332991) Receives Food And Drug Administration Breakthrough Therapy Designation For Potential Treatment Of Patients With Breast Cancer Wednesday, April 10, 2013 8:00 am EDT The U. S. Food and Drug Administration granted accelerated approval to palbociclib (IBRANCE, Pfizer, Inc.) for use in combination with letrozole for the treatment of postmenopausal women with estrogen receptor (ER)- positive, human epidermal growth factor receptor 2 (HER2)-negative advanced breast cancer as initial endocrine-based therapy for their metastatic disease. February 3, 2015
- 31. OTHER TUMOR TYPES IN CLINICAL STUDY 31 Development Continues •Various Stages (I-II) • Mantle Cell Lymphoma • Refractory GIST • NSCLC • Glioblastoma • Liver
- 32. Timeline September 2002 1996 Project Initiated PD0332991enters Development DISCOVERY December 2003 IND September 2004 FIH PRECLINICAL SAFETY ASSESSMENT PHASE I Multipleongoing PhaseI/IItrials June 2007 2011 1stPhaseI datapublished PHASE I/II Multiple Myeloma (+Velcade and Dex) Mantle Cell Lymphoma MCL (+Velcade) Non-Hodgkins Lymphoma Liposarcoma Glioblastoma Multiforme Breast Cancer (+Letrozole) Solid Tumors 2013 2014 FDADeclares BreakthroughStatus PFEannounces ItwillsubmitNDA PHASE III 2015 FDAGrantsAccelerated ApprovalforPalbociclib
- 33. Summary • Selective CDK4/6 inhibition was hypothesized to present a safer approach to treating Rb-positive tumors with a deregulated cell cycle • PD 0332991 was discovered as a result of a concerted effort to identify CDK4/6-selective kinase inhibitors by employing a number of chemical tool compounds throughout the process. • PD 0332991 was advanced to human clinical trials on the basis of broad-based antitumor efficacy vs Rb-positive human xenograft tumors in mouse models and its preclinical safety profile • FDA has granted breakthrough status to the compound for the treatment of breast cancer. • Additional studies ongoing for treatment of other malignancies.
- 34. CONTACT US J. A. Cornicelli, Ph.D., F.A.H.A. 251 Ballardvale Street Wilmington, MA 01887 askcharlesriver@crl.com www.criver.com 877.CRIVER.1