the path from chemical tool to approvable drug
TRANSCRIPT
THE PATH FROM CHEMICAL TOOL TO APPROVABLE DRUGAn Integrated Approach to Drug Discovery
EVERY STEP OF THE WAY
EVERY STEP OF THE WAY
The Path to a Clinical Candidate
‘END TO END’ INTEGRATED DRUG DISCOVERY
3 EVERY STEP OF THE WAY
B/DMolecular Biology
Cell LineGeneration
FBDDStructuralBiology
CRISPRAdenoviral
Platform Human 1o Cells
Functional Genomics
Library Design
Analytical &Purification
ProcessChemistryCADD
Synthetic Chemistry
Formulation
Pharmaceutics
Chemo-genomics
D/PS/I
Safety Assessment
Safety Pharmacology
Non-GLP/GLP Toxicology
Anatomic &Clinical Pathology
Imaging
Animal ModelDevelopment
Large Animal Efficacy Models
DiscoveryPathology
In Vivo Efficacy
In Vivo Validation
PK/PD
Dose to Human Predictions
ADME
Bioanalysis
Targets Clinical Candidate
Pharmacologyin vitro/in vivo
Hit Finding:HTS, HCS
IND EnablingStudies
MedicinalChemistry
BiomarkerDevelopment
Target Discovery
& Validation
DP DP
Ch
Discovery Pathway
Chemistry
Biology/ Discovery Technologies
DMPK/Pharmacology/Safety/ In vivo models
4 EVERY STEP OF THE WAY
Are we there yet?
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
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
Inhibition of Cyclin-Dependent Kinase-4: A Targeted Approach to Cancer Therapy
A Case Study in Cancer Drug DiscoveryPeter Toogood, David Fry, W. R. Leopold
CASE STUDYRoad from Tool to Approved Drug
Cyclin DCdk4/6
Cdk2Cyclin E
E2F/DP1
Rb
The Eukaryotic Cell Cycle
Growth FactorsG0
PP
G1
SG2
M
Transcription stops,DNA replication starts
E2F/DP1
Rb
PPRb
PP E2F/DP1 Transcription
E2F/DP1P
PPRb
PPCyclin A
PPRb
PP
PPRb
PP
Cdk1Cyclin 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 ACdk2
HDAC
HDAC
RATIONALE FOR TARGETING CDK4/6Beginning 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
N
N N OHN
N
N N OHN2
6
26
8
PD 0168553
Cdk4/D IC50 > 10 µMCdk1/B IC50 > 10 µMCdk2/A IC50 > 10 µMCdk2/E IC50 > 10 µMPDGFr IC50 = 0.184 µMFGFr IC50 = 1.479 µMcSrc IC50 = 1.754 µM
PD 0166447
Cdk4/D IC50 = 0.620 µMCdk1/B IC50 = 1.015 µMCdk2/A IC50 = 0.129 µMCdk2/E IC50 = 0.410 µMPDGFr IC50 = 1.785 µMFGFr IC50 = 3.295 µMcSrc IC50 = 21.50 µM
S/T Kinases
Tyr-Kinases
IDENTIFICATION OF NOVEL INHIBITORSTransitioning From Tyr Kinase Inhibition to Ser/Thr Kinase Inhbition
Crystal Structure of PD 0172803 Bound to Cdk2 0.00 0.10 0.20 0.30
40 nM
20 nM10 nM
control
1 / [ATP](mM)
1Rate of pRb
phosphorylation
Increasing[Inhibitor]
0.5
1.0
1.5
2.0
INHIBITORS ARE ATP-COMPETITIVEUnderstanding Mechanism
Electron donating, polar substituents provide optimal
potency and physical properties
Cycloalkyl groups preferred over alkyl- or aryl-
substituents
Are any substituentstolerated here?
M. Barvian et al. J. Med. Chem. 2000, 43, 4606.
N
N NHN O
N
NH
56
28
PD 0205606
INITIAL OPTIMIZATION FOR POTENCYA Rational Synthetic Strategy
N
N NHN O
N
NH
2 8
0205606
Cdk4/D:Cdk2/A:
FGFr:HCT-116:
IC50 (µM)0.0060.0150.0810.138
PD 0205606 is a potent Cdk inhibitor with reasonable solubility (0.12 mg/mL)
But….
CL = 142 mL/min/KgAUC (PO) = 145 ng/hr/mLt 1/2(PO) = 1.6 h
Loss of selectivity!
Poor Pharmacokinetics!
THE FIRST CHEMICAL TOOL
PD 0202020
Cdk4/D IC50 = 0.720 µM Cdk1/B IC50 = 0.193 µMCdk2/A IC50 = 0.012 µMCdk2/E IC50 = 0.218 µMPDGFr IC50 = 10.82 µMFGFr IC50 = 3.900 µMcSrc 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
Op450
Proposed biotransformation
ENGINEERING OUT METABOLIC LIABILITY
N
N NHN O
N
NH
56 N
N NHN O
N
NH
0205606Cdk4/D IC50 = 0.006 µMCdk2/A IC50 = 0.024 µMFGFr IC50 = 0.081 µMPDGFr IC50 = 0.5 µM
0217048 (R = H)Cdk4/D IC50 = 0.014 µMCdk2/A IC50 > 5 µMFGFr IC50 = 4.23 µMPDGFr 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 POSITIONC5-Methylation Affords Cdk4 Selectivity
N
N NHN O
N
NH
O
PD 0326562
Enzyme IC50 (µM)
Cdk4/DCdk1/BCdk2/ACdk2/EFGFrPDGFrVEGFrLck
MDA-MB-435
0.002>50.231.153.393.101.691.744
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 ?
0174413Cdk4/D:Cdk2/A:
0.210 µM0.012 µM
0204661Cdk4/D:Cdk2/A:
0.092 µM0.002 µM
0205783Cdk4/D:Cdk2/A:
0.145 µM5.010 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 TOXICITYA Re-examination of the Pharmacophore
N
N NHN O
N
NH
2 8
6 BrN
N NHN O
N
N
NH
Br
Rat PKCL = 53 mL/min/KgIV t1/2 = 2 h F = 10%
Enzyme IC50 (µM)
Cdk4/DCdk1/BCdk2/ACdk2/EFGFrPDGFr
0.016>5>5>51.164.36
0325056 causes a clean G1 arrest up to 10.0 µM
0221933 0325056
DISCOVERY OF EXQUISITE SELECTIVITY OF CDK4
PD Number
022193303250560324275033272503265620332991
Cdk4/D
0.0020.0160.0060.1230.0020.011
Cdk1/B
0.281>53.530->5>5
Cdk2/A
0.042>50.556>50.23>5
FGFr
0.0801.160.535-3.39>5
Cdk2/E
0.172>53.500-1.15>5
PDGFr
0.0564.360.300-3.10> 5
Potency and selectivity for Cdk4: IC50 (µM)
1 2 3
Structure
112233
A
CHNCHNCHN
N
N NHN O
A
N
NH
BrN
N NHN O
A
N
NH
O
N
N NHN O
A
N
NH
O
THE SELECTIVITY EFFECT IS GENERAL
Protein Kinase IC50 (mM)
Cdk4/D1Cdk4/D3Cdk6/D2Cdk2/E2Cdk2/ACdk1/BCdk5/p25Gsk3βEGFrFGFrPDGFrVEGFrLckInsulin ReceptorAMPKChk1
0.0110.0090.015>10>10>10>10>10>10>10>10>10>10>10>10>10
Protein Kinase IC50 (mM)
CK-1SGKC-srcJNKMAPKAP-K1aMKKp70S6KPDK1PHKSAPK3MAPK2p70S6KPKAPKBPKCDYRK 1A
>10>10>10>108.0>10>10>10>10>10>10>10>10>10>102.0
PD0332991 IS SELECTIVE FOR CDK4/6 VS OTHER KINASES
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 PD0332991Dependence on Rb
Biological Effect
Rb phos. (Ser780)Cellular ProliferationInitiate G1 arrest
IC50 (µM)
0.0660.0320.040
Data obtained using MDA-MB-435 and MDA-MB-453 breast carcinoma cell lines
RELATING TARGET MODULATION TO BIOLOGICAL EFFECTSTarget Biomarker Correlates with Biological Effects
1
10
100
1000
10000
0 5 10 15 20 25 30Time (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)
Conc
entr
atio
n (n
g/m
L) t1/2 = 2.8 hF = 56%
PD 0332991 IS ORALLY BIOAVAILABLEWell Behaved PK Profile
Days Post Tumor Implant20 30 40 50
Med
ian
Tum
or M
ass
(mg)
150
200
250
300
400
500
600
700
800900
1500
100
1000
control36 mg/kg58 mg/kg93 mg/kg150 mg/kg240 mg/kg
Rx days:12-39Route: OralSchedule: Q1DDuration of therapy
PD0332991 INHIBITS TUMOR GROWTH IN VIVOMDA-MB-435 Breast Cancer Line
Days Post Tumor Implant20 25 30 35 40 45 50 55 60 65 70 75 80 85
Med
ian
Tum
or M
ass
(mg)
60
708090
150
200
250
300
400
500
600
700800900
100
1000
control12.5 mg/kg37.5 mg/kg75 mg/kg150 mg/kg
Rx days:18-31Route: OralSchedule: Q1D
Limit of palpation
Duration of therapy
PD0332991 INHIBITS TUMOR GROWTH IN VIVOColo-205
Med
ian
Tum
or M
ass
(mg)
Days Post Implant
Daily PO Therapy
MDA-MB-468
60708090
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 TUMORSPredicted by Hypothesis
DEVELOPMENT OF PD 0332991 FOR CLINICAL USEObserved Clinical Toxicity is Exaggerated Pharmacology
PD 0332991 was nominated for development in 2002An Investigational New Drug application was filed in 2003Phase 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 0332991O’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)
CLINICAL ACTIVITY AGAINST ER-POSITIVE BREAST CANCERPromising 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 CancerSlamon 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
PHASE I CLINICAL DATA IN MULTIPLE MYELOMAOther 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 carfilzomibtherapies and a stem cell transplant
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 CancerWednesday, 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
OTHER TUMOR TYPES IN CLINICAL STUDY
31
Development Continues
•Various Stages (I-II)• Mantle Cell Lymphoma• Refractory GIST• NSCLC• Glioblastoma• Liver
Timeline
September 2002
1996
Project Initiated PD 0
3329
91 e
nter
s De
velo
pmen
tDISCOVERY
December 2003
IND
September2004
FIH
PRECLINICALSAFETY ASSESSMENT
PHASE I
Mul
tiple
ong
oing
Phas
e I/
II tr
ials
June2007
2011
1stPh
ase
Ida
ta p
ublis
hed
PHASE I/II
Multiple Myeloma (+Velcade and Dex)Mantle Cell Lymphoma
MCL (+Velcade)Non-Hodgkins Lymphoma
LiposarcomaGlioblastoma MultiformeBreast Cancer (+Letrozole)
Solid Tumors
2013 2014
FDA
Decl
ares
Br
eakt
hrou
gh S
tatu
s
PFE
anno
unce
sIt
will
subm
it N
DA
PHASE III
2015
FDA
Gra
nts A
ccel
erat
edAp
prov
al fo
r Pal
boci
clib
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.
CONTACT USJ. A. Cornicelli, Ph.D., F.A.H.A.
251 Ballardvale StreetWilmington, MA01887
www.criver.com
877.CRIVER.1