Antireplication Agents: CDK Inhibitors and Telomerase Inhibitors

Richard S. Finn, MD

Assistant Professor of Medicine Geffen School of Medicine at UCLA

Faculty Disclosure

Richard Finn, MD, has disclosed that he has received fees for non-CME services from Genentech.

Cyclin D Kinases and Cancer

 CDKs are a subgroup of seine/ threonine kinases  In general very small proteins (34-40 kDa)   Bind to activating proteins: cyclins Without cyclins, CDKs have little kinase activity  Play a key role in regulating cell cycle progression through all phases of the cell cycle  Various cyclin/ CDK complexes act at different parts of the cell cycle  Temporal and quantitative regulation  Negative regulation by cyclin dependent kinase inhibitors (CKI)  INK 4 family (p15, p16, 18, p19  Cip/ Kip family (p21, p27, p57)  Alterations in CDKs are uncommon, compared with cyclin dysregulation  Altered regulation/ expression in many malignaices  Cyclin D1 amplification has been described in various malignancies, including breast cancer, with variable prognostic significance  t(11;14) mantle cell lymphoma  Rb loss, a well known oncogenic event 1. Finn RS, et al. Breast Cancer Res. 2009;11:R77. 2. Lundberg AS, et al Eur J Cancer. 1999;35:1886-1894.3. Buckley MF, et al. Oncogene. 1993;8:2127-2133. 4. Dickson C, et al. Cancer Lett. 1995;90:43-50.

Cyclin D Kinases and Cancer

Vermullen Cell Proliferation

Rb as Master-Regulator of the R Point

p16(INK4a)

Modified from Figure 8.19 .

The Biology of Cancer. © Garland Science 2007.

CDK/ Cyclin Inhibitors

 Targeting CDK  >50 inhibitors described  Generally in several classes – Purine analogoues (ie roscovitine)  Selicicilib (CYC202)  CDK2/E CDk 2/A CDK7 CDK 9 – Pyrimidine analgoues  Dinaciclib (SCH727965)  CDK 2, CDk5, CDK 1, CDK 9 – – – Flavonoids (flavopiridol) Indolinones Staurosporine  Targeting Cyclin  cyclin expression modulators (ON013105)

CDK/ Cyclin Inhibitors

 Selicicilib (CYC202) – Phase II  Nasopharyngeal  APPRAISE Study  Randomized Phase II, NSCLCA 3 rd line or greater vs BSC  Dinaciclib (SCH727965)  Phase II – Melanoma – AML  ON013105  Phase I- mantle cell lymphoma

PD 0332991: Background

  The compound  Potent, selective, reversible inhibitor of CDK4,6 Small molecule Oral agent   The opportunity   Potential first in class Potential impact on hematopoietic and solid tumors Potential use in pediatric indications Single-agent and combination approaches under investigation Finn RS, et al. Breast Cancer Res. 2009;11:R77.

O N N H N N N O N + N H 2 O S O O OH PD 332991

Rb as Master-Regulator of the R Point

Target of PD 0332991 p16(INK4a) Inactivates Rb and allows progression

Modified from Figure 8.19 .

The Biology of Cancer. © Garland Science 2007.

Sorlie et al PNAS 2001

Human Breast Cancer Cell Line Panel Can Recapitulate the Molecular Heterogeneity of Clinical Disease

51 Human Breast Cell Lines

25 Luminal 26 Non-luminal 10 ER positive Normal HER-2 9 ER positive HER-2 amplified 6 ER negative HER-2 amplified 4 Non-malignant 13 Basal/Progenitor 9 Mesenchymal 1 HER-2 Amplified 1 HER-2 Amplified

PD-0332991: CDK 4/6 Inhibitor: Breast Panel 1000 900 800 700 600 500 400 300 200 100 0

MB -1 75 ZR 75 -3 0 C A MA -1 MB 13 4 H C C U 20 A 2 C C -8 93 EF M1 9 SU M1 90 EF M1 92 A MB -3 H 61 C C 15 H 00 C C 14 19 H C C 38 MB -4 15 MC F-1 U 0A A C C -8 H 12 C C 22 18 ZR 75 MD -1 A MB 45 3 18 4A 1 T4 7D MC F7 B T-2 0 A MB MD 43 5 B T4 74 SK B R 3 K PL H -1 C C 11 MD 43 A MB 23 H 1 C C 13 95 SU M-2 25 H S5 78 T 18 4B U 5 A C C 73 2 C A L -5 1 B T5 C 49 O LO 82 4 D U 44 H 75 C C 11 H 87 C C 15 H 69 C C 18 H 06 C C 19 H 37 C C 19 54 H C C 70 MB -4 36 MB 15 MD A 7 MB 46 8

Subtype Luminal HER2 Amplified Immortalized Non-luminal/post EMT Non-luminal

Finn RS Breast Can Res 2009

PD 0332991: Cell Cycle Analysis

MCF7 %

100 90 80 70 60 50 40 30 20 10 0 G0/G1 S G2

EFM192A

100 90 80 70 60 50 40 30 20 10 0 G0/G1 S

Sensitive lines

G2 100 90 80 70 60 50 40 30 20 10 0 G0/G1

HCC 1419

S G2

HCC 1937 HCC 1187 %

50 40 30 20 10 0 100 90 80 70 60 50 40 30 20 10 0 100 90 80 70 60 G0/G1 S G2 G0/G1 S

Resistant lines

Finn RS, et al. Breast Cancer Res. 2009;11:R77.

G2 50 40 30 20 10 0 100 90 80 70 60 G0/G1

MDA MB 468

S G2

PD 0332991: Effects on Phosphorylation on Retinoblasoma Gene Product

30’ A. Total pRb 60’ 12 hr 24 hr 48 hr B. Phospho-Rb (serine 780) 0 30’ 60’ 12 hr 24 hr 48 hr MCF7 Time 0 MB453

Sensitive

T47D EFM19 HCC1187 HCC1954 CAL 51

Resistant

Finn RS, et al. Breast Cancer Res. 2009;11:R77.

Hypothesis: Patient Selection in Breast Cancer Population

Elevated

Cyclin D1 RB

Decreased

p16

Gauthier ML, et al. Cancer Cell. 2007;12:479-491.

MCF7

100 80 60 40 20 0 Tam PD 10000 100

EFM19 T47D

100 80 60 40 20 0 Tam PD 100 5000 50 80 60 40 20 Combo PD-2991 Tamoxifen 0 Tam PD 5000 50 5000 50 2500 1250 25 12.5

Concentration (nM)

625 6.25

2500 25 1250 12.5

Concentration (nM)

625 6.25

2500 25 1250 12.5

Concentration (nM)

625 6.25

312 3.125

312 3.125

312 3.125

2 1 0 1000 100 2 5000 50 2500 1250 25 12.5

Concentration (nM)

625 6.25

312.5

3.125

1 0 Tam PD 5000 50 2 1 2500 25 1250 12.5

Concentration (nM)

625 6.25

312 3.125

0 Tam PD 5000 50 2500 25 1250 12.5

Concentration (nM )

625 6.25

312 3.125

TRIO 18/A5481003: Phase I/II Study of Letrozole in Combinations With PD-0332991 in Postmenopausal ER+ Advanced Breast Cancer

  Phase I complete Randomized phase II accruing ClincailTrials.gov. NCT00721409.

TRIO 18: Phase I Patient Summary

Slamon DJ, et al. ASCO 2010. Abstract 3060.

TRIO 18: Most Common AEs (N = 12)

PT, n

Neutropenia Fatigue Leukopenia Nausea Diarrhea Anemia Cough Decreased appetite Dyspnea Hot flush Nasal congestion Arthralgia Back pain Creatinine increased

Grade 1

0 6 0 5 4 2 2 3 3 2 2 1 2 0

Grade 2

1 2 3 0 0 1 1 0 0 1 1 1 0 1

Grade 3

7 0 2 0 0 0 0 0 0 0 0 0 0 1

Grade 4

2

0 1 0 0 0 0 0 0 0 0 0 0 0

Total

10 8 6 5 4 3 3 3 3 3 3 2 2 2 Slamon DJ, et al. ASCO 2010. Abstract 3060.

TRIO 18: Most Common Treatment Related AEs (N = 12)

PT, n

Neutropenia Fatigue Leukopenia Nausea Anemia Decreased appetite Diarrhea Hot flush Dyspnea Headache Thrombocytopenia

Grade 1

2 2 1 1 2 3 3 0 4 0 5

Grade 2

1 0 1 1 1 0 0 1 2 3 0

Grade 3

0 0 0 0 0 0 0 7 0 2 0

Grade 4

0 0 0 0 0 0 0 2 0 1 0

Total

3 2 2 2 3 3 3 10 6 6 5 Slamon DJ, et al. ASCO 2010. Abstract 3060.

TRIO 18: Phase I Summary

 Phase I (N = 12) • MTD: PD 0332991 125 mg QD (schedule 3/1) in combination with letrozole 2.5 mg QD • 3 DLTs: – – 2 patients with grade 4 neutropenia 1 patient with 5 doses held due to elevated creatinine deemed treatment related • • No treatment-related SAEs No discontinuations due to AEs – Common treatment-related AEs: neutropenia, leukopenia, fatigue • No febrile neutropenia • • • No drug-drug interaction Efficacy: 3 PRs and 9 SDs (PR: 33%; CBR: 67%) Median duration of treatment: 12 mos (range: 2-21+) • Currently 6 patients active Slamon DJ, et al. ASCO 2010. Abstract 3060.

Hypothesis: Biomarkers Predictive of PD 0332991 Sensitivity

Desired biomarker profile:  ER+, HER2 – Wild-type Rb1 – Plus • Amplified cyclin D1/CCND1 • OR Inactivated CDKN2A/p16 INK4a Finn RS, et al. Breast Cancer Res. 2009;11:R77.

Phase II Study Design (Part I, Completed)

ER+, HER2- breast cancer

Stratification Factors:



Disease site

−

Visceral vs bone only vs other



Disease-free interval

−

> 12 vs ≤ 12 mos

R A N D O M I Z A T I O N

N = 60 1:1

Arm A

PD 0332991 125 mg/day (Schedule 3/1) + Letrozole 2.5 mg/day

Arm B

Letrozole 2.5 mg/day

Primary endpoint: PFS ClincailTrials.gov. NCT00721409.

Phase II Study Design (Part II, Ongoing)

ER+, HER2- breast cancer

Biomarker Selection



CCND1 amp



And/or loss of p16

R A N D O M I Z A T I O N

N = 150 1:1

Arm A

PD 0332991 125 mg/day (Schedule 3/1) + Letrozole 2.5 mg/day

Arm B

Letrozole 2.5 mg/day

Primary endpoint: PFS ClincailTrials.gov. NCT00721409.

Similar Observations in Other Histologies

   Ovarian cancer [1] Glioblastoma [2,3] Multiple myeloma [4] 1. Konecny GE, et al. Clin Cancer Res. 2011;17:1591-1602. 2. Michaud K. Cancer Res. 2010;70:3228 3238. 3. Wiedemeyer WR, et al. Proc Natl Acad Sci U S A. 2010;107:11501-11506. 4. Menu E, et al. Cancer Res. 2008;68:5519-5523.

Telomerase and Cancer

   Telomerase: reverse transcriptase that adds DNA repeats (TTAGGG) to the 3’ end of DNA strands (telomere region) – Consists of 2 molecules each of telomerase reverse transcriptase (TERT), telomerase RNA (TERC), and dyskerin (DKC1) Protects DNA from genomic damage/loss during replication In cancer, cells that lose telomeres become unstable, accumulate genetic damage, and eventually undergo apoptosis  Activation of telomerase can prevent the apoptosis event and cause cells to become immortalized – Telomerase is activated in 90% of cancer (but not somatic cells)  Hypotheses: block telomerase, induce telomere shortening, genetic instability and cell death Shay JW, et al. Human Mol Gen. 2001;10:677-685.

Telomerase MOA

Harley Nat rev Cancer 2008

Telomerase and Cancer

 Various potential methods of silencing telomerase – Oligonucleotides • • Target the template region (activation site) of telomerase Imetelstat (GRN163L): 13 mer oligonucleotide, not antisense but a direct telomerase inhibitor [1] – Vaccines • Dendritic cell –based (GRNVAC1) • • Use hTERT pulsed autologous dendritic cells Phase I: generally well tolerated, achieved levels felt to be sufficient for hTERT inactivation [2] • Phase II study in AML as consolidation • Nondendritic based • Phase I in solid tumors (V934/V935) [4] • GV1001 peptide with temozolomide for melanoma [5] 1. Herbert BS, et al. Oncogene. 2005;24:5262-5268. 2. Su Z, et al. J Immunol. 2005;174:3798-3807. 3. ClinicalTrials.gov. NCT00510133. 4. ClinicalTrials.gov. NCT00753415. 5. ClinicalTrials.gov. NCT01247623.

Conclusions

    Newer targets in molecular oncology include the CDK pathway and telomerase Ongoing clinical studies (TRIO18) with PD 0332991, an oral, small molecule kinase inhibitor, in breast cancer aim at validating laboratory science that identified ER+ breast cancer as being susceptible to CDK 4/6 inhibition  To date, predictable and manageable toxicity has been seen with this class of agent Studies with other CDK targeted agents and Cyclin targeted agents are ongoing  MCL should be “proof of concept” Telomerase inhibitors are moving to the clinic and hold promise but predictors of response will be necessary for successful clinical development  Including vaccine strategies and short-oligos