Targeting CDK4/6 in

Breast Cancer

Dr. Iskandar Ali, Sp.B(K)Onk

1
Cell Cycle Checkpoints Regulate Cell Growth

 The cell cycle governs cell

growth and proliferation1
Check-
point
 Cell cycle checkpoints ensure
the fidelity of each phase before
Check- Mitosis
point and cell continuing to the next phase1
division
 Malignant tumors can form
M G0
Rapid
G2 G1 Growth when cells progress through the
growth
S cell cycle unchecked1
DNA  Preventing cell division by
Check-
replication
point inhibiting the cell cycle is a
Check-
strategy for anticancer therapy
point
 The cyclin D–CDK4/6–INK4–Rb
signaling pathway regulates the
G1/S checkpoint

1. Gabrielli B, et al. Front Pharmacol 2012;3:9. 2

Paring of CDK/Cyclins & cyclin fluctuation
Different CDKs and cyclins act during the cell cycle progression

3
Weinberg et al. The Biology of Cancer, 2nd edition 3
Cell Cycle Progression and Cyclin-Dependent Kinases

G0

M Mitogenic
G2
+ signals

Cyclin A-CDK1 Cyclin D-CDK4 - INK4 family

proteins
Cyclin D-CDK6
(eg, p16)
G1
R point

Cyclin E-CDK2
- Growth inhibitory
Cyclin A-CDK2 signals

Otto T, et al. Nat Rev Cancer. 2017;17:93-115. 4

CDK4/6 Controls Cell Cycle Progression1–4

Signal

p21 p53
Mitogenic signaling pathways CDK4/6
stimulate synthesis of D-type Cyclin D–CDK4/6
cyclins, which bind to and Cyclin D p16 activity is inhibited
activate CDK4/6 by p16 and p21

Active cyclin D–CDK4/6

complexes are transported
into the nucleus
P P
P
P P CDK4/6
E2F activates
Rb
Cyclin D Rb binds to and
transcription of
inhibits the E2F
genes necessary Phosphorylation of Rb by transcription factor,
for cell cycle cyclin D–CDK4/6 decouples forming a
progression Rb from E2F Rb transcriptionally
E2F X E2F
repressive complex

The primary
Summary !
action of CDK4/6 is to
phosphorylate Rb protein and
promote cell cycle progression

1. Shapiro GI. J Clin Oncol 2006;23:1770–1783; 2. Lange CA, Yee D. Endocr Relat Cancer 2011;18:C19–C24;
3. Graf F, et al. Mini Revs Med Chem 2010;10:527–539; 4. Malumbres M, et al. Nat Rev Cancer 2009;9:153–166. 5
Cyclin D-CDK4/6 Activity Required for
Tumor Development/Maintenance in Preclinical Models

• CDK4/6 inhibition induces growth arrest or senescence in

o ER-positive breast cancer cell lines
o Melanoma cell lines

• Cyclin D1 gene inactivation induces senescence in mouse model of

HER2-driven breast cancer
o Cyclin D3 gene inactivation induces apoptosis in mouse model of
Notch1-driven T-ALL
o CDK4 gene inactivation induces senescence in mouse model of KRAS-
driven NSCLC

Finn RS, et al. Breast Cancer Res. 2009;11:R77. Anders L, et al. Cancer Cell. 2011;20:620-634.
Choi YJ, et al. Cancer Cell. 2012;22:438-451. Puyol M, et al. Cancer Cell. 2010;18:63-73. 6
Cyclin D–CDK4/6 Disruption in Cancer

 Increased cyclin D–CDK4/6 activity occurs in cancer through several

mechanisms, including:1–4

Overexpression/
amplification/translocation
of cyclin D genes

Activating mutation Loss of CDK4/6

or amplification inhibitors, such as
of CDK4/6 INK4 proteins

 Activation of cyclin D-CDK4/6 results in:5

– Hyperphosphorylation of Rb
– Inactivation of the growth inhibitory function of Rb
– Release of Rb from E2F
– Unchecked S phase entry and cell cycle progression

1. Shapiro GI. J Clin Oncol 2006;23:1770–1783; 2. Malumbres M, et al. Nat Rev Cancer 2009;9:153–166;
3. Ortega S, et al. Biochim Biophys Acta 2002;1602:73–87; 4. Dean JL, et al. Oncogene 2010;29:4018–4032;
5. Lange CA, Yee D. Endocr Relat Cancer 2011;18:C19–C24. 7
Rb+ Cancers Are Dependent on CDK4/6

 Rb loss also results in a disrupted

cell cycle;1 however, deficiency in
Rb is associated with:2,3 CDK4/6

– Evolution of a Cyclin D

CDK4/6-independent state
– Resistance to CDK4/6 inhibition
Rb–
 Most (> 90%) ER+ breast cancers
express functional Rb4 Rb

 Rb+ cancers driven by oncogenic X E2F

signaling pathways upstream of
cyclin D are expected to be Cell cycle
dependent on CDK4/6 activity for progression

cell proliferation

ER+, estrogen receptor-positive; Rb–, Rb-negative; Rb+, Rb-positive.

1. Shapiro GI. J Clin Oncol 2006;23:1770–1783; 2. Dean JL, et al. Oncogene 2010;29:4018–4032; 8
3. Ertel A, et al. Cell Cycle 2010;9:4153–4163; 4. Abraham R, et al. AACR 2014 [Abstract SY34-03].
Signaling Pathways Converge at Cyclin D

 Multiple different signaling pathways can activate cyclin D–CDK4/6

through regulation of cyclin D1,2

NF-κB STATs
ER/PgR/AR MAPKs

Pl3K/AKT/mTOR
Wnt/b-catenin
CDK4/6
Cyclin D

 Mitogenic signaling through ER and HER2 requires cyclin D1

– CCND1 (encoding cyclin D1) is a direct ER-target gene that is required for
estrogen-dependent cell proliferation3,4
– Cyclin D1-deficient mice are resistant to HER2-induced breast cancers5
– ER+/HER2+ cell lines are the most sensitive to CDK4/6 inhibition6
HER2, human epidermal growth factor receptor 2.
1. Lange CA, Yee D. Endocr Relat Cancer 2011;18:C19–C24; 2. Witzel II, et al. Biochem Soc Trans 2010;38:217–222;
3. Lukas J, et al. Mol Cell Biol 1996;16:6917–6925; 4. Prall OW, et al. J Steroid Biochem Mol Biol 1998;65:169–174; 9
5. Yu Q, et al. Nature 2001;411:1017–1021; 6. Finn RS, et al. Breast Cancer Res 2009;11:R77.
 Cell Cycle Control and Estrogen Signaling

M
G2 • Preclinical evidence suggests that
E2F1-3
pRb
o In AI resistance models, ER
drives a CDK4/E2F-dependent
Cyclin D1
transcriptional program
CDK4 o CDK4/6 inhibition reduces cell
S proliferation in both ER-
Cyclin D1
dependent and ER-independent,
CDK6
AI-resistant breast cancer models
E2F1-3 Cyclin E
o Dysregulation of the cell cycle
CDK2
P P P P may lead to endocrine resistance
P P

pRb

G1

AI, aromatase inhibitor; CDK, cyclin-dependent kinase; ER, estrogen receptor; PI3K, phosphatidylinositol-3 kinase; Rb, retinoblastoma protein.

van den Heuvel S, Dyson NJ. Nat Rev Mol Cell Biol. 2008;9(9):713-724; Osborne CK, Schiff R. Annu Rev Med. 2011;62:233-247. 10
CDK4/6 and Resistance to Endocrine Therapy

 Endocrine (hormonal) therapy is the predominant front-line

treatment for patients with HR+ breast cancer; however:1
– Not all patients respond (de novo resistance)
– Patients who do respond eventually relapse (acquired resistance)2

 Resistance to endocrine therapy can be the result of activated

cyclin D–CDK4/63,4

 CDK4/6 inhibition may overcome endocrine resistance

CDK4/6 inhibition blocks

cell cycle progression in
endocrine-resistant breast
cancer cells5

HR+, hormone receptor-positive.

1. Cardoso F, et al. Ann Oncol 2014;25:1871–1888; 2. Johnston SR, et al. Clin Cancer Res 2010;16:1979–1987;
3. Thangavel C, et al. Endocr Relat Cancer 2011;18:333–345; 4. Lundgren K, et al. Breast Cancer Res 11
2012;14:R57; 5. Finn RS, et al. Breast Cancer Res 2009;11:R77.
CDK4/6 Pathway Disruption in Breast Cancer

Inactivating alterations of TP53

Amplification of CDK4 and CDK6
HER2 occur in 84% of basal and ≈27% of
Estrogen occurs in 16% and 17% of breast
non-basal tumors1
cancer, respectively1
p21 p53
PI3K CDK4/6
AKT
mTOR
Cyclin D p16 Loss of p16 occurs
in ≈49% of breast
cancer2
ER Amplification of CCND1 occurs in
35% of breast cancer1

P P
P
P P CDK4/6
Rb
Cyclin D

Rb
E2F X E2F

1. TCGA, Nature 2012;490:61–70; 2. Geradts J, Wilson PA. Am J Pathol 1996:149:15–20. 12

 Action of Different Agents

Alpelisib is a PI3K Buparlisib is a pan Everolimus is an Ribociclib is a

alpha inhibitor PI3K inhibitor mTOR inhibitor CDK4/6 inhibitor

Alpelisib
Buparlisib Everolimus Ribociclib
Estrogen HER2
Letrozole is a
nonsteroidal p53
p21
aromatase inhibitor PI3K CDK4/6
AKT
mTOR
Letrozole Cyclin D p16

Fulvestrant ER

Fulvestrant is an
ER downregulator
P P
P
P P CDK4/6
Rb
Cyclin D

Rb
E2F X E2F

13
 Biomarkers Development

Biomarkers are measured values that are associated with disease outcome,
which may be predictive or prognostic

Predictive biomarkers note if

the outcome of treatment is Prognostic biomarkers can
different for patients with a give information about
marker than for patients outcome regardless of
without the marker treatment received

Ballman KV. J Clin Oncol. 2015;33(33):3968-3973. 14

Examples of Biomarkers in Breast Cancer

Biomarkers in Routine Investigational

Clinical Use1,2 Biomarkers1
Pathway Candidate Drug Pathway Candidate Drug

ESR1 Steroid hormone Not known

HR+, HER2– disease signaling

ER/PgR Steroid Endocrine-based PIK3CA PI3K/AKT/mTOR PI3K inhibitor

hormone therapies
signaling
Cyclin D1 Cell proliferation CDK4/6 inhibitor

Ki67 Cell growth Multiple

Germline Predisposition (predictive of
response to
BRCA1/2 DNA repair PARP inhibitor therapy)

AKT, protein kinase B; BC, breast cancer; CDK, cyclin-dependent kinase; ER, estrogen receptor;
HER2–, human epidermal growth factor receptor 2-negative; mTOR, mammalian target of rapamycin;
PARP, poly ADP ribose polymerase; PgR, progesterone receptor; PI3K, phosphatidylinositol 3-kinase.

1. Arnedos M, et al. Nat Rev Clin Oncol. 2015;12(12):693-704; 2. Couch FJ, et al. Science. 2014;343(6178):1466-1470. 15
 Status of Biomarkers of CDK4/6 Inhibitor
Sensitivity and Primary Resistance

Sensitivity Biomarkers Tumor Types Resistance Biomarkers Tumor Types

Accepted in the clinic Accepted in the clinic
 ER positive HR+ MBC  RB loss Multiple
Potential  p16 overexpression HPV-positive
 Cyclin D1 overexpression MCL, esophageal cancers (eg.
 CDK4 overexpression Liposarcoma, GBM cervical cancer,
 p16 loss Melanoma, others HNSCC)
Potential
 Cyclin E Uterine (40%)
overexpression Bladder (15%)
Breast (15%)
Ovarian (20%)
Prostate (20%)
 E2F overexpression Bladder (30%)

Knudsen ES, et al. Trends in Cancer. 2017;3:39-55. 16

Ongoing Trials Evaluating CDK4/6 Inhibitors
Using Patient (Biomarker) Selection

Trial Tumor Type Selection Criteria Treatment

NCI-MATCH Cyclin D1-3

Multiple Palbociclib
(NCT02465060) amplification

CDK4
amplification/mutation
CDK6
SIGNATURE
Multiple amplification/mutation Ribociclib
(NCT02187783)
Cyclin D1, D3
amplification
p16 mutation

17
Combination Therapy: Rationale

Growth factor Cell membrane

RTK (eg, EGFR, HER2)

RAS
Buparlisib PI3K
RAF
MEK
Everolimus mTOR
P
Rb
P

Rb E2F Nucleus
Cyclin D ↑ D
CDK4/
CDK6
X
AI G1-S transition
Fulvestrant ER E2F gene expression
Palbociclib
Tamoxifen Ribociclib
Abemaciclib

O’Leary B, et al. Nat Rev Clin Oncol. 2016;13:417-430.

Overview of Trials With CDK4/6 Inhibition
in HR+, HER2– Breast Cancer

Primary Disease • Palbociclib

HR+ BC Adjuvant therapy recurrence – PALOMA-1/2
• Ribociclib
Endocrine • Palbociclib Endocrine – MONALEESA-2

First-line
therapy
+ CDKi – PENELOPE-B + CDKi – MONALEESA-3
– PALLAS – MONALEESA-7
• Abemaciclib
– MONARCH-2
– MONARCH-3
De novo
metastatic First-line therapy Disease Subsequent treatment
HR+ BC progression
Endocrine • Palbociclib
– PALOMA-1/2
Endocrine • Palbociclib
+ CDKi + CDKi
• Ribociclib – PALOMA-3
– MONALEESA-2 – PEARL
– MONALEESA-3 • Ribociclib
– MONALEESA-7 – MONALEESA-3
• Abemaciclib • Abemaciclib
– MONARCH-2 – MONARCH-2
– MONARCH-3

BC, breast cancer; CDKi, cyclin-dependent kinase inhibitor; HER2–, human epidermal growth factor receptor-2–negative; 19
HR+, hormone receptor-positive.
Summary

• Cell cycle pathway disruption is a hallmark of cancer

- CDK4/6 controls cell cycle progression
- Dysregulation of the cell cycle may lead to endocrine resistance

• The cyclin D–CDK4/6–INK4–Rb pathway is a rational target for the

treatment of breast cancer
- Selective CDK4/6 inhibitors show promise in a broad group of cancers, including
breast cancer

• Preclinical and clinical studies are evaluating biomarkers that may

help identify patients who may benefit from CDK4/6 inhibitor therapy
• Combination regimens may improve anticancer activity and overcome
resistance
- Selective CDK4/6 inhibitors, in combination with endocrine therapy, have been
clinically tested in patients with HR+/HER2- MBC

20