Checkpoint Blockade in Cancer Immunotherapy Prof. James ...The screen versions of these slides have...

Checkpoint Blockade

in Cancer Immunotherapy

Prof. James P. Allison

The screen versions of these slides have full details of copyright and acknowledgements 1

1

James P. Allison, Ph.D.

Howard Hughes Medical Institute

Memorial Sloan-Kettering Cancer Center

Checkpoint Blockade in Cancer Immunotherapy

2

Checkpoint BlockadeIn Cancer Immunotherapy

NSRRLLQSDYMNMTPRRPGLTRKPYQPYAPPp85, GRB2

PP2A

p85

KMLKKRSPLTTGYVKMPPTEPEDEKQFQPYFPNP

PP2A?AP-2

CD28

CTLA-4

5 months Post-RxPre-Rx

3

Two Signals Are Required for Activation of Naïve T Cells

“Professional” APC(Dendritic cells, etc.)

Ag-presenting cell

IL-2 secretionProliferation

Effector Function

TCR

CD28

Ag/MHC

B7

No IL-2Anergy?

Non-hematopoietic cellsTumor cells

Ag-presenting cell

Checkpoint Blockade




4

Tissue macrophage

B-cell

ICOSL

CD40

Germinal centre development

Antibody class switch

ICOSL

CD80

CD86

CTLA-4ICOS

CD28

TCR

CD40L

CD30

4.1BB

OX40

Peptide

MHC

CD40

CD153

4.1BBL

OX40L

Activation of APC

Regulating cytokines (TH1, TH2,

Treg)

T-cell toleranceTH1 polarisation

T-cell

Increase anti-apoptotic proteins

Regulate effector persistence

?

PD-1

PD-L1

PD-L2

??Arrest cells in G0/G1

Reduced cytokines

Costimulatory signals

Dendritic cell

Peptide

MHC

5

B7-1,2 Binding

Half-life

Cellular location

Functional Asymmetry of CD28 and CTLA-4

V

V

SS

SS

SS

CTLA-4

Constitutive Induced by TCR

CD28Costimulatory Inhibitory

Low avidity High avidity

>8 hrs

Expression

1-11/2 hrs

Surface, polarized

by TCR signals

Intracellular, translocated

by TCR signals

6

Peptide/MHC

CD28

TCR

No Proliferation

Anergy?

T Cell

Dynamic Integration of TCR and Costimulatory Signals

Antigen presenting cell

Cdk4

Cdk6Cyclin D3etc.

B7-1,2

Bcl-xL,γ

IL-2

p27kip

Activation,

Initiation

CTLA-4

InhibitionRestrictedProliferation

Checkpoint Blockade




7

Tubulin CD28 Overlay

Tubulin CTLA-4 Overlay

Localization of CD28 and CTLA-4 in Migrating T Cells

8

Localization of CD28 and CTLA-4to the T Cell-APC Interface

CD28 CTLA-4

~ 5 minutes

9

Do B7-1 or/and B7-2 Dimerize in the APC Membrane?

=> Fusions to “reconstituted” YFP

L myc B7-1 L50 N -YFP

L FLAG B7-1 L50 C -YFP

L myc B7-2 L50 N -YFP

L FLAG L50 C -YFPB7-2

Checkpoint Blockade




10

B7-1flag-C-YFP-Cy3B7-1myc-N-YFP-Cy5 YFP signal

B7-1-N-YFP and B7-1-C-YFP May Form Dimers, While B7-2-N-YFP

and B7-2-C-YFP Appear Monomeric

B7-2flag-C-YFP-Cy3B7-2myc-N-YFP-Cy5 YFP signal

11

CD28

0 20 40 60 80 100

WT

CTLA-4

0 20 40 60 80

WT

APCB7-2

(Based on S.J. Davis, S.J. Nathenson, and others)

CD28 and CTLA-4 Signaling Complexes Differ in Stoichiometry

B7-2

T cellCD28 CD28

B7-1

B7-2

CD28

B7-1

CTLA-4

APC

T cell

CTLA-4 CTLA-4

B7-1

B7-1 KO

B7-2 KO

B7-1 KO

B7-2 KO

12CTLA-4 B7-2CD28TCR B7-1

Checkpoint Blockade




13

CD28 and CTLA-4 Cytoplasmic Tails Control Signaling and Cellular Localization

NSRRLLQSDYMNMTPRRPGLTRKPYQPYAPARDFAAYRPP

GRB2CDCDCDCD28282828

PP2A,

KMLKKRSPLTTGYVKMPPTEPEDEKQFQPYFPN

CTLACTLACTLACTLA----4444PP2A?

�PI3K Binding�Internalization by clathrin-coated pits

�Ubiquitination and lysosomal targeting

P p85

p85

AP-2

14

StrongAgonist(MCC)

WeakAgonist(T102S)

DAPI/Tub./CTLA-4

DAPI/mIg/CTLA-4Tubulin mIg CTLA-4 Interface

Strong Agonist Peptides Are More Efficient at Localizing CTLA-4

to the Immunological Synapse

T

T

APC

APC

15

Strong agonist

More inhibition

Weak agonist

Less inhibition

APC

T-cell

CD28 and CTLA-4 Trafficking During T Cell/APC Interactions

MHC

B7

CTLA-4

CD28

TCR w/PTKs

Nucleus

MTOC/Golgi

Checkpoint Blockade




16

Differential Regulation of T Cell ResponsesAccording to Quality of TCR-pMHC Interaction

CFSE LabelLN T Cells

Anti-CTLA-4or

Control Mab

MCC (Strong Agonist: 50 nM KD)or

T102S (Weak Agonist: >400 nM KD)

Recipient

5C.C7 RAG-/-

(or 5C.C7 RAG-/-CTLA4-/-)

4 Days

5C.C7 cells in draining LN

Proliferation

Donor

17

CTLA-4 Preferentially Inhibits the Best-Fit Response

18TCR Affinity

T cell Stimulation

With CTLA-4Without CTLA-4

Threshold

Expansive Capacity

Clone

Attenuation

T cell Stimulation

Population

TCR Affinity

Models for Biological Role of CTLA-4

Checkpoint Blockade




19

TC

R A

vid

ity

Lymph NodeCompetition for APC

With CTLA-4

No CTLA-4

CTLA-4 Broadens Responding Repertoire by Constraining “Best Fit” T cells

Expansion

20

Biological Role of CTLA-4

T cell Stimulation

Attenuation

TCR Affinity

Expansive Capacity

# of Responders

Population

Clone

21

APC

Tumor

APC

IL-2

Attenuated

or Terminat edProliferation

CTLA-4 BlockadeEnhances Tumor-Specific Immune Responses

Unrestrained

Proliferation

� Necrotic Death � Chemotherapy� Irradiation� Hormone therapy� Anti-angiogenesis

� Vaccines

Checkpoint Blockade




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Anti-CTLA-4 Induces Regression of Transplantable Colon Carcinoma

0

50

100

150

200

0 3 6 9 12 15 18 21 24 27 30 33 35 38

Days After Tumor Injection

Ave

rage

T

umor

S

ize

(mm2)

Anti-CD28

Anti-CTLA-4

No Rx

Rx

23

Anti-CTLA-4 and GM-CSF Tumor Cell Vaccine Synergize to Eradicate Established B16 Melanoma

0 2416 20 2812 32 36

Days After Tumor Injection

Ave

rage

T

umor

S

ize

(mm2) No Rx

GM-VaccineAnti-CTLA-4

Both0

100

200

300

400

24

Skin and Hair Depigmentation FollowingRejection of B16 Melanoma

challenge

vaccination

rejected B16-F10 lung metastases

rejected day 8 tumor

CD4 depleted

vaccination

rejected day 0 tumor

Checkpoint Blockade




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No treatment

ααααCTLA-4/GVax Increases Number of Tumor Infiltrating T Cells

Foxp3CD4CD8

αCTLA-4/GVax

Foxp3CD4CD8

26

TCR

T cell

+ +

Anti-CTLA-

4 mAb

-

Cytotoxic T

cell

Regulatory

T cell

Inhibitory

cytokines

Perforin

Tumor

Dendritic cell

MHC

CD86

CD80

CD28

CTLA-4

CTLA-4

(a)

(b)

(c)

(d)

27

Treg in Lymph Nodes Expand,When Exposed to Anti-CTLA-4 In Vivo

# CD4+CD25+ # CD4+Foxp3+

2.5x106

2.0x106

1.5x106

1.0x106

5.0x105

0.0016

Ab

solu

te n

umb

er

Of

CD

4+ C

D2

5+

2.5x106

2.0x106

1.5x106

1.0x106

5.0x105

0.0011

Ab

solu

te n

umb

er

Of

CD

4+ F

oxp3

+

Checkpoint Blockade




28

Treg Activity Is Not Affected by Chronic Exposure to Anti-CTLA-4 In Vitro

29

Anti-CTLA-4 Does Not Block Treg Activity In Vitro

30

CD4/Foxp3 CD8/Foxp3

ααααCTLA-4/GVax Increases Teff/Treg Ratio in Tumor

Checkpoint Blockade




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Effective Combinations of Anti-CTLA-4Against Poorly Immunogenic Tumors

Immunotherapies

� Gvax: B16 melanoma, TRAMP CaP

� Peptide-pulsed (mugp100) DCs: B16 melanoma

� DNA vaccine (huTRP2): B16 Melanoma

� Prior depletion of CD25+ cells + vaccine: B16 melanoma

Conventional therapies

� Chemotherapy (cisplatin): Mammary carcinoma

� Local Irradiation: Mammary carcinoma

� Androgen deprivation: TRAMP CaP

� Surgical reduction: TRAMP CaP

Anything that kills tumor cells or primes T cells

32

Chimeric Murine CTLA-4 (Human Exon 2) Transgene

Regulated normally in transgenic mice:� Expressed only after activation,

not in naïve T cellsRescues phenotype in CTLA-4 -/- mice:

� No polyclonal T cell activation� Normal life span

17 Kb mouse genomic fragment with human exon 2 (ectodomain)

Exons 1 2 (Hu) 3 4

STATxOct-1 Ik-2 Oct-1AP-1

33

Effect of Anti-CTLA-4 on MC38 Tumor Growthin Mice Expressing Human CTLA-4

Day

0

100

200

300

400

500

600

700

800

900

Mean Tum

or Volum

e [m

m3]

6 8 10 12 14 16 18 20 22 24

Anti-Human CTLA-4 (147)

Anti-Human CTLA-4 (10D1) Anti-Mouse CTLA-4 (9H10)

Checkpoint Blockade




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MDX-010(Ipilumumab)

� Fully human IgG1 monoclonal antibody to human CTLA-4

created by Medarex

� Blocks binding of CTLA-4 to CD80 and CD86

� Augments immune responses in primate models

� Co-developed by Medarex and Bristol-Myers Squibb

in multiple cancer indications

� Phase III study in metastatic melanoma ongoing

� Phase II studies in renal cell carcinoma, prostate cancer,

ovarian cancer, and others

35

Clinical Response - Melanoma

Baseline and 5 months post-MDX-010 treatment CT scans of patient with metastatic melanoma (1 month status post dendritic cell vaccine)

Pre-treatment

Lung Mass

Pleural

Effusion

5 months Post-treatment

Residual

Mass

The patient continues without relapse at last reported follow-up visit

who experienced regression of all known sites of disease

36

MDX-010-05 Study DesignHLA-A2+, Stage IV Melanoma

(Rosenberg and colleagues, NCI)

Phan et al., PNAS. 2003 Jul;100(14):8372-8377

MDX-010 Dose Levels

3 mg/kg – 14 patients

3, 1, 1, 1 mg/kg – 27 patients

Follow Up

Q3mo x 1 year, q6mo x 2 yr, then annually

Option to retreat

Checkpoint Blockade




37

Complete Responder: Patient 11

Experienced complet e resolution of 2 subcutaneous nodules,

31 lung metastases and 0.5 cm brain metastasis

Complete Response:

Response associated with drug-related hypopituitar ism –

Responded to medical therapy

38

CTLA-4 Blockade in Metastatic Melanoma& Renal Cancer(Surgery Branch, NCI)

136

20 (15%)

44 (32%)

40+, 37+, 37+, 32+, 31+, 15+, 14+, 13+, 12+, 11+, 10+, 7+, 4+, 4+, 7, 7, 7, 5, 4, 4

(13 of 19 ongoing)

Total (number of patients)

Objective responses

Grade III/IV immune breakthrough events

Melanoma Renal

Duration (months)

(As of Oct. 2005)

7 (12%)

20 (33%)

15+, 11+, 6+, 18, 12, 8, 8

(3 of 7 ongoing)

61

39

Reversible Immune Mediated Toxicity Associated with Objective Tumor Responses

Sources: Phan et al., PNAS. 2003 Jul;100(14):8372-8377 and Medarex unpublished data

� Drug related inflammatory reactions consistent

with immune mediated toxicity

� Adverse events:

� Rash, colitis, hepatitis and hypopituitarism

� Resolved after symptomatic treatment

and/or corticosteroids and discontinuation

of MDX-010, except hypophysitis

� Objective responses are durable

Checkpoint Blockade




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MDX-010-020: Pivotal Phase III Trial

� 750 patients (HLA-A2+) with Stage III or Stage IV

metastatic melanoma

� Failed, relapsed or unable to tolerate IL-2, DTIC

or temozola mi de

� Blinded, randomized at 3:1:1

� MDX-010/MDX-1379 combination (450 patients)

� MDX-1379 alone (150 patients)

� MDX-010 alone (150 patients)

� MDX-010 (3 mg/kg) q 3 weeks x 4 doses

41

Autologous GVAX Followed by MDX-010in Melanoma and Ovarian Cancer

(Steve Hodi & Glenn Dranoff (DFCI)

� Anti-tumor effects with minimal adverse events

� Tumor specific effects of CTLA-4 blockade?

� Optimal timing of antibody in relation to prior GVAX?

42

Pt. MEL 15

July 2004 September 2005September 2004

GVAX: May to June 2004MDX-010: August 2004 to present q 2 mos

Toxicities limited to Grade 1 skin rash

Checkpoint Blockade




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Ovarian GVAX and Anti-CTLA-4 Ab

VAX

0

1000

2000

3000

4000

5000

6000

3/2

5/0

3

5/2

5/0

3

7/2

5/0

3

9/2

5/0

3

11/2

5/0

3

1/2

5/0

4

3/2

5/0

4

5/2

5/0

4

7/2

5/0

4

9/2

5/0

4

11/2

5/0

4

1/2

5/0

5

3/2

5/0

5

5/2

5/0

5

7/2

5/0

5

9/2

5/0

5

MDX-010

44

3/6/067/9/03

Reduction in Tumor Nodules Following MDX-010 Rx

45

“Lupus-Like” Rash in Ovarian Cancer

A B

C D

Checkpoint Blockade




46

0

1000

2000

3000

4000

5000

6000

3/2

5/0

3

5/2

5/0

3

7/2

5/0

3

9/2

5/0

3

11/2

5/0

3

1/2

5/0

4

3/2

5/0

4

5/2

5/0

4

7/2

5/0

4

9/2

5/0

4

11/2

5/0

4

1/2

5/0

5

3/2

5/0

5

5/2

5/0

5

7/2

5/0

5

9/2

5/0

5

MDX-010

Ovarian GVAX and Anti-CTLA-4 Ab

VAX

47

Contributors

Dept Medical Oncology

Tanja de Gruijl

Sinéad Lougheed

Helen Gall

Bob Pinedo

Beppe Giaccone

Winald Gerritsen

Fons van den Eertwegh

Dept Pathology

Saskia Santegoets

Anita Stam

Petra Scholten

Erik Hooijberg

Mary von Blomberg

Rik Scheper

Natalie Sacks

Kristen Hege

Shirley Clift

Karin Jooss

David Rhodes

Sayeh Morali

Israel Lowy

Steven Fischkoff

Elizabeth Levy

CELL GE�ESYSCELL GE�ESYS

48

GVAX Immunotherapy (CG1940/CG8711) + Ipilimumab (MDX-010: Anti-CTLA-4) for HRPC

VUmc Cancer Center Amsterdam

CG 1940 and CG8711 vaccinations every 2 weeks for a total of 13 vaccinations

MDX-010 every 4 weeks for 6 infusions

Follow-up visit 1 four weeks after the last

vaccination

Quarterly follow-up every 12 weeks

for progression and survival

Checkpoint Blockade




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PSA Curves – Dose Level 3 (3 mg/kg)

a:

b:

c:

13Mar 06: SAE - Hypophysitis (7 mo)

03Feb 06: Hypophysitis (5 mo)

09Feb 06: SAE – Hypophysitis (5 mo)

a

b

c

Pt 7 Pt 8

Pt 9

010

2030

4050

6070

8090

100

6/7/05

7/7/05

8/7/05

9/7/05

10/7/05

11/7/05

12/7/05

1/7/06

2/7/06

3/7/06

0

10

20

30

40

50

60

6/20/05

7/20/05

8/20/05

9/20/05

10/20/05

11/20/05

12/20/05

1/20/06

2/20/06

3/20/06

0510

152025

3035

404550

6/9/05

7/9/05

8/9/05

9/9/05

10/9/05

11/9/05

12/9/05

1/9/06

2/9/06

3/9/06

4/9/06

5015Sept05 29Mar06

Bone Scan Improvement in Patient 8 (3 mg/kg)

51

Checkpoint Blockade WorksCheckpoint Blockade WorksCheckpoint Blockade WorksCheckpoint Blockade Works(at least for CTLA-4)

Checkpoint Blockade




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� What distinguishes responders from non-responders?

� Are adverse events an inherent consequence of the therapeutic mechanism?

� Will short-term usage with vaccines/conventionaltherapies minimize adverse events?

� What are the best vaccines to be used combinatorially?

� What are the best conventional therapies to be used combinatorially?

53

Allison Lab Members

Emily CorseElisa CardenasMike Curran

Karl PeggsTsvet PenchevaVirginia Pedicord

Peter SavageAlejandro SepulvedaSergio Quezada

Becky WaitzJoyce WeiKatie Wojnoonski

Xingxing Zang

Former Colleagues

Cynthia ChambersMax KrummelAndy Hurwitz

Andrea van ElsasDana LeachJackson Egen

Mike KuhnsP’ng LokeMarcella Fasso

John Engelhardt

Collaborators

Eugene Kwon (Mayo)Norm Greenberg (FHCC)Kees Melief (Leiden)

Alan Korman (Medarex)

Glenn Dranoff (DFCI)Steve Rosenberg (NCI)

54

Maria ChoiKatie WojnoonskiVirginia PedicordXingxing ZangMike CurranJoyce Wei

Tsvet PenchevaPeter SavageCenk SumenBecky WaitzAlejandro Sepulveda

Elisa CardenasKarl PeggsJohn EngelhardtSergio QuezadaEmi ly CorseTyler

Checkpoint Blockade




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Checkpoint Blockade in Cancer Immunotherapy Prof. James ...The screen versions of these slides have...

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