Chimeric Antigen Receptor (CAR) T cell Therapy for B cell Cancers

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Chimeric Antigen Receptor (CAR)

T cell Therapy for B cell Cancers

13th_{International Congress on Targeted Anticancer Therapies (TAT)}

Paris France March 2, 2015 Renier Brentjens MD PhD Associate Member Leukemia Service

Chief, Cellular Therapeutics Center Department of Medicine

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Renier Brentjens MD PhD

•Stockholder: Juno Therapeutics (scientific co-founder)

•Royalties: Juno Therapeutics •Honoraria: none

•Reserch Funding: Juno Therapeutics •Consultant fees: Juno Therapeutics

•Discussion of off-label drug use: Tocilizumab

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Generation of a tumor targeted chimeric

antigen receptor (CAR)

V_H V_L 5’ LTR α-tumor scFv CD8 ζ chain 3’ LTR ψ SD SA α-TAA mAb TCR complex

CAR retroviral vector α-TAA scFv—CD8-ζ

(4)

TAA CAR VH VL 5’ LTR αTAA scFv CD8 ζ chain 3’ LTR ψ SD SA

Generation of TAA-targeted

T cells for treatment of Cancer

αTAA scFv CD8 CD3 ζ 1. Construct a chimeric antigen receptor (CAR) SFG-CAR 2. Subclone CAR gene into a retroviral vector (SFG) 3. Transduce and expand patient T cells ex vivo 4. Infuse transduced T cells to eradicate TAA+_{tumor cells}

TAA

Native TCR

(5)

• HLA-independent antigen recognition,

therefore universal application

• Active in both CD4

+

and CD8

+

T cells

• Target antigens include proteins,

carbohydrates and glycolipids

• Rapid generation of tumor specific T cells

• Minimal risk of autoimmunity or GvHD

• A living drug, single infusion

Advantages of CAR T cell

therapy

(6)

Expression of CD19 and other B

cell markers on B lineage cells

preB-ALL

B cell lymphomas

and leukemias myelomas

Stem Cell pro B _{pre B} _{immature B} _{mature B} _{plasma cell} CD19

CD22 CD20

(7)

(8)

Clinical trials using CD19

targeted T cells in relapsed B cell

ALL

(9)

Eligibility and Treatment Schema

• Adult patients are eligible (>18 year old).

• Patients must have B-ALL refractory, relapsed, MRD+, or in CR1. Ph+, extramedullary disease, prior history of CNS

leukemia, and/or

relapsed after prior allo- stem cell transplant are all eligible.

(10)

Sci Transl Med. 2013 Mar 20;5(177):177ra38

Patient characteristics and treatment

outcomes

(11)

Rapid tumor elimination and recovery

of normal bone marrow after 19-28z

CAR T cell therapy

(12)

Summary of Clinical

Outcomes

Number of Patients, N=27

Overall CR Rate

MRD Negative CR Rate

24/27 (89%)

21/24 (88%)

(13)

Overall Survival

Median Survival: 8.5 months

Survival Rate at 6 mo.: 57% (95% CI, 40-79)

Median Survival: 10.8 vs. 8.5 months Survival Rate at 6 mo.: 68% vs. 67%

0 12 24 36

0 50 100

Months After CAR T Cell Infusion

O v e ra ll S u rv iv a l

Survival proportions: Survival of Allo-SCT post-CAR Responders

Allo-SCT Post-CAR No SCT Post-CAR Post-CAR Allo-SCT vs. No SCT (N=24) P=0.5204 0 12 24 36 0 50 100

Months After CAR T Cell Infusion

O v e ra ll S u rv iv a l

Survival Curve for All Patients_{All Patients}

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Adverse Events: Cytokine Release

Syndrome

• Cytokine release syndrome (CRS)

– fevers, hypotension, hypoxia, malaise

• Neurologic changes*

– confusion, speech disorders, obtundation, seizure-like activities

• There is a strong correlation between sCRS and pre-T cell disease burden

• 0/10 sCRS in MRD patients

(15)

Maude et al N Engl J Med. 2014, 371:1507-17

UPenn studies of relapsed B-ALL

• 25 pediatric and 5 adult relapsed or

refractory B-ALL patients treated

• 19-4-1BBz CAR design

• 90% CR

• 6 month EFS 67%

• 6 month OSR 78%

(16)

Lee et al Lancet. Published online October 13, 2014

NCI studies of relapsed B-ALL

• 20 pediatric and young adult relapsed or

refractory B-ALL patients treated.

• 19-28z CAR design

• 70% CR (14/20)

• 60% MRD- CR

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Clinical trials using CD19

targeted T cells in low grade B

cell malignancies (CLL)

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Kalos et al Sci Trans Med 2011

UPenn clinical trial results

Patient Prior Chemotherapy Conditioning

Chemotherapy Response 1 Fludarabine, Rituximab, Alemtuzumab, R-CVP, Lenolidomide, PCR Bendamustine CR (3+ years) 2 Alemtuzumab Bendamustine/Rituximab PR (7 months) 3 Rituximab/Fludarabine, Rituximab/Bendamustine. Alemtuzumab Pentostatin/Cytoxan CR (3+ years)

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Updated UPenn Trials in CLL

(ASH 2013)

• Abstract 4162

– CD19 CAR T cells treating relapsed/refractory CLL

• Utilizing a 4-1BBz CAR construct, 14 CLL patients treated

• 3/14 patients obtained CR (21%), 5/14 patients obtained PR (36%), 6/14 patients with no response (43%)

• 6/14 patients with persistent detectable CAR T cells (5-35 months) • No CR patients with reported relapsed disease

• No dose response reported

• Abstract 873

– Dose randomized dose optimization trial of CLL patients with either high or low dose CAR T cell infusions

• Utilizing a 4-1BBz CAR construct, 27 CLL patients treated

• Patients randomized to either low dose (5 x 107_{CAR T cells) or high dose (5}

x 108_{CAR T cells)}

• No dose response benefit seen in these treated patients • Overall response rate (CR + PR) was 40%

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MSKCC clinical trial results: CLL

Cyclophosphamide

No

Cyclophosphamide

(22)

CAR questions

• What is the etiology of differential

responses by CAR T cell therapy to

relapsed B-ALL versus CLL?

• What is the role of bulky disease and CAR

T cell anti-tumor efficacy?

• The role of the hostile tumor

micro-environment and CAR T cell function

• How to build a better T cell?

• How can we extrapolate this technology to

other (solid) tumors?

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The hostile tumor microenvironment

The tumor microenvironment contains multiple

inhibitory factors designed to potentially

suppress effector T cells.

– CD4+_CD25hi_FoxP3+_{regulatory T cells (Tregs)}

– MDSCs – TAMs

– Expression of inhibitory ligands by tumor (PD-L1) – Tumor secretion of T cell suppressive cytokines

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19z1

+

Tregs abrogate anti-tumor

efficacy of 19z1

+

effector T cells

A * 0 20 40 60 80 100 0 10 20 30 40 50 100 Time (d) % Sur viva l 1928z Teff alone 1928z Teff + 19z1 Treg 19z1 Treg alone Pz1 Teff alone p < 0.001 (n=10) (n=10) (n=10) 0 20 40 60 80 100 0 10 20 30 40 50 100 Time (d) % Survival 1:16 Tregs 1:8 Tregs 1928z Teff alone 1:1 Tregs 1:4 Tregs Pz1 Teff alone (n=4) B (n=10) (n=10) (n=10) (n=10) p < 0.001 (n=10) p = 0.02

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The solution?

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Armored CAR T cells:

IL-12 secreting CAR

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IL-12

• A heterodimeric cytokine secreted by activated APCs, neutrophils and macrophages.

• Induces Th1 CD4+_{T cell response enhancing IL-2 and}

IFN-γ secretion

• Enhances T cell clonal expansion and effector function in concert with TCR signaling (signal 1) and CD28

co-stimulation (signal 2), serving as a signal 3. • Avoids/reverses T cell anergy

• May overcome Treg mediated effector T cell inhibition • Recruits and activates NK cells

• Clinical trials in cancer using systemic IL-12 therapy has been limited by severe inflammatory side effects

(29)

(30)

(31)

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IL-12 secreting CAR T cells in vivo

efficacy

(33)

IL-12 secreting CAR T cells are

(34)

Syngeneic EL4(hCD19) tumor model

mCD19-/-_hCD19 +/-mCD19-/-_hCD19 +/-IV injection EL4(hCD19) Harvest splenocytes IV injection Retroviral transduction with chimeric receptor Assess T cell homing to tumor Assess T cell eradication of tumor Assess T cell proliferation in vivo Assess long- term survival of T cells

Assess the efficacy of suicide vectors Assess memory T cell response to rechallenge with tumor 53%

Determine the side effects of therapy

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Pe

rcent

Su

rvival

Days since tumor cell injection

Lymphodepletion enhances anti-tumor

efficacy of 19z1

+

T cells

100 ₁₀1 ₁₀2 ₁₀3 ₁₀4 100 101 102 103 104 1Élive cells FL1-H: FL1-Height F L 2 -H : h C D 1 9 P E 24.3 0.11 0.063 75.5 24.3% 100 101 102 103 104 100 101 102 103 104 6Élive cells FL1-H: mCD19 F L 2 -H : h C D 1 9 2.07 0.02 0.29 97.6 2.07% 0 20 40 60 80 100 0 10 20 30 40 50 60 Time (d) % S u rv iv a l Cytoxan + 19z1 No Cytoxan + 19z1 Cytoxan Pz1

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Pre-cytoxan Post-cytoxan 0 2 4 6 8 10 F o xP 3 + c e lls i n p e ri p h e ra l b lo o d ( % ) * A B

Cyclophosphamide lymphodepletion reduces

Tregs and induces IL-12 and IFNγ secretion

Pegram et al Blood 2012 0 20 40 60 80 100 120 140 0 1 2 5 7 9

Time post-cytoxan (days)

se ru m c o n ce n tr a ti o n ( pg/ m l) IL-12 IFNγ

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19z1IRESIL-12 modified T cells secrete biologically active IL-12 and exhibit enhanced targeted cytotoxic function and resistance

to Tregs 0 10 20 30 40 19mzIRESIL-12 19mz E:T ratio S p e c if ic L y s is ( % ) 1:1 2.5:1 5:1 10:1 * * * * A B C _D Pegram et al Blood 2012

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A B

Syngeneic IL-12 secreting CD19 targeted

T cells induce B cell aplasias and tumor

eradication

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Complete eradication of ID8(MUC-CD) ovarian tumors in mice with MUC16 targeted T cells expressing IL-12

ID8(MUC-CD) lysis by CAR+ T cells

0 20 40 60 80 100 20 10 5 2.5 E : T Ratio Ly s is (% ) 19m28z 19m28zIresIL12 4H11m28z 4H11m28zIresIL12 A B C D 0 20 40 60 80 100 120 140 T N F -a ( p g /m l) ID8(MUC-CD) 19m28mz 19m28mzIresIL12 4H11delIresIL12 4H11mz 4H11m28mz 4H11m28mzIresIL12 0 10000 20000 30000 40000 50000 60000 70000 IF N -g ( p g /m l) ID8(MUC-CD) 19m28mz 19m28mzIresIL12 4H11delIresIL12 4H11mz 4H11m28mz 4H11m28mzIresIL12

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Enhanced CM phenotype, enhanced cytotoxicity, enhanced persistence Resistance to Treg and TGFβ inhibition NK cell Recruitment and activation IL-12 secretion IL-12 secretion Targeted tumor cytotoxicity Targeted tumor cytotoxicity Targeted tumor cytotoxicity Tumor cell NK cell Activated TIL Anergic TIL IL-12 secretion Reversal of anergy CAR-IRES IL-12

IL-12 genetically modified T cells:

Armored CAR T cells

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Conclusions

• Autologous CD19 targeted CAR modified T cells have demonstrated very promising anti-tumor efficacy in B cell ALL with more modest responses in patients with low grade B cell malignancies.

• Etiologies of CAR T cell resistance may be related to the hostile tumor microenvironment.

• Application of CAR T cell therapy for low grade B cell malignancies as well as moving forward towards application to solid tumor malignancies

requires “armored” CAR T cells designed to both overcome the hostile

tumor microenvironment and exhibit enhanced anti-tumor efficacy and long term persistence.

• Variations of “armored CAR” T cells appear to have enhanced anti-tumor efficacy based on pre-clinical anti-tumor models.

• Future studies using “armored” CAR T cell technology will focus on translation of these armored CAR T cells to the clinical setting both in the context hematological as well as solid tumor malignancies.

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Renier Brentjens Hollie Pegram Mythili Koneru Swarish Rafiq Swati Pendeharkar James Lee Yan Nikhamin Jae Park Kevin Curran Peter Chang Michel Sadelain Marco Davila Michael Gong

Jean Baptiste Latouche

Leukemia Service David Scheinberg Jae Park Mark Frattini Peter Maslak Mark Heaney Joe Jurcic Nicole Lamanna Marco Davila Dan Douer

Cell Therapy and Cell Engineering Facility

(Isabelle Riviere, Director)

R&D, Manufacturing Xiuyan Wang (Dan Hollyman) Jolanta Stefanski Malgorzata Olszewska Oriana Borquez-Ojeda Clare Taylor Teresa Wasielewska Jinrong Qu QA/QC Shirley Bartido (Mark Przybylowski) James Hosey Domenick Pirraglia Vanessa Capacio Clinical Research Yvette Bernal Funding

CA59350 (MS) ; P30 CA-008748 (CT); 3RO1CA138738-02S1(RJB); Alliance for Cancer Gene Therapy ; Terry Fox Run for Cancer Research; William H. Goodwin and Alice Goodwin, and the Commonwealth Cancer Foundation for Research and the ETC of MSKCC; Damon Runyon Clinical Investigator Award (RJB); William Lawrence & Blanche Hughes Foundation (RJB); CLL-Global Research Foundation (RJB)

Lymphoma Service Craig Moskowitz Ariela Noy GYN service Samith Sandadi Stephen Lee Roisin O’Clearbhail Adult BMT Service Sergio Geralt Craig Sauter Department of Clinical Laboratories Lillian Reich David Wuest Kathy Smith Biostatistics Glenn Heller

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