NY-ESO-1 Specific T Cell Therapy for Patients with Metastatic Sarcoma

Seth M. Pollack, MD

Associate in Clinical Research, Fred Hutchinson Cancer Research Center

Acting Instructor, University of Washington

Cancer immunology review:

Sarcoma Cell

Tumor specific proteins are composed of peptides

CD8+ T cell

Tumor specific T cells recognize

peptides displayed in the MHC

complex..

MHC complex

Different HLA types:

EFFECTOR CELLS

Endogenous

Non-specific

LAK

TIL

Specific

CD8

CD4

Genetically modified

Re-directed specifity

TCR

CAR

Enhanced Function

Pollack SM and Yee C Innate Immune Regulation and Cancer Immunotherapy Springer 2012

EFFECTOR CELLS

Endogenous

Non-specific

LAK

TIL

Specific

CD8

CD4

Genetically modified

Re-directed specifity

TCR

CAR

Enhanced Function

Pollack SM and Yee C Innate Immune Regulation and Cancer Immunotherapy Springer 2012

Robbins et al., JCO 2011

Cancer Testis Antigens are Promising Targets

• In adults, only expressed in testis (some are also expressed in placenta)

• Expressed in many cancers

• Epigenetically regulated***

• Unknown biologic function

•Includes NY-ESO-1, LAGE-1, PRAME, MAGE-A3A: MAGE-A4 in tests

B: MAGE-C1 in fetal ovaryC: MAGE-A in trophoblastic placentaD: NY-ESO-1 in bladder cancer

Simpson AJ Nat Rev Cancer, 2005

NY-ESO-1 Is a Cancer Testis Antigen Frequently Expressed Target In Synovial Sarcoma

• NY-ESO stained 20/25 patients. • Staining homogonous in 14/20 patients

• 4 of the 5 patients without NY-ESO expression were biphasic phenotype (usually ssx-1)

• Targeted in the NCI Surgery Branch Study

Jungbluth AA et al., International Journal of Cancer, 2001

Computed tomography scans demonstrating tumor regression.

Robbins P F et al. JCO 2011;29:917-924

©2011 by American Society of Clinical Oncology

Question: Are there other sarcoma subtypes that frequently express NY-ESO-1?

• We examined 25 Myxoid Liposarcoma tumors, all expressed NY-ESO-1 (100%)

• >70% of cases, homogenous

• In all but 2 case, patients would qualify for trials of NY-ESO-1 directed therapy based on staining.

• MRCL cell lines can be lysed using NY-ESO-1 specific effectors

• No other disease (including synovial sarcoma) expresses NY-ESO-1 with this frequency

Myxoid/ Round Cell Liposarcomas Always Express NY-ESO-1, Usually Homogenously

Pollack et al., Cancer 2012

Question: If Cancer Testis Antigens are epigenetically regulated, can sarcomas be induced to express them?

Decitabine induces NY-ESO-1 and MAGE-A3 in tumors from lung

cancer patients

• Patients on a phase I trial of decitabine for lung cancer

• Serial biopsies performed

• Analyzed retrospectively for CT Antigen Expression

• Increased expression following treatment

•This increase was sustained months after treatment

Schrump D S et al. Clin Cancer Res 2006;12:5777-5785

A minority of chondrosarcoma tumors express NY-ESO-1/LAGE-1.

•36% of tumors expressed either NY-ESO-1 or LAGE-1 at a level that might be targeted – could this be improved?

Pollack et al. Plos One, 2012

JJ FS 105KC SW13530.0001

0.0010.01

0.11

10

JJ FS 105KC SW13530.0001

0.0010.01

0.11

10

JJ FS 105KC SW13530.0001

0.0010.01

0.11

10

A. NY-ESO-1 Expression in Chondrosarcoma Cell Lines Relative to Mel375

B. PRAME Expression in Chondrosarcoma Cell Lines Relative to Mel375

C. LAGE-1s Expression in Chondrosarcoma Cell Lines Relative to JJ

Untreated 5-Aza-dC

Decitabine (5-Aza-dC) Treatment Increases mRNA Expression of NY-ESO-1, LAGE-1s and PRAME by qPCR.

Pollack et al. Plos One, 2012

10:1 20:1 40:1 MART-1-5.00%

0.00%

5.00%

10.00%

15.00%

20.00%

25.00%

A. Percentage Lysis of FS Using NY-ESO-1/LAGE-1s Specific Effectors

FS FS + 5-Aza-dC

10:1 20:1 40:1 MART-10.00%

15.00%

30.00%

45.00%

60.00%

B. Percentage Lysis of JJ Using NY-ESO-1/LAGE-1s Specific Effectors

JJ JJ+5-Aza-dC

10:1 20:1 40:1 MART-1-5.00%

0.00%

5.00%

10.00%

15.00%

20.00%

25.00%

C. Percentage Lysis of FS Using PRAME Specific Effectors

FS FS + 5-Aza-dC

10:1 20:1 40:1 MART-10.00%5.00%

10.00%15.00%20.00%25.00%30.00%35.00%

D. Percentage Lysis of JJ Using PRAME Specific Effectors

JJ JJ+5-Aza-dC

Lysis of the FS and JJ Cell Lines With and Without Decitabine (5-Aza-dC) Using NY-ESO-1/LAGE-1s and PRAME Specific Effector T

Cells

Pollack et al. Plos One, 2012

Question: Can we isolate and expand NY-ESO-1 specific T cells from sarcoma patients with NY-ESO-1 expressing tumors?

Generation of Autologous NY-ESO-1 Specific T Cells

Step 1: Leukapharesis

Step 2: Generation of dendritic cells

CD25 depletion (to reduce regulatory T cells)

Step 3: Stimulation of T cells using peptide pulsed dendritic cells

Step 4: Clinical grade sorting

Rapid Expansion

Examples of Clinical Grade Products

Sort and Expand

Sort and Expand

NY-ESO-1 Tetramer

CD8

+ 3 additional wells <1% tetramer +

Patient 2537-S02

Patient 2537-S04

+ 1 additional well <1% tetramer +

T2 un-

pulsed

T2 pulsed A3750.00%

20.00%

40.00%

60.00%

80.00%

100.00%

T2 un-

pulsed

T2 pulsed A3750.00%

10.00%20.00%30.00%40.00%50.00%60.00%70.00%80.00%90.00%

100.00%

tetramer

Peptide Titration With NY-ESO-1 Specific Effectors:

Pros and cons of autologous vs. virally transfected NY-ESO-1 specificity?

FHCRC Protocol #2537

• Autologous NY-ESO-1 specific T cell therapy • Metastatic or unresectable Synovial Sarcoma

and Myxoid/round cell liposarcoma • Patient must have HLA type A*0201• 2 patients have been treated• 4 additional patients have T cell products

frozen • All finalized products kill tumor in vitro.• Lymphodelpetion with Cyclophosphamide as

is considered standard

-3 2 7 12 17 22 27 320

5

10

15

20

25

Day (infusion-day 0)

Adoptively transferred NY-ESO-1 Specific T cells Persist Following Infusion (from first patient treated)

% T

etra

mer

+ (%

of C

D8+

cel

ls)

6 wks 8 wks 10 wks

Prior to infusion 4 weeks post- infusion

Elimination of some small nodules

Pre-treatment Post-treatment (10 weeks)

Response in Large Right Upper Lobe Tumor

But the response was mixed …

• By IHC, few CD3+ cells could be seen in the tumor.• We expanded these few cells and a very small percentage were tetramer positive (a lower percentage than was in the peripheral blood). • These few cells could be sorted and expanded and remained functional.

NY-ESO-1 Tetramer

T2 unpulse

d

T2+N

Y-ESO

A375 tumor

526 tumor

0%10%20%30%40%50%60%70%80%

Sorted and Expanded Cells from Tumor Still Kill Tumor in

vitro

4/9/2012 7/2/2012

NY-ESO-1 Remains Post-Tx (20x)

Conclusion

• NY-ESO-1 is generally homogenously expressed in Myxoid/ Round Cell Liposarcoma tumors

• NY-ESO-1/LAGE-1s and PRAME can be up-regulated in Chondrosarcoma cell lines

• NY-ESO-1 specific T cells can be isolated and expanded from patients with NY-ESO-1 expressing sarcomas

Future Directions

• Expand to different HLA types, cancer testis antigens and sarcoma subtypes.

• Explore mechanisms of immune evasion in sarcoma tumors.

THANK YOU, SARC!!!

Acknowledgements:

FHCRC Immunology

Program:

Eric Farrar

Ivy Lai

Dawn Stief

Heather Sloan

Mentors: Cassian Yee, Stan Riddell

Robin Jones

Funding:

• SARC Career Development Award• Doug and Maggie Walker Foundation• Gilman Sarcoma Foundation

The SCCA Sarcoma Group

Chappie Conrad

Janet Eary

Doug Hawkins

Darin Davidson

Elizabeth Loggers

Gabrielle Kane

Edward Kim

Benjamin Hoch

Gary Mann

Venu Pillarisetty

Jennifer Hammilton

MSKCC/LudwigAchim JungbluthSacha Gnjatic