Multiple chimeric antigen receptors successfully target chondroitin sulfate proteoglycan 4 in several different cancer histologies and cancer stem cells

Journal for ImmunoTherapy of Cancer

Richard A Morgan²  Steven A Rosenberg⁴  Soldano Ferrone³  Howard A Fine¹  Shannon F Rosati⁴  Daniel Abate-Daga⁴  Stephen M Hewitt⁵  Eric Tran⁴  William R Burns⁴  Kiran H Lagisetty⁴  Zhili Zheng⁴  Rachel E Beard⁴ 

[1] Division of Hematology and Medical Oncology, New York University Langone Medical Center, New York, New York, USA;Current address: Bluebird bio, 150 Second St, Cambridge, MA 02141, USA;Department of Surgery, Massachusetts General Hospital, 55 Fruit Street, Boston, MA 02114, USA;Surgery Branch, Center for Cancer Research, National Cancer Institute, 10 Center Drive, Building 10 Hatfield CRC, Rm 3-5930, 20892-1201 Bethesda, MD, USA;Laboratory of Pathology, Center for Cancer Research, National Cancer Institute, 10 Center Drive, Building 10 Hatfield CRC, MSC 4605, 20892-1201 Bethesda, MD, USA

关键词: Glioblastoma;    Melanoma;    Cancer stem cells;    Chimeric antigen receptor;    CSPG4;    Immunotherapy;   

Others  :  1139876
DOI  :  10.1186/2051-1426-2-25

 received in 2014-02-25, accepted in 2014-06-20,  发布年份 2014

【 摘 要 】

Background

The development of immunotherapy has led to significant progress in the treatment of metastatic cancer, including the development of genetic engineering technologies that redirect lymphocytes to recognize and target a wide variety of tumor antigens. Chimeric antigen receptors (CARs) are hybrid proteins combining antibody recognition domains linked to T cell signaling elements. Clinical trials of CAR-transduced peripheral blood lymphocytes (PBL) have induced remission of both solid organ and hematologic malignancies. Chondroitin sulfate proteoglycan 4 (CSPG4) is a promising target antigen that is overexpressed in multiple cancer histologies including melanoma, triple-negative breast cancer, glioblastoma, mesothelioma and sarcoma.

Methods

CSPG4 expression in cancer cell lines was assayed using flow cytometry (FACS) and reverse-transcription PCR (RT-PCR). Immunohistochemistry was utilized to assay resected melanomas and normal human tissues (n = 30) for CSPG4 expression and a reverse-phase protein array comprising 94 normal tissue samples was also interrogated for CSPG4 expression. CARs were successfully constructed from multiple murine antibodies (225.28S, TP41.2, 149.53) using second generation (CD28.CD3ζ) signaling domains. CAR sequences were cloned into a gamma-retroviral vector with subsequent successful production of retroviral supernatant and PBL transduction. CAR efficacy was assayed by cytokine release and cytolysis following coculture with target cell lines. Additionally, glioblastoma stem cells were generated from resected human tumors, and CSPG4 expression was determined by RT-PCR and FACS.

Results

Immunohistochemistry demonstrated prominent CSPG4 expression in melanoma tumors, but failed to demonstrate expression in any of the 30 normal human tissues studied. Two of 94 normal tissue protein lysates were positive by protein array. CAR constructs demonstrated cytokine secretion and cytolytic function after co-culture with tumor cell lines from multiple different histologies, including melanoma, breast cancer, mesothelioma, glioblastoma and osteosarcoma. Furthermore, we report for the first time that CSPG4 is expressed on glioblastoma cancer stem cells (GSC) and demonstrate that anti-CSPG4 CAR-transduced T cells recognize and kill these GSC.

Conclusions

The functionality of multiple different CARs, with the widespread expression of CSPG4 on multiple malignancies, suggests that CSPG4 may be an attractive candidate tumor antigen for CAR-based immunotherapies using appropriate technology to limit possible off-tumor toxicity.

【 授权许可】

   
2014 Beard et al.; licensee BioMed Central Ltd.

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【 参考文献 】

• [1]Rosenberg SA, Restifo NP, Yang JC, Morgan RA, Dudley ME: Adoptive cell transfer: a clinical path to effective cancer immunotherapy. Nat Rev Cancer 2008, 8:299-308.
• [2]Johnson LA, Morgan RA, Dudley ME, Cassard L, Yang JC, Hughes MS, Kammula US, Royal RE, Sherry RM, Wunderlich JR, Lee CC, Restifo NP, Schwarz SL, Cogdill AP, Bishop RJ, Kim H, Brewer CC, Rudy SF, VanWaes C, Davis JL, Mathur A, Ripley RT, Nathan DA, Laurencot CM, Rosenberg SA: Gene therapy with human and mouse T-cell receptors mediates cancer regression and targets normal tissues expressing cognate antigen. Blood 2009, 114:535-546.
• [3]Robbins PF, Morgan RA, Feldman SA, Yang JC, Sherry RM, Dudley ME, Wunderlich JR, Nahvi AV, Helman LJ, Mackall CL, Kammula US, Hughes MS, Restifo NP, Raffeld M, Lee CC, Levy CL, Li YF, El-Gamil M, Schwarz SL, Laurencot C, Rosenberg SA: Tumor regression in patients with metastatic synovial cell sarcoma and melanoma using genetically engineered lymphocytes reactive with NY-ESO-1. J Clin Oncol 2011, 29:917-924.
• [4]Morgan RA, Chinnasamy N, Abate-Daga D, Gros A, Robbins PF, Zheng Z, Dudley ME, Feldman SA, Yang JC, Sherry RM, Phan GQ, Hughes MS, Kammula US, Miller AD, Hessman CJ, Stewart AA, Restifo NP, Quezado MM, Alimchandani M, Rosenberg AZ, Nath A, Wang T, Bielekova B, Wuest SC, Akula N, McMahon FJ, Wilde S, Mosetter B, Schendel DJ, Laurencot CM, et al.: Cancer regression and neurological toxicity following anti-MAGE-A3 TCR gene therapy. J Immunother 2013, 36:133-151.
• [5]Parkhurst MR, Yang JC, Langan RC, Dudley ME, Nathan DA, Feldman SA, Davis JL, Morgan RA, Merino MJ, Sherry RM, Hughes MS, Kammula US, Phan GQ, Lim RM, Wank SA, Restifo NP, Robbins PF, Laurencot CM, Rosenberg SA: T cells targeting carcinoembryonic antigen can mediate regression of metastatic colorectal cancer but induce severe transient colitis. Mol Ther 2011, 19:620-626.
• [6]Cartellieri M, Bachmann M, Feldmann A, Bippes C, Stamova S, Wehner R, Temme A, Schmitz M: Chimeric antigen receptor-engineered T cells for immunotherapy of cancer. J Biomed Biotechnol 2010, 2010:956304.
• [7]Jena B, Dotti G, Cooper LJ: Redirecting T-cell specificity by introducing a tumor-specific chimeric antigen receptor. Blood 2010, 116:1035-1044.
• [8]Park TS, Rosenberg SA, Morgan RA: Treating cancer with genetically engineered T cells. Trends Biotechnol 2011, 29:550-557.
• [9]Park JR, Digiusto DL, Slovak M, Wright C, Naranjo A, Wagner J, Meechoovet HB, Bautista C, Chang WC, Ostberg JR, Jensen MC: Adoptive transfer of chimeric antigen receptor re-directed cytolytic T lymphocyte clones in patients with neuroblastoma. Mol Ther 2007, 15:825-833.
• [10]Kochenderfer JN, Dudley ME, Feldman SA, Wilson WH, Spaner DE, Maric I, Stetler-Stevenson M, Phan GQ, Hughes MS, Sherry RM, Yang JC, Kammula US, Devillier L, Carpenter R, Nathan DA, Morgan RA, Laurencot C, Rosenberg SA: B-cell depletion and remissions of malignancy along with cytokine-associated toxicity in a clinical trial of anti-CD19 chimeric-antigen-receptor-transduced T cells. Blood 2012, 119:2709-2720.
• [11]Brentjens RJ, Davila ML, Riviere I, Park J, Wang X, Cowell LG, Bartido S, Stefanski J, Taylor C, Olszewska M, Borquez-Ojeda O, Qu J, Wasielewska T, He Q, Bernal Y, Rijo IV, Hedvat C, Kobos R, Curran K, Steinherz P, Jurcic J, Rosenblat T, Maslak P, Frattini M, Sadelain M: CD19-targeted T cells rapidly induce molecular remissions in adults with chemotherapy-refractory acute lymphoblastic leukemia. Sci Transl Med 2013, 5:177ra138.
• [12]Kochenderfer JN, Rosenberg SA: Treating B-cell cancer with T cells expressing anti-CD19 chimeric antigen receptors. Nat Rev Clin Oncol 2013, 10(5):267-276.
• [13]Kochenderfer JN, Wilson WH, Janik JE, Dudley ME, Stetler-Stevenson M, Feldman SA, Maric I, Raffeld M, Nathan DA, Lanier BJ, Morgan RA, Rosenberg SA: Eradication of B-lineage cells and regression of lymphoma in a patient treated with autologous T cells genetically engineered to recognize CD19. Blood 2010, 116:4099-4102.
• [14]Porter DL, Levine BL, Kalos M, Bagg A, June CH: Chimeric antigen receptor-modified T cells in chronic lymphoid leukemia. New Engl J Med 2011, 365:725-733.
• [15]Grupp SA, Kalos M, Barrett D, Aplenc R, Porter DL, Rheingold SR, Teachey DT, Chew A, Hauck B, Wright JF, Milone MC, Levine BL, June CH: Chimeric antigen receptor-modified T cells for acute lymphoid leukemia. New Engl J Med 2013, 368:1509-1518.
• [16]Wang X, Wang Y, Yu L, Sakakura K, Visus C, Schwab JH, Ferrone CR, Favoino E, Koya Y, Campoli MR, McCarthy JB, DeLeo AB, Ferrone S: CSPG4 in cancer: multiple roles. Curr Mol Med 2010, 10:419-429.
• [17]Campoli M, Ferrone S, Wang X: Functional and clinical relevance of chondroitin sulfate proteoglycan 4. Adv Cancer Res 2010, 109:73-121.
• [18]Price MA, Colvin Wanshura LE, Yang J, Carlson J, Xiang B, Li G, Ferrone S, Dudek AZ, Turley EA, McCarthy JB: CSPG4, a potential therapeutic target, facilitates malignant progression of melanoma. Pigment Cell Melanoma Res 2011, 24:1148-1157.
• [19]Wang X, Osada T, Wang Y, Yu L, Sakakura K, Katayama A, McCarthy JB, Brufsky A, Chivukula M, Khoury T, Hsu DS, Barry WT, Lyerly HK, Clay TM, Ferrone S: CSPG4 protein as a new target for the antibody-based immunotherapy of triple-negative breast cancer. J Natl Cancer Inst 2010, 102:1496-1512.
• [20]Svendsen A, Verhoeff JJ, Immervoll H, Brogger JC, Kmiecik J, Poli A, Netland IA, Prestegarden L, Planaguma J, Torsvik A, Kjersem AB, Sakariassen PO, Heggdal JI, Van Furth WR, Bjerkvig R, Lund-Johansen M, Enger PO, Felsberg J, Brons NH, Tronstad KJ, Waha A, Chekenya M: Expression of the progenitor marker NG2/CSPG4 predicts poor survival and resistance to ionising radiation in glioblastoma. Acta Neuropathol 2011, 122:495-510.
• [21]Rivera Z, Ferrone S, Wang X, Jube S, Yang H, Pass HI, Kanodia S, Gaudino G, Carbone M: CSPG4 as a target of antibody-based immunotherapy for malignant mesothelioma. Clin Cancer Res 2012, 18:5352-5363.
• [22]Burns WR, Zhao Y, Frankel TL, Hinrichs CS, Zheng Z, Xu H, Feldman SA, Ferrone S, Rosenberg SA, Morgan RA: A high molecular weight melanoma-associated antigen-specific chimeric antigen receptor redirects lymphocytes to target human melanomas. Cancer Res 2010, 70:3027-3033.
• [23]Noronha EJ, Wang X, Desai SA, Kageshita T, Ferrone S: Limited diversity of human scFv fragments isolated by panning a synthetic phage-display scFv library with cultured human melanoma cells. J Immunol 1998, 161:2968-2976.
• [24]Kusama M, Kageshita T, Chen ZJ, Ferrone S: Characterization of syngeneic antiidiotypic monoclonal antibodies to murine anti-human high molecular weight melanoma-associated antigen monoclonal antibodies. J Immunol 1989, 143:3844-3852.
• [25]Temponi M, Gold AM, Ferrone S: Binding parameters and idiotypic profile of the whole immunoglobulin and Fab’ fragments of murine monoclonal antibody to distinct determinants of the human high molecular weight-melanoma associated antigen. Cancer Res 1992, 52:2497-2503.
• [26]REMBRANDT home page http://rembrandt.nci.nih.gov webcite
• [27]Lee J, Kotliarova S, Kotliarov Y, Li A, Su Q, Donin NM, Pastorino S, Purow BW, Christopher N, Zhang W, Park JK, Fine HA: Tumor stem cells derived from glioblastomas cultured in bFGF and EGF more closely mirror the phenotype and genotype of primary tumors than do serum-cultured cell lines. Cancer Cell 2006, 9:391-403.
• [28]Campoli MR, Chang CC, Kageshita T, Wang X, McCarthy JB, Ferrone S: Human high molecular weight-melanoma-associated antigen (HMW-MAA): a melanoma cell surface chondroitin sulfate proteoglycan (MSCP) with biological and clinical significance. Crit Rev Immunol 2004, 24:267-296.
• [29]Benassi MS, Pazzaglia L, Chiechi A, Alberghini M, Conti A, Cattaruzza S, Wassermann B, Picci P, Perris R: NG2 expression predicts the metastasis formation in soft-tissue sarcoma patients. J Orthop Res 2009, 27:135-140.
• [30]Wang X, Katayama A, Wang Y, Yu L, Favoino E, Sakakura K, Favole A, Tsuchikawa T, Silver S, Watkins SC, Kageshita T, Ferrone S: Functional characterization of an scFv-Fc antibody that immunotherapeutically targets the common cancer cell surface proteoglycan CSPG4. Cancer Res 2011, 71:7410-7422.
• [31]Mittelman A, Chen ZJ, Liu CC, Hirai S, Ferrone S: Kinetics of the immune response and regression of metastatic lesions following development of humoral anti-high molecular weight-melanoma associated antigen immunity in three patients with advanced malignant melanoma immunized with mouse antiidiotypic monoclonal antibody MK2-23. Cancer Res 1994, 54:415-421.
• [32]Quan WD Jr, Dean GE, Spears L, Spears CP, Groshen S, Merritt JA, Mitchell MS: Active specific immunotherapy of metastatic melanoma with an antiidiotype vaccine: a phase I/II trial of I-Mel-2 plus SAF-m. J Clin Oncol 1997, 15:2103-2110.
• [33]Mittelman A, Chen GZ, Wong GY, Liu C, Hirai S, Ferrone S: Human high molecular weight-melanoma associated antigen mimicry by mouse anti-idiotypic monoclonal antibody MK2-23: modulation of the immunogenicity in patients with malignant melanoma. Clin Cancer Res 1995, 1:705-713.
• [34]Sadelain M, Brentjens R, Riviere I: The basic principles of chimeric antigen receptor design. Canc Discov 2013, 3:388-398.
• [35]Song DG, Ye Q, Poussin M, Harms GM, Figini M, Powell DJ Jr: CD27 costimulation augments the survival and antitumor activity of redirected human T cells in vivo. Blood 2012, 119:696-706.
• [36]Geldres C, Savoldo B, Hoyos V, Caruana I, Zhang M, Yvon E, Del Vecchio M, Creighton CJ, Ittmann M, Ferrone S, Dotti G: T lymphocytes redirected against the chondroitin sulfate proteoglycan-4 control the growth of multiple solid tumors both in vitro and in vivo. Clin Cancer Res 2014, 20:962-971.
• [37]Clarke MF, Dick JE, Dirks PB, Eaves CJ, Jamieson CH, Jones DL, Visvader J, Weissman IL, Wahl GM: Cancer stem cells–perspectives on current status and future directions: AACR Workshop on cancer stem cells. Cancer Res 2006, 66:9339-9344.
• [38]Lamers CH, Sleijfer S, van Steenbergen S, van Elzakker P, van Krimpen B, Groot C, Vulto A, den Bakker M, Oosterwijk E, Debets R, Gratama JW: Treatment of metastatic renal cell carcinoma with CAIX CAR-engineered T cells: clinical evaluation and management of on-target toxicity. Mol Ther 2013, 21:904-912.
• [39]Morgan RA, Yang JC, Kitano M, Dudley ME, Laurencot CM, Rosenberg SA: Case report of a serious adverse event following the administration of T cells transduced with a chimeric antigen receptor recognizing ERBB2. Mol Ther 2010, 18:843-851.
• [40]Zhao Y, Wang QJ, Yang S, Kochenderfer JN, Zheng Z, Zhong X, Sadelain M, Eshhar Z, Rosenberg SA, Morgan RA: A herceptin-based chimeric antigen receptor with modified signaling domains leads to enhanced survival of transduced T lymphocytes and antitumor activity. J Immunol 2009, 183:5563-5574.
• [41]Zheng Z, Chinnasamy N, Morgan RA: Protein L: a novel reagent for the detection of chimeric antigen receptor (CAR) expression by flow cytometry. J Transl Med 2012, 10:29. BioMed Central Full Text
• [42]Morgan RA, Johnson LA, Davis JL, Zheng Z, Woolard KD, Reap EA, Feldman SA, Chinnasamy N, Kuan CT, Song H, Zhang W, Fine HA, Rosenberg SA: Recognition of glioma stem cells by genetically modified T cells targeting EGFRvIII and development of adoptive cell therapy for glioma. Hum Gene Ther 2012, 23:1043-1053.