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

Rachel E. Beard, Zhili Zheng, Kiran H. Lagisetty, William Burns, Eric Tran, Stephen M. Hewitt, Daniel Abate-Daga, Shannon F. Rosati, Howard A. Fine, Soldano Ferrone, Steven A. Rosenberg, Richard A. Morgan

Research output: Contribution to journalArticle

Abstract

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.

Original languageEnglish (US)
Article number25
JournalJournal for ImmunoTherapy of Cancer
Volume2
Issue number1
DOIs
StatePublished - Aug 19 2014
Externally publishedYes

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Antigen Receptors
Neoplastic Stem Cells
Histology
Glioblastoma
Neoplasms
Melanoma
Protein Array Analysis
Mesothelioma
Neoplasm Antigens
Lymphocytes
Coculture Techniques
Immunotherapy
Reverse Transcription
Immunohistochemistry
chondroitin sulfate proteoglycan 4
Triple Negative Breast Neoplasms
Cytokines
Technology
Cell Line
Polymerase Chain Reaction

Keywords

  • Cancer stem cells
  • Chimeric antigen receptor
  • CSPG4
  • Glioblastoma
  • Immunotherapy
  • Melanoma

ASJC Scopus subject areas

  • Oncology
  • Immunology and Allergy
  • Cancer Research
  • Molecular Medicine
  • Pharmacology
  • Immunology

Cite this

Multiple chimeric antigen receptors successfully target chondroitin sulfate proteoglycan 4 in several different cancer histologies and cancer stem cells. / Beard, Rachel E.; Zheng, Zhili; Lagisetty, Kiran H.; Burns, William; Tran, Eric; Hewitt, Stephen M.; Abate-Daga, Daniel; Rosati, Shannon F.; Fine, Howard A.; Ferrone, Soldano; Rosenberg, Steven A.; Morgan, Richard A.

In: Journal for ImmunoTherapy of Cancer, Vol. 2, No. 1, 25, 19.08.2014.

Research output: Contribution to journalArticle

Beard, RE, Zheng, Z, Lagisetty, KH, Burns, W, Tran, E, Hewitt, SM, Abate-Daga, D, Rosati, SF, Fine, HA, Ferrone, S, Rosenberg, SA & Morgan, RA 2014, 'Multiple chimeric antigen receptors successfully target chondroitin sulfate proteoglycan 4 in several different cancer histologies and cancer stem cells', Journal for ImmunoTherapy of Cancer, vol. 2, no. 1, 25. https://doi.org/10.1186/2051-1426-2-25
Beard, Rachel E. ; Zheng, Zhili ; Lagisetty, Kiran H. ; Burns, William ; Tran, Eric ; Hewitt, Stephen M. ; Abate-Daga, Daniel ; Rosati, Shannon F. ; Fine, Howard A. ; Ferrone, Soldano ; Rosenberg, Steven A. ; Morgan, Richard A. / Multiple chimeric antigen receptors successfully target chondroitin sulfate proteoglycan 4 in several different cancer histologies and cancer stem cells. In: Journal for ImmunoTherapy of Cancer. 2014 ; Vol. 2, No. 1.
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AU - Zheng, Zhili

AU - Lagisetty, Kiran H.

AU - Burns, William

AU - Tran, Eric

AU - Hewitt, Stephen M.

AU - Abate-Daga, Daniel

AU - Rosati, Shannon F.

AU - Fine, Howard A.

AU - Ferrone, Soldano

AU - Rosenberg, Steven A.

AU - Morgan, Richard A.

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KW - Cancer stem cells

KW - Chimeric antigen receptor

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KW - Glioblastoma

KW - Immunotherapy

KW - Melanoma

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