Rapid generation of NY-ESO-1-specific CD4⁺ T_HELPER1 cells for adoptive T-cell therapy

Abstract

Tumor-associated antigens such as NY-ESO-1 are expressed in a variety of solid tumors but absent in mature healthy tissues with the exception of germline cells. The immune system anti-cancer attack is mediated by cell lysis or induction of growth arrest through paralysis of tumor cells, the latter of which can be achieved by tumor-specific CD4⁺, IFNγ-producing T_Helper type 1 (T_H1) cells. Translation of these immune-mediated mechanisms into clinical application has been limited by availability of immune effectors, as well as the need for complex in vitro protocols and regulatory hurdles. Here, we report a procedure to generate cancer-testis antigen NY-ESO-1-targeting CD4⁺ T_H1 cells in vitro for cancer immunotherapy in the clinic. After in vitro sensitization by stimulating T cells with protein-spanning, overlapping peptide pools of NY-ESO-1 in combination with IL-7 and low dose IL-2, antigen-specific T cells were isolated using IFNγ capture technique and subsequently expanded with IL-2, IL-7 and IL-15. Large numbers of NY-ESO-1-specific CD4⁺ T cells with a T_H1 cytokine profile and lower numbers of cytokine-secreting CD8⁺ T cells could be generated from healthy donors with a high specificity and expansion potential. Manufactured CD4⁺ T cells showed strong specific T_H1-responses with IFNγ⁺, TNFα⁺, IL-2⁺ and induced cell cycle arrest and apoptosis in tumor cells. The protocol is GMP-grade and approved by the regulatory authorities. The tumor-antigen specific CD4⁺ T_H1 lymphocytes can be adoptively transferred as a T-cell therapy to boost anticancer immunity and this novel cancer treatment approach is applicable to both T cells from healthy allogeneic donors as well as to autologous T cells derived from cancer patients.

Keywords:

• adoptive T-cell transfer
• CD4+ T_HELPER1 cells
• cell cycle arrest
• immunotherapy
• polyfunctional T cells

Abbreviations:

• ACT, adoptive T cell transfer
• CTL, cytotoxic T lymphocyte
• DC, dendritic cell
• GMP, good manufacturing practice
• NK, natural killer
• TIL, tumor-infiltrating lymphocyte
• T_H1, T_Helper1
• Treg, regulatory T cell

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.

Acknowledgments

We thank Sylvia Klein, Christiane Braun, Adelheid Hauler, Marie-Luise Schwesinger, Roksana Wojcik and Sarah Bühler for their excellent technical assistance and Marco Sterk for kindly providing regulatory T cells as positive controls. We thank the Department of Pathology, University of Tübingen for kindly performing NY-ESO-1 immunhistochemistry of tumor cell lines and the Institute of Transfusion Medicine, University Hospital Tübingen for HLA typing and Miltenyi Biotec for providing CliniMACS materials free of charge.

Funding

This work was supported by the Deutsche Forschungsgemeinschaft (SFB685), the German Federal Ministry of Science and Education (BMBF, iVAC-ALL) and Fortüne-program (University of Tübingen, 1892–0–0 and 1977–0–1), the European Social Fund in Baden-Württemberg and the Institutional Strategy of the University of Tübingen (Deutsche Forschungsgemeinschaft, ZUK 63).