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In vitro synthesis of primary specific anti-breast cancer antibodies by normal human peripheral blood mononuclear cells

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Abstract

In this study, we developed a unique in vitro model to mimic the endogenous tumor microenvironment to understand the effect of immunotherapy with activated T-cells (ATC) armed with anti-CD3 × anti-Her2 bispecific antibody (aATC) on antibody response by naive immune cells. This model contained a co-culture of naïve peripheral blood mononuclear cells (PBMC), breast cancer cells (SK-BR-3), ATC or aATC and CpG ODNs. Culture supernatants were tested at various time points for anti-SK-BR-3 antibodies by ELISA, Western blot and flow cytometry. PBMC cocultured with non-irradiated aATC or irradiated (*) aATC showed significant increases in anti-tumor antibody production at day 14 (P < 0.0001) in the presence of CpG-ODN compared to unstimulated PBMC cultures (n = 9). Antibody specificity was confirmed by ELISA, Western blot and flow cytometry. Co-cultures containing *aATC and CpG showed significantly enhanced levels of IgG2 (P < 0.001) and cytokines that promote IgG2 synthesis including IL-13 (P < 0.02), IFNγ (P < 0.01) and GM-CSF (P < 0.05) compared to unstimulated PBMC control (n = 3). We show that aATC targeting and lysis of tumor cells induces an anti-tumor antibody response in our in vitro model. This model provides a unique opportunity to evaluate the interactions of T-cells, B-cells, and antigen-presenting cells leading to specific anti-tumor antibody responses.

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Acknowledgments

These studies were funded in part by R01 CA 092344 (L.G.L.), R01 CA 140412 (L.G.L), 5P39 CA 022453 from National Cancer Institute, Translational Grants #6066-06 and #6092-09 from the Leukemia and Lymphoma Society (L.G.L), Susan G. Komen Foundation Translational Grant #BCTR0707125 (L.G.L), and Michigan Cell Therapy Center for Excellence Grant from the State of Michigan #1819, and startup funds from the Barbara Ann Karmanos Cancer Institute.

Conflict of interest

The authors have no financial conflict of interest. L.G.L. is a founder of Transtarget, Inc.

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  1. Bone Marrow Transplantation and Immunotherapy Program, Departments of Oncology, Medicine, Immunology and Microbiology, Wayne State University School of Medicine, 731 Hudson Webber Cancer Research Center, 4100 John R., Detroit, MI, 48201, USA

    Archana Thakur, Oxana Norkina & Lawrence G. Lum

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  1. Archana Thakur
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  2. Oxana Norkina
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Correspondence to Archana Thakur.

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Thakur, A., Norkina, O. & Lum, L.G. In vitro synthesis of primary specific anti-breast cancer antibodies by normal human peripheral blood mononuclear cells. Cancer Immunol Immunother 60, 1707–1720 (2011). https://doi.org/10.1007/s00262-011-1056-9

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