Unique characteristics of autoantibodies targeting MET in patients with breast and lung cancer
Michal Navon, Noam Ben-Shalom, Maya Dadiani, Michael Mor, Ron Yefet, Michal Bakalenik-Gavry, Dana Chat, Nora Balint-Lahat, Iris Barshack, Ilan Tsarfaty, Einav Nili Gal-Yam, Natalia T. Freund
Michal Navon, Noam Ben-Shalom, Maya Dadiani, Michael Mor, Ron Yefet, Michal Bakalenik-Gavry, Dana Chat, Nora Balint-Lahat, Iris Barshack, Ilan Tsarfaty, Einav Nili Gal-Yam, Natalia T. Freund
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Research Article Immunology

Unique characteristics of autoantibodies targeting MET in patients with breast and lung cancer

Abstract

The presence of B cells in tumors is correlated with favorable prognosis and efficient response to immunotherapy. While tumor-reactive antibodies have been detected in several cancer types, identifying antibodies that specifically target tumor-associated antigens remains a challenge. Here, we investigated the antibodies spontaneously elicited during breast and lung cancer that bind the cancer-associated antigen MET. We screened patients with lung (n = 25) and breast (n = 75) cancer and found that 13% had antibodies binding to both the recombinant ectodomain of MET, and the ligand binding part of MET, SEMA. MET binding in the breast cancer cohort was significantly correlated with hormone receptor–positive status. We further conducted immunoglobulin sequencing of peripheral MET-enriched B cells from 6 MET-reactive patients. The MET-enriched B cell repertoire was found to be polyclonal and prone to non-IgG1 subclass. Nine monoclonal antibodies were cloned and analyzed, and these exhibited MET binding, low thermostability, and high polyreactivity. Among these, antibodies 87B156 and 69B287 effectively bound to tumor cells and inhibited MET-expressing breast cancer cell lines. Overall, our data demonstrate that some patients with breast and lung cancer develop polyreactive antibodies that cross-react with MET. These autoantibodies have a potential contribution to immune responses against tumors.

Authors

Michal Navon, Noam Ben-Shalom, Maya Dadiani, Michael Mor, Ron Yefet, Michal Bakalenik-Gavry, Dana Chat, Nora Balint-Lahat, Iris Barshack, Ilan Tsarfaty, Einav Nili Gal-Yam, Natalia T. Freund

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Figure 4

MET-binding antibodies are polyreactive.

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MET-binding antibodies are polyreactive. (A) Comparison between denatura...
(A) Comparison between denaturation midpoint (left) and cumulant radius (right) of MET-binding mAbs (n = 8) and RBD-binding mAbs (n = 7). (B) Heatmap representing binding to non-MET proteins as detected by ELISA. Each column represents 1 mAb, and each row represents a single antigen. Color code is given on the right of the figure. mGO.53 is an isotype nonbinding control and TAU-2303 is an anti-RBD mAb that is not polyreactive (35). (C) Correlation between MET binding and polyreactive score, calculated by the average of all non-MET antigens at OD650. (D) Binding of 69B287, 87B156, their predicted germline versions, and germline-mutated chimeric versions to MET as detected by ELISA. (E) Heatmap representing binding by ELISA of 69B287, 87B156, their predicted germline versions, and germline-mutated chimeric versions to non-MET proteins. (F) Calculated AUC of the confluence measured by Incucyte of CAL-51, MDA-MB-468, and HCC70 breast cancer cell lines over time, incubated with each mAb (87B156, 69B287, and mGO.53), with 4–6 replicates for each condition. Data presented as mean ± SD. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 by 1-way ANOVA with Tukey’s multiple-comparison post hoc test. (G) Quantification of chemotactic cell migration and invasion. A total of 5000 CAL-51 cells were seeded in the upper chamber of each well in a 96-well chemotaxis microplate and treated with 100 μM mAbs (87B156, 69B287, and mGO.53). EGF (1 μg/mL) was added to the lower chamber of all wells as a chemoattractant. Cell migration was monitored using live imaging, and the number of migrated CAL-51 cells was quantified. Migration counts were normalized to the initial cell number, with 3 replicates for each condition.