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IL-15 and IL-2 increase Cetuximab-mediated cellular cytotoxicity against triple negative breast cancer cell lines expressing EGFR

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Abstract

Triple negative breast cancer (TNBC) patients are not likely to benefit from anti-estrogen or anti-HER2 therapy and this phenotype is associated with a more aggressive clinical course and worse clinical outcome. Taking into account the limited treatment possibilities in TNBC, the aim of the present work was to study a potential therapy based on Cetuximab-mediated immune activity by natural killer (NK) cells. We performed in vitro studies on human breast cancer (BC) cell lines, IIB-BR-G, and the in vivo metastatic variant IIB-BR-G MT. The immunohistochemical analysis showed a TNBC phenotype with high but different levels of EGFR expression on each cell line, measured by flow cytometry. DNA sequencing showed that both cell lines have a mutated K-RAS status, 38 G > A at codon 13. Consequently, Cetuximab did not inhibit cellular proliferation or induce apoptosis. We investigated if Cetuximab could trigger immune mechanisms, and we determined that both cell lines treated with 1 μg/ml Cetuximab were susceptible to antibody dependent cellular cytotoxicity (ADCC), mediated by peripheral blood mononuclear cells (PBMC). At 50:1 effector:target ratio, lytic activity was 34 ± 2% against IIB-BR-G and 27 ± 6% against IIB-BR-G MT cells. PBMC pretreatment with IL-2 allowed reaching 65 ± 3% of Cetuximab-mediated ADCC against IIB-BR-G and 63 ± 6.5% against IIB-BR-G MT. Furthermore, IL-15 pretreatment increased the ADCC up to 71 ± 3% in IIB-BR-G and 79 ± 3.5% in IIB-BR-G MT. We suggest that NK cells are the effectors present in PBMC since they were able to induce ADCC at lower effector:target ratios. Besides, IL-2- and mainly IL-15-induced upregulation of NK activating receptors CD16 and NKG2D and enhanced IFN-γ production. EGFR-expressing TNBC could be killed by Cetuximab-mediated ADCC at clinically achievable concentrations. IL-15 could advantageously replace IL-2 in most of its immunologic activities, stimulating the ability to produce IFN-γ, and paralleling the up-regulation of activating receptors.

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Acknowledgments

This work was supported with funds from Fundación Sales, Fundación P. Mosoteguy, Fundación Cáncer (FUCA), Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT) and Fundación María Calderón de la Barca. M. Bianchini, M.M. Barrio, J. Mordoh, and E.M. Levy are members of CONICET. We are grateful to Dr. Norberto W. Zwirner for kindly providing us IL-15 and Biochemist Fernanda Reynoso for performing PCR for K-RAS analysis and to Ms. María Luisa Poljak for revising this manuscript. We also thank the Servicio de Hemoterapia del Instituto Médico Especializado A. Fleming for providing healthy donors blood samples.

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Authors and Affiliations

  1. Centro de Investigaciones Oncológicas, Fundación Cáncer and Instituto Alexander Fleming, Cramer 1180 (1426), Buenos Aires, Argentina

    M. P. Roberti, M. M. Barrio, Y. S. Rocca, J. M. Arriaga, M. Bianchini, J. Mordoh & E. M. Levy

  2. Hospital Interzonal General de Agudos Eva Perón, Av Ricardo Balbin 3200 KM 18, San Martin, Argentina

    A. I. Bravo

  3. IIBBA CONICET, Fundación Instituto Leloir, Av. Patricias Argentinas 435 (1405), Buenos Aires, Argentina

    J. Mordoh

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  1. M. P. Roberti
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Roberti, M.P., Barrio, M.M., Bravo, A.I. et al. IL-15 and IL-2 increase Cetuximab-mediated cellular cytotoxicity against triple negative breast cancer cell lines expressing EGFR. Breast Cancer Res Treat 130, 465–475 (2011). https://doi.org/10.1007/s10549-011-1360-2

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