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

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.

Anti-EGFR Antibody Cetuximab Enhances the Cytolytic Activity of Natural Killer Cells toward Osteosarcoma

“…Cetuximabinduced ADCC was comparable with the maximal killing achieved by IL-15-activated NK cells. In contrast to some other models (32,34,40,41), we did not observe an additive effect of cetuximab on the lysis by cytokine (IL-15)-activated NK cells.…”

“…Finally, cetuximab may induce complement-dependent cytotoxicity or cytolytic effects by immune cells via ADCC (13,32,38,39,43). An advantage of cetuximab-mediated ADCC is that it would be independent of the EGFR mutation status (34,38) and persistently activated EGFR signaling pathways (13,40).…”

KRAS mutation confers resistance to antibody‐dependent cellular cytotoxicity of cetuximab against human colorectal cancer cells

“…[15][16][17]23 Consistent with these reports, a remarkable effect of cetuximab was not achieved even in KRAS-wild-type cells (Fig. 1c).…”

Interleukin-15 in the treatment of cancer

“…In particular, IL-15 is being incorporated in vaccines [96]. Furthermore, IL-15 is very effective in activating NK cells and monocytes suggesting that it may increase antibodydependent cellular cytotoxicity mediated by antineoplastic monoclonal antibodies [97][98][99].…”