title: 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.
authors.affiliation: Centro de Investigaciones Oncológicas, Fundación Cáncer and Instituto Alexander Fleming, Cramer 1180 (1426), Buenos Aires, Argentina.
authors.forename: M P
authors.initials: MP
authors.lastname: Roberti
authors.forename: M M
authors.initials: MM
authors.lastname: Barrio
authors.forename: A I
authors.initials: AI
authors.lastname: Bravo
authors.forename: Y S
authors.initials: YS
authors.lastname: Rocca
authors.forename: J M
authors.initials: JM
authors.lastname: Arriaga
authors.forename: M
authors.initials: M
authors.lastname: Bianchini
authors.forename: J
authors.initials: J
authors.lastname: Mordoh
authors.forename: E M
authors.initials: EM
authors.lastname: Levy
mesh_terms: D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D001483:Base Sequence; D001943:Breast Neoplasms; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D000068818:Cetuximab; D018920:Coculture Techniques; D004252:DNA Mutational Analysis; D066246:ErbB Receptors / Q000378:metabolism*; D005260:Female; D058851:GPI-Linked Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D019409:Interleukin-15 / Q000494:pharmacology* / Q000502:physiology; D007376:Interleukin-2 / Q000494:pharmacology* / Q000502:physiology; D007694:Killer Cells, Natural / Q000187:drug effects / Q000378:metabolism; D055655:NK Cell Lectin-Like Receptor Subfamily K / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D011518:Proto-Oncogene Proteins / Q000235:genetics; D016283:Proto-Oncogene Proteins p21(ras); D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism; D011960:Receptors, Estrogen / Q000378:metabolism; D017452:Receptors, IgG / Q000235:genetics / Q000378:metabolism; D011980:Receptors, Progesterone / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects; D018631:ras Proteins / Q000235:genetics
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D000068818:Cetuximab; D066246:ErbB Receptors; D058851:GPI-Linked Proteins; D007371:Interferon-gamma; D019409:Interleukin-15; D007376:Interleukin-2; D055655:NK Cell Lectin-Like Receptor Subfamily K; D011518:Proto-Oncogene Proteins; D016283:Proto-Oncogene Proteins p21(ras); D018719:Erb-b2 Receptor Tyrosine Kinases; D011960:Receptors, Estrogen; D017452:Receptors, IgG; D011980:Receptors, Progesterone; D018631:ras Proteins; C428200:FCGR3B protein, human; C508594:IL2 protein, human; C527085:KLRK1 protein, human; C117307:KRAS protein, human; C508053:ERBB2 protein, human