Centro de Investigaciones Oncológicas, Fundación Cáncer and Instituto Alexander Fleming, Cramer 1180 (1426), Buenos Aires, Argentina.
Breast Cancer Res Treat. 2011 Nov;130(2):465-75. doi: 10.1007/s10549-011-1360-2. Epub 2011 Feb 10.
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.
三阴性乳腺癌（TNBC）患者不太可能从抗雌激素或抗 HER2 治疗中获益，这种表型与更具侵袭性的临床过程和更差的临床结局相关。考虑到 TNBC 的治疗选择有限，本研究的目的是研究一种基于西妥昔单抗介导的自然杀伤（NK）细胞免疫活性的潜在治疗方法。我们对人乳腺癌（BC）细胞系 IIB-BR-G 和体内转移变体 IIB-BR-G MT 进行了体外研究。免疫组织化学分析显示，两种细胞系均表现出 TNBC 表型，通过流式细胞术测量，EGFR 表达水平较高，但存在差异。DNA 测序显示，两种细胞系均具有突变型 K-RAS 状态，38 号密码子 G > A。因此，西妥昔单抗不能抑制细胞增殖或诱导细胞凋亡。我们研究了西妥昔单抗是否能触发免疫机制，并确定两种细胞系用 1μg/ml 的西妥昔单抗处理后均易受抗体依赖性细胞毒性（ADCC）的影响，这种 ADCC 由外周血单核细胞（PBMC）介导。在 50:1 的效应细胞：靶细胞比值下，对 IIB-BR-G 的裂解活性为 34±2%，对 IIB-BR-G MT 的裂解活性为 27±6%。PBMC 用 IL-2 预处理后，针对 IIB-BR-G 的西妥昔单抗介导的 ADCC 活性可达 65±3%，针对 IIB-BR-G MT 的 ADCC 活性可达 63±6.5%。此外，IL-15 预处理可使 ADCC 分别提高至 IIB-BR-G 的 71±3%和 IIB-BR-G MT 的 79±3.5%。我们认为 NK 细胞是 PBMC 中存在的效应细胞，因为它们能够在较低的效应细胞：靶细胞比值下诱导 ADCC。此外，IL-2 和主要是 IL-15 诱导 NK 激活受体 CD16 和 NKG2D 的上调，并增强 IFN-γ 的产生。在临床可达到的浓度下，表达 EGFR 的 TNBC 可通过西妥昔单抗介导的 ADCC 杀伤。IL-15 可以在大多数免疫活性方面有利地替代 IL-2，刺激产生 IFN-γ的能力，并与激活受体的上调相平行。
