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Abstract：
Melanoma is a lethal skin cancer, and the risk of developing it is increased by exposure to ultraviolet (UV) radiation. The production of cytokines such as interleukin-15 (IL-15), induced by the exposure of skin cells to UV rays, could also promote melanoma development. The aim of this study is to investigate the possible role of Interleukin-15/Interleukin-15 Receptor α (IL-15/IL-15Rα) complexes in melanoma development.
The expression of IL-15/IL-15Rα complexes by melanoma cells was evaluated both ex vivo and in vitro by tissue microarray, PCR, and flow cytometry. The presence of the soluble complex (sIL-15/IL-15Rα) in the plasma of metastatic melanoma patients was detected using an ELISA assay. Subsequently, we investigated the impact of natural killer (NK) cell activation after rIL-2 starvation followed by exposure to the sIL-15/IL-15Rα complex. Finally, by analyzing public datasets, we studied the correlation between IL-15 and IL-15Rα expressions and melanoma stage, NK and T-cell markers, and overall survival (OS).
Analysis of a melanoma tissue microarray shows a significant increase in the number of IL-15+ tumor cells from the benign nevi to metastatic melanoma stages. Metastatic melanoma cell lines express a phorbol-12-myristate-13-acetate (PMA)-cleavable membrane-bound IL-15 (mbIL-15), whereas cultures from primary melanomas express a PMA-resistant isoform. Further analysis revealed that 26% of metastatic patients present with consistently high plasmatic levels of sIL-15/IL-15Rα. When the recombinant soluble human IL-15/IL-15Rα complex is added to briefly starved rIL-2-expanded NK cells, these cells exhibit strongly reduced proliferation and levels of cytotoxic activity against K-562 and NALM-18 target cells. The analysis of public gene expression datasets revealed that high IL-15 and IL-15Rα intra-tumoral production correlates with the high levels of expression of CD5+ and NKp46+ (T and NK markers) and significantly correlates with a better OS in stages II and III, but not in stage IV.
Membrane-bound and secreted IL-15/IL-15Rα complexes are continuously present during progression in melanoma. It is notable that, although IL-15/IL-15Rα initially promoted the production of cytotoxic T and NK cells, at stage IV promotion of the development of anergic and dysfunctional cytotoxic NK cells was observed. In a subgroup of melanoma metastatic patients, the continuous secretion of high amounts of the soluble complex could represent a novel NK cell immune escape mechanism.
The expression of IL-15/IL-15Rα complexes by melanoma cells was evaluated both ex vivo and in vitro by tissue microarray, PCR, and flow cytometry. The presence of the soluble complex (sIL-15/IL-15Rα) in the plasma of metastatic melanoma patients was detected using an ELISA assay. Subsequently, we investigated the impact of natural killer (NK) cell activation after rIL-2 starvation followed by exposure to the sIL-15/IL-15Rα complex. Finally, by analyzing public datasets, we studied the correlation between IL-15 and IL-15Rα expressions and melanoma stage, NK and T-cell markers, and overall survival (OS).
Analysis of a melanoma tissue microarray shows a significant increase in the number of IL-15+ tumor cells from the benign nevi to metastatic melanoma stages. Metastatic melanoma cell lines express a phorbol-12-myristate-13-acetate (PMA)-cleavable membrane-bound IL-15 (mbIL-15), whereas cultures from primary melanomas express a PMA-resistant isoform. Further analysis revealed that 26% of metastatic patients present with consistently high plasmatic levels of sIL-15/IL-15Rα. When the recombinant soluble human IL-15/IL-15Rα complex is added to briefly starved rIL-2-expanded NK cells, these cells exhibit strongly reduced proliferation and levels of cytotoxic activity against K-562 and NALM-18 target cells. The analysis of public gene expression datasets revealed that high IL-15 and IL-15Rα intra-tumoral production correlates with the high levels of expression of CD5+ and NKp46+ (T and NK markers) and significantly correlates with a better OS in stages II and III, but not in stage IV.
Membrane-bound and secreted IL-15/IL-15Rα complexes are continuously present during progression in melanoma. It is notable that, although IL-15/IL-15Rα initially promoted the production of cytotoxic T and NK cells, at stage IV promotion of the development of anergic and dysfunctional cytotoxic NK cells was observed. In a subgroup of melanoma metastatic patients, the continuous secretion of high amounts of the soluble complex could represent a novel NK cell immune escape mechanism.
摘要：
■黑色素瘤是一种致命的皮肤癌,并且由于暴露于紫外线（UV）辐射而增加了开发它的风险。细胞因子如白细胞介素-15（IL-15）的产生，由皮肤细胞暴露于紫外线诱导，也可以促进黑色素瘤的发展。这项研究的目的是研究白细胞介素15/白细胞介素15受体α（IL-15/IL-15Rα）复合物在黑色素瘤发展中的可能作用。
■通过组织微阵列在离体和体外评估黑素瘤细胞对IL-15/IL-15Rα复合物的表达，PCR,和流式细胞术。使用ELISA测定法检测转移性黑色素瘤患者血浆中可溶性复合物（sIL-15/IL-15Rα）的存在。随后，我们调查了rIL-2饥饿后暴露于sIL-15/IL-15Rα复合物后自然杀伤（NK）细胞活化的影响。最后,通过分析公共数据集，我们研究了IL-15和IL-15Rα表达与黑色素瘤分期的相关性，NK和T细胞标记，总生存率（OS）。
黑素瘤组织微阵列的分析显示从良性痣到转移性黑素瘤阶段的IL-15+肿瘤细胞数量显著增加。转移性黑色素瘤细胞系表达佛波醇-12-肉豆蔻酸酯-13-乙酸酯（PMA）可切割的膜结合IL-15（mbIL-15），而来自原发性黑色素瘤的培养物表达PMA抗性亚型。进一步分析显示，26％的转移性患者始终存在高血浆水平的sIL-15/IL-15Rα。当将重组可溶性人IL-15/IL-15Rα复合物添加到短暂饥饿的rIL-2扩增的NK细胞中时，这些细胞表现出强烈降低的增殖和对K-562和NALM-18靶细胞的细胞毒活性水平。对公开基因表达数据集的分析显示，高IL-15和IL-15Rα在肿瘤内的产生与CD5+和NKp46+（T和NK标记）的高水平表达相关，并且与II期和III期更好的OS显著相关。但不是在第四阶段.
膜结合和分泌的IL-15/IL-15Rα复合物在黑素瘤的发展过程中持续存在。值得注意的是,虽然IL-15/IL-15Rα最初促进细胞毒性T和NK细胞的产生，在IV期，观察到无反应性和功能失调的细胞毒性NK细胞的发育。在黑色素瘤转移患者的亚组中，大量可溶性复合物的持续分泌可能代表了一种新的NK细胞免疫逃逸机制。
■通过组织微阵列在离体和体外评估黑素瘤细胞对IL-15/IL-15Rα复合物的表达，PCR,和流式细胞术。使用ELISA测定法检测转移性黑色素瘤患者血浆中可溶性复合物（sIL-15/IL-15Rα）的存在。随后，我们调查了rIL-2饥饿后暴露于sIL-15/IL-15Rα复合物后自然杀伤（NK）细胞活化的影响。最后,通过分析公共数据集，我们研究了IL-15和IL-15Rα表达与黑色素瘤分期的相关性，NK和T细胞标记，总生存率（OS）。
黑素瘤组织微阵列的分析显示从良性痣到转移性黑素瘤阶段的IL-15+肿瘤细胞数量显著增加。转移性黑色素瘤细胞系表达佛波醇-12-肉豆蔻酸酯-13-乙酸酯（PMA）可切割的膜结合IL-15（mbIL-15），而来自原发性黑色素瘤的培养物表达PMA抗性亚型。进一步分析显示，26％的转移性患者始终存在高血浆水平的sIL-15/IL-15Rα。当将重组可溶性人IL-15/IL-15Rα复合物添加到短暂饥饿的rIL-2扩增的NK细胞中时，这些细胞表现出强烈降低的增殖和对K-562和NALM-18靶细胞的细胞毒活性水平。对公开基因表达数据集的分析显示，高IL-15和IL-15Rα在肿瘤内的产生与CD5+和NKp46+（T和NK标记）的高水平表达相关，并且与II期和III期更好的OS显著相关。但不是在第四阶段.
膜结合和分泌的IL-15/IL-15Rα复合物在黑素瘤的发展过程中持续存在。值得注意的是,虽然IL-15/IL-15Rα最初促进细胞毒性T和NK细胞的产生，在IV期，观察到无反应性和功能失调的细胞毒性NK细胞的发育。在黑色素瘤转移患者的亚组中，大量可溶性复合物的持续分泌可能代表了一种新的NK细胞免疫逃逸机制。
