Requirement of KISS1 secretion for multiple organ metastasis suppression and maintenance of tumor dormancy
Requirement of KISS1 secretion for multiple organ metastasis suppression and maintenance of tumor dormancy
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DOI：
10.1093/jnci/djk053
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发表时间：
2007-02-21
影响因子：
10.3
通讯作者：
Welch, Danny R.
中科院分区：
文献类型：
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作者：
Nash, Kevin T.;Phadke, Pushkar A.;Welch, Danny R.
背景KISS 1蛋白在不阻断原位肿瘤生长的情况下抑制几种肿瘤模型的转移，但其机制仍不清楚。由于其在人类性成熟中的作用，KISS 1蛋白被分泌并加工成kisspeptins，其与G蛋白偶联受体GPR 54结合。方法将含有内FLAG表位的KISS 1（KFM）或不含信号序列（KFM Δ SS）转染到不表达内源性KISS 1的C8161. 9人黑色素瘤细胞中，并检测KISS 1的表达。收集来自C8161.9（KFM）和C8161.9（KFM Delta SS）细胞的全细胞裂解物和条件培养基，并通过免疫沉淀和酶联免疫吸附测定分析吻肽。使用实时逆转录聚合酶链反应测量GPR 54水平。评价了C8161.9（KFM）和C8161.9（KFM Delta SS）细胞的条件培养基刺激表达GPR 54的中国仓鼠卵巢细胞（CHO-G）和C8161.9细胞中钙动员的能力。在注射用增强型绿色荧光蛋白标记的C8161.9（KFM）或C8161.9（KFM Delta SS）细胞的无胸腺小鼠（每组10只）中监测转移。结果在C8161.9（KFM）和C8161.9（KFM Delta SS）细胞的全细胞裂解物中分别检测到全长KFM和KFM Delta SS，但仅在C8161.9（KFM）细胞的条件培养液中检测到kisspeptins。在体内，C8161.9（KFM）而不是C8161.9（KFM Delta SS）细胞被抑制转移到肺、眼、肾和骨，小鼠存活率有相应的差异（中值> 120天对42天）。C8161.9（KFM）细胞接种小鼠肺，但未形成肉眼可见的转移。来自C8161.9（KFM）而非C8161.9（KFI Delta SS）的条件培养基刺激CHC-G细胞中的钙动员。C8161.9（KFM）细胞的条件培养液不能刺激C8161.9细胞表达GPR 54，而C8161.9细胞中GPR 54表达较低。在C8161.9细胞（其转移扩散被KFM抑制）中GPR 54表达的缺乏表明转移抑制不是通过该受体介导的。这些结果暗示了另一种KISS 1受体和/或旁分泌信号的存在。这些发现提高了可溶性KISS 1、kisspeptins或模拟物可用于维持肿瘤休眠的可能性，从而治疗已经扩散的肿瘤细胞（即，微转移）的合法目标。
Background The KISS1 protein suppresses metastasis of several tumor models without blocking orthotopic tumor growth, but the mechanism remains elusive. For its role in human sexual maturation, KISS1 protein is secreted and processed to kisspeptins, which bind to the G protein-coupled receptor GPR54. We tested the hypothesis that KISS1 secretion is required for metastasis suppression via GPR54.Methods KISS1 containing an internal FLAG epitope with (KFM) or without (KFM Delta SS) a signal sequence was transfected into C8161.9 human melanoma cells, which do not express endogenous KISS1. Whole-cell lysates and conditioned medium from C8161.9(KFM) and C8161.9(KFM Delta SS) cells were collected and analyzed for kiss-peptins by immunoprecipitation and enzyme-linked immunosorbent assay. GPR54 levels were measured using real-time reverse transcription-polymerase chain reaction. The ability of conditioned medium from C8161.9(KFM) and C8161.9(KFM Delta SS) cells to stimulate calcium mobilization in GPR54-expressing Chinese hamster ovary cells (CHO-G) and in C8161.9 cells was evaluated. Metastasis was monitored in athymic mice (groups of 10 per experiment) that were injected with C8161.9(KFM) or C8161.9(KFM Delta SS) cells labeled with enhanced green fluorescent protein. Survival of mice injected with C8161.9 or C8161.9(KFM) cells was analyzed by Kaplan-Meier methods.Results Full-length KFM and KFM Delta SS were detected in whole-cell lysates of C8161.9(KFM) and C8161.9(KFM Delta SS) cells, respectively, but kisspeptins were detected only in conditioned medium of C8161.9(KFM) cells. In vivo, C8161.9(KFM), but not C8161.9(KFM Delta SS), cells were suppressed for metastasis to lung, eye, kidney, and bone, with corresponding differences in mouse survival (median > 120 versus 42 days). C8161.9(KFM) cells seeded mouse lungs but did not form macroscopic metastases. Conditioned medium from C8161.9(KFM), but not C8161.9(KFI Delta SS), cells stimulated calcium mobilization in CHC-G cells. GPR54 expression was low in C8161.9 cells, which were not stimulated by conditioned medium from C8161.9(KFM) cells.Conclusions KISS1 secretion was required for multiple organ metastasis suppression and for maintenance of disseminated cells in a dormant state. The absence of GPR54 expression in C8161.9 cells (whose metastatic spread was suppressed by KFM) suggests that metastasis suppression is not mediated through this receptor. The results imply the existence of another KISS1 receptor and/or paracrine signaling. The findings raise the possibility that soluble KISS1, kisspeptins, or mimetics could be used to maintain tumor dormancy, rendering treatment of already disseminated tumor cells (i.e., micrometastases) a legitimate target.