Department of Ophthalmology, University of Lübeck, Lübeck, Germany.
Department of Pathology, University of Lübeck, Lübeck, Germany.
Invest Ophthalmol Vis Sci. 2014 Jun 26;55(7):4395-404. doi: 10.1167/iovs.14-14512.
Despite successful local tumor control, uveal melanoma (UM) patients may develop lethal metastases. To reliably identify circulating melanoma cells (CMC) in UM patients, we set out to test a new immunomagnetic enrichment assay and screened UM patients for the presence of CMC. We also determined whether we could find CMC in culture; for example, for future drug testing.
A dual-immunomagnetic enrichment assay using antibodies against two melanoma markers (NKI/C3 and NKI/beteb) was used to determine the presence of UM cells in blood. The sensitivity of the assay was determined by spiking normal blood with 92.1 cells (concentration range, 1-10(4) cells/mL). Isolated cells were characterized by immunocytochemistry directly after immunoenrichment and after a 2-week culture. The presence of CMC was determined in the peripheral blood of 31 patients with UM, and results were compared to clinical prognostic factors at the time of presentation.
The CMC were detected in 93.5% (n = 29 of 31) of the patients with primary nonmetastatic UM at a median density of 3.5 cells/10 mL blood (range, 0-10.2 cells), as well as in blood cultures. No significant association was observed between the presence or number of CMC and any clinical prognostic factors.
The improved dual-immunoenrichment assay enabled the detection of intact and viable CMC in the majority of UM patients. We also were able to identify CMC after short-term culturing. Molecular characterization of the CMC rather than the prevalence of these cells is expected to provide relevant information on the individual risk of metastasis.
尽管局部肿瘤控制成功，葡萄膜黑色素瘤（UM）患者仍可能发生致命转移。为了可靠地鉴定 UM 患者循环中的黑色素瘤细胞（CMC），我们着手测试一种新的免疫磁珠富集检测方法，并筛选 UM 患者 CMC 的存在情况。我们还确定是否可以在培养物中找到 CMC；例如，用于未来的药物测试。
使用针对两种黑色素瘤标志物（NKI/C3 和 NKI/beteb）的双免疫磁珠富集检测方法来确定血液中 UM 细胞的存在。通过将 92.1 个细胞（浓度范围为 1-104 个细胞/ml）掺入正常血液中来确定检测方法的灵敏度。直接在免疫富集后和 2 周培养后对分离的细胞进行免疫细胞化学特征分析。在 31 例原发性非转移性 UM 患者的外周血中检测到 CMC，并将结果与发病时的临床预后因素进行比较。
在中位数为 3.5 个细胞/10ml 血液（范围 0-10.2 个细胞）的情况下，在 93.5%（31 例中有 29 例）的原发性非转移性 UM 患者中检测到 CMC，以及在血液培养物中也检测到 CMC。未观察到 CMC 的存在或数量与任何临床预后因素之间存在显著相关性。
改良的双重免疫富集检测方法使大多数 UM 患者中完整和存活的 CMC 得以检测。我们还能够在短期培养后鉴定 CMC。CMC 的分子特征而不是这些细胞的流行率预计将为个体转移风险提供相关信息。
