Peripheral blood tyrosinase messenger RNA detection and survival in malignant melanoma
Peripheral blood tyrosinase messenger RNA detection and survival in malignant melanoma
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DOI：
10.1093/jnci/88.9.590
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发表时间：
1996-05-01
期刊：
影响因子：
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通讯作者：
Atzpodien, J
中科院分区：
文献类型：
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作者：
Kunter, U;Buer, J;Atzpodien, J
背景：最广泛接受的恶性黑色素瘤预后评估标准是组织病理学和临床表现。目前没有可用的实验室测试提供额外的预后信息。最近有人提出，基于逆转录和聚合酶链反应 (RT-PCR) 的外周血中酪氨酸酶信使 RNA (mRNA) 的检测可能有助于循环肿瘤细胞的早期检测，因为酪氨酸酶被认为是黑素细胞特异性标记物。目的：为了进一步评估这一潜在标志物的临床相关性，我们检查了不同疾病阶段的恶性黑色素瘤患者的外周血样本中是否存在酪氨酸酶 mRNA。方法：使用 RNAzol A 方法从 64 名恶性黑色素瘤患者、5 名健康对照受试者和 4 名其他癌症患者的肝素化外周血细胞中提取总细胞 RNA。为了分析酪氨酸酶 mRNA，按照 Smith 等人之前的描述进行 RT-PCR；使用从健康对照受试者的外周血连续稀释的人黑色素瘤细胞（SK-mel 1 和 SK-mel 3 细胞系）中提取的 RNA 来测试该测定的灵敏度。另外两种人类黑色素瘤细胞系（SK-mel 30 和 RPMI-7951）用作 RT-PCR 检测酪氨酸酶 mRNA 的阳性对照。使用 Kaplan-Meier 估计构建总体患者生存曲线。结果：通过 RT-PCR 检测，所有四种已建立的黑色素瘤细胞系均检测到了酪氨酸酶 mRNA。 64 名恶性黑色素瘤患者中，有 9 名被发现外周血细胞中可检测到酪氨酸酶 mRNA（酪氨酸酶阳性患者）。 16 名局限性原发性黑色素瘤患者的外周血细胞中未检测到酪氨酸酶 mRNA。在 48 名患有转移性疾病的患者中，所有 27 名没有表现出疾病进展证据的患者均为酪氨酸酶阴性。值得注意的是，所有九名酪氨酸酶阳性患者均出现内脏转移，并在采样时发现疾病进展。九名酪氨酸酶阳性患者中，有四名有时检测呈阴性，但没有疾病进展的证据；一名患者在病情稳定后呈阴性，三名患者在疾病进展时酪氨酸酶转录物呈阳性。测试时，9 名酪氨酸酶阳性患者的生存概率显着低于 23 名患有类似疾病但未检测到酪氨酸酶 mRNA 的患者（双侧；P 小于或等于 0.05）。结论：本研究结果表明，RT-PCR 检测外周血酪氨酸酶 mRNA 可能是预测恶性黑色素瘤患者肿瘤进展和不良临床结果的有用预后标志物。
Background: The most widely accepted criteria for the evaluation of prognosis of malignant melanoma are histopathologic and clinical presentation. No currently available laboratory tests provide additional prognostic information. It has recently been suggested that reverse transcription and polymerase chain reaction (RT-PCR)-based detection of tyrosinase messenger RNA (mRNA) in peripheral blood might be useful in the early detection of circulating tumor cells, since tyrosinase is thought to be a melanocyte-specific marker. Purpose: To further evaluate the clinical relevance of this potential marker, we examined peripheral blood samples from patients with malignant melanoma in different stages of disease for the presence of tyrosinase mRNA. Methods: Total cellular RNA was extracted from heparinized peripheral blood cells from 64 patients with malignant melanoma, from five healthy control subjects, and from four patients with other cancers using the RNAzol A method. For analysis of tyrosinase mRNA, RT-PCR was performed as previously described by Smith et al.; the sensitivity of this assay was tested using RNA extracted from human melanoma cells (SK-mel 1 and SK-mel 3 cell lines) serially diluted with peripheral blood obtained from healthy control subjects. Two additional human melanoma cell lines (SK-mel 30 and RPMI-7951) served as positive controls for RT-PCR detection of tyrosinase mRNA. Overall patient survival curves were constructed using Kaplan-Meier estimates. Results: Tyrosinase mRNA was detected by RT-PCR assay of all four of the established melanoma cell lines tested. Nine of the 64 patients with malignant melanoma were found to have detectable tyrosinase mRNA in their peripheral blood cells (tyrosinase-positive patients). The 16 patients with localized primary melanoma did not have detectable tyrosinase mRNA in their peripheral blood cells. Among the 48 patients with metastatic disease, all 27 patients who exhibited no evidence of disease progression were tyrosinase negative. Notably, all nine tyrosinase-positive patients had visceral metastases and were found to exhibit disease progression at the time of the sampling. Four of the nine tyrosinase-positive patients were also found to test negative at times without evidence of progressive disease; one patient became negative after achieving stable disease and three became positive for tyrosinase transcripts on disease progression. The probability of survival from time of sampling was significantly lower in the nine tyrosinase-positive patients when tested versus the 23 patients with comparable disease but without detectable tyrosinase mRNA (two-sided; P less than or equal to .05). Conclusions: The results of this study demonstrate that the tyrosinase mRNA in peripheral blood by RT-PCR may be a useful prognostic marker for predicting tumor progression and poor clinical outcome in patients with malignant melanoma.