Detection of mutated BRAFV600E variant in circulating DNA of stage III-IV melanoma patients
Detection of mutated BRAFV600E variant in circulating DNA of stage III-IV melanoma patients
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DOI：
10.1002/ijc.22598
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发表时间：
2007-06-01
影响因子：
6.4
通讯作者：
Rodolfo, Monica
中科院分区：
文献类型：
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作者：
Daniotti, Maria;Vallacchi, Viviana;Rodolfo, Monica
BRAFV 600 E是皮肤黑色素瘤中最具代表性的体细胞点突变，因此为该疾病提供了独特的分子标记。开发有效的方法检测其在患者的游离循环DNA可能会导致诊断和预后工具的改进。为此，我们在一项初步研究中评估了BRAFV 600 E是否代表黑色素瘤患者血浆/血清中的可检测标志物。从15名健康供体和41名不同临床阶段的黑色素瘤患者的血清或血浆中提取循环游离DNA，并在手术前或手术后随访期间获得。定量分析显示，与对照组相比，患者的循环游离DNA水平较高，在术前和IV期获得的样本中检测到的水平最高。比较了四种不同的PCR方法在野生型DNA中扩增几个BRAFV 600 E拷贝的能力。在12例患者的循环DNA中可检测到BRAFV 600 E，而在对照组中均未检测到BRAFV 600 E;仅1种PCR方法可重复扩增BRAFV 600 E。从IV期的8/13例患者和III期的4/24例患者中获得阳性样本，但在I-II期的4例患者中未获得阳性样本;一半的阳性样本在术前获得，一半在随访时获得。对20例患者的循环DNA和相关肿瘤之间的对应关系进行了检查，发现IV期患者存在相关性。总之，这种方法可以用于监测疾病的第四阶段黑色素瘤患者，但它似乎不能令人满意的早期检测黑色素瘤。(c)2007 Wiley-Liss，Inc.
BRAFV600E is the most represented somatic point mutation in cutaneous melanoma, thus providing a unique molecular marker for this disease. The development of efficient methods for its detection in free circulating DNA of patients may lead to the improvement of diagnostic and prognostic tools. With this aim, we evaluated whether BRAFV600E represents a detectable marker in the plasma/serum from melanoma patients in a pilot study. Circulating cell-free DNA was extracted from the serum or plasma of 15 healthy donors and 41 melanoma patients at different clinical stages and obtained either presurgery or after surgery during follow-up. Quantitative analysis showed higher levels of circulating free DNA in patients compared to controls, with the highest levels detected in samples obtained presurgery and at stage IV. Four different PCR methods were compared for their capacity to amplify a few copies of BRAFV600E in wild-type DNA. BRAFV600E was detectable in circulating DNA of 12 patients and in none of the controls; only 1 PCR method reproducibly amplified BRAFV600E. Positive samples were obtained from 8/13 patients at stage IV and from 4/24 patients at stage III, but not in 4 patients at stage I-II; half of the positives were obtained presurgery and half at follow-up. Correspondence between circulating DNA and related tumors were examined for 20 patients, and a correlation was found for stage IV patients. In conclusion, this method can be utilized for monitoring the disease in stage IV melanoma patients but it appears unsatisfactory for the early detection of melanoma. (c) 2007 Wiley-Liss, Inc.