Dopierala Justyna, Damato Bertil E, Lake Sarah L, Taktak Azzam F G, Coupland Sarah E
Department of Pathology, School of Cancer Studies, University of Liverpool, Liverpool, UK.
Invest Ophthalmol Vis Sci. 2010 Oct;51(10):4898-905. doi: 10.1167/iovs.09-5004. Epub 2010 May 19.
To determine intratumor genetic heterogeneity in uveal melanoma (UM) by multiplex ligation-dependent probe amplification (MLPA) in formalin-fixed, paraffin-embedded (FFPE) tumor tissues.
DNA was extracted from whole tumor sections and from two to nine different areas microdissected from 32 FFPE UMs. Thirty-one loci on chromosomes 1, 3, 6, and 8 were tested with MLPA for copy number changes. The tumor was considered heterogeneous at a locus if (1) the difference in dosage quotients (DQs) of any two areas was 0.2 or more, and (2) the DQs of the areas belonged to different ranges.
Comparison of MLPA data obtained from microdissected areas of the UMs showed heterogeneity in 1 to 26 examined loci in 24 (75%) tumors, with only 25% of the tumors being homogeneous. Intratumor heterogeneity of 3p12.2, 6p21.2, and 8q11.23 was most common, occurring in >30% of the UMs. Gains of chromosome 3 were observed in four UMs, with three of these tumors showing the highest degree of heterogeneity. Copy number variation was associated with differences in tumor cell type, but not with differences in tumor pigmentation or reactive inflammation. UMs with genetic heterogeneity across multiple sample sites showed equivocal MLPA results when the whole tumor section was examined. These results suggest that different clones dilute MLPA results.
Heterogeneity of chromosomal abnormalities of chromosomes 1, 3, 6, and 8 is present in most UMs. This heterogeneity causes equivocal MLPA results. One random tumor sample may not be representative of the whole tumor and, therefore, may be insufficient for prognostic testing.
通过对福尔马林固定、石蜡包埋（FFPE）肿瘤组织进行多重连接依赖探针扩增（MLPA）来确定葡萄膜黑色素瘤（UM）的肿瘤内基因异质性。
从32例FFPE UM的整个肿瘤切片以及从两到九个不同区域显微切割提取DNA。使用MLPA检测1、3、6和8号染色体上的31个位点的拷贝数变化。如果（1）任何两个区域的剂量商（DQ）差异为0.2或更大，且（2）这些区域的DQ属于不同范围，则认为该肿瘤在某个位点存在异质性。
对UM显微切割区域获得的MLPA数据进行比较显示，24例（75%）肿瘤中1至26个检测位点存在异质性，只有25%的肿瘤是同质的。3p12.2、6p21.2和8q11.23的肿瘤内异质性最为常见，在>30%的UM中出现。在4例UM中观察到3号染色体的增益，其中3例肿瘤表现出最高程度的异质性。拷贝数变异与肿瘤细胞类型的差异相关，但与肿瘤色素沉着或反应性炎症的差异无关。当检查整个肿瘤切片时，多个样本位点存在基因异质性的UM显示出模糊的MLPA结果。这些结果表明不同克隆稀释了MLPA结果。
大多数UM中存在1、3、6和8号染色体的染色体异常异质性。这种异质性导致MLPA结果模糊。一个随机的肿瘤样本可能不代表整个肿瘤，因此可能不足以进行预后检测。
