Center for Preclinical Research, The Department of Methodology, Medical University of Warsaw, 1B Banacha Str., 02-097 Warsaw, Poland.
Doctoral School, Medical University of Warsaw, 61 Zwirki and Wigury Str., 02-091 Warsaw, Poland.
Int J Mol Sci. 2021 Apr 9;22(8):3886. doi: 10.3390/ijms22083886.
Melanoma tumors are the most heterogeneous of all tumor types. Tumor heterogeneity results in difficulties in diagnosis and is a frequent cause of failure in treatment. Novel techniques enable accurate examination of the tumor cells, considering their heterogeneity. The study aimed to determine the somatic variations among high and low proliferating compartments of melanoma tumors. In this study, 12 archival formalin-fixed paraffin-embedded samples of previously untreated primary cutaneous melanoma were stained with Ki-67 antibody. High and low proliferating compartments from four melanoma tumors were dissected using laser-capture microdissection. DNA was isolated and analyzed quantitatively and qualitatively. Libraries for amplicon-based next-generation sequencing (NGS) were prepared using NEBNext Direct Cancer HotSpot Panel. NGS detected 206 variants in 42 genes in melanoma samples. Most of them were located within exons (135, 66%) and were predominantly non-synonymous single nucleotide variants (99, 73.3%). The analysis showed significant differences in mutational profiles between high and low proliferation compartments of melanoma tumors. Moreover, a significantly higher percentage of variants were detected only in high proliferation compartments (39%) compared to low proliferation regions (16%, < 0.05). Our results suggest a significant functional role of genetic heterogeneity in melanoma.
黑色素瘤肿瘤是所有肿瘤类型中异质性最高的。肿瘤异质性导致诊断困难，是治疗失败的常见原因。新技术能够准确检查肿瘤细胞，考虑到其异质性。本研究旨在确定黑色素瘤肿瘤中高增殖和低增殖区室之间的体细胞变异。在这项研究中，对 12 个未经治疗的原发性皮肤黑色素瘤的存档福尔马林固定石蜡包埋样本进行了 Ki-67 抗体染色。使用激光捕获显微切割术从四个黑色素瘤肿瘤中分离出高增殖和低增殖区室。提取 DNA 并进行定量和定性分析。使用 NEBNext Direct Cancer HotSpot Panel 制备用于基于扩增子的下一代测序 (NGS) 的文库。NGS 在黑色素瘤样本中的 42 个基因中检测到 206 个变体。它们大多数位于外显子内（135，66%），主要是非同义单核苷酸变异（99，73.3%）。分析表明，黑色素瘤肿瘤高增殖和低增殖区室之间的突变谱存在显著差异。此外，与低增殖区（16%，<0.05）相比，仅在高增殖区室（39%）中检测到的变异百分比显著更高。我们的结果表明遗传异质性在黑色素瘤中具有重要的功能作用。
