Table2_Extracellular vesicle DNA from human melanoma tissues contains cancer-specific mutations.xlsx

Liquid biopsies are promising tools for early diagnosis and residual disease monitoring in patients with cancer, and circulating tumor DNA isolated from plasma has been extensively studied as it has been shown to contain tumor-specific mutations. Extracellular vesicles (EVs) present in tumor tissues carry tumor-derived molecules such as proteins and nucleic acids, and thus EVs can potentially represent a source of cancer-specific DNA. Here we identified the presence of tumor-specific DNA mutations in EVs isolated from six human melanoma metastatic tissues and compared the results with tumor tissue DNA and plasma DNA. Tumor tissue EVs were isolated using enzymatic treatment followed by ultracentrifugation and iodixanol density cushion isolation. A panel of 34 melanoma-related genes was investigated using ultra-sensitive sequencing (SiMSen-seq). We detected mutations in six genes in the EVs (BRAF, NRAS, CDKN2A, STK19, PPP6C, and RAC), and at least one mutation was detected in all melanoma EV samples. Interestingly, the mutant allele frequency was higher in DNA isolated from tumor-derived EVs compared to total DNA extracted directly from plasma DNA, supporting the potential role of tumor EVs as future biomarkers in melanoma.

液体活检（Liquid Biopsies）是极具应用前景的癌症患者早期诊断与残留病监测工具，从血浆中分离得到的循环肿瘤DNA（circulating tumor DNA）因被证实携带肿瘤特异性突变，已得到广泛研究。肿瘤组织中的细胞外囊泡（Extracellular Vesicles, EVs）携带有蛋白质、核酸等肿瘤源性分子，因此EVs有望成为癌症特异性DNA的潜在来源。本研究从6例人类转移性黑色素瘤组织中分离得到EVs，并检测到其中存在肿瘤特异性DNA突变，同时将检测结果与肿瘤组织DNA及血浆DNA进行了对比分析。本研究采用酶解法结合超速离心与碘克沙醇密度梯度垫层分离法，完成肿瘤组织EVs的分离纯化。随后采用超灵敏测序技术（SiMSen-seq），对靶向覆盖34个黑色素瘤相关基因的基因panel进行了检测分析。研究人员在EVs的6个基因（BRAF、NRAS、CDKN2A、STK19、PPP6C及RAC）中均检测到突变，且所有黑色素瘤EV样本均至少检出1处突变。值得注意的是，肿瘤源性EVs分离得到的DNA中，突变等位基因频率高于直接从血浆中提取的总DNA，这一结果进一步证实了肿瘤EVs有望成为黑色素瘤未来的生物标志物。