男性局部高危前列腺癌的准确风险分类直接影响治疗管理决策和患者预后。有广泛的风险评估和分类。然而,每种方法在区分惰性前列腺癌和侵袭性前列腺癌方面都有显著的局限性.循环肿瘤细胞(CTC)可能提供另一种来源,除了组织活检,为了实现针对个体患者的临床评估,仅仅因为CTC可以更精确地揭示肿瘤来源和种系特异性的遗传信息,而不是通过单一诊断活检获得的信息。在这项研究中,我们将基于过滤的CTC分离技术与前列腺癌CTC免疫分型相结合,以鉴定前列腺癌CTC.接下来,我们在激光显微切割和单细胞全外显子组测序(WES)前对来自10例局部高危前列腺癌患者样本的21个CTC和4个淋巴细胞进行了3-D端粒分析.局部高风险前列腺癌患者CTC呈现大量端粒信号,信号强度较低(端粒短)。为了捕获高风险前列腺癌CTC的遗传多样性/异质性,我们进行了全外显子组测序.我们确定了202,241个单核苷酸变体(SNV)和137,407个插入缺失(indel),其中不到10%的遗传变异在编码区域内。遗传变异(SNV+indel)和拷贝数改变(CNAs)谱是高度异质的,并且观察到患者内CTC变异。通路富集分析显示在9个端粒维持通路(患者3、5、6和7)中存在遗传变异,包括端粒维持的重要基因,称为端粒重复结合因子2(TRF2)。使用PharmGKB数据库,我们确定了9个与多西他赛疗效相关的遗传变异.总共48个SNV可以影响24种已知癌症药物的药物反应。基因集富集分析(GSEA)(患者1、3、6和8)确定了在11种不同途径中存在CNAs,包括DNA损伤修复(DDR)途径。总之,单细胞方法(WES和3-D端粒谱分析)显示可用于揭示CTC异质性。DDR途径突变已被确立为癌症治疗的靶途径。然而,在局部高危患者中发现的频繁的CNA扩增可能在前列腺癌的治疗抵抗中起关键作用.
Accurate risk classification of men with localized high-risk prostate cancer directly affects treatment management decisions and patient outcomes. A wide range of risk assessments and classifications are available. However, each one has significant limitations to distinguish between indolent and aggressive prostate cancers. Circulating tumor cells (CTCs) may provide an alternate additional source, beyond tissue biopsies, to enable individual patient-specific clinical assessment, simply because CTCs can reveal both tumor-derived and germline-specific genetic information more precisely than that gained from a single diagnostic biopsy. In this study, we combined a filtration-based CTC isolation technology with prostate cancer CTC immunophenotyping to identify prostate cancer CTCs. Next, we performed 3-D telomere profiling prior to laser microdissection and single-cell whole-exome sequencing (WES) of 21 CTCs and 4 lymphocytes derived from 10 localized high-risk prostate cancer patient samples. Localized high-risk prostate cancer patient CTCs present a high number of telomere signals with lower signal intensities (short telomeres). To capture the genetic diversity/heterogeneity of high-risk prostate cancer CTCs, we carried out whole-exome sequencing. We identified 202,241 single nucleotide variants (SNVs) and 137,407 insertion-deletions (indels), where less than 10% of these genetic variations were within coding regions. The genetic variation (SNVs + indels) and copy number alteration (CNAs) profiles were highly heterogeneous and intra-patient CTC variation was observed. The pathway enrichment analysis showed the presence of genetic variation in nine telomere maintenance pathways (patients 3, 5, 6, and 7), including an important gene for telomere maintenance called telomeric repeat-binding factor 2 (TRF2). Using the PharmGKB database, we identified nine genetic variations associated with response to docetaxel. A total of 48 SNVs can affect drug response for 24 known cancer drugs. Gene Set Enrichment Analysis (GSEA) (patients 1, 3, 6, and 8) identified the presence of CNAs in 11 different pathways, including the DNA damage repair (DDR) pathway. In conclusion, single-cell approaches (WES and 3-D telomere profiling) showed to be useful in unmasking CTC heterogeneity. DDR pathway mutations have been well-established as a target pathway for cancer therapy. However, the frequent CNA amplifications found in localized high-risk patients may play critical roles in the therapeutic resistance in prostate cancer.