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Genomic Analysis of Localized High-Risk Prostate Cancer Circulating Tumor Cells at the Single-Cell Level.

ABSTRACT: 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.

SUBMITTER: Rangel-Pozzo A

PROVIDER: S-EPMC7466090 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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Genomic Analysis of Localized High-Risk Prostate Cancer Circulating Tumor Cells at the Single-Cell Level.

Rangel-Pozzo Aline A Liu Songyan S Wajnberg Gabriel G Wang Xuemei X Ouellette Rodney J RJ Hicks Geoffrey G GG Drachenberg Darrel D Mai Sabine S

Cells 20200808 8

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 C ...[more]

PMID: 32784507

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Project description:PurposeUsing nonenrichment-based, potentially more sensitive Epic Sciences circulating tumor cell (CTC) platform, we sought to detect and characterize CTCs in untreated, high-risk localized prostate cancer and to evaluate their clinical implication.MethodsBetween 2012 and 2015, blood samples were prospectively collected from patients with National Comprehensive Cancer Network high-risk localized prostate cancer undergoing either radiotherapy (XRT) plus androgen deprivation therapy or radical prostatectomy (RP) with curative intent. Samples were analyzed with the Epic Sciences platform with 4J,6-diamidino-2-phenylindole, CD45, cytokeratin (CK), and androgen receptor (AR) N-terminal staining. CTC counts were correlated with biochemical recurrence (BCR).ResultsA diversity of CTC subtypes, including CK-positive, CK-negative, AR-positive, and CTC clusters, were observed in 73.3% (33 of 45) of patients with evaluable data. The median follow-up was 14.2 months (range, 0.5 to 43.7 months). BCR occurred more frequently in the RP group than XRT (15 of 26 v one of 19), with most patients in the XRT group continuing to receive androgen deprivation therapy. A higher proportion of metastatic events were observed in the RP group (five of 26 v one of 19). In the RP group, BCR and development of metastases were associated with a higher total number of CTCs, AR-positive CTCs, and CTC phenotypic heterogeneity. One patient who developed BCR and metastases quickly after RP had diverse phenotypical CTC subtypes, and single-cell genomic analyses of all detectable CTCs confirmed common prostate cancer copy number alterations and PTEN loss.ConclusionCTCs can be identified in most patients with high-risk localized prostate cancer before definitive therapy using the Epic Sciences platform. If confirmed in a larger cohort with longer follow-up, phenotypic and genomic characterization of CTCs pretherapy may provide an additional means of risk stratifying patients with newly diagnosed high-risk disease and potentially help identify patients who could require multimodal therapy.

Project description:BackgroundAlthough circulating tumor cells (CTCs) are considered as a surrogate marker in monitoring disease progression and treatment response in late stage prostate cancer (PCa), its clinical impact in localized PCa remains unclear, indicating the limitation that is simply based on cell count. This perspective observational study aimed to detect the epithelial-to-mesenchymal transition (EMT) subtypes of CTCs in localized PCa and analyze their clinical relevance and application in predicting PCa stages before surgery compared with the Partin table.Patients and methodsBetween August 2017 and April 2019, 80 newly diagnosed localized PCa patients were enrolled in the study. Peripheral blood samples (5 mL) were collected prior to surgery. The CanPatrolTM CTC enrichment technique, a size-based isolation method, was used to detect the EMT CTCs. Clinical relevance of the CTCs was analyzed with Spearman's rank correlation test. Models to predict pathological were built with multivariate logistic regression. Receiver operating characteristic (ROC) curve and area under the curve (AUC) analysis were performed to evaluate the accuracy of the prediction model.ResultsCTCs were detected in 55% of all patients. The biophenotypic CTCs were most valuable and closely correlated with PSA, Gleason score, D'Amico risk classification, and pathological stage in localized PCa. The mesenchymal subtype was rare in this population but associated with seminal vesicle invasion, while the epithelial subtype had limited clinical significance. In addition, the biophenotypic CTCs combined with traditional clinical variables were analyzed by multivariate logistic regression to predict organ-confined disease before surgery, of which the AUC reached 0.818 and was superior to the Partin table 2017 in our cohort.ConclusionThis study highlights the clinical impact of the biophenotypic CTCs in localized PCa, which was most closely related to clinical variables and could help to predict pathology outcomes before surgery.

Project description:Carotenoids are a class of nutrients with antioxidant properties that have been purported to protect against cancer. However, the reported associations between carotenoids and prostate cancer have been heterogeneous and lacking data on interactions with nucleotide sequence variations and genomic biomarkers.To examine the associations between carotenoid levels and the risk of high-grade prostate cancer, also considering antioxidant-related genes and tumor instability.We measured plasma levels of carotenoids and genotyped 20 single nucleotide polymorphisms (SNP) in SOD1, SOD2, SOD3, XRCC1, and OGG1 among 559 men with non-metastatic prostate cancer undergoing radical prostatectomy. We performed copy number analysis in a subset of these men (n?=?67) to study tumor instability assessed as Fraction of the Genome Altered (FGA). We examined associations between carotenoids, genotypes, tumor instability and risk of high-grade prostate cancer (Gleason grade???4?+?3) using logistic and linear regression.Circulating carotenoid levels were inversely associated with the risk of high-grade prostate cancer; odds ratios (OR) and 95% confidence intervals (CI) comparing highest versus lowest quartiles were: 0.34 (95% CI: 0.18-0.66) for ?-carotene, 0.31 (95% CI: 0.15-0.63) for ?-carotene, 0.55 (0.28-1.08) for lycopene and 0.37 (0.18-0.75) for total carotenoids. SNPs rs25489 in XRCC1, rs699473 in SOD3 and rs1052133 in OGG1 modified these associations for ?-carotene, ?-carotene and lycopene, respectively (P???0.05). The proportion of men with a high degree of FGA increased with Gleason Score (P?<?0.001). Among men with Gleason score ??3?+?4, higher lycopene levels were associated with lower FGA (P?=?0.04).Circulating carotenoids at diagnosis, particularly among men carrying specific somatic variations, were inversely associated with risk of high-grade prostate cancer. In exploratory analyses, higher lycopene level was associated with less genomic instability among men with low-grade disease which is novel and supports the hypothesis that lycopene may inhibit progression of prostate cancer early in its natural history.

Project description:The number of circulating tumor cells (CTCs) in metastatic prostate cancer patients provides prognostic and predictive information. However, it is the molecular characterization of CTCs that offers insight into the biology of these tumor cells in the context of personalized treatment.We developed a novel approach to isolate CTCs away from hematopoietic cells with high purity, enabling genomic analysis of these cells. The isolation protocol involves immunomagnetic enrichment followed by fluorescence activated cell sorting (IE/FACS). To evaluate the feasibility of isolation of CTCs by IE/FACS and downstream genomic profiling, we conducted a pilot study in patients with metastatic castration resistant prostate cancer (CRPC). Twenty (20) sequential CRPC patients were assayed using CellSearch™. Twelve (12) patients positive for CTCs were subjected to immunomagnetic enrichment and fluorescence activated cell sorting (IE/FACS) to isolate CTCs. Genomic DNA of CTCs was subjected to whole genome amplification (WGA) followed by gene copy number analysis via array comparative genomic hybridization (aCGH).CTCs from nine (9) patients successfully profiled were observed to have multiple copy number aberrations including those previously reported in primary prostate tumors such as gains in 8q and losses in 8p. High-level copy number gains at the androgen receptor (AR) locus were observed in 7 (78%) cases. Comparison of genomic profiles between CTCs and archival primary tumors from the same patients revealed common lineage. However, high-level copy number gains in the AR locus were observed in CTCs, but not in the matched archival primary tumors.We developed a new approach to isolate prostate CTCs without significant leukocyte admixture, and to subject them to genome-wide copy number analysis. Our assay may be utilized to explore genomic events involved in cancer progression, e.g. development of castration resistance and to monitor therapeutic efficacy of targeted therapies in clinical trials in a relatively non-invasive manner.

Dataset Information

Genomic Analysis of Localized High-Risk Prostate Cancer Circulating Tumor Cells at the Single-Cell Level.

Publications

Genomic Analysis of Localized High-Risk Prostate Cancer Circulating Tumor Cells at the Single-Cell Level.

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