RESUMO
Previous studies have demonstrated focal but limited molecular similarities between circulating tumor cells (CTCs) and biopsies using isolated genetic assays. We hypothesized that molecular similarity between CTCs and tissue exists at the single cell level when characterized by whole genome sequencing (WGS). By combining the NanoVelcro CTC Chip with laser capture microdissection (LCM), we developed a platform for single-CTC WGS. We performed this procedure on CTCs and tissue samples from a patient with advanced prostate cancer who had serial biopsies over the course of his clinical history. We achieved 30X depth and ≥ 95% coverage. Twenty-nine percent of the somatic single nucleotide variations (SSNVs) identified were founder mutations that were also identified in CTCs. In addition, 86% of the clonal mutations identified in CTCs could be traced back to either the primary or metastatic tumors. In this patient, we identified structural variations (SVs) including an intrachromosomal rearrangement in chr3 and an interchromosomal rearrangement between chr13 and chr15. These rearrangements were shared between tumor tissues and CTCs. At the same time, highly heterogeneous short structural variants were discovered in PTEN, RB1, and BRCA2 in all tumor and CTC samples. Using high-quality WGS on single-CTCs, we identified the shared genomic alterations between CTCs and tumor tissues. This approach yielded insight into the heterogeneity of the mutational landscape of SSNVs and SVs. It may be possible to use this approach to study heterogeneity and characterize the biological evolution of a cancer during the course of its natural history.
Assuntos
Biomarcadores Tumorais/genética , Perfilação da Expressão Gênica/métodos , Genômica/métodos , Células Neoplásicas Circulantes/química , Neoplasias da Próstata/genética , Sequência de Bases , Biópsia , Separação Celular , Cromossomos Humanos , Análise Mutacional de DNA , Regulação Neoplásica da Expressão Gênica , Rearranjo Gênico , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Microdissecção e Captura a Laser , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/secundário , Masculino , Dados de Sequência Molecular , Mutação , Nanotecnologia , Células Neoplásicas Circulantes/patologia , Análise de Sequência com Séries de Oligonucleotídeos , Fenótipo , Polimorfismo de Nucleotídeo Único , Valor Preditivo dos Testes , Neoplasias da Próstata/sangue , Neoplasias da Próstata/patologia , Neoplasias da Próstata/terapia , Fatores de TempoRESUMO
Circulating tumor cells (CTCs) are one of the most crucial topics in rare cell biology and have become the focus of a significant and emerging area of cancer research. While CTC enumeration is a valid biomarker in prostate cancer, the current FDA-approved CTC technology is unable to detect CTCs in a large portion of late stage prostate cancer patients. Here we introduce the NanoVelcro CTC Chip, a device composed of a patterned silicon nanowire substrate (SiNW) and an overlaid polydimethylsiloxane (PDMS) chaotic mixer. Validated by two institutions participating in the study, the NanoVelcro Chip assay exhibits very consistent efficiency in CTC-capture from patient samples. The utilized protocol can be easily replicated at different facilities. We demonstrate the clinical utility of the NanoVelcro Chip by performing serial enumerations of CTCs in prostate cancer patients after undergoing systemic therapy. Changes in CTC numbers after 4-10 weeks of therapy were compared with their clinical responses. We observed a statistically significant reduction in CTCs counts in the clinical responders. We performed long-term follow up with serial CTC collection and enumeration in one patient observing variations in counts correlating with treatment response. This study demonstrates the consistency of the NanoVelcro Chip assay over time for CTC enumeration and also shows that continuous monitoring of CTC numbers can be employed to follow responses to different treatments and monitor disease progression.
