Dataset Information

CRISPR/Cas9-derived models of ovarian high grade serous carcinoma targeting Brca1, Pten and Nf1, and correlation with platinum sensitivity.

ABSTRACT: Transplantable murine models of ovarian high grade serous carcinoma (HGSC) remain an important research tool. We previously showed that ID8, a widely-used syngeneic model of ovarian cancer, lacked any of the frequent mutations in HGSC, and used CRISPR/Cas9 gene editing to generate derivatives with deletions in Trp53 and Brca2. Here we have used one ID8 Trp53 -/- clone to generate further mutants, with additional mutations in Brca1, Pten and Nf1, all of which are frequently mutated or deleted in HGSC. We have also generated clones with triple deletions in Trp53, Brca2 and Pten. We show that ID8 Trp53 -/-;Brca1 -/- and Trp53 -/-;Brca2 -/- cells have defective homologous recombination and increased sensitivity to both platinum and PARP inhibitor chemotherapy compared to Trp53 -/-. By contrast, loss of Pten or Nf1 increases growth rate in vivo, and reduces survival following cisplatin chemotherapy in vivo. Finally, we have also targeted Trp53 in cells isolated from a previous transgenic murine fallopian tube carcinoma model, and confirmed that loss of p53 expression in this second model accelerates intraperitoneal growth. Together, these CRISPR-generated models represent a new and simple tool to investigate the biology of HGSC, and the ID8 cell lines are freely available to researchers.

SUBMITTER: Walton JB

PROVIDER: S-EPMC5715106 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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CRISPR/Cas9-derived models of ovarian high grade serous carcinoma targeting Brca1, Pten and Nf1, and correlation with platinum sensitivity.

Walton Josephine B JB Farquharson Malcolm M Mason Susan S Port Jennifer J Kruspig Bjorn B Dowson Suzanne S Stevenson David D Murphy Daniel D Matzuk Martin M Kim Jaeyeon J Coffelt Seth S Blyth Karen K McNeish Iain A IA

Scientific reports 20171204 1

Transplantable murine models of ovarian high grade serous carcinoma (HGSC) remain an important research tool. We previously showed that ID8, a widely-used syngeneic model of ovarian cancer, lacked any of the frequent mutations in HGSC, and used CRISPR/Cas9 gene editing to generate derivatives with deletions in Trp53 and Brca2. Here we have used one ID8 Trp53 <sup>-/-</sup> clone to generate further mutants, with additional mutations in Brca1, Pten and Nf1, all of which are frequently mutated or ...[more]

PMID: 29203787

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Project description:Expression of the progesterone receptor (PR) has been reported to influence survival outcomes in patients with ovarian high-grade serous carcinoma (HGSC). In the present study, we attempted to investigate the association among PR and its isoforms' expression, platinum sensitivity, and survival in ovarian HGSC. This retrospective study reviewed ovarian HGSC patients who received surgery followed by adjuvant chemotherapy. We analyzed total PR and PR isoform-B (PR-B) expression by immunohistochemical staining and quantified using the H-score. Then, we compared platinum sensitivity and survival outcomes between those patients with weak and strong PR-B expression. Cisplatin viability assays were carried out in ovarian HGSC cell lines (OC-3-VGH and OVCAR-3) with different PR-B expression. Among 90 patients, 49 and 41 patients were considered to have platinum-sensitive and platinum-resistant disease, respectively. Pearson's correlation model showed that the H-score of total PR correlated positively with PR-B (r = 0.813). The PR-B H-score of tumors was significantly higher in the platinum-sensitive group (p = 0.004). Multivariate analysis revealed that the PR-B H-score and optimal debulking status were independent factors predicting platinum sensitivity. When compared with strong PR-B expression, patients with weak PR-B had significantly poorer progression-free (p = 0.021) and cancer-specific survival (p = 0.046). In a cell model, cisplatin-resistant OC-3-VGH cells expressed a lower level of PR-B than wild-type cells. Overexpression of PR-B or progesterone could increase cisplatin sensitivity in both OC-3-VGH and OVCAR-3 cells via the mechanism of promoting cisplatin-related apoptosis. When compared to weak PR-B, ovarian HGSC patients with a strong PR-B expression had a better chance of platinum sensitivity and survival, and this finding was compatible with our experimental results. Progesterone seemed to be a platinum sensitizer, but the value of adding progesterone in the treatment of ovarian HGSC should be further investigated.

Project description:BackgroundDespite an early response to platinum-based chemotherapy in advanced stage high-grade serous ovarian cancer (HGSOC), the majority of patients will relapse with drug-resistant disease. Aberrant epigenetic alterations like DNA methylation are common in HGSOC. Differences in DNA methylation are associated with chemoresponse in these patients. The objective of this study was to identify and validate novel epigenetic markers of chemoresponse using genome-wide analysis of DNA methylation in extreme chemoresponsive HGSOC patients.MethodsGenome-wide next-generation sequencing was performed on methylation-enriched tumor DNA of two HGSOC patient groups with residual disease, extreme responders (≥18 months progression-free survival (PFS), n = 8) and non-responders (≤6 months PFS, n = 10) to platinum-based chemotherapy. DNA methylation and expression data of the same patients were integrated to create a gene list. Genes were validated on an independent cohort of extreme responders (n = 21) and non-responders (n = 31) using pyrosequencing and qRT-PCR. In silico validation was performed using publicly available DNA methylation (n = 91) and expression (n = 208) datasets of unselected advanced stage HGSOC patients. Functional validation of FZD10 on chemosensitivity was carried out in ovarian cancer cell lines using siRNA-mediated silencing.ResultsIntegrated genome-wide methylome and expression analysis identified 45 significantly differentially methylated and expressed genes between two chemoresponse groups. Four genes FZD10, FAM83A, MYO18B, and MKX were successfully validated in an external set of extreme chemoresponsive HGSOC patients. High FZD10 and MKX methylation were related with extreme responders and high FAM83A and MYO18B methylation with non-responders. In publicly available advanced stage HGSOC datasets, FZD10 and MKX methylation levels were associated with PFS. High FZD10 methylation was strongly associated with improved PFS in univariate analysis (hazard ratio (HR) = 0.43; 95% CI, 0.27-0.71; P = 0.001) and multivariate analysis (HR = 0.39; 95% CI, 0.23-0.65; P = 0.003). Consistently, low FZD10 expression was associated with improved PFS (HR = 1.36; 95% CI, 0.99-1.88; P = 0.058). FZD10 silencing caused significant sensitization towards cisplatin treatment in survival assays and apoptosis assays.ConclusionsBy applying genome-wide integrated methylome analysis on extreme chemoresponsive HGSOC patients, we identified novel clinically relevant, epigenetically-regulated markers of platinum-sensitivity in HGSOC patients. The clinical potential of these markers in predictive and therapeutic approaches has to be further validated in prospective studies.

Dataset Information

CRISPR/Cas9-derived models of ovarian high grade serous carcinoma targeting Brca1, Pten and Nf1, and correlation with platinum sensitivity.

Publications

CRISPR/Cas9-derived models of ovarian high grade serous carcinoma targeting Brca1, Pten and Nf1, and correlation with platinum sensitivity.

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