CRISPR/Cas9 mediated knockout of rb1 and rbl1 leads to rapid and penetrant retinoblastoma development in Xenopus tropicalis

Retinoblastoma is a pediatric eye tumor in which bi-allelic inactivation of the Retinoblastoma 1 (RB1) gene is the initiating genetic lesion. Although recently curative rates of retinoblastoma have increased, there are at this time no molecular targeted therapies available. This is, in part, due to the lack of highly penetrant and rapid retinoblastoma animal models that facilitate rapid identification of targets that allow therapeutic intervention. Different mouse models are available, all based on genetic deactivation of both Rb1 and Retinoblastoma-like 1 (Rbl1), and each showing different kinetics of retinoblastoma development. Here, we show by CRISPR/Cas9 techniques that similar to the mouse, neither rb1 nor rbl1 single mosaic mutant Xenopus tropicalis develop tumors, whereas rb1/rbl1 double mosaic mutant tadpoles rapidly develop retinoblastoma. Moreover, occasionally presence of pinealoblastoma (trilateral retinoblastoma) was detected. We thus present the first CRISPR/Cas9 mediated cancer model in Xenopus tropicalis and the first genuine genetic non-mammalian retinoblastoma model. The rapid kinetics of our model paves the way for use as a pre-clinical model. Additionally, this retinoblastoma model provides unique possibilities for fast elucidation of novel drug targets by triple multiplex CRISPR/Cas9 gRNA injections (rb1 + rbl1 + modifier gene) in order to address the clinically unmet need of targeted retinoblastoma therapy

preoperative c reactive protein and thrombocyte count as potential markers for longterm survival in ovarian cancer

n/

Tisotumab Vedotin in Combination with Carboplatin, Pembrolizumab, or Bevacizumab in Recurrent or Metastatic Cervical Cancer:Results from the innovaTV 205/GOG-3024/ENGOT-cx8 Study

PURPOSE Tissue factor is highly expressed in cervical carcinoma and can be targeted by tisotumab vedotin (TV), an antibody-drug conjugate. This phase Ib/II study evaluated TV in combination with bevacizumab, pembrolizumab, or carboplatin for recurrent or metastatic cervical cancer (r/mCC). METHODS This open-label, multicenter study (ClinicalTrials.gov identifier: NCT03786081) included dose-escalation arms that assessed dose-limiting toxicities (DLTs) and identified the recommended phase II dose (RP2D) of TV in combination with bevacizumab (arm A), pembrolizumab (arm B), or carboplatin (arm C). The dose-expansion arms evaluated TV antitumor activity and safety at RP2D in combination with carboplatin as first-line (1L) treatment (arm D) or with pembrolizumab as 1L (arm E) or second-/third-line (2L/3L) treatment (arm F). The primary end point of dose expansion was objective response rate (ORR). RESULTS A total of 142 patients were enrolled. In dose escalation (n = 41), no DLTs were observed; the RP2D was TV 2 mg/kg plus bevacizumab 15 mg/kg on day 1 once every 3 weeks, pembrolizumab 200 mg on day 1 once every 3 weeks, or carboplatin AUC 5 on day 1 once every 3 weeks. In dose expansion (n = 101), the ORR was 54.5% (n/N, 18/33; 95% CI, 36.4 to 71.9) with 1L TV + carboplatin (arm D), 40.6% (n/N, 13/32; 95% CI, 23.7 to 59.4) with 1L TV + pembrolizumab (arm E), and 35.3% (12/34; 19.7 to 53.5) with 2L/3L TV + pembrolizumab (arm F). The median duration of response was 8.6 months, not reached, and 14.1 months, in arms D, E, and F, respectively. Grade ≥3 adverse events (≥15%) were anemia, diarrhea, nausea, and thrombocytopenia in arm D and anemia in arm F (none ≥15%, arm E).CONCLUSION TV in combination with bevacizumab, carboplatin, or pembrolizumab demonstrated manageable safety and encouraging antitumor activity in treatment-naive and previously treated r/mCC.</p

Tisotumab Vedotin in Combination With Carboplatin, Pembrolizumab, or Bevacizumab in Recurrent or Metastatic Cervical Cancer: Results From the innovaTV 205/GOG-3024/ENGOT-cx8 Study.

PURPOSE: Tissue factor is highly expressed in cervical carcinoma and can be targeted by tisotumab vedotin (TV), an antibody-drug conjugate. This phase Ib/II study evaluated TV in combination with bevacizumab, pembrolizumab, or carboplatin for recurrent or metastatic cervical cancer (r/mCC). METHODS: This open-label, multicenter study (ClinicalTrials.gov identifier: NCT03786081) included dose-escalation arms that assessed dose-limiting toxicities (DLTs) and identified the recommended phase II dose (RP2D) of TV in combination with bevacizumab (arm A), pembrolizumab (arm B), or carboplatin (arm C). The dose-expansion arms evaluated TV antitumor activity and safety at RP2D in combination with carboplatin as first-line (1L) treatment (arm D) or with pembrolizumab as 1L (arm E) or second-/third-line (2L/3L) treatment (arm F). The primary end point of dose expansion was objective response rate (ORR). RESULTS: A total of 142 patients were enrolled. In dose escalation (n = 41), no DLTs were observed; the RP2D was TV 2 mg/kg plus bevacizumab 15 mg/kg on day 1 once every 3 weeks, pembrolizumab 200 mg on day 1 once every 3 weeks, or carboplatin AUC 5 on day 1 once every 3 weeks. In dose expansion (n = 101), the ORR was 54.5% (n/N, 18/33; 95% CI, 36.4 to 71.9) with 1L TV + carboplatin (arm D), 40.6% (n/N, 13/32; 95% CI, 23.7 to 59.4) with 1L TV + pembrolizumab (arm E), and 35.3% (12/34; 19.7 to 53.5) with 2L/3L TV + pembrolizumab (arm F). The median duration of response was 8.6 months, not reached, and 14.1 months, in arms D, E, and F, respectively. Grade ≥3 adverse events (≥15%) were anemia, diarrhea, nausea, and thrombocytopenia in arm D and anemia in arm F (none ≥15%, arm E). CONCLUSION: TV in combination with bevacizumab, carboplatin, or pembrolizumab demonstrated manageable safety and encouraging antitumor activity in treatment-naive and previously treated r/mCC

Curative effect of second curettage for treatment of gestational trophoblastic disease - Results of the Belgian registry for gestational trophoblastic disease.

peer reviewedOBJECTIVE: We assessed the curative effect of a second curettage in patients with persistent hCG serum levels after first curettage for a gestational trophoblastic disease (GTD). STUDY DESIGN: This prospective observational study used the data of the Belgian register for GTD between July 2012 and January 2017. We analysed the data of patients who underwent a second curettage. We included 313 patients in the database. Primary endpoints were need for second curettage and chemotherapy. RESULTS: Thirty-seven patients of the study population (12 %) underwent a second curettage. 20 had persistent human chorionic gonadotropin hormone (hCG) elevation before second curettage. Of them, 9 patients (45 %) needed no further treatment afterwards. Eleven patients (55 %) needed further chemotherapy. Nine (82 %) were cured with single-agent chemotherapy and 2 patients (18 %) needed multi-agent chemotherapy. Of the 37 patients, patients with hCG levels below 5000 IU/L undergoing a second curettage were cured without chemotherapy in 65 % versus 45 % of patients with hCG level more than 5000 IU/L. Of the ten patients with a hCG level below 1000 IU/L, eight were cured without chemotherapy. CONCLUSIONS: Patients with post-mole gestational trophoblastic neoplasia can benefit from a second curettage to avoid chemotherapy, especially when the hCG level is lower than 5000 IU/L

No evidence that genetic variation in the myeloid-derived suppressor cell pathway influences ovarian cancer survival

BACKGROUND: The precise mechanism by which the immune system is adversely affected in cancer patients remains poorly understood, but the accumulation of immune suppressive/pro-tumorigenic myeloid-derived suppressor cells (MDSCs) is thought to be one prominent mechanism contributing to immunologic tolerance of malignant cells in epithelial ovarian cancer (EOC). To this end, we hypothesized genetic variation in MDSC pathway genes would be associated with survival after EOC diagnoses. METHODS: We measured the hazard of death due to EOC within 10 years of diagnosis, overall and by invasive subtype, attributable to SNPs in 24 genes relevant in the MDSC pathway in 10,751 women diagnosed with invasive EOC. Versatile Gene-based Association study (VEGAS) and the Admixture Likelihood method (AML), were used to test gene and pathway associations with survival. RESULTS: We did not identify individual SNPs that were significantly associated with survival after correction for multiple testing (p<3.5 x 10-5), nor did we identify significant associations between the MDSC pathway overall, or the 24 individual genes and EOC survival. CONCLUSIONS: In this well-powered analysis, we observed no evidence that inherited variations in MDSC-associated SNPs, individual genes, or the collective genetic pathway contributed to EOC survival outcomes. IMPACT: Common inherited variation in genes relevant to MDSCs were not associated with survival in women diagnosed with invasive EOC

Methylome analysis of extreme chemoresponsive patients identifies novel markers of platinum sensitivity in high-grade serous ovarian cancer

Background: Despite 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. Methods: Genome-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 ( Results: Integrated 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. Conclusions: By 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

Germline polymorphisms in an enhancer of PSIP1 are associated with progression-free survival in epithelial ovarian cancer.

Women with epithelial ovarian cancer (EOC) are usually treated with platinum/taxane therapy after cytoreductive surgery but there is considerable inter-individual variation in response. To identify germline single-nucleotide polymorphisms (SNPs) that contribute to variations in individual responses to chemotherapy, we carried out a multi-phase genome-wide association study (GWAS) in 1,244 women diagnosed with serous EOC who were treated with the same first-line chemotherapy, carboplatin and paclitaxel. We identified two SNPs (rs7874043 and rs72700653) in TTC39B (best P=7x10-5, HR=1.90, for rs7874043) associated with progression-free survival (PFS). Functional analyses show that both SNPs lie in a putative regulatory element (PRE) that physically interacts with the promoters of PSIP1, CCDC171 and an alternative promoter of TTC39B. The C allele of rs7874043 is associated with poor PFS and showed increased binding of the Sp1 transcription factor, which is critical for chromatin interactions with PSIP1. Silencing of PSIP1 significantly impaired DNA damage-induced Rad51 nuclear foci and reduced cell viability in ovarian cancer lines. PSIP1 (PC4 and SFRS1 Interacting Protein 1) is known to protect cells from stress-induced apoptosis, and high expression is associated with poor PFS in EOC patients. We therefore suggest that the minor allele of rs7874043 confers poor PFS by increasing PSIP1 expression.This project has been supported by a grant from Cancer Australia. The Mayo Clinic GWAS was supported by R01CA114343 (Haplotype-based genome screen for ovarian cancer loci). The Ovarian Cancer Association Consortium is supported by a grant from the Ovarian Cancer Research Fund thanks to donations by the family and friends of Kathryn Sladek Smith. The AOCS was supported by the U.S. Army Medical Research and Materiel Command under DAMD17-01-1-0729, the National Health and Medical Research Council (NHMRC) of Australia (grants 400281, 400413), Cancer Council Victoria, Cancer Council Queensland, Cancer Council New South Wales, Cancer Council South Australia, The Cancer Foundation of Western Australia, and Cancer Council Tasmania. G. Chenevix-Trench is a Senior Principal Research fellow of the NHMRC. Y. Lu is funded by NHMRC grant 496675, S. MacGregor is supported by an NHMRC career development award, S. Edwards and J. French are supported by Fellowships from the National Breast Cancer Foundation (NBCF) Australia. The QIMR Berghofer groups were supported by NHMRC project grants (1051698 to SM and 1058415 to SLE and JDF) and a Weekend to End Women’s Cancer Research Grant (to SLE). A deFazio is funded by the University of Sydney Cancer Research Fund and A deFazio and PR Harnett are funded by the Cancer Institute NSW through the Sydney-West Translational Cancer Research Centre. B. Gao is supported by NHMRC and Cancer Institute NSW scholarship. KBM and MO’R are funded by CR-UK. The Bavarian study (BAV) was supported by ELAN Funds of the University of Erlangen-Nuremberg. HSK would like to thank Ira Schwaab for her tireless work on sample preparation. The Belgian study (BEL) was funded by Nationaal Kankerplan and we would like to thank Gilian Peuteman, Thomas Van Brussel and Dominiek Smeets for technical assistance. The Japanese study (JPN) was funded by a Grant-in-Aid for the Third Term Comprehensive 10-Year Strategy for Cancer Control from the Ministry of Health, Labour and Welfare. The International Collaborative Ovarian Neoplasm study (ICON)7 trial team would like to thank the Medical Research Council (MRC) Clinical Trial Unit (CTU) at the University of London (UCL), the ICON7 Translational Research Sub-group, and the University of Leeds for their work on the coordination of samples and data from the ICON7 trial. The LAX study (Women’s Cancer Program) was supported by the American Cancer Society Early Detection Professorship (120950-SIOP-06-258-06-COUN) and Entertainment Industry Foundation. Funding for MALOVA (MAL) was provided by research grant RO1 CA 61107 from the National Cancer Institute, Bethesda, MD; research grant 94 222 52 from the Danish Cancer Society, Copenhagen, Denmark; and the Mermaid I project. The Mayo Clinic study (MAYO) was supported by R01 CA122443, P50 CA136393. The Oregon study (ORE) was funded by the Sherie Hildreth Ovarian Cancer Research Fund and the OHSU Foundation. We would like to thank all members of Scottish Gynaecological Clinical Trials group and the SCOTROC1 investigators. SCOTROC1 (SRO) was funded by Cancer Research UK, and the SCOTROC biological studies were supported by Cancer Research UK (grant C536/A6689). RSH receives support from NIH/NIGMS grant K08GM089941, NIH/NCI grant R21 CA139278, NIH/NIGMS grant UO1GM61393, University of Chicago Cancer Center Support Grant (#P30 CA14599) and Breast Cancer SPORE Career Development Award.This is the final version of the article. It first appeared from Impact Journals via http://dx.doi.org/10.18632/oncotarget.704

rs495139 in the TYMS-ENOSF1 Region and Risk of Ovarian Carcinoma of Mucinous Histology

Thymidylate synthase (TYMS) is a crucial enzyme for DNA synthesis. TYMS expression is regulated by its antisense mRNA, ENOSF1. Disrupted regulation may promote uncontrolled DNA synthesis and tumor growth. We sought to replicate our previously reported association between rs495139 in the TYMS-ENOSF1 3' gene region and increased risk of mucinous ovarian carcinoma (MOC) in an independent sample. Genotypes from 24,351 controls to 15,000 women with invasive OC, including 665 MOC, were available. We estimated per-allele odds ratios (OR) and 95% confidence intervals (CI) using unconditional logistic regression, and meta-analysis when combining these data with our previous report. The association between rs495139 and MOC was not significant in the independent sample (OR = 1.09; 95% CI = 0.97-1.22; p = 0.15; N = 665 cases). Meta-analysis suggested a weak association (OR = 1.13; 95% CI = 1.03-1.24; p = 0.01; N = 1019 cases). No significant association with risk of other OC histologic types was observed (p = 0.05 for tumor heterogeneity). In expression quantitative trait locus (eQTL) analysis, the rs495139 allele was positively associated with ENOSF1 mRNA expression in normal tissues of the gastrointestinal system, particularly esophageal mucosa (r = 0.51, p = 1.7 x 10(-28)), and nonsignificantly in five MOC tumors. The association results, along with inconclusive tumor eQTL findings, suggest that a true effect of rs495139 might be small.Peer reviewe