Essential role of PLD2 in hypoxia-induced stemness and therapy resistance in ovarian tumors.

BACKGROUND: Hypoxia in solid tumors is an important source of chemoresistance that can determine poor patient prognosis. Such chemoresistance relies on the presence of cancer stem cells (CSCs), and hypoxia promotes their generation through transcriptional activation by HIF transcription factors. METHODS: We used ovarian cancer (OC) cell lines, xenograft models, OC patient samples, transcriptional databases, induced pluripotent stem cells (iPSCs) and Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq). RESULTS: Here, we show that hypoxia induces CSC formation and chemoresistance in ovarian cancer through transcriptional activation of the PLD2 gene. Mechanistically, HIF-1α activates PLD2 transcription through hypoxia response elements, and both hypoxia and PLD2 overexpression lead to increased accessibility around stemness genes, detected by ATAC-seq, at sites bound by AP-1 transcription factors. This in turn provokes a rewiring of stemness genes, including the overexpression of SOX2, SOX9 or NOTCH1. PLD2 overexpression also leads to decreased patient survival, enhanced tumor growth and CSC formation, and increased iPSCs reprograming, confirming its role in dedifferentiation to a stem-like phenotype. Importantly, hypoxia-induced stemness is dependent on PLD2 expression, demonstrating that PLD2 is a major determinant of de-differentiation of ovarian cancer cells to stem-like cells in hypoxic conditions. Finally, we demonstrate that high PLD2 expression increases chemoresistance to cisplatin and carboplatin treatments, both in vitro and in vivo, while its pharmacological inhibition restores sensitivity. CONCLUSIONS: Altogether, our work highlights the importance of the HIF-1α-PLD2 axis for CSC generation and chemoresistance in OC and proposes an alternative treatment for patients with high PLD2 expression.

Mirvetuximab Soravtansine in FRα-Positive, Platinum-Resistant Ovarian Cancer.

BACKGROUND: Mirvetuximab soravtansine-gynx (MIRV), a first-in-class antibody-drug conjugate targeting folate receptor α (FRα), is approved for the treatment of platinum-resistant ovarian cancer in the United States. METHODS: We conducted a phase 3, global, confirmatory, open-label, randomized, controlled trial to compare the efficacy and safety of MIRV with the investigator's choice of chemotherapy in the treatment of platinum-resistant, high-grade serous ovarian cancer. Participants who had previously received one to three lines of therapy and had high FRα tumor expression (≥75% of cells with ≥2+ staining intensity) were randomly assigned in a 1:1 ratio to receive MIRV (6 mg per kilogram of adjusted ideal body weight every 3 weeks) or chemotherapy (paclitaxel, pegylated liposomal doxorubicin, or topotecan). The primary end point was investigator-assessed progression-free survival; key secondary analytic end points included objective response, overall survival, and participant-reported outcomes. RESULTS: A total of 453 participants underwent randomization; 227 were assigned to the MIRV group and 226 to the chemotherapy group. The median progression-free survival was 5.62 months (95% confidence interval [CI], 4.34 to 5.95) with MIRV and 3.98 months (95% CI, 2.86 to 4.47) with chemotherapy (P<0.001). An objective response occurred in 42.3% of the participants in the MIRV group and in 15.9% of those in the chemotherapy group (odds ratio, 3.81; 95% CI, 2.44 to 5.94; P<0.001). Overall survival was significantly longer with MIRV than with chemotherapy (median, 16.46 months vs. 12.75 months; hazard ratio for death, 0.67; 95% CI, 0.50 to 0.89; P = 0.005). During the treatment period, fewer adverse events of grade 3 or higher occurred with MIRV than with chemotherapy (41.7% vs. 54.1%), as did serious adverse events of any grade (23.9% vs. 32.9%) and events leading to discontinuation (9.2% vs. 15.9%). CONCLUSIONS: Among participants with platinum-resistant, FRα-positive ovarian cancer, treatment with MIRV showed a significant benefit over chemotherapy with respect to progression-free and overall survival and objective response. (Funded by ImmunoGen; MIRASOL ClinicalTrials.gov number, NCT04209855.).

Clinical and molecular signature of survival and resistance to olaparib plus pegylated liposomal doxorubicin in platinum-resistant ovarian cancer: a stratified analysis from the phase II clinical trial ROLANDO, GEICO-1601.

OBJECTIVE: To determine the potential prognostic value of clinical and molecular biomarkers in the survival of patients with platinum-resistant ovarian cancer treated with olaparib and pegylated liposomal doxorubicin. METHODS: ROLANDO was a single-arm phase II trial that included patients with high-grade serous or endometrioid tumors and at least one previous platinum-resistant recurrence regardless of BRCA status. Patients received 6 cycles of pegylated liposomal doxorubicin every 28 days plus olaparib 300 mg twice daily. followed by olaparib 300 mg twice daily; monotherapy until progression or unacceptable toxicity. Prognostic factors including previous lines (and platinum-containing ones), BRCA mutation status, previous bevacizumab, CA-125 levels, and the neutrophil/lymphocyte ratio, lymphocyte/monocyte ratio, and platelet/lymphocyte ratio calculated at inclusion were analyzed through a multivariate logistic regression and factor analysis of mixed data. RESULTS: Thirty-one patients were included. Median age was 57 years (range 43-75), Eastern Cooperative Oncolgy Group performance status 0/1: 32.3%/67.7% and BRCA mutated: 16.1%. Prior treatment lines were >2 lines: 14 (45.2%) patients, ≥2 platinum lines: 21 patients (67.7%) and previous bevacizumab 19 (61.3%) patients. CA-125 was >2 upper limit normal in 24 (77.4%) patients. A high neutrophil/lymphocyte ratio was associated with worse overall survival by univariate/multivariate regression model (HR=11.18; 95% CI 1.1 to 114.5; p=0.042). No other factors were associated with overall survival in the multivariate model. A multifactorial signature based on clinical and molecular baseline characteristics was capable of defining six patient clusters. Three of these clusters had significantly better prognosis, with a median overall survival of 21.3 months (95% CI 12.2 to not reached). CONCLUSIONS: High neutrophil/lymphocyte ratio at platinum-resistant relapse indicated poor prognosis in patients treated with olaparib plus pegylated liposomal doxorubicin. A multifactorial clinical signature was more precise than single variables for implying the prognosis and may help in therapeutic assignment after further validation in large prospective cohorts.

Case-Control Analysis of the Impact of Anemia on Quality of Life in Patients with Cancer: A Qca Study Analysis.

Anemia is a common condition in cancer patients and is associated with a wide variety of symptoms that impair quality of life (QoL). However, exactly how anemia affects QoL in cancer patients is unclear because of the inconsistencies in its definition in previous reports. We aimed to examine the clinical impact of anemia on the QoL of cancer patients using specific questionnaires. We performed a post-hoc analysis of a multicenter, prospective, case-control study. We included patients with cancer with (cases) or without (controls) anemia. Participants completed the European Organization for Research and Treatment of Cancer Quality of Life questionnaire version 3.0 (EORTC QLQ-C30) and Euro QoL 5-dimension 3-level (EQ-5D-3L) questionnaire. Statistically significant and clinically relevant differences in the global health status were examined. From 2015 to 2018, 365 patients were included (90 cases and 275 controls). We found minimally important differences in global health status according to the EORTC QLQ-C30 questionnaire (case vs. controls: 45.6 vs. 58%, respectively; mean difference: -12.4, p < 0.001). Regarding symptoms, cancer patients with anemia had more pronounced symptoms in six out of nine scales in comparison with those without anemia. In conclusion, cancer patients with anemia had a worse QoL both clinically and statistically.

Downregulation of MYPT1 increases tumor resistance in ovarian cancer by targeting the Hippo pathway and increasing the stemness.

BACKGROUND: Ovarian cancer is one of the most common and malignant cancers, partly due to its late diagnosis and high recurrence. Chemotherapy resistance has been linked to poor prognosis and is believed to be linked to the cancer stem cell (CSC) pool. Therefore, elucidating the molecular mechanisms mediating therapy resistance is essential to finding new targets for therapy-resistant tumors. METHODS: shRNA depletion of MYPT1 in ovarian cancer cell lines, miRNA overexpression, RT-qPCR analysis, patient tumor samples, cell line- and tumorsphere-derived xenografts, in vitro and in vivo treatments, analysis of data from ovarian tumors in public transcriptomic patient databases and in-house patient cohorts. RESULTS: We show that MYPT1 (PPP1R12A), encoding myosin phosphatase target subunit 1, is downregulated in ovarian tumors, leading to reduced survival and increased tumorigenesis, as well as resistance to platinum-based therapy. Similarly, overexpression of miR-30b targeting MYPT1 results in enhanced CSC-like properties in ovarian tumor cells and is connected to the activation of the Hippo pathway. Inhibition of the Hippo pathway transcriptional co-activator YAP suppresses the resistance to platinum-based therapy induced by either low MYPT1 expression or miR-30b overexpression, both in vitro and in vivo. CONCLUSIONS: Our work provides a functional link between the resistance to chemotherapy in ovarian tumors and the increase in the CSC pool that results from the activation of the Hippo pathway target genes upon MYPT1 downregulation. Combination therapy with cisplatin and YAP inhibitors suppresses MYPT1-induced resistance, demonstrating the possibility of using this treatment in patients with low MYPT1 expression, who are likely to be resistant to platinum-based therapy.

New markers for human ovarian cancer that link platinum resistance to the cancer stem cell phenotype and define new therapeutic combinations and diagnostic tools.

BACKGROUND: Ovarian cancer is the leading cause of gynecologic cancer-related death, due in part to a late diagnosis and a high rate of recurrence. Primary and acquired platinum resistance is related to a low response probability to subsequent lines of treatment and to a poor survival. Therefore, a comprehensive understanding of the mechanisms that drive platinum resistance is urgently needed. METHODS: We used bioinformatics analysis of public databases and RT-qPCR to quantitate the relative gene expression profiles of ovarian tumors. Many of the dysregulated genes were cancer stem cell (CSC) factors, and we analyzed its relation to therapeutic resistance in human primary tumors. We also performed clustering and in vitro analyses of therapy cytotoxicity in tumorspheres. RESULTS: Using bioinformatics analysis, we identified transcriptional targets that are common endpoints of genetic alterations linked to platinum resistance in ovarian tumors. Most of these genes are grouped into 4 main clusters related to the CSC phenotype, including the DNA damage, Notch and C-KIT/MAPK/MEK pathways. The relative expression of these genes, either alone or in combination, is related to prognosis and provide a connection between platinum resistance and the CSC phenotype. However, the expression of the CSC-related markers was heterogeneous in the resistant tumors, most likely because there were different CSC pools. Furthermore, our in vitro results showed that the inhibition of the CSC-related targets lying at the intersection of the DNA damage, Notch and C-KIT/MAPK/MEK pathways sensitize CSC-enriched tumorspheres to platinum therapies, suggesting a new option for the treatment of patients with platinum-resistant ovarian cancer. CONCLUSIONS: The current study presents a new approach to target the physiology of resistant ovarian tumor cells through the identification of core biomarkers. We hypothesize that the identified mutations confer platinum resistance by converging to activate a few pathways and to induce the expression of a few common, measurable and targetable essential genes. These pathways include the DNA damage, Notch and C-KIT/MAPK/MEK pathways. Finally, the combined inhibition of one of these pathways with platinum treatment increases the sensitivity of CSC-enriched tumorspheres to low doses of platinum, suggesting a new treatment for ovarian cancer.

A Case-Control Analysis of the Impact of Venous Thromboembolic Disease on Quality of Life of Patients with Cancer: Quality of Life in Cancer (Qca) Study.

Although there is published research on the impact of venous thromboembolism (VTE) on quality of life (QoL), this issue has not been thoroughly investigated in patients with cancer-particularly using specific questionnaires. We aimed to examine the impact of acute symptomatic VTE on QoL of patients with malignancies. This was a multicenter, prospective, case-control study conducted in patients with cancer either with (cases) or without (controls) acute symptomatic VTE. Participants completed the EORTC QLQ-C30, EQ-5D-3L, PEmb-QoL, and VEINES-QOL/Sym questionnaires. Statistically significant and clinically relevant differences in terms of global health status were examined. Between 2015 and 2018, we enrolled 425 patients (128 cases and 297 controls; mean age: 60.2 ± 18.4 years). The most common malignancies were gastrointestinal (23.5%) and lung (19.8%) tumors. We found minimally important differences in global health status on the EQ-5D-3L (cases versus controls: 0.55 versus 0.77; mean difference: -0.22) and EORTC QLQ-C30 (47.7 versus 58.4; mean difference: -10.3) questionnaires. There were minimally important differences on the PEmb-QoL questionnaire (44.4 versus 23; mean difference: -21.4) and a significantly worse QoL on the VEINES-QOL/Sym questionnaire (42.7 versus 51.7; mean difference: -9). In conclusion, we showed that acute symptomatic VTE adversely affects the QoL of patients with malignancies.

Gene expression profile predictive of response to chemotherapy in metastatic colorectal cancer.

Fluoropyrimidine-based chemotherapy (CT) has been the mainstay of care of metastatic colorectal cancer (mCRC) for years. Response rates are only observed, however, in about half of treated patients, and there are no reliable tools to prospectively identify patients more likely to benefit from therapy. The purpose of our study was to identify a gene expression profile predictive of CT response in mCRC. Whole genome expression analyses (Affymetrix GeneChip HG-U133 Plus 2.0) were performed in fresh frozen tumor samples of 37 mCRC patients (training cohort). Differential gene expression profiles among the two study conditions (responders versus non-responders) were assessed using supervised class prediction algorithms. A set of 161 differentially expressed genes in responders (23 patients; 62%) versus non-responders (14 patients; 38%) was selected for further assessment and validation by RT-qPCR (TaqMan Low Density Arrays (TLDA) 7900 HT Micro Fluidic Cards) in an independent multi-institutional cohort (53 mCRC patients). Seven of these genes were confirmed as significant predictors of response. Patients with a favorable predictive signature had significantly greater response rate (58% vs. 13%, p = 0.024), progression-free survival (61% vs. 13% at 1 year, HR = 0.32, p = 0.009) and overall survival (32 vs. 16 months, HR = 0.21, p = 0.003) than patients with an unfavorable gene signature. This is the first study to validate a gene-expression profile predictive of response to CT in mCRC patients. Larger and prospective confirmatory studies are required, however, in order to successfully provide oncologists with adequate tools to optimize treatment selection in routine clinical practice.

MiR-107 and miR-99a-3p predict chemotherapy response in patients with advanced colorectal cancer.

BACKGROUND: MicroRNAs (miRNAs) are involved in numerous biological and pathological processes including colorectal cancer (CRC). The aim of our study was to evaluate the ability of miRNA expression patterns to predict chemotherapy response in a cohort of 78 patients with metastatic CRC (mCRC). METHODS: We examined expression levels of 667 miRNAs in the training cohort and evaluated their potential association with relevant clinical endpoints. We identified a miRNA profile that was analysed by RT-qPCR in an independent cohort. For a set of selected miRNAs, bioinformatic target predictions and pathway analysis were also performed. RESULTS: Eight miRNAs (let-7 g*, miR-107, miR-299-5p, miR-337-5p, miR-370, miR-505*, miR-889 and miR-99a-3p) were significant predictors of response to chemotherapy in the training cohort. In addition, overexpression of miR-107, miR-337-5p and miR-99a-3p, and underexpression of miR-889, were also significantly associated with improved progression-free and/or overall survival. MicroRNA-107 and miR-99a-3p were further validated in an independent cohort as predictive markers for chemotherapy response. In addition, an inverse correlation was confirmed in our study population between miR-107 levels and mRNA expression of several potential target genes (CCND1, DICER1, DROSHA and NFKB1). CONCLUSIONS: MiR-107 and miR-99a-3p were validated as predictors of response to standard fluoropyrimidine-based chemotherapy in patients with mCRC.

Spinophilin loss correlates with poor patient prognosis in advanced stages of colon carcinoma.

PURPOSE: The genomic region 17q21 is frequently associated with microsatellite instability and LOH in cancer, including gastric and colorectal carcinomas. This region contains several putative tumor suppressor genes, including Brca1, NM23, prohibitin, and spinophilin (Spn, PPP1R9B, neurabin II). The scaffold protein Spn is one of the regulatory subunits of phosphatase-1 (PP1) that targets PP1 to distinct subcellular locations and couples PP1 to its target. Thus, Spn may alter cell-cycle progression via the regulation of the phosphorylation status of the retinoblastoma protein, a direct target of PP1. Therefore, we analyzed whether Spn levels were reduced in colorectal carcinomas and whether Spn levels correlated with prognosis or response to therapy. EXPERIMENTAL DESIGN: By means of immunohistochemistry or quantitative PCR, we studied the levels of Spn in stages II, III, and IV colorectal carcinoma tumors and correlated to other clinicopathologic features as well as prognosis or response to therapy. RESULTS: Spn was lost in a percentage of human gastric, small intestine, and colorectal carcinomas. In patients with colorectal carcinoma, tumoral Spn downregulation correlated with a more aggressive histologic phenotype (poorer tumor differentiation and higher proliferative Ki67 index). Consistent with this observation, lower Spn protein expression levels were associated with faster relapse and poorer survival in patients with stage III colorectal carcinoma, particularly among those receiving adjuvant fluoropyrimidine therapy. We validated this result in an independent cohort of patients with metastatic colorectal carcinoma treated with standard chemotherapy. Although patients that achieved an objective tumor response exhibited Spn levels similar to nontumoral tissue, nonresponding patients showed a significant reduction in Spn mRNA levels. CONCLUSIONS: Our data suggest that Spn downregulation contributes to a more aggressive biologic behavior, induces chemoresistance, and is associated with a poorer survival in patients with advanced stages of colorectal carcinoma.

PDGFRα/ß and VEGFR2 polymorphisms in colorectal cancer: incidence and implications in clinical outcome.

BACKGROUND: Angiogenesis plays an essential role in tumor growth and metastasis, and is a major target in cancer therapy. VEGFR and PDGFR are key players involved in this process. The purpose of this study was to assess the incidence of genetic variants in these receptors and its potential clinical implications in colorectal cancer (CRC). METHODS: VEGFR2, PDGFRα and PDGFRß mutations were evaluated by sequencing their tyrosine kinase domains in 8 CRC cell lines and in 92 samples of patients with CRC. Correlations with clinicopathological features and survival were analyzed. RESULTS: Four SNPs were identified, three in PDGFRα [exon 12 (A12): c.1701A>G; exon 13 (A13): c.1809G>A; and exon 17 (A17): c.2439+58C>A] and one in PDGFRß [exon 19 (B19): c.2601A>G]. SNP B19, identified in 58% of tumor samples and in 4 cell lines (LS174T, LS180, SW48, COLO205), was associated with higher PDGFR and pPDGFR protein levels. Consistent with this observation, 5-year survival was greater for patients with PDGFR B19 wild type tumors (AA) than for those harboring the G-allele genotype (GA or GG) (51% vs 17%; p=0.073). Multivariate analysis confirmed SNP B19 (p=0.029) was a significant prognostic factor for survival, independent of age (p=0.060) or TNM stage (p<0.001). CONCLUSIONS: PDGFRß exon 19 c.2601A>G SNP is commonly encountered in CRC patients and is associated with increased pathway activation and poorer survival. Implications regarding its potential influence in response to PDGFR-targeted agents remain to be elucidated.