bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract B13: Utilizing metabolic reprogramming to regulate fibroblast phenotype and reduce radiation fibrosis
Authors: Zhao Xiao , Psarianos Pamela , Goldstein David , Gilbert Ralph , Witterick Ian , Ailles Laurie , Haibe-Kains Benjamin , Liu Fei-Fei
Published: September 2020

Introduction: Radiation fibrosis affects up to half of all patients undergoing radiotherapy for head and neck cancer treatment. It is characterized by excess extracellular matrix deposition, leading to tissue dysfunction, distortion, hardening, and pain. There are currently no effective treatments for radiation fibrosis. A fundamental challenge has been to understand and modulate the function of fibroblasts, a key mediator in extracellular matrix production and degradation. Recent evidence has demonstrated that metabolic regulation plays an important role in altering cell behavior in cancer biology and immunology. However, little is known about how metabolic changes affect fibroblast phenotype.

Hypothesis: We propose that metabolic reprogramming may be an effective strategy to modulate fibroblast function and reduce radiation fibrosis.

Results: An interplay between fatty acid oxidation and glycolysis was found to be a convergence point directly governing fibroblast behavior. It was demonstrated that manipulating the balance between FAO and glycolysis in fibroblasts induced either an extracellular matrix anabolic or catabolic phenotype. Specifically, anabolic fibroblasts relied on a fatty acid oxidation to glycolysis shift to produce extracellular matrix components. Reversal of this metabolic shift generated a catabolic fibroblast, which downregulated extracellular matrix synthesis and upregulated extracellular matrix lysosomal degradation. We further uncovered that CD36, a multifunctional fatty acid transporter, was a crucial link connecting fibroblast metabolism with extracellular matrix regulation, as its depletion completely inhibited collagen-1 internalization and degradation. Finally, through metabolic reprogramming using fibroblasts expressing high levels of CD36, but not CD36 knockout fibroblasts, metabolic balance could be restored, and in turn extracellular matrix accumulation was reduced in murine radiation fibrosis.

Conclusions: We have uncovered that a fibroblast’s phenotype can be differentiated and regulated based on its metabolic signature. These findings have significant implications for drug development and for future cellular therapies to reduce radiation fibrosis.

Citation Format: Xiao Zhao, Pamela Psarianos, David Goldstein, Ralph Gilbert, Ian Witterick, Laurie Ailles, Benjamin Haibe-Kains, Fei-Fei Liu. Utilizing metabolic reprogramming to regulate fibroblast phenotype and reduce radiation fibrosis [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; 2019 Apr 29-30; Austin, TX. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_2):Abstract nr B13.
Abstract B17: Functional characterization of HPV-human fusion transcripts in oropharyngeal cancer
Authors: Pinatti Lisa M. , Walline Heather M. , Carey Thomas E.
Published: September 2020

Human papillomavirus (HPV) genomic integration is frequently seen in oropharyngeal squamous cell carcinoma (OPSCC). Integration into cellular genes can cause the generation of HPV-host fusion transcripts, which have been associated with worse survival in OPSCC patients, but their functional consequences are unknown, as HPV alone is capable of transformation. The forced expression of fusion transcripts versus HPV transcripts in spontaneously immortalized human oral keratinocytes (NOKSI) could demonstrate their effects on cell behavior. Our research focuses on characterizing HPV integration sites and fusion transcripts in cell lines and tumors, followed by in vitro analysis of fusion transcript function. By Detection of Integrated Papillomavirus Sequences (DIPS-PCR) and RT-PCR, we have identified integration events and fusion transcripts in a subset of samples in both intergenic and genic regions of the genome. We began our analysis with two fusion transcripts, one reading from HPV16 E6*-E7-E1 into the tumor suppressor gene TP63 from UM-SCC-47 and the other reading from HPV16 E6*-E4 into the solute carrier gene SLC47A2 from UM-SCC-104. The full fusion transcripts and associated controls were cloned into lentiviral vectors, and stable keratinocyte populations expressing these transcripts were selected. These cultures were subjected to proliferation, migration, and invasion assays. Subsequent in-depth molecular characterization and in vivo studies will help clarify their effects on cancer formation and metastasis. These studies will advance our understanding of how fusion transcripts lead to worse patient survival and help develop new therapies for these patients.

Citation Format: Lisa M. Pinatti, Heather M. Walline, Thomas E. Carey. Functional characterization of HPV-human fusion transcripts in oropharyngeal cancer [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; 2019 Apr 29-30; Austin, TX. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_2):Abstract nr B17.
Abstract B35: Characterization of exosomes extracted from primary tumor samples of HPV+/- oropharyngeal cancer
Authors: Wu Vivian F. , Dai Xiang Guo , Chopp Michael
Published: September 2020

Objective: To determine if distinctions in exosomal cargo contribute to differences in EMT between HPV-related and HPV-unrelated oropharyngeal disease and can provide a signature for prediction of tumor aggressiveness and metastatic potential.

Subjects and Methods: Specimen from patients with HPV+/- OPSCC were collected according to an IRB-approved protocol including primary tumor, lymph node metastasis, distant metastasis, and serum when available. Exosomes isolated from HPV+ and HPV- tumors were analyzed with miARma-seq and proteomics. These same exosomes were cocultured with well-characterized HPV+ and HPV- cell lines as well as fibroblasts and normal keratinocytes. Tumor aggressiveness, EMT, and potential for metastasis were measured utilizing wound healing scratch assays and transwell invasion assay techniques based on the Boyden Chamber migration assay. Confirmation of epithelial to mesenchymal transition was evaluated by Western blot and qPCR using a panel of well-established EMT markers (e-cadherin, n-cadherin, vimentin, snail, slug, twist, ZEB1/2).

Results: Tumors from HPV+ patients demonstrate exosomal content enriched in EMT markers compared to HPV-negative patient cultures. Exosomes extracted from primary tumor and lymph node metastasis of the same patient demonstrate differential expression of markers of EMT. Exosomes extracted from lymph node and cocultured with fibroblasts demonstrate increased invasion and migration.

Conclusions: Exosomes can be extracted from primary patient tumor cultures of both primary tumor and lymph node metastasis. These exosomes demonstrate differential expression of markers of EMT. Exosomal contents can be exploited for biomarker discovery to aide in prognostication.

Citation Format: Vivian F. Wu, Xiang Guo Dai, Michael Chopp. Characterization of exosomes extracted from primary tumor samples of HPV+/- oropharyngeal cancer [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; 2019 Apr 29-30; Austin, TX. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_2):Abstract nr B35.
Abstract B40: Trends in lymph node yield for neck dissection and oral cavity carcinoma survival
Authors: Patel Punam , Kedarisetty Suraj , Lango Miriam
Published: September 2020

Objectives: Lymph node yields (LNYs) following neck dissection have been associated with differences in oral cavity cancer survival. The objective of this study was to evaluate temporal trends in LNYs in neck dissections for oral cavity carcinoma.

Methods: Data were obtained from the Surveillance, Epidemiology, and End Results (SEER 18) database. Inclusion criteria included diagnosis of malignant oral cavity cancer (tongue and floor of mouth) squamous cell carcinomas diagnosed from January 1, 2006-December 31, 2016. Patients with AJCC 6th Edition Stages 1-4a who underwent surgery were included. All-cause age-adjusted mortality for tongue and floor of mouth cancers during this period was assessed using SEER*Stat.

Results: The median LNY among 8,580 patients in the sample was 25 LNs (Interquartile range 14-39). LNY statistically increased over time from 22 LNs in 2004 to 27 in 2014 (p < 0.0001). In contrast, there was no change in the median number of positive LNs during the same period (median 2 positive LNs in 2004 and 2014). There was no significant difference in lymph node yield based on sex, region, insurance status, or relative location to a metropolitan center. After adjusting for patient age, sex, race, marital status, regional income, and percent current smokers in the county of residence, tumor subsite and TNM stage, LNY less than 18 was independently associated with a 44% greater risk of death from any cause, compared with greater LNYs (HR 1.44, 95% CI 1.29-1.60, p=.0001). During this period, all-cause age-adjusted mortality declined from 469.7 per 100,000 (468.2-471.2) to 429.0/100,000 (428.2-430.8).

Conclusion: LNY following neck dissection for oral tongue and floor of mouth cancers increased significantly over a decade, during which survival also improved. The cause of improved LNY is not known but may nevertheless be contributing to declining mortality in patients with oral tongue and floor of mouth cancers.

Citation Format: Punam Patel, Suraj Kedarisetty, Miriam Lango. Trends in lymph node yield for neck dissection and oral cavity carcinoma survival [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; 2019 Apr 29-30; Austin, TX. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_2):Abstract nr B40.
Abstract IA10: Targeting glutamine metabolism as a means of enhancing immunotherapy for cancer
Authors: Powell Jonathan , Leone Robert , Oh Min-Hee , Rais Rana , Majer Pavel , Slusher Barbara
Published: September 2020

Tumors possess specialized metabolic programming that facilitates their rapid growth. We have developed a novel glutamine antagonist that not only demonstrates single-agent efficacy in a variety of mouse tumor models but also synergizes with anti-PD-1, adoptive cellular therapy (ACT), and A2aR antagonism to overcome resistance to immunotherapy and promote durable cures. Our data demonstrate that targeting tumor glutamine metabolism alters the tumor microenvironment by making it less acidic, less hypoxic, and more nutrient replete. These alterations lead to enhanced endogenous antitumor responses (in the absence of additional immunotherapy). Further analysis of tumor-infiltrating lymphocyte (TIL) reveals markedly increased infiltration of CD8+ T cells with increased proliferative index (Ki67) and the generation of robust memory phenotypes in the treated mice. Further, RNA sequencing data revealed that CD8+ TIL from vehicle-treated mice showed significantly enhanced expression of apoptotic transcriptional programs compared with TIL from treated mice. Additionally, targeting glutamine metabolism leads to decreased MDSC with an increase in inflammatory TAMs. Furthermore, we observe a downregulation of IDO and a marked decrease in kynurenine in the tumors of treated mice. These changes in macrophage subsets also correlated with decreased metastasis. Overall our findings suggest a novel “metabolic” approach to enhancing immunotherapy for cancer.

Citation Format: Jonathan Powell, Robert Leone, Min-Hee Oh, Rana Rais, Pavel Majer, Barbara Slusher. Targeting glutamine metabolism as a means of enhancing immunotherapy for cancer [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; 2019 Apr 29-30; Austin, TX. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_2):Abstract nr IA10.
Abstract PR02: Mutational analysis of head and neck squamous cell carcinoma stratified by smoking status
Authors: Ghasemi Farhad , Prokopec Stephenie D. , MacNeil Danielle , Mundi Neil , Howlett Christopher , Stecho William , Fung Kevin , Yoo John , Winquist Eric , Gameiro Steven , Sahovaler Axel , Plantinga Paul , Mymryk Joe S. , Barrett John W. , Boutros Paul C. , Nichols Anthony C.
Published: September 2020

Smoking has historically been recognized as a negative prognostic factor in head and neck squamous cell carcinoma (HNSCC). This study aimed to assess the mutational differences between heavy smokers (>20 pack-years) and never smokers among the HNSCC patients within The Cancer Genome Atlas (TCGA). Single nucleotide variation (SNV) and copy number aberration (CNA) differences between heavy smokers and never smokers were compared within HPV-positive (n=67) and negative (n=431) TCGA HNSCC cohorts, and the impact of these mutations on survival was assessed. No genes were differentially mutated between smoking and never smoking patients with HPV-positive tumors. By contrast, in HPV-negative tumors, NSD1 and COL1A11 were found to be more frequently mutated in heavy smokers, while CASP8 was more frequently altered in never smokers. HPV-negative patients with NSD1 mutations experienced significantly improved overall survival compared with NSD1 wild-type patients. This improved prognosis was validated in an independent cohort of 77 oral cavity cancers and a meta-analysis that included two additional datasets (688 total patients, hazard ratio for death 0.44, 95% CI 0.30-0.65). NSD1 mutations are more common in HPV-negative heavy smokers and define a cohort with favorable prognosis, and may represent a clinically useful biomarker to guide treatment deintensification for HPV-negative patients.

Citation Format: Farhad Ghasemi, Stephenie D. Prokopec, Danielle MacNeil, Neil Mundi, Christopher Howlett, William Stecho, Kevin Fung, John Yoo, Eric Winquist, Steven Gameiro, Axel Sahovaler, Paul Plantinga, Joe S. Mymryk, John W. Barrett, Paul C. Boutros, Anthony C. Nichols. Mutational analysis of head and neck squamous cell carcinoma stratified by smoking status [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; 2019 Apr 29-30; Austin, TX. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_2):Abstract nr PR02.
Abstract PR07: WEE1 inhibition triggers premature mitotic entry in HPV16-E6-expressing cells independent of p53 inactivation
Authors: Diab Ahmed , Swanger Jherek , Gem Hakan , Galloway Denise , Mendez Eduardo , Sidorova Julia , Clurman Bruce
Published: September 2020

Inhibition of the protein kinase WEE1 induces cancer cell death by forcing cells arrested in S-phase directly into mitosis. However, in the absence of S-phase perturbations, WEE1 inhibition is typically insufficient to trigger premature mitosis. Here, we show that WEE1 inhibition alone drives premature mitotic entry in HPV+ head and neck cancer cells. We also observed unscheduled mitotic entry in E6- and E6E7-expressing HPV-negative UMSCC74a cells upon WEE1 inhibition, but not in empty vector controls. Unlike E6 expression, p53 depletion alone was not sufficient to cooperate with WEE1 inhibition to effect forced mitotic entry. In vitro kinase assays showed elevated CDK1 activity in E6 and E6E7 cells compared to empty vector controls, particularly after WEE1 inhibition. We find that addiction of E6E7 cells to cell cycle inhibitor p21 causes the AZD1775-mediated CDK hyperactivation, therefore intensifying genotoxic replication stress. WEE1 inhibition therefore exacerbates E6E7-associated replication stress leading to slowing of replication forks, excess single-stranded DNA, and elevated RPA loading. RPA depletion further sensitized E6E7 cells to WEE1 inhibition-induced ssDNA accumulation, DNA damage, and cell death, implying a role for RPA in limiting WEE1i-induced replication stress in E6E7-expressing cells. Hallmarks of replication stress and DNA damage persisted in E6E7-expressing cells after recovery from WEE1 inhibition. These observations collectively explain the unique hypersensitivity of HPV+ head and neck tumors to WEE1i.

Citation Format: Ahmed Diab, Jherek Swanger, Hakan Gem, Denise Galloway, Eduardo Mendez, Julia Sidorova, Bruce Clurman. WEE1 inhibition triggers premature mitotic entry in HPV16-E6-expressing cells independent of p53 inactivation [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; 2019 Apr 29-30; Austin, TX. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_2):Abstract nr PR07.
Abstract A18: Evaluating the potential to repurpose statins for ovarian cancer therapy
Authors: Kroeger Paul T. , Drapkin Ronny
Published: September 2020

The recent success of PARP inhibitors in patients with BRCA1/2 mutant ovarian tumors highlights the importance of identifying druggable vulnerabilities in this disease. The fact that not all ovarian cancer patients will benefit from PARP inhibitors or experience durable responses places great emphasis on the need to identify additional therapies. Repurposing existing FDA-approved drugs offers an opportunity to accelerate the progress of drug development. Statins are drugs primarily prescribed for hypercholesterolemia and have been scrutinized for decades. Some studies demonstrated, through epidemiologic approaches and experimental studies, that statins have anticancer properties. We hypothesized that statins can be repositioned as part of a therapeutic regimen for HGSOC. Here, we demonstrate that certain ovarian cancer cell lines are exquisitely sensitive to treatment with statins. In addition, we show that the sensitivity to statins is an on-target effect by rescuing with downstream members of the mevalonate pathway. Our ongoing studies reveal that HGSOC cells that are sensitive to stain treatment manifest an amplification of chromosome 1q21. Within this genomic region are the S100 proteins, previously identified as potential biomarkers in several cancers, as well as the antiapoptotic protein MCL1. Using reverse phase protein arrays (RPPA) we found that MCL1 levels were altered in statin-sensitive cell lines, while there was no such change in non-sensitive lines. After identifying MCL1 as a potential marker of response, Western blot analysis demonstrated a dose-dependent decrease in the protein level of MCL1 with increasing statin concentration. Additionally, a short, proapoptotic splice variant of MCL1 was produced upon statin treatment, but only in the sensitive lines. Mechanistically, we find that phospho-YAP (inactive) is upregulated after statin treatment. Using a syngeneic cell line system, we were able to demonstrate that overexpression of YAP increases sensitivity to statins. Interestingly, a constitutively active form of YAP (nonphosphorylatable) remains insensitive, suggesting the importance of the phosphorylation event itself. This data corroborated previous research that suggested that multiple different statins decrease nuclear YAP and suggest a mechanism for statin sensitivity. Given its role as an oncogene, the apparent ability of statins to deactivate YAP and downregulate MCL1 steers statins in a promising direction for their repositioning for future therapeutic use.

Citation Format: Paul T. Kroeger, Jr., Ronny Drapkin. Evaluating the potential to repurpose statins for ovarian cancer therapy [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A18.
Abstract A25: Withaferin A ameliorates ovarian cancer-induced cachexia through regulation of NF-kB/NLRP3 signaling
Authors: Straughn Alex R. , Ratajczak Mariusz Z. , Kakar Sham S.
Published: September 2020

A majority of ovarian cancer patients exhibit clinical symptoms of cachexia, which accounts for up to 30% of cancer deaths depending on the oncologic setting. Withaferin A is a steroidal lactone with anti-inflammatory properties and is known to inhibit the growth of ovarian cancer and reduce metastasis through targeting cancer cells and cancer stem cells. The purpose of this study was to investigate whether Withaferin A could ameliorate the cachectic phenotype induced by ovarian cancer. We believe that treatment with Withaferin A could rescue body composition and muscle strength changes. The ovarian cancer cell line A2780 was xenografted into severely immunodeficient NSG mice via an intraperitoneal (i.p.) injection. Control mice received an i.p. injection of sterile saline. Following an engraftment period, mice were treated with vehicle or Withaferin A via i.p. injection every 3 days until the end of the study. Body weight and grip strength were assessed on a weekly basis. DEXA scan was performed post mortem, followed by collection/sectioning/staining of tumor samples and various skeletal muscle tissue. Withaferin A treatment significantly reduced the mortality rate associated with ovarian cancer. Treatment with Withaferin A inhibited an accumulation of adipose tissue within the thoracoabdominal region, associated with ovarian cancer progression. Low doses of Withaferin A were protective for forelimb and total limb grip strength, whereas higher doses were initially protective but resulted in a significant reduction. Further, Withaferin A ameliorated the reduction in myofibrillar cross-sectional area induced by ovarian cancer. Additionally, treatment with Withaferin A completely inhibited the slow-to-fast myofiber-type conversion seen in the settings of cancer cachexia. Lastly, using immunohistochemical staining and qPCR, we demonstrate that Withaferin A causes a downregulation of NF-kB-dependent proinflammatory cytokines and inhibits activation of the NLRP3 inflammasome, which are known to mediate cachectic signaling. Collectively, we provide preliminary data indicating that in ovarian cancer-induced cachexia, body composition changes and weakening of the skeletal muscle can be ameliorated through a downregulation of NF-kB-NLRP3 signaling.

Citation Format: Alex R. Straughn, Mariusz Z. Ratajczak, Sham S. Kakar. Withaferin A ameliorates ovarian cancer-induced cachexia through regulation of NF-kB/NLRP3 signaling [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A25.
Abstract A26: Molecular analysis of short- vs. long-term ovarian cancer survivors
Authors: Stur Elaine , Dal Molin Graziela Zibetti , Wu Sherry , Baggerly Keith , Tworoger Shelley , Levine Douglas , Karlan Beth , Moore Kathleen , Lutgendorf Susan , Sood Anil
Published: September 2020

Background: High-grade serous ovarian carcinomas (HGSOC) comprise the majority of ovarian cancer cases and usually present at diagnosis with late stages of disease. Despite having similar histology, patients with HGSOC often experience highly variable outcomes. The underlying determinants for long-term (LT- ≥10 years) versus short-term (ST- <3 years) survival are largely unknown. The present study sought to identify molecular predictors of LT survival for women with HGSOC. To achieve this, we used deep molecular analysis of tumors from LT and ST survival patients of HGSOC.

Methods: For the discovery tumor cohort, RNA sequencing was conducted on twenty-four (24) frozen samples HGSOC tumors (12 from LT and 12 from ST survivors). A validation cohort included 29 samples (15 from LT and 14 from ST survivors). In vitro studies were carried out using OVCAR8 and SKOV3 cell lines.

Results: From RNAseq analysis, a total of 11 genes were differentially expressed between ST and LT survivors (fold change >2, p < 1 × 10-5, FDR <0.01). A secondary screen was used to eliminate genes that were expressed below threshold levels (fold change >2). Three genes were overexpressed in LT survivors; one had sufficient expression for future therapeutic benefit—TMEM62. In the subsequent validation cohort, TMEM62 had the most significant result of the initial genes of interest, with qRT-PCR data demonstrating an average 3.0-fold higher expression of TMEM62 mRNA in LT versus ST survivors. Next, OVCAR8 and SKOV3 ovarian cancer cells were transfected with TMEM62 using a bacterial plasmid; increased expression of TMEM62 led to decreased proliferation (OVCAR8 73%; SKOV3: 33%), invasion (OVCAR8: 73%; SKOV3: 33%), and a slight decrease in migration (about 2%) in both cell lines. Using a protein array, a decrease in chemokines in SKOV3 cells (MIF, Serpin E1, IL6, CXCL7, IL8, Midkine, and CXCL1) was observed, suggesting that increased TMEM62 expression may lead to reduced tumor inflammatory markers.

Conclusion: Our results demonstrate that TMEM62 may be a novel treatment target for HGSOC given its much higher expression in those patients who achieve long-term survival. These findings may have implications for biomarker and intervention strategies to help improve patient outcomes.

Citation Format: Elaine Stur, Graziela Zibetti Dal Molin, Sherry Wu, Keith Baggerly, Shelley Tworoger, Douglas Levine, Beth Karlan, Kathleen Moore, Susan Lutgendorf, Anil Sood. Molecular analysis of short- vs. long-term ovarian cancer survivors [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A26.
Abstract A37: PAX8 drives ovarian cancer angiogenesis through interaction with SOX17
Authors: Chaves-Moreira Daniele , Mitchell Marilyn A. , Rendi Jessica , Nameki Robbin , Brady Donita C. , Sidoli Simone , Garcia Benjamin , Morin Patrice J. , Lawrenson Kate , Drapkin Ronny
Published: September 2020

A major problem in the treatment of ovarian cancer is the heterogeneity among ovarian tumors. DNA and RNA sequencing studies have demonstrated both intertumoral and intratumoral genetic variation. Despite efforts to elucidate common signaling pathways among various ovarian cancer subtypes, few have led to meaningful patient stratification or individualized targeted molecular therapies. A theoretical new approach to the treatment of ovarian cancer would be to target the signaling pathways that are essential to the development of the HGSOC progenitor cells, the secretory epithelium of the fallopian tube (FT). PAX8, a transcription factor that identifies and sustains high-grade serous ovarian carcinomas (HGSOC), is also the main regulator of FT development. We investigated whether blocking the ability of PAX8 to influence its gene targets, either by interrupting PAX8 protein-protein interactions or by inhibiting the products of PAX8-driven signaling, has the potential to eliminate the primary growth stimulus for high-grade serous tumors. Herein, we have identified SOX17 as a bona fide PAX8-interacting partner and elucidated their collaborative impact on HGSOC. We observed that PAX8 and SOX17 are master regulators of HGSOC identity, as both were found binding in super-enhancer regions regulating most of the same set of genes. Ontology analysis after PAX8 or SOX17 loss showed alteration in three main pathways: cell cycle, apoptosis, and angiogenesis. Most remarkably, we discovered that PAX8 and SOX17 regulate angiogenesis in vitro and in vivo by suppressing SERPINE1, which enables VEGFR2 signaling pathway. These results broaden our understanding of the roles of PAX8 and SOX17 in the regulation of angiogenesis and suggest novel therapeutic strategies that could overcome ovarian cancer heterogeneity and resistance.

Citation Format: Daniele Chaves-Moreira, Marilyn A. Mitchell, Jessica Rendi, Robbin Nameki, Donita C. Brady, Simone Sidoli, Benjamin Garcia, Patrice J. Morin, Kate Lawrenson, Ronny Drapkin. PAX8 drives ovarian cancer angiogenesis through interaction with SOX17 [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A37.
Abstract A42: Potassium channel activity unveils ovarian cancer vulnerability: From signaling to precision medicine
Authors: Lopes Daniela Fukushiro , Hegel Alexandra , Senyuk Vitaly , Russo Angela , Liotta Margaret , Potkul Ronald , Beeson Craig , Brurgette Joanna , Gentile Saverio
Published: September 2020

Decades of studies on ion channels have vastly demonstrated the critical functions of these proteins in many physiologic and pathologic conditions. Also, studies on ion channels have provided for an extraordinary pharmacopeia of useful compounds, often with selective actions and minimal side effects. Nevertheless, the function of ion channels in controlling cancer biology is still unknown and underexplored. Our research demonstrates that the Kir6.2/SUR2 potassium ion channel can be a prognostic factor in gynecologic cancers. Here we present data showing that the Kir6.2/SUR2 channel activity control biochemical pathways underlying proliferation and metabolism in gynecologic cancers, including endometrioid and high-grade serous ovarian cancer. Use of activators of the Kir6.2/SUR2 produced an arrest of the cell cycle, a significant mitochondrial dysfunction that is associated with alteration of the oxidative cellular state and DNA damage in cisplatin-resistant ovarian cancer. Also, our in vivo experiments show the antigrowth effects of pharmacologically targeting Kir6.2/SUR2 on ovarian cancer with FDA-approved molecules. This study offers the opportunity to consider the therapeutic benefits of repurposing drugs targeting the Kir6.2/SUR2 channel for a treatment of gynecologic cancers that is efficient and safe.

Citation Format: Daniela Fukushiro Lopes, Alexandra Hegel, Vitaly Senyuk, Angela Russo, Margaret Liotta, Ronald Potkul, Craig Beeson, Joanna Brurgette, Saverio Gentile. Potassium channel activity unveils ovarian cancer vulnerability: From signaling to precision medicine [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A42.
Abstract A47: Preclinical efficacy of CPI-1688, a novel EZH2 inhibitor, in epithelial ovarian cancer with alterations in the SWI/SNF chromatin remodeling complex
Authors: Magno Elizabeth , Karakashev Sergey , Mertz Jennifer , Trojer Patrick , Zhang Rugang
Published: September 2020

Epithelial ovarian cancer is a paradigm for exploring precision medicine in the context of genetic alteration in the SWI/SNF chromatin remodeling complex. For example, the ARID1A subunit of the SWI/SNF complex is mutated in more than 50% of clear cell ovarian carcinomas. In addition, CARM1 is amplified/overexpressed in ~20% of high-grade serous ovarian cancer. CARM1 is an arginine methyltransferase that asymmetrically dimethylates protein substrates on arginine residues. CARM1 regulates the SWI/SNF genome-wide distribution by post-transcriptionally methylating SWI/SNF core subunit BAF155. EZH2 is the catalytic subunit of the polycomb repressive complex that typically suppresses its gene expression through generating lysine 27 trimethylation on histone H3 (H3K27me3). Our previous studies established that inhibition of the methyltransferase activity of EZH2 is synthetically lethal with ARID1A mutation or CARM1 overexpression due to the antagonistic roles played by the SWI/SNF and EZH2/PRC2 complexes. Here we report the preclinical efficacy of CPI-1688, a highly selective second-generation EZH2 methyltransferase inhibitor, in ARID1A mutated clear-cell ovarian cancer cells and CARM1 overexpressed high-grade serous ovarian cancer cells. Notably, the antitumor effects conferred by CPI-1688 correlate with ARID1A mutation or CARM1 expression status. In addition, at the pharmacodynamic levels, the observed efficacy correlates with a reduction in H3K27me3 both in vitro and in vivo. This suggests that the observed antitumor effects are due to on-target inhibition of EZH2 methyltransferase activity. Finally, this observed selectivity correlates with reactivation of EZH2 target tumor suppressor genes. Together, these results indicate that CPI-1688 is a promising new therapeutic for ovarian cancer with ARID1A mutation or CARM1 overexpression.

Citation Format: Elizabeth Magno, Sergey Karakashev, Jennifer Mertz, Patrick Trojer, Rugang Zhang. Preclinical efficacy of CPI-1688, a novel EZH2 inhibitor, in epithelial ovarian cancer with alterations in the SWI/SNF chromatin remodeling complex [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A47.
Abstract A67: Understanding HIF1A-mediated therapy in clear cell ovarian cancer
Authors: Price Colles , Gill Stanley , Ho Zandra V. , Davidson Shawn M. , Merkel Erin , McFarland James M. , Leung Lisa , Tang Andrew , Kost-Alimova Maria , Tsherniak Aviad , Jonas Oliver , Vazquez Francisca , Hahn William C.
Published: September 2020

Previously, we have used an integrated approach to use genome-scale screening in RNAi and CRISPR-Cas9 to identify several putative cancer dependencies. From these putative dependencies, we discovered EGLN1 was a significant and selective dependency in ovarian cancer and EGLN1 dependency was further enriched in clear-cell ovarian cancers. Genetic knockout and small-molecule inhibition of EGLN1 reduced cell proliferation and increased apoptosis in vitro and in vivo. We found the function of EGLN1 as a cancer dependency is through its canonical role as a negative HIF1A regulator and knockout of HIF1A, but not HIF2A, rescues genetic knockout and small-molecule inhibition of EGLN1. Genetic and small-molecule suppression of VHL, downstream of EGLN1 in the degradation of HIF1A, similarly reduces proliferation and increases apoptosis, and is rescued by HIF1A knockout. Further, we found that forced expression of a mutated HIF1A that blocks EGLN1-dependent hydroxylation reduces proliferation and promotes apoptosis. Using RNA-sequencing, we identified several putative downstream targets of HIF1A and confirmed that HIF1A directly binds and drives transcription of negative MYC regulator MXI1. Further, we found that knockout of MXI1 reduced the effects of EGLN1-mediated suppression, indicating HIF1A-driven MYC suppression as one mechanism underlying EGLN1 dependency. Further understanding of HIF1A-mediated suppression of ovarian cancer progression will be critical to potential small-molecule therapy.

Citation Format: Colles Price, Stanley Gill, Zandra V. Ho, Shawn M. Davidson, Erin Merkel, James M. McFarland, Lisa Leung, Andrew Tang, Maria Kost-Alimova, Aviad Tsherniak, Oliver Jonas, Francisca Vazquez, William C. Hahn. Understanding HIF1A-mediated therapy in clear cell ovarian cancer [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A67.
Abstract A70: Studying the signaling pathways of tumorigenic epithelial fallopian tube subpopulations during early tumorigenesis
Authors: Russo Angela , Dean Matthew , Lantvit Dan , Burdette Joanna E.
Published: September 2020

Survival rates for high-grade serous ovarian carcinoma (HGSOC) patients have not substantially improved over the past decades, mainly because the disease is only detected at the metastatic stage. The findings of early premalignant lesions in the fallopian tube, but not in the ovary, suggested that HGSOCs may originate from the fallopian tube epithelium (FTE) and metastasize to the ovary. Studying the mechanisms of HGSOC genesis from the correct cell of origin is critical to the early detection of the disease. Our main goal is to determine how genetic and molecular alterations contribute to the early events of HGSOC genesis from FTE. Deletion of PTEN alone in FTE is sufficient to lead to tumor formation and peritoneal dissemination, suggesting that loss of PTEN may play a pivotal role in the early events of the tumorigenic cascade. Herein we show that loss of PTEN in the FTE upregulates markers of tumor-initiating cells (TIC) such as WNT4, LGR5, ALDH1, c-Kit, CD44, and Dll4. We also discovered that the TIC markers are confined in a specific population of cells with increased diameter that we call TICHigh as compared to a smaller population that we call TICLow. In fact, when these two populations are separated by flow cytometry-based sorting, the TICHigh retains all the TIC markers. In addition, using a limiting dilution assay, we found that the TICHigh subpopulation forms tumor at a faster rate in vivo and colonize more the ovary ex vivo. In addition, TICHigh forms bigger spheroids and are more resistant to chemotherapeutic agents, suggesting that more targeted therapy may be developed for these stem-like, tumorigenic subpopulations. We also show that loss of PAX2 is mediating the TICHigh phenotype and that activation of AKT is not sufficient to induce the TIC phenotype whereas p70S6K activation is required. These findings suggest that therapies to re-express PAX2 may prevent development of HGSOC. Together, these findings provide a novel model to study the mechanism of fallopian tube tumor initiation and invasion to the ovary mediated by loss of PTEN, which may help to define early events of human ovarian carcinogenesis.

Citation Format: Angela Russo, Matthew Dean, Dan Lantvit, Joanna E. Burdette. Studying the signaling pathways of tumorigenic epithelial fallopian tube subpopulations during early tumorigenesis [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A70.
Abstract A73: BCL-XL dependency in a subset of ovarian clear-cell carcinoma cell lines
Authors: Stover Elizabeth H. , Gelbard Maya K. , Matulonis Ursula A. , Meyerson Matthew
Published: September 2020

Clear-cell ovarian carcinoma is a rare histologic subtype of epithelial ovarian cancers. Clear-cell ovarian cancers tend to be less responsive to platinum-based chemotherapy compared to high-grade serous ovarian cancers. In a dataset of large-scale CRISPR knockout and RNAi screens of cancer cell lines (depmap.org), a subset of ovarian clear-cell carcinoma cell lines were sensitive to knockout or knockdown of BCL2L1, encoding antiapoptotic protein BCL-XL. Most clear-cell ovarian cancer cell lines also express BCL2L1 mRNA and protein. We therefore tested a panel of ovarian cancer cell lines for response to BH3 mimetic drugs, which inhibit different members of the BCL-2 family of antiapoptotic proteins. Initial data showed that a subset of ovarian clear-cell carcinoma cell lines was sensitive to single-agent BCL-XL inhibition with A1331852 and dual BCL-2/BCL-XL inhibition with ABT-263. In contrast, these cell lines were relatively insensitive to single-agent BCL-2 inhibition or MCL1 inhibition. We are currently applying BH3 profiling, a functional assay for apoptotic priming, to test the response of ovarian clear-cell carcinoma cell lines to BCL-XL inhibition using a BCL-XL specific peptide, HRK. Collectively, these studies suggest that ovarian clear-cell carcinomas may be sensitive to BCL-XL inhibition, either alone or in combination with chemotherapy.

Citation Format: Elizabeth H. Stover, Maya K. Gelbard, Ursula A. Matulonis, Matthew Meyerson. BCL-XL dependency in a subset of ovarian clear-cell carcinoma cell lines [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr A73.
Abstract B25: SMARCA4/BRG1 and AP-1 co-regulate an epithelial-like signature in small-cell carcinoma of ovary, hypercalcemic type (SCCOHT)
Authors: Orlando Krystal A. , Raab Jesse R. , Douglas Amber K. , Abudu Aierken , Wang Yemin , Negri Gian Luca , Colborne Shane , Morin Gregg B. , Lang Jessica D. , Hendricks William P.D. , Raupach Elizabeth A. , Pirrotte Patrick , Huntsman David G. , Trent Jeffrey M. , Parker Joel S. , Weissman Bernard E.
Published: September 2020

Mutations in SMARCA4 (BRG1), one of the two mutually exclusive ATPases of the SWI/SNF chromatin remodeling complex, occur in >95% of small cell carcinomas of the ovary, hypercalcemic type (SCCOHT), a rare and aggressive form of ovarian cancer. Because of its apparent role as a driver for SCCOHT, we performed integrative multi-omic analyses in a SCCOHT cell line +/- BRG1 re-expression to identify its role in SCCOHT tumorigenesis. After re-expression, BRG1 was recruited to both distal and promoter regions. We also observed increased chromatin accessibility at distal sites enriched for transcription factor binding motifs for AP-1 family members. Of interest, BRG1 re-expression induced an epithelial-like gene and protein expression concomitant with enrichment of AP-1 motifs at the TSS of highly upregulated epithelial genes. To determine the biologic relevance of these changes at AP-1 binding sites, we used a dominant negative AP-1 cell line to demonstrate that the necessity of AP-1 DNA binding activity and BRG1 re-expression for the protein expression of epithelial genes. Our study demonstrates that BRG1 loss may drive SCCOHT tumorigenesis by diminishing an epithelial-like gene and protein signature of its cell of origin driven by altered AP-1 binding.

Citation Format: Krystal A. Orlando, Jesse R. Raab, Amber K. Douglas, Aierken Abudu, Yemin Wang, Gian Luca Negri, Shane Colborne, Gregg B. Morin, Jessica D. Lang, William P.D. Hendricks, Elizabeth A. Raupach, Patrick Pirrotte, David G. Huntsman, Jeffrey M. Trent, Joel S. Parker, Bernard E. Weissman. SMARCA4/BRG1 and AP-1 co-regulate an epithelial-like signature in small-cell carcinoma of ovary, hypercalcemic type (SCCOHT) [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr B25.
Abstract B30: The effect of OCP use on the incidence of precancerous p53 lesions in fallopian tube fimbria
Authors: Greening Kendall , Karnezis Anthony , Cochrane Dawn , Farnell David , Hoang Lien , Hanley Gillian , Huntsman David
Published: September 2020

Background: High-grade serous ovarian carcinoma (HGSOC) accounts for >70% of ovarian cancer-related deaths and is the most common ovarian cancer histotype, most originating from precancerous p53 lesions in the fallopian tube (FT) fimbria. Use of oral contraceptive pills (OCPs) for 5 years or more is associated with a >40% reduction in risk of HGSOC, but the mechanism is unknown. We hypothesize that OCP use reduces the incidence of p53 lesions. Our preliminary data show higher incidence of p53 lesions in post- compared to premenopausal women; therefore we aim to quantify p53 lesions in postmenopausal women who previously did or did not use OCPs. This will provide insight into the protective effects of OCPs against HGSOC.

Preliminary Results: We determined the presence of p53 lesions by immunohistochemistry (IHC) in FT of women up to 40 years old (n=27) and >60 years old (n=24) who underwent salpingectomies for noncancer reasons. p53 lesions were identified in 3/27 cases of the younger cohort (11%) and in 10/24 of the older cohort (42%). Thus, we conclude an increased incidence of p53 lesions in older compared to younger women.

Proposed Design: IHC for p53 will be performed on FT fimbria of women >55 years old who received salpingectomy for noncancer reasons. Based on an assumed reduction in p53 lesions of 35% in women who used OCPs for 5 years or more compared to nonusers (25 vs. 42%), analysis of 190 cases from each group will provide >80% power (p<0.05). Cases will be identified through Population Data BC and blind analysis by p53 IHC will be performed at the Vancouver General Hospital. Data will be flowed back to Pop Data BC to compare to OCP data.

Conclusion: Our preliminary study found that 42% of postmenopausal women had p53 lesions, informing this study design. The study registered through this abstract will be the first to examine the impact of OCPs on the earliest known precursors of HGSOC.

Citation Format: Kendall Greening, Anthony Karnezis, Dawn Cochrane, David Farnell, Lien Hoang, Gillian Hanley, David Huntsman. The effect of OCP use on the incidence of precancerous p53 lesions in fallopian tube fimbria [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr B30.
Abstract B33: Reproductive factors and risk of ovarian cancer: The Singapore Chinese Health Study
Authors: Lei Ming , Adambekov Shalkar , Wang Renwei , Yuan Jianmin , Lopa Samia , Linkov Faina
Published: September 2020

Background: Ovarian cancer (OC) is the deadliest gynecologic cancer, with varying incidence across geographic regions. Asian population is underrepresented in etiologic and epidemiologic investigations in the area of ovarian cancer. We explored association between ovarian cancer incidence and several lifestyle, dietary, reproductive, and hormonal risk factors in the Singapore Chinese cohort study.

Methods: A population of 34,028 Singaporean Chinese women aged 45-74 were enrolled in the Singapore Chinese Health Study between 1993 to 1998, and followed through Dec. 31, 2010. Data on demographics, tobacco use, dietary and reproductive factors were collected. Cases of OC were determined via linkage to Singapore Cancer Registry. Poisson regression model was performed and hazard ratios (HR) and 95% confidence intervals (CI) were estimated for each variable. Established risk factors and variables showing a significant result in univariate analysis were included in the multivariate analysis.

Results: 128 cases of OC were identified. No dietary factors were associated with OC incidence in this population, nor was tobacco/alcohol use. In univariate analysis, significant results included later age of menopause (RR=2.38, 95%CI 1.23-4.30), later age of first birth (RR=0.43, 95%CI 0.17-0.93) and longer period of menstruating (RR=1.48, 95%CI 1.02-2.18), parity (RR=0.37, 95%CI 0.22-0.65), and oral contraceptives use (RR=0.54, 95% CI 0.33-0.84). In multivariate analysis, higher number of children was associated with decreased ovarian cancer risk (1-2, 3-4, 5+ vs nulliparity: RR=0.45, 95%CI 0.26-0.8; RR= 0.35, 95%CI 0.20-0.51 RR= 0.32, 95%CI 0.18-0.60, respectively).

Conclusions: Parity, contraceptive use, and later age of first birth decreased the risk of OC incidence, while later age at menopause and longer menstruating period increased ovarian cancer risk in Singaporean Chinese cohort. This finding is consistent with findings in Western populations. Future studies are needed to determine risk factors associated with ovarian cancer histologic types and identify markers of elevated risk, especially in Asian populations.

Citation Format: Ming Lei, Shalkar Adambekov, Renwei Wang, Jianmin Yuan, Samia Lopa, Faina Linkov. Reproductive factors and risk of ovarian cancer: The Singapore Chinese Health Study [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr B33.