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Systematic approaches for accurate repurposing of targeted therapies are needed. We developed and aimed to biologically validate our therapy predicting tool (TPT) for the repurposing of targeted therapies for specific tumor types by testing the role of Bromodomain and Extra-Terminal motif inhibitors (BETi) in inhibiting BRD4 function and downregulating Notch3 signaling in ovarian cancer.Utilizing established ovarian cancer preclinical models, we carried out in vitro and in vivo studies with clinically relevant BETis to determine their therapeutic effect and impact on Notch3 signaling.Treatment with BETis or siRNA-mediated BRD4 knockdown resulted in decreased cell viability, reduced cell proliferation, and increased cell apoptosis in vitro. In vivo studies with orthotopic mouse models demonstrated that treatment with BETi decreased tumor growth. In addition, knockdown of BRD4 with doxycycline-inducible shRNA increased survival up to 50% (P < 0.001). Treatment with either BETis or BRD4 siRNA decreased Notch3 expression both in vitro and in vivo. BRD4 inhibition also decreased the expression of NOTCH3 targets, including HES1. Chromatin immunoprecipitation revealed that BRD4 was present at the NOTCH3 promoter.Our findings provide biological validation for the TPT by demonstrating that BETis can be an effective therapeutic agent for ovarian cancer by downregulating Notch3 expression.The TPT could rapidly identify candidate drugs for ovarian or other cancers along with novel companion biomarkers.

Raw RPPA data for OVCAR 5.

EP-100 is a synthetic lytic peptide that specifically targets the gonadotropin-releasing hormone receptor on cancer cells. To extend the utility of EP-100, we aimed to identify effective combination therapies with EP-100 for ovarian cancer and explore potential mechanisms of this combination. A series of in vitro (MTT assay, immunoblot analysis, reverse-phase protein array, comet assay, and immunofluorescence staining) and in vivo experiments were carried out to determine the biological effects of EP-100 alone and in combination with standard-of-care drugs. EP-100 decreased the viability of ovarian cancer cells and reduced tumor growth in orthotopic mouse models. Of five standard drugs tested (cisplatin, paclitaxel, doxorubicin, topotecan, and olaparib), we found that the combination of EP-100 and olaparib was synergistic in ovarian cancer cell lines. Further experiments revealed that combined treatment of EP-100 and olaparib significantly increased the number of nuclear foci of phosphorylated histone H2AX. In addition, the extent of DNA damage was significantly increased after treatment with EP-100 and olaparib in comet assay. We performed reverse-phase protein array analyses and identified that the PI3K/AKT pathway was inhibited by EP-100, which we validated with in vitro experiments. In vivo experiment using the HeyA8 mouse model demonstrated that mice treated with EP-100 and olaparib had lower tumor weights (0.06 ± 0.13 g) than those treated with a vehicle (1.19 ± 1.09 g), EP-100 alone (0.62 ± 0.78 g), or olaparib alone (0.50 ± 0.63 g). Our findings indicate that combining EP-100 with olaparib is a promising therapeutic strategy for ovarian cancer.

Supplementary Table S1. The IC50s of EP-100 against ovarian cancer cells after 4-, 24-, and 72-h treatment.

File contains synthesis of CN210, sequences for siRNAs, supplementary tables 1 to 3, supplementary figures 1 through 8 and figure legends.

Raw RPPA data for OVCAR 4.

EP-100 is a synthetic lytic peptide that specifically targets the gonadotropin-releasing hormone receptor on cancer cells. To extend the utility of EP-100, we aimed to identify effective combination therapies with EP-100 for ovarian cancer and explore potential mechanisms of this combination. A series of in vitro (MTT assay, immunoblot analysis, reverse-phase protein array, comet assay, and immunofluorescence staining) and in vivo experiments were carried out to determine the biological effects of EP-100 alone and in combination with standard-of-care drugs. EP-100 decreased the viability of ovarian cancer cells and reduced tumor growth in orthotopic mouse models. Of five standard drugs tested (cisplatin, paclitaxel, doxorubicin, topotecan, and olaparib), we found that the combination of EP-100 and olaparib was synergistic in ovarian cancer cell lines. Further experiments revealed that combined treatment of EP-100 and olaparib significantly increased the number of nuclear foci of phosphorylated histone H2AX. In addition, the extent of DNA damage was significantly increased after treatment with EP-100 and olaparib in comet assay. We performed reverse-phase protein array analyses and identified that the PI3K/AKT pathway was inhibited by EP-100, which we validated with in vitro experiments. In vivo experiment using the HeyA8 mouse model demonstrated that mice treated with EP-100 and olaparib had lower tumor weights (0.06 ± 0.13 g) than those treated with a vehicle (1.19 ± 1.09 g), EP-100 alone (0.62 ± 0.78 g), or olaparib alone (0.50 ± 0.63 g). Our findings indicate that combining EP-100 with olaparib is a promising therapeutic strategy for ovarian cancer.

Systematic approaches for accurate repurposing of targeted therapies are needed. We developed and aimed to biologically validate our therapy predicting tool (TPT) for the repurposing of targeted therapies for specific tumor types by testing the role of Bromodomain and Extra-Terminal motif inhibitors (BETi) in inhibiting BRD4 function and downregulating Notch3 signaling in ovarian cancer. Utilizing established ovarian cancer preclinical models, we carried out in vitro and in vivo studies with clinically relevant BETis to determine their therapeutic effect and impact on Notch3 signaling. Treatment with BETis or siRNA-mediated BRD4 knockdown resulted in decreased cell viability, reduced cell proliferation, and increased cell apoptosis in vitro. In vivo studies with orthotopic mouse models demonstrated that treatment with BETi decreased tumor growth. In addition, knockdown of BRD4 with doxycycline-inducible shRNA increased survival up to 50% (P < 0.001). Treatment with either BETis or BRD4 siRNA decreased Notch3 expression both in vitro and in vivo. BRD4 inhibition also decreased the expression of NOTCH3 targets, including HES1. Chromatin immunoprecipitation revealed that BRD4 was present at the NOTCH3 promoter. Our findings provide biological validation for the TPT by demonstrating that BETis can be an effective therapeutic agent for ovarian cancer by downregulating Notch3 expression. The TPT could rapidly identify candidate drugs for ovarian or other cancers along with novel companion biomarkers.

The standard treatment for high-grade serous ovarian cancer is primary debulking surgery followed by chemotherapy. The extent of metastasis and invasive potential of lesions can influence the outcome of these primary surgeries. Here, we explored the underlying mechanisms that could increase metastatic potential in ovarian cancer. We discovered that FABP4 (fatty acid binding protein) can substantially increase the metastatic potential of cancer cells. We also found that miR-409-3p regulates FABP4 in ovarian cancer cells and that hypoxia decreases miR-409-3p levels. Treatment with DOPC nanoliposomes containing either miR-409-3p mimic or FABP4 siRNA inhibited tumor progression in mouse models. With RPPA and metabolite arrays, we found that FABP4 regulates pathways associated with metastasis and affects metabolic pathways in ovarian cancer cells. Collectively, these findings demonstrate that FABP4 is functionally responsible for aggressive patterns of disease that likely contribute to poor prognosis in ovarian cancer., In ovarian cancer, metastatic phenotype may impact surgical outcomes. Here, the authors show miR-409-3p regulates FABP4 which can increase metastatic potential of ovarian cancer, and treatment with DOPC nanoliposomes containing either miR-409--3p mimic or FABP4 siRNA inhibits tumor progression in mice.

Background: Posaconazole can cause pseudohyperaldosteronism via inhibition of 11-beta hydroxylase and 11-beta-hydroxysteroid dehydrogenase type 2 (1). The accumulation of 11-deoxycorticosterone and increased cortisol-to-cortisone ratio in the kidney causes apparent mineralocorticoid excess. The effect of posaconazole on the glucocorticoid axis is less established. Clinical Case: A 56-year-old Hispanic man with a history of chronic septic arthritis of the left ankle from Coccidioides presented with 3 months of malaise, nausea, weight loss of 30 pounds, and recurrent hypokalemia. He was recently switched from long-term fluconazole therapy to posaconazole around the time his symptoms began. His initial labs at our hospital were notable for low potassium (2.9 mmol/L, nl 3.5–5.5 mmol/L) and a random cortisol of 5.8 mcg/dL (nl ≥2.0 mcg/dL). A Cosyntropin stimulation test revealed elevated ACTH (168 pg/mL, nl 7.2–63.3 pg/mL) with minimal rise of cortisol from 4.6 to 7.2 mcg/dL at 1 hour after Cosyntropin administration (nl ≥18 mcg/dL at 1-hour post-Cosyntropin). His plasma renin activity was below detection ( Conclusion: Pseudohyperaldosteronism with glucocorticoid deficiency requiring hydrocortisone treatment has thus far not been reported with posaconazole. Our case shows that posaconazole may lead to true primary glucocorticoid deficiency that can persist after discontinuation of posaconazole and reversal of pseudohyperaldosteronism. Reference: (1) Sanchez-Niño MD, Ortiz A. Unravelling drug-induced hypertension: Molecular mechanisms of aldosterone-independent mineralocorticoid receptor activation by posaconazole. Clin Kidney J 2018;11(5):688–90.

Anti-vascular endothelial growth factor (VEGF) therapy has demonstrated efficacy in treating human metastatic cancers, but therapeutic resistance is a practical limitation and most tumors eventually become unresponsive. To identify microenvironmental factors underlying the resistance of cancer to antiangiogenesis therapy, we conducted genomic analyses of intraperitoneal ovarian tumors in which adaptive resistance to anti-VEGF therapy (B20 antibody) developed. We found that expression of the microseminoprotein, prostate-associated (MSMP) gene was substantially upregulated in resistant compared with control tumors. MSMP secretion from cancer cells was induced by hypoxia, triggering MAPK signaling in endothelial cells to promote tube formation in vitro. Recruitment of the transcriptional repressor CCCTC-binding factor (CTCF) to the MSMP enhancer region was decreased by histone acetylation under hypoxic conditions in cancer cells. MSMP siRNA, delivered in vivo using the DOPC nanoliposomes, restored tumor sensitivity to anti-VEGF therapy. In ovarian cancer patients treated with bevacizumab, serum MSMP concentration increased significantly only in non-responders. These findings imply that MSMP inhibition combined with the use of antiangiogenesis drugs may be a new strategy to overcome resistance to antiangiogenesis therapy.

// Yasmin A. Lyons 1 , Sunila Pradeep 1 , Sherry Y. Wu 1 , Monika Haemmerle 1 , Jean M. Hansen 1 , Michael J. Wagner 1 , Alejandro Villar-Prados 1 , Archana S. Nagaraja 1 , Robert L. Dood 1 , Rebecca A. Previs 1 , Wei Hu 1 , Yang Zhao 4 , Duncan H. Mak, 7 Zhilan Xiao 5 , Brenda D. Melendez 5 , Gregory A. Lizee 5 , Imelda Mercado-Uribe 6 , Keith A. Baggerly 4 , Patrick Hwu 5 , Jinsong Liu 6 , Willem W. Overwijk 5 , Robert L. Coleman 1 and Anil K. Sood 1,2,3 1 Departments of Gynecologic Oncology and Reproductive Medicine, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 2 Center for RNAi and Non-Coding RNA, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 3 Department of Cancer Biology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 4 Department of Bioinformatics and Computational Biology, Division of Quantitative Sciences, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 5 Department of Melanoma Medical Oncology-Research, Division of Cancer Medicine, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 6 Department of Pathology, Division of Pathology and Laboratory Medicine, Section of Gynecologic Pathology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA 7 Department of Leukemia, The University of Texas MD Anderson Cancer Center, Houston, TX, USA Correspondence to: Anil K. Sood, email: // Keywords : adaptive resistance, anti-VEGF therapy, tumor associated macrophages, tumor microenvironment, CSF1R inhibition Received : April 18, 2017 Accepted : July 20, 2017 Published : August 24, 2017 Abstract Anti-angiogenesis therapy has shown clinical benefit in patients with high-grade serous ovarian cancer (HGSC), but adaptive resistance rapidly emerges. Thus, approaches to overcome such resistance are needed. We developed the setting of adaptive resistance to anti-VEGF therapy, and performed a series of in vivo experiments in both immune competent and nude mouse models. Given the pro-angiogenic properties of tumor-associated macrophages (TAMs) and the dominant role of CSF1R in macrophage function, we added CSF1R inhibitors following emergence of adaptive resistance to anti-VEGF antibody. Mice treated with a CSF1R inhibitor (AC708) after anti-VEGF antibody resistance had little to no measurable tumor burden upon completion of the experiment while those that did not receive a CSF1R inhibitor still had abundant tumor. To mimic clinically used regimens, mice were also treated with anti-VEGF antibody and paclitaxel until resistance emerged, and then a CSF1R inhibitor was added. The addition of a CSF1R inhibitor restored response to anti-angiogenesis therapy, resulting in 83% lower tumor burden compared to treatment with anti-VEGF antibody and paclitaxel alone. Collectively, our data demonstrate that the addition of a CSF1R inhibitor to anti-VEGF therapy and taxane chemotherapy results in robust anti-tumor effects.

SUMMARY Despite wide use of anti-vascular endothelial growth factor (VEGF) therapy for many solid cancers, most individuals become resistant to this therapy, leading to disease progression. Therefore, new biomarkers and strategies for blocking adaptive resistance of cancer to anti-VEGF therapy are needed. As described here, we demonstrate that cancer-derived small extracellular vesicles package increasing quantities of VEGF and other factors in response to anti-VEGF therapy. The packaging process of VEGF into small extracellular vesicles (EVs) is mediated by the tetraspanin CD63. Furthermore, small EV-VEGF (eVEGF) is not accessible to anti-VEGF antibodies and can trigger intracrine VEGF signaling in endothelial cells. eVEGF promotes angiogenesis and enhances tumor growth despite bevacizumab treatment. These data demonstrate a mechanism where VEGF is partitioned into small EVs and promotes tumor angiogenesis and progression. These findings have clinical implications for biomarkers and therapeutic strategies for ovarian cancer., Graphical abstract, In brief Ma et.al report that cancer-cell-derived small EVs contain increasing amounts of VEGF (eVEGF) and contribute to resistance to anti-VEGF therapy (AVT). CD63 is a potential mediator that regulates packaging of VEGF into small EVs. eVEGF can trigger intracrine VEGF signaling in endothelial cells and promote angiogenesis despite AVT.

EP-100 is a synthetic lytic peptide that specifically targets the gonadotropin-releasing hormone receptor on cancer cells. To extend the utility of EP-100, we aimed to identify effective combination therapies with EP-100 for ovarian cancer and explore potential mechanisms of this combination. A series of in vitro (MTT assay, immunoblot analysis, reverse-phase protein array, comet assay, and immunofluorescence staining) and in vivo experiments were carried out to determine the biological effects of EP-100 alone and in combination with standard-of-care drugs. EP-100 decreased the viability of ovarian cancer cells and reduced tumor growth in orthotopic mouse models. Of five standard drugs tested (cisplatin, paclitaxel, doxorubicin, topotecan, and olaparib), we found that the combination of EP-100 and olaparib was synergistic in ovarian cancer cell lines. Further experiments revealed that combined treatment of EP-100 and olaparib significantly increased the number of nuclear foci of phosphorylated histone H2AX. In addition, the extent of DNA damage was significantly increased after treatment with EP-100 and olaparib in comet assay. We performed reverse-phase protein array analyses and identified that the phosphoinositide 3-kinase/AKT pathway was inhibited by EP-100, which we validated with in vitro experiments. In vivo experiment using the HeyA8 mouse model demonstrated that mice treated with EP-100 and olaparib had lower tumor weights (0.06 ± 0.13 g) than those treated with a vehicle (1.19 ± 1.09 g), EP-100 alone (0.62 ± 0.78 g), or olaparib alone (0.50 ± 0.63 g). Our findings indicate that combining EP-100 with olaparib is a promising therapeutic strategy for ovarian cancer.

Cell adhesion and apicobasal polarity together maintain epithelial tissue organization and homeostasis. Loss of adhesion has been described as a prerequisite for the epithelial to mesenchymal transition. However, what role misregulation of apicobasal polarity promotes tumor initiation and/or early progression remains unclear. We find that human low-grade endometrial cancers are associated with disrupted localization of the apical polarity protein Par3 and Ezrin while, the adhesion molecule E-cadherin remains unchanged, accompanied by decreased Notch signaling, and altered Notch receptor localization. Depletion of Par3 or Ezrin, in a cell-based model, results in loss of epithelial architecture, differentiation, increased proliferation, migration and decreased Notch signaling. Re-expression of Par3 in endometrial cancer cell lines with disrupted Par3 protein levels blocks proliferation and reduces migration in a Notch dependent manner. These data uncover a function for apicobasal polarity independent of cell adhesion in regulating Notch-mediated differentiation signals in endometrial epithelial cells.

TP53 is the most commonly mutated gene in cancer. Certain mutations can produce gain-of-function (GOF) phenotypes, resulting in pathogenic effects beyond the loss of p53 function. Recent study suggested that tumor cells could affect the p53 status of stromal cells while the mechanisms remains unclear. Exosomes have emerged as an important mode of extracellular communication and a major player in cancer pathogenesis, by promoting pro-tumor microenvironment. We hypothesized that packaging of mutated p53 protein into exosomes and dissemination via exosomes could impact a variety of stromal cell processes, producing a pro-tumorigenic microenvironment. In this study, we carried out a series of in vitro (immunoblot analysis, Q-PCR, Gene array, Co-Immunoprecipitation and lentiviral transfection) and in vivo (subcutaneous mouse models) experiments to determine the presence of mutant p53 in exosomes and its biological significance in tumor microenvironment. We demonstrated that gain-of-function (GOF) p53 protein is packaged into p53 mutant cancer cell-derived exosomes and transferred to fibroblasts. Further studies showed that this process is facilitated by the enhanced stability and consequent intracellular concentration of GOF p53. The packaging of GOF p53 into exosomes is selectively bound by HSP90 chaperone proteins. Inhibition of HSP90 blocks the packaging of GOF p53, but not wild-type into exosomes. Uptake of GOF p53-containing exosomes by fibroblasts results in conversion to a cancer-associated phenotype. In vivo study have shown that GOF p53 exosomes-induced cancer-associated fibroblasts promote tumor growth in a colon cancer cells (HT29 cells) xenograft mouse model. These findings reveal a crucial role of exosomes in modifying tumor microenvironment and hold great consequence in broadening our understanding of tumor biology. Citation Format: Shaolin Ma, Michael H. McGuire, Elaine Stur, Emine Bayraktar, Alejandro Villar-Prados, Sherry Y. Wu, Akira Yokoi, Santosh K. Dasari, Cristian Rodriguez-Aguayo, Jinsong Liu, Gabriel Lopez-Berestein, Anil K. Sood. Exosomal gain-of-function p53 protein promotes cancer-associated phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-311.

Exosomes are small extracellular vesicles secreted from all living cells; its contents are highly varied, including proteins, small RNAs and recently discovered, genomic DNA (gDNA). The presence of nuclear content in exosomes suggests that different cell compartments can contribute to exosome cargo; however, little is known about this sub population of nuclear derived exosomes (nExo). Here, we analyzed nExo isolated from ovarian cancer cells and characterized them to explore potential clinical relevance such as cancer detection and therapeutic response. Exosomes were isolated from ovarian cancer cell lines, normal human fallopian epithelial tube cells and bio-fluids from mouse and human samples. To determine the purity of exosomes, nanoparticle tracking system, immunoblotting assay and cryo-transmission electron microscopy were used. To analyze the content of exosomes, protein mass spectrometry and imaging flow cytometry analysis was performed. First, we determined that mass spectrometry analyses for ovarian cancer cell exosomes revealed that 12.5% of proteins are derived from the cell nucleus sub-population. Additionally, using a novel imaging flow cytometry technique which enables us to identify single exosome and to quantify them, we determined that 10% of exosomes carried gDNA. Conversely, normal cells secreted only 0.2% of nExo. Then we hypothesized that the prevalence of gDNA is affected by genotoxic drugs. Treating cells with either the PARP inhibitor olaparib or topoisomerase inhibitor topotecan increased the proportion of nExo secreted from ovarian cancer cells. In addition, PARP proteins exist in ovarian cancer cell exosomes and 80.4 % of the exosome co-localized with gDNA. Using in vivo preclinical models, serum from ovarian cancer bearing mice contained a higher number of nExo than non-tumor controls, and this population was increased in response to treatment of genotoxic drugs. Furthermore, plasma and ascites from ovarian cancer patients contain around 0.3 % of nExo, which include nuclear proteins and gDNA. We demonstrate that nExo are predominantly secreted from cancer cells and could serve as an important biomarker. Although the subpopulation is relatively rare, their presence in bio-fluids isolated from cancer patients holds the potential of serving as novel biomarkers for early detection and therapeutic response to genotoxic agents. Citation Format: Akira Yokoi, Alejandro Villar-Prados, Paul Allen Oliphint, Theresa J. O'Halloran, Jason Roszik, Anil K. Sood. A novel approach for liquid biopsy by using nExo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-232.

Advances in cancer research have led to the rapid development of novel therapies designed to target specific molecular pathways altered in tumors. However, reliable high-throughput approaches for matching novel drugs to specific tumor types are needed. The potential of systematically identifying and repurposing currently existing targeted therapies designed for a specific cancer to treat other malignancies could hasten the availability of therapeutic options for patients. To address this problem, we have developed an integrative computational therapy-predicting algorithm. This algorithm incorporates data from The Cancer Genome Atlas (TCGA) as well as molecular and survival correlations to identify early-stage antitumor drugs that can be repurposed for other malignancies by targeting novel molecular pathways. To biologically validate our bioinformatics tool, we chose ovarian cancer as a model, which is currently the most lethal gynecologic malignancy afflicting women in the U.S. Our analysis predicted that bromodomain inhibitors, which inhibit bromodomain-containing proteins such as BRD4 and have proved effective in various types of leukemias, would provide a survival benefit for ovarian cancer patients. Furthermore, our analysis revealed that BRD4 inhibition could target the Notch3 pathway, which is known to play an important role in ovarian cancer pathogenesis and is associated with worse patient survival and chemoresistance. We hypothesize that inhibition of BRD4 is an effective therapeutic target in ovarian cancer by downregulating the Notch3 pathway. Inhibition of BRD4 by either using the small-molecule CPI203 or siRNA transfection resulted in a decrease in both Notch3 transcription and protein levels in a panel of Notch3 overexpressing ovarian cancer cell lines. Given that BRD4 regulates gene transcription, we performed chromatin immunoprecipitation (ChIP) and observed that BRD4 was present in the Notch3 gene promoter, but not in the promoter of other Notch genes, such as NOTCH2. To determine the therapeutic efficacy of BRD4 inhibition, we conducted in vitro and in vivo experiments using both CPI203 and siRNA approaches. BRD4 inhibition decreased cell viability, as demonstrated by both 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 2-D colony formation assays in ovarian cancer cell lines. The biologic mechanism of this is mainly by inhibiting cell proliferation, as determined by EdU incorporation assays. Utilizing the OVCAR5 orthotopic tumor model, we determined that treatment with CPI203 significantly decreased tumor weight in mice (p = 0.0468). Additionally, sustained downregulation of BRD4 in OVCAR 5 tumors using a shRNA doxycycline inducible system significantly increases survival by 57% (p = 0.0008). Our current findings suggest that by inhibiting BRD4 in ovarian cancer, we can directly target Notch3 simultaneously and provide both a positive therapeutic and survival effect in ovarian cancer. Completion of this work will provide biologic validation of our Therapy Forecasting Tool by delineating a systematic and accurate approach in identifying and repurposing currently existing targeted therapies. This allows for the expansion of our approach for other tumor types by targeting novel downstream signaling pathways with the ultimate goal of improving patient survival. Citation Format: Alejandro Villar-Prados, Sherry Y. Wu, Jason Roszik, Margaret I. Engelhardt, Anil K. Sood. Systematic approach for identifying and validating novel therapies and targets for ovarian cancer. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr B60.

The role of Human Papillomavirus (HPV) in colorectal carcinogenesis remains elusive. Based on the high incidence of HPV-associated malignancies among Puerto Rican Hispanics, this study aimed to assess the prevalence of HPV infection and viral integration in colorectal tissues in order to evaluate its putative role in colorectal cancer (CRC). In this case-control study, the prevalence of HPV infection in CRC (casesn= 45) and normal colon mucosa from cancer-free subjects (controlsn= 36) was assessed by a nested PCR strategy. HPV-16 genotyping was performed in HPV-positive tissues and the physical status of the HPV-16 genome was determined by E2 detection. HPV was detected in 19 of 45 (42.2%) CRC cases (mean age 61.1 ± 10.7 years, 24 males) and in 1 of 36 (2.8%) controls (mean age 60.9 ± 9.6 years, 24 males) with an OR = 25.58 (95% CI 3.21 to 203.49). HPV-16 was detected in 63.2% of the HPV-positive colorectal tumors; genome integration was observed in all HPV-16 positive cases. This is the first report showing the high prevalence of HPV infections in Caribbean Hispanic colorectal tumors. Despite evidence of HPV integration into the host genome, further mechanistic analysis examining HPV oncoprotein expression and the putative role of these oncoproteins in colorectal carcinogenesis is warranted.

Advances in cancer research have led to the rapid development of novel drug therapies designed for optimal personalized patient care. Despite this surge of treatments, there is still a challenge in accurately predicting the most effective targeted therapy for a specific cancer. Additionally, the potential of systematically identifying and re-purposing currently existing therapeutics designed for one specific cancer to treat other malignancies remains understudied. To address this problem, our group has developed an integrative computational therapy-forecasting algorithm. This algorithm incorporates data from The Cancer Genome Atlas (TCGA) as well as molecular and survival correlations to identify candidate anti-tumor drugs that can be re-purposed for other malignancies by targeting novel pathways. Our computational analysis predicted that bromodomain inhibitors, which inhibit bromodomain-containing proteins such as BRD4, would target the Notch3 pathway in high-grade serous ovarian carcinoma (HGSC). Upregulation of Notch3 plays a crucial role in HGSC tumorigenesis and is associated with worse patient survival. Current Notch3 targeted therapies are not effective, thus designing therapeutic strategies to target Notch3 are critical for HGSC. We hypothesize that inhibition of BRD4 is an effective therapeutic target in HGSC by downregulating the Notch3 pathway. Current in vitro results demonstrated that BRD4 inhibition either by using chemical inhibitors or siRNA results in a decrease in HGSC cell viability by both MTT and 2-D colony formation assays. Furthermore, inhibition of BRD4 resulted in a decrease in both Notch3 transcription and protein levels. Given that BRD4 regulates gene transcription, we performed chromatin immunoprecipitation (ChIP) and observed that BRD4 was present in the Notch3 gene promoter. These findings suggest that by inhibiting BRD4 in HGSC, we can target Notch3 and biologically validates our initial in sillico analysis. The successful completion of this project could have major implications for development of personalized therapies not only for HGSC but for other cancers as well. Note: This abstract was not presented at the meeting. Citation Format: Alejandro Villar-Prados, Sherry Y. Wu, Jason Roszik, Anil K. Sood, Margaret I. Engelhardt. Forecasting novel therapies by understanding the role BRD4 in regulating the Notch3 signaling in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1528. doi:10.1158/1538-7445.AM2017-1528