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Background To explore the role of heparin-binding proteins released from glycocalyx and neutrophils in the mechanism of Sepsis-induced coagulopathy development. The therapeutic mechanism of Heparin in septic coagulation caused by Hparin-binding Protein and glycocalyx was studied by comparing the changes of Hparin-binding Protein and glycocalyx metabolites before and after heparin administration.Methods Patients diagnosed with sepsis from November 2021 to November 2022 were selected and prospectively studied. Basic clinical data were collected and main indicators were detected for enrolled patients. Hparin-binding Protein and glycocalyx metabolites(HA、HS、SDC-1) were determined by ELISA kit. The main observation indexes were the changes of Hparin-binding Protein and glycocalyx metabolites in two groups before and after heparin administration.Results A total of 148 subjects were screened, and 80 met inclusion and exclusion criteria. Of these, 24 were in the SIC( Sepsis-induced coagulopathy) group and 56 were in the non-SIC group. The levels of HBP(Hparin-binding Protein) and glycocalyx metabolites in SIC group were higher than those in non-SIC group. After heparin treatment, the levels of HBP in SIC and non-sic groups were increased, and the levels of glycocalyx metabolites were decreased. Compared with the non-SIC group, patients in the SCI group had more significant increase in HBP level and decrease in glycocalyx metabolites.Conclusion Plasma HBP may affect blood coagulation and endothelial function on glycocalyx. Heparin may indirectly protects the glycocalyx by replacing the HBP that binds to the glycocalyx.

CRM1 inhibitors have demonstrated antitumor effects in ovarian and other cancers; however, rational combinations are largely unexplored. We performed a high-throughput drug library screen to identify drugs that might combine well with selinexor in ovarian cancer. Next, we tested the combination of selinexor with the top hit from the drug screen in vitro and in vivo. Finally, we assessed for mechanisms underlying the identified synergy using reverse phase protein arrays (RPPA). The drug library screen assessing 688 drugs identified olaparib (a PARP inhibitor) as the most synergistic combination with selinexor. Synergy was further demonstrated by MTT assays. In the A2780luc ip1 mouse model, the combination of selinexor and olaparib yielded significantly lower tumor weight and fewer tumor nodules compared with the control group (P < 0.04 and P < 0.03). In the OVCAR5 mouse model, the combination yielded significantly fewer nodules (P = 0.006) and markedly lower tumor weight compared with the control group (P = 0.059). RPPA analysis indicated decreased expression of DNA damage repair proteins and increased expression of tumor suppressor proteins in the combination treatment group. Collectively, our preclinical findings indicate that combination with selinexor to expand the utility and efficacy of PARP inhibitors in ovarian cancer warrants further exploration.

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.

Supplementary data

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.

Revised Supplementary Data Demonstrating All Supplementary Figures 1-6 and Supplementary Table 1

Here, we examined the role of EP-100 [luteinizing hormone-releasing hormone (LHRH) ligand joined to a lytic peptide], improving the efficacy of immune checkpoint blockade. LHRH-R–positive murine ovarian cancer cells (ID8, IG10, IF5, and 2C12) were sensitive to EP-100 and were specifically killed at low micromolar levels through LHRH-R. EP-100 increased PD-L1 levels on murine ovarian cancer cells. In vivo syngeneic mouse models (ID8 and IG10) demonstrated that single-agent EP-100 reduced tumor volume, tumor weight, and ascites volume. The greatest reductions in tumor and ascites volume were observed with the combination of EP-100 with an anti–PD-L1 antibody. Immune profiling analysis showed that the population of CD8+ T cells, natural killer cells, dendritic cells, and macrophages were significantly increased in tumor and ascitic fluid samples treated with anti–PD-L1, EP-100, and the combination. However, monocytic myeloid suppressor cells, B cells, and regulatory T cells were decreased in tumors treated with anti–PD-L1, EP-100, or the combination. In vitro cytokine arrays revealed that EP-100 induced IL1α, IL33, CCL20, VEGF, and Low-density lipoprotein receptor (LDLR) secretion. Of these, we validated increasing IL33 levels following EP-100 treatment in vitro and in vivo; we determined the specific biological role of CD8+ T-cell activation with IL33 gene silencing using siRNA and Cas9-CRISPR approaches. In addition, we found that CD8+ T cells expressed very low level of LHRH-R and were not affected by EP-100. Taken together, EP-100 treatment had a substantial antitumor efficacy, particularly in combination with an anti–PD-L1 antibody. These results warrant further clinical development of this combination.

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.

Purpose: VEGF-targeted therapies have modest efficacy in cancer patients, but acquired resistance is common. The mechanisms underlying such resistance are poorly understood.Experimental Design: To evaluate the potential role of immune cells in the development of resistance to VEGF blockade, we first established a preclinical model of adaptive resistance to anti-VEGF therapy. Additional in vitro and in vivo studies were carried out to characterize the role of macrophages in such resistance.Results: Using murine cancer models of adaptive resistance to anti-VEGF antibody (AVA), we found a previously unrecognized role of macrophages in such resistance. Macrophages were actively recruited to the tumor microenvironment and were responsible for the emergence of AVA resistance. Depletion of macrophages following emergence of resistance halted tumor growth and prolonged survival of tumor-bearing mice. In a macrophage-deficient mouse model, resistance to AVA failed to develop, but could be induced by injection of macrophages. Downregulation of macrophage VEGFR-1 and VEGFR-3 expression accompanied upregulation of alternative angiogenic pathways, facilitating escape from anti-VEGF therapy.Conclusions: These findings provide a new understanding of the mechanisms underlying the modest efficacy of current antiangiogenesis therapies and identify new opportunities for combination approaches for ovarian and other cancers. Clin Cancer Res; 23(22); 7034–46. ©2017 AACR.

S Figure 1 (A) C57BL/6 mice that received intra-peritoneal injections of IG10 cells were randomly assigned to one of two groups (control or B20 treatment). S Figure 2 (A) Bar graph represents the CD11b+/F4/80+ macrophages in tumor tissue by FACS analysis. S Figure 3 Anti-VEGF therapyââ,¬â€œresistant macrophages have heightened viability, migration, and invasion. S Figure 4 (A) Bar graph represents the percentage of viability of B20 treated macrophages. S Figure 5. VEGFR1-positive macrophages increase the effectiveness of anti-VEGF therapy.

Table S1

CRM1 inhibitors have demonstrated antitumor effects in ovarian and other cancers; however, rational combinations are largely unexplored. We performed a high-throughput drug library screen to identify drugs that might combine well with selinexor in ovarian cancer. Next, we tested the combination of selinexor with the top hit from the drug screen in vitro and in vivo. Finally, we assessed for mechanisms underlying the identified synergy using reverse phase protein arrays (RPPA). The drug library screen assessing 688 drugs identified olaparib (a PARP inhibitor) as the most synergistic combination with selinexor. Synergy was further demonstrated by MTT assays. In the A2780luc ip1 mouse model, the combination of selinexor and olaparib yielded significantly lower tumor weight and fewer tumor nodules compared with the control group (P < 0.04 and P < 0.03). In the OVCAR5 mouse model, the combination yielded significantly fewer nodules (P = 0.006) and markedly lower tumor weight compared with the control group (P = 0.059). RPPA analysis indicated decreased expression of DNA damage repair proteins and increased expression of tumor suppressor proteins in the combination treatment group. Collectively, our preclinical findings indicate that combination with selinexor to expand the utility and efficacy of PARP inhibitors in ovarian cancer warrants further exploration.

Background Nivolumab, an anti-programmed cell death protein 1 antibody, is commonly used as an immune checkpoint inhibitor in various cancers. Various adverse events are associated with these therapies, including hepatitis, dermatitis, and myocarditis. Myocarditis is a relatively rare but potentially fatal immune-mediated adverse reaction. Case presentation We report a case of colon cancer in a 56-year-old Chinese patient with lung and liver metastasis who developed fulminant myocarditis by nivolumab and survived with the support of extracorporeal membrane oxygenation. After six cycles (within 3 months) of nivolumab treatment, the patient developed chest tightness and was hospitalized. A diagnosis of fulminant myocarditis associated with immunotherapy was confirmed based on the clinical manifestations and laboratory examinations. He recovered well and was discharged on day 45 after management with extracorporeal membrane oxygenation, intravenous methylprednisolone, and immunoglobulin. Conclusions This case illustrates a severe cardiovascular complication of immunotherapy, strongly suggesting the necessity of close monitoring for outpatient usage of nivolumab. Additionally, our experience provided an efficient management strategy of extracorporeal membrane oxygenation in terms of life-threatening conditions.

The global coronavirus disease 2019 epidemic is still in a pandemic state. Aging population with underlying diseases is prone to become severe, and have a higher mortality. The treatment capacity of the critical care department directly determines the treatment success rate of critical illness. At present, there is still a certain gap between domestic and foreign countries in intensive care unit (ICU), which is not only in the allocation of medical staff, but also in the beds and settings. The current medical model cannot fully meet the needs of development. The experience and lessons of many major public health emergencies suggested that "dual track of peace and war" approach in discipline construction of critical care is the best medical model. Following the concept of "combination of peace and war", strengthening the discipline construction of critical care department in municipal and district designated hospitals, allocating reasonable standard ICU, step-down ICU and combat readiness ICU, establishing rapid response team, and strengthening regular training and scientific management may be the key measures to deal with the epidemic.

Patients with high-grade serous ovarian cancer (HGSC) who have no visible residual disease (R0) after primary surgery have the best clinical outcomes, followed by patients who undergo neoadjuvant chemotherapy (NACT) and have a response enabling interval cytoreductive surgery. Clinically useful biomarkers for predicting these outcomes are still lacking. Extracellular vesicles (EVs) have been recognized as liquid biopsy-based biomarkers for early cancer detection and disease surveillance in other disease settings. In this study, we performed extensive molecular characterization of serum-derived EVs and correlated the findings with therapeutic outcomes in patients with HGSC. Using EV-DNA whole-genome sequencing and EV-RNA sequencing, we identified distinct somatic EV-DNA alterations in cancer-hallmark genes and in ovarian cancer genes, as well as significantly altered oncogenic pathways between the R0 group and NACT groups. We also found significantly altered EV-RNA transcriptomic variations and enriched pathways between the groups. Taken together, our data suggest that the molecular characteristics of EVs could enable prediction of patients with HGSC who could undergo R0 surgery or respond to chemotherapy.

BACKGROUND: Single disciplinary management of patients with vertigo and dizziness is an important challenge for most physicians in China. OBJECTIVE: To assess the impact of a new paradigm of practice (Clinic for Vertigo and Dizziness, CVD) performed by a multidisciplinary team (MDT) on diagnostic spectrum, medical costs, and patient satisfaction. DESIGN: Retrospective before-after study. PARTICIPANTS: Sample of 29,793 patients with vertigo/dizziness as primary complaint. MEASURES: Changes in diagnostic spectrum, medical costs, and patient satisfaction before and after the establishment of a CVD based on a 4-year database in three tertiary hospitals in northwestern China. KEY RESULTS: The most common diagnoses of patients with vertigo and dizziness were Meniere’s disease (25.77%), cervical disease (25.00%), cerebral vascular disease (13.96%), vestibular syndrome (10.57%), and other etiologies (6.34%) before the CVD establishment. In contrast, after the CVD establishment, the most common diagnoses were BPPV (23.92%), vestibular migraine (15.83%), Meniere’s disease (14.22%), CSD/PPPD (11.61%), and cerebral vascular diseases (4.45%). Extended implementation of a structured questionnaire for vertigo/dizziness and vestibular-oriented examinations (nystagmus, positional tests, HINTS) at the CVD resulted in a remarkable decline in the utility of CT/MRI and X-ray examination (p

Background Adaptor proteins such as growth factor receptor-bound protein-2 (Grb2) play important roles in cancer cell signaling. In the present study, we examined the biological effects of liposomal antisense oligodeoxynucleotide that blocks Grb2 expression (L-Grb2) in gynecologic cancer models. Materials and methods Murine orthotopic models of ovarian (OVCAR5 and SKOV3ip1) and uterine (Hec1a) cancer were used to study the biological effects of L-Grb2 on tumor growth. In vitro experiments (cell viability assay, Western blot analysis, siRNA transfection, and reverse phase protein array) were carried out to elucidate the mechanisms and potential predictors of tumor response to L-Grb2. Findings Treatment with L-Grb2 decreased tumor growth and metastasis in orthotopic models of ovarian cancer (OVCAR5, SKOV3ip1) by reducing angiogenesis and increasing apoptosis at a dose of 15 mg/kg with no effect on mouse body weight. Treatment with L-Grb2 and paclitaxel led to the greatest decrease in tumor weight (mean ± SEM, 0.17 g ± 0.10 g) compared with that in control mice (0.99 g ± 0.35 g). We also observed a reduction in tumor burden after treatment with L-Grb2 and the anti-VEGF antibody B-20 (86% decrease in tumor weight compared with that in controls). Ovarian cancer cells with ErbB2 amplification (OVCAR8 and SKOV3ip1) were the most sensitive to Grb2 downregulation. Reverse phase protein array analysis identified significant dysregulation of metabolites (LDHA, GAPDH, and TCA intermediates) in ovarian cancer cells after Grb2 downregulation. Interpretation L-Grb2 has therapeutic efficacy in preclinical models of ovarian and uterine cancer. These findings support further clinical development of L-Grb2.

Background: The two main causes of body inflammation caused by lipopolysaccharide (LPS) -induced acute lung injury (ALI) are chronic inflammation and oxidant / antioxidant imbalances. Hydrogen (H2), a gaseous molecule without known toxicity, could react with hydroxyl radical to remove the reactive oxygen species (ROS). Both in vivo and vitro studies support the protective effect of hydrogen on injuries caused by oxidative stress and inflammations. Methods: In vivo study, mice were randomly divided into three groups: H2 control group, LPS group and LPS+H2 group. The mice were euthanized at the indicated time points and the specimens were collected. The 72h survival rates, cytokines contents, pathological changes, expression of Toll-like receptor 4(TLR4) and oxidative stress indicators were observed. In vitro study, according to different culture conditions, RAW 264.7 mouse macrophages were divided into the following groups: PBS group, LPS group and LPS+H2 group. The cell Viability, intracellular ROS, cytokines and expression of TLR4 and nuclear factor kappa-B (NF-κB) were observed. Results: In vivo, the 72h survival rate of hydrogen inhalation mice was increased to 80%, and H2 significantly lighten LPS-induced lung damages and decreased inflammatory cytokine release. Besides, H2 showed remarked anti-oxidative activity to reduce the MDA and NO contents in lung. In vitro, H2 down-regulated the levels of ROS, NO, TNF-α, IL-6 and IL-1β in LPS-stimulated macrophages. Second, H2 inhibited the expression of TLR4 and the activation of nuclear factor kappa-B (NF-κB). In summary, these findings supported that H2 attenuates LPS-induced inflammatory responses, which may be mediated by TLR4-NF-κB pathway.

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.

BackgroudPatients with sepsis complicated by anemia have a higher risk of mortality. It is clinically important to study the risk factors associated with the prognosis of this disease. The aim of this study was to establish a predictive model of mortality during hospitalization by extracting clinical data from the Medical Information Mart for Intensive Care III (MIMIC-III) database. MethodsThe clinical data of patients with sepsis complicated by anemia in the MIMIC-III database were retrospectively analyzed. Indexes were screened by stepwise logistic regression (LR), and machine learning predictive models such as Decision Tree (DT), Random Forests (RF), and eXtreme Gradient Boosting (XGBoost) were developed and compared, identifying advantages and disadvantages of each model. ResultsA total of 13,547 patients with sepsis complicated by anemia were included in the study, among which 1,827 died during hospitalization and 11,720 were still alive at discharge. The preliminary stepwise regression model selected 20 clinical indexes, including Elixhauser comorbidity index, maximum blood urea nitrogen (BUN), and maximum hemoglobin reduction. The predictive models showed good discriminative ability (area under the receiver operating characteristic curve [AUROC]:LR, 0.777; DT, 0.726; RF, 0.788; XGBoost, 0.815) and goodness of fit (area under the precision-recall curve [AUPRC]: LR, 0.350; DT, 0.290; RF, 0.400; XGBoost, 0.428). The Shapley Additive exPlanation (SHAP) values in the XGBoost model showed that Elixhauser comorbidity index, maximum BUN, maximum hemoglobin reduction, ventilator use within 24 hours of admission, and age were significant features for predicting in-hospital mortality in patients with sepsis complicated by anemia. ConclusionsThe XGBoost model had better discrimination ability and goodness of fit when compared with other models. Machine learning algorithms have significant practical value in the development of an early warning system for patients with sepsis complicated by anemia.

BACKGROUND: Our pilot clinical study of EphA2 inhibitor (dasatinib) plus paclitaxel and carboplatin showed interesting clinical activity in endometrial cancer with manageable toxicity. However, the underlying mechanisms of dasatinib resistance in uterine cancer are unknown. Here, we investigated potential mechanisms underlying resistance to EphA2 inhibitors in uterine cancer and examined the anti-tumor activity of EphA2 inhibitors alone and in combination with a MEK inhibitor. METHODS: We evaluated the antitumor activity of EphA2 inhibitors plus a MEK inhibitor using in vitro and in vivo orthotopic models of uterine cancer. RESULTS: EphA2 inhibitor induced MAPK in dasatinib-resistant uterine cancer cells (HEC-1A and Ishikawa) and BRAF/CRAF heterodimerization in HEC-1A cells. EphA2 inhibitor and trametinib significantly increased apoptosis in cancer cells resistant to EphA2 inhibitors compared with controls (p

In the field of cognitive radio, communication emitter recognition plays an important role. The traditional methods need to determine expert feature in advance, which leads to inefficiency in some case. One is that the emitters may not be observed in the training and we need to classify much larger new classes examples without knowledge of the label. Therefore, this paper proposes a semi-supervised method for extracting RF fingerprint based on supervised Convolutional Neural Networks and unsupervised clustering. Experimental results demonstrate that the features extracted from CNN can distinguish emitters and the proposed method has a great performance in classifying unknown types of emitters.

Objective: To study the incidence and risk factors of postoperative clinical delirium in critically ill elderly patients in order to devise strategies for prevention and treatment.Methods: We collected clinical data from 451 critically ill patients in the intensive care unit from four hospitals in Shanghai Pudong New Area. Postoperative delirium was measured using the Confusion Assessment Method for the ICU (CAM-ICU), and then the incidence of postoperative delirium was calculated. We used univariate analysis and multivariate logistic regression analysis to determine the risk factors associated with postoperative delirium in critically ill elderly patients.Results: Among the 451 critically ill patients, 56 had delirium after operation. The incidence of postoperative delirium was 12.4%. Univariate analysis showed that the incidence of postoperative delirium was significantly higher (P Conclusion: Adavanced age, COPD history, emergency surgery, postoperative hypertension, use of methylprednisolone, and no postoperative analgesia are risk factors for postoperative delirium in critically ill elderly patients. Medical staff should be educated in evaluating delirium in postoperative patients and conducting risk assessment in order to prevent and treat the disease earlier, and reduce the incidence of delirium.

Objective: The main purpose of this paper is to investigate sleep quality in the withdrawal of medical members dispatched to control the Corona Virus Disease 2019(COVID-19) outbreak in Wuhan, Hubei province, China. Methods: Forty-seven medical members (including twenty medical members treating mild COVID-19, seventeen medical members treating severe COVID-19 and ten logistics team members) completed questionnaire using Pittsburgh Sleep Quality Index. Pittsburgh Sleep Quality Index (PSQI) was used to evaluate the sleep quality of the medical members. Results: A total of forty-seven medical members participated in the sleep quality survey. The PSQI total scores are 5.6±4.3, 11.0±5.0 and 3.4±2.0 in treating mild COVID-19, treating severe COVID-19 and logistics team members, respectively. Medical members treating patients with severe COVID-19 had significantly higher PSQI total scores than those who facing up to the patients with mild COVID-19 and logistics team members. (PConclusions: Findings indicate that medical members treating patients with severe COVID-19 had worse sleep quality than who facing up to the patients with mild COVID-19 and logistics team members.

Here, we examined the role of EP-100 [luteinizing hormone-releasing hormone (LHRH) ligand joined to a lytic peptide], improving the efficacy of immune checkpoint blockade. LHRH-R–positive murine ovarian cancer cells (ID8, IG10, IF5, and 2C12) were sensitive to EP-100 and were specifically killed at low micromolar levels through LHRH-R. EP-100 increased PD-L1 levels on murine ovarian cancer cells. In vivo syngeneic mouse models (ID8 and IG10) demonstrated that single-agent EP-100 reduced tumor volume, tumor weight, and ascites volume. The greatest reductions in tumor and ascites volume were observed with the combination of EP-100 with an anti–PD-L1 antibody. Immune profiling analysis showed that the population of CD8+ T cells, natural killer cells, dendritic cells, and macrophages were significantly increased in tumor and ascitic fluid samples treated with anti–PD-L1, EP-100, and the combination. However, monocytic myeloid suppressor cells, B cells, and regulatory T cells were decreased in tumors treated with anti–PD-L1, EP-100, or the combination. In vitro cytokine arrays revealed that EP-100 induced IL1α, IL33, CCL20, VEGF, and Low-density lipoprotein receptor (LDLR) secretion. Of these, we validated increasing IL33 levels following EP-100 treatment in vitro and in vivo; we determined the specific biological role of CD8+ T-cell activation with IL33 gene silencing using siRNA and Cas9-CRISPR approaches. In addition, we found that CD8+ T cells expressed very low level of LHRH-R and were not affected by EP-100. Taken together, EP-100 treatment had a substantial antitumor efficacy, particularly in combination with an anti–PD-L1 antibody. These results warrant further clinical development of this combination.

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.