Immunotherapy Of Ovarian Cancer Research Articles

Identification of a Novel Tumor Microenvironment Prognostic Signature for Advanced-Stage Serous Ovarian Cancer.

Simple SummaryThe expression of tumor microenvironment-related genes is known to be correlated with ovarian cancer patients’ prognosis. Immunotherapeutic targets are in part located in this complex cluster of cells and soluble factors. In our study, we constructed a prognostic 11-gene signature for advanced serous ovarian cancer from tumor microenvironment-related genes through lasso regression. The established risk score can quantify the prognosis of ovarian cancer patients more accurately and is able to predict the putative biological response of cancer samples to a programmed death ligand 1 blocking immunotherapy. This might empower the role of immunotherapy in ovarian cancer through its usage in future study protocols.(1) Background: The tumor microenvironment is involved in the growth and proliferation of malignant tumors and in the process of resistance towards systemic and targeted therapies. A correlation between the gene expression profile of the tumor microenvironment and the prognosis of ovarian cancer patients is already known. (2) Methods: Based on data from The Cancer Genome Atlas (379 RNA sequencing samples), we constructed a prognostic 11-gene signature (SNRPA1, CCL19, CXCL11, CDC5L, APCDD1, LPAR2, PI3, PLEKHF1, CCDC80, CPXM1 and CTAG2) for Fédération Internationale de Gynécologie et d’Obstétrique stage III and IV serous ovarian cancer through lasso regression. (3) Results: The established risk score was able to predict the 1-, 3- and 5-year prognoses more accurately than previously known models. (4) Conclusions: We were able to confirm the predictive power of this model when we applied it to cervical and urothelial cancer, supporting its pan-cancer usability. We found that immune checkpoint genes correlate negatively with a higher risk score. Based on this information, we used our risk score to predict the biological response of cancer samples to an anti-programmed death ligand 1 immunotherapy, which could be useful for future clinical studies on immunotherapy in ovarian cancer.

Immunotherapy in ovarian cancer: we are not there yet.

Immunotherapy in ovarian cancer: we are not there yet.

Ovarian Cancer Immunotherapy and Personalized Medicine.

Ovarian cancer response to immunotherapy is limited; however, the evaluation of sensitive/resistant target treatment subpopulations based on stratification by tumor biomarkers may improve the predictiveness of response to immunotherapy. These markers include tumor mutation burden, PD-L1, tumor-infiltrating lymphocytes, homologous recombination deficiency, and neoantigen intratumoral heterogeneity. Future directions in the treatment of ovarian cancer include the utilization of these biomarkers to select ideal candidates. This paper reviews the role of immunotherapy in ovarian cancer as well as novel therapeutics and study designs involving tumor biomarkers that increase the likelihood of success with immunotherapy in ovarian cancer.

Ovarian cancer modulates the immunosuppressive function of CD11b+Gr1+ myeloid cells via glutamine metabolism

ObjectiveImmature CD11b + Gr1+ myeloid cells that acquire immunosuppressive capability, also known as myeloid-derived suppressor cells (MDSCs), are a heterogeneous population of cells that regulate immune responses. Our study's objective was to elucidate the role of ovarian cancer microenvironment in regulating the immunosuppressive function of CD11b+Gr1+ myeloid cells. MethodsAll studies were performed using the intraperitoneal ID8 syngeneic epithelial ovarian cancer mouse model. Myeloid cell depletion and immunotherapy were carried out using anti-Gr1 mAb, gemcitabine treatments, and/or anti-PD1 mAb. The treatment effect was assessed by a survival curve, in situ luciferase-guided imaging, and histopathologic evaluation. Adoptive transfer assays were carried out between congenic CD45.2 and CD45.1 mice. Immune surface and intracellular markers were assessed by flow cytometry. ELISA, western blot, and RT-PCR techniques were employed to assess the protein and RNA expression of various markers. Bone marrow-derived myeloid cells were used for ex-vivo studies. ResultsThe depletion of Gr1+ immunosuppressive myeloid cells alone and in combination with anti-PD1 immunotherapy inhibited ovarian cancer growth. In addition to the adoptive transfer studies, these findings validate the role of immunosuppressive CD11b+Gr1+ myeloid cells in promoting ovarian cancer. Mechanistic investigations showed that ID8 tumor cells and their microenvironments produced recruitment and regulatory factors for immunosuppressive CD11b+Gr1+ myeloid cells. CD11b+Gr1+ myeloid cells primed by ID8 tumors showed increased immunosuppressive marker expression and acquired an energetic metabolic phenotype promoted primarily by increased oxidative phosphorylation fueled by glutamine. Inhibiting the glutamine metabolic pathway reduced the increased oxidative phosphorylation and decreased immunosuppressive markers’ expression and function. Dihydrolipoamide succinyl transferase (DLST), a subunit of α-KGDC in the TCA cycle, was found to be the most significantly elevated gene in tumor-primed myeloid cells. The inhibition of DLST reduced oxidative phosphorylation, immunosuppressive marker expression and function in myeloid cells. ConclusionOur study shows that the ovarian cancer microenvironment can regulate the metabolism and function of immunosuppressive CD11b + Gr1+ myeloid cells and modulate its immune microenvironment. Targeting glutamine metabolism via DLST in immunosuppressive myeloid cells decreased their activity, leading to a reduction in the immunosuppressive tumor microenvironment. Thus, targeting glutamine metabolism has the potential to enhance the success of immunotherapy in ovarian cancer.

Associations between DNA damage and PDL‐1 expression in ovarian cancer, a potential biomarker for clinical response

Despite the majority of ovarian cancer patients achieving a complete clinical response with first-line therapies, nearly 80% will recur within 24 months. There is an urgent need for novel therapeutic strategies to treat ovarian cancer effectively. Renewed interest in immunotherapy in ovarian cancer has shown that tumor-infiltrating lymphocytes are associated with high-grade serous, clear cell, and endometrioid histology. As a result, immune checkpoint inhibitors have emerged as promising treatments for ovarian cancer. Programmed death ligand-1 (PD-L1) is expressed by cancer cells to evade host surveillance. Its interactions with Programmed death-1 (PD-1) negatively regulate CD8+ T-cells to control inflammatory responses and maintain self-tolerance. Blockade of PD-1/PDL-1 restores anti-tumor cell immunity and has been successfully used to treat some cancers, including non-small cell lung cancer and melanoma. The use of PD-1/PDL-1 blockade in ovarian cancer has been limited by our lack of understanding of the mechanisms that control PD-L1 expression. Emerging evidence has suggested that DNA repair defects and genomic instability influence the expression of PDL-1. However, little is known about the relationship between genomic instability and PDL-1 expression in ovarian cancer, despite the high prevalence of DNA repair defects reported. We hypothesized that genomic instability increases the expression of PD-L1; therefore, PD-L1 expression levels will correlate with high levels of DNA damage in the tumor genome. To test this hypothesis, we have measured DNA repair defects and genomic instability in ovarian tumors using Repair Assisted Damage Detection (RADD). RADD measures persistent DNA lesion and strand breaks within tumor cells using a cocktail of DNA repair enzymes. Lesion sites are then tagged with nucleotide analog and fluorescently labeled for detection. DNA repair defects are revealed by the presence of DNA damage within the tumor genome. In parallel, we also measured PDL-1 expression within these tumor cells using immunohistochemistry. Tumors were categorized and stratified based on the basal levels of DNA damage, and PDL-1 expression was correlated to the type and quantity of DNA damage present in each tumor sample. Using this method, we have established that differences in DNA damage and PD-L1 expression exist among ovarian tumor grade, stage, and pathology. Using different enzyme cocktails in our RADD protocol, we were also able to establish that oxidative lesions are primarily retained within the genome of ovarian tumors. Correlations in DNA damage and PD-L1 expression suggest that oxidative lesions influence the expression of PD-L1 in ovarian cancers. This study establishes a potential biomarker for assessing the utility of PD-L1 inhibitors in ovarian tumors.

CD122-SELECTIVE IL-2 COMPLEXES TREAT OVARIAN CARCINOMAS, INDUCE TREG FRAGILITY AND PROMOTE T CELL STEM CELLS

Abstract Regulatory T cells (Treg) engage IL-2 by high affinity CD25, and anti-tumor effector T cells (Teff) use intermediate affinity CD122. We studied IL-2 complexes (IL-2c) that selectively activate CD122+ Teff over CD25+ Tregs. We found IL-2c but not aPD-L1 potently inhibits ID8agg, an aggressive mouse ovarian cancer (OC) model. IL-2c decreased ascites Treg functional markers (CD25, GranzymeB) while upregulating them on Teffs. IL-2c inhibited Treg suppressive function in ascites but not TDLN. Ascites Tregs after IL-2c showed a fragile phenotype (increased PD-1, T-bet, and IFNg with maintained FoxP3) known to contribute to better immunotherapy response. These data suggest IL-2c inhibits Treg in the ascites through inducing fragile Tregs. Neither CD8+ T cells nor gd T cells is required for IL-2c induced Treg fragility. Studies testing other populations are ongoing. CD8+TCF-1+T cell stem cells (TCSC) improve immune checkpoint blockade in many cancer types. In ID8agg tumors, IL-2c induces CD8+TCF-1+ TCSC with tumor inhibitory capacity when transferred into Rag KO mice. tSNE analysis revealed IL-2c-induced TCSC are CXCR5+PD-1− express CCR2 CXCR3 and produce TNFa, while aPD-L1 induced TCSC are CXCR5+ and PD-1+, consistent with previous reports. These data suggest IL-2c induces beneficial CD8+TCSC distinct from aPD-L1. Although aPD-L1 is not efficacious alone in ID8agg, it significantly promoted IL-2c efficacy. Mice treated with IL-2c+αPD-L1 are resistant to tumor re-challenge even 6 weeks after final dose, suggesting durable immune memory. In sum, IL-2c is a novel OC immunotherapy that targets distinct immune pathways simultaneously, including inhibiting Tregs, boosting TCSCs, and inducing immune memory when combined with aPD-L1.

Bispecific T-Cell Engaging Antibodies Against MUC16 Demonstrate Efficacy Against Ovarian Cancer in Monotherapy and in Combination With PD-1 and VEGF Inhibition.

Immunotherapy for ovarian cancer is an area of intense investigation since the majority of women with relapsed disease develop resistance to conventional cytotoxic therapy. The paucity of safe and validated target antigens has limited the development of clinically relevant antibody-based immunotherapeutics for this disease. Although MUC16 expression is almost universal in High Grade Serous Ovarian Cancers, engagement of the shed circulating MUC16 antigen (CA-125) presents a theoretical risk of systemic activation and toxicity. We designed and evaluated a series of bispecific tandem single-chain variable fragments specific to the retained portion of human MUC16 ectodomain (MUC16ecto) and human CD3. These MUC16ecto- BiTEDs retain binding in the presence of soluble MUC16 (CA-125) and show cytotoxicity against a panel of ovarian cancer cells in vitro. MUC16ecto- BiTEDs delay tumor progression in vivo and significantly prolong survival in a xenograft model of ovarian peritoneal carcinomatosis. This effect was significantly enhanced by antiangiogenic (anti-VEGF) therapy and immune checkpoint inhibition (anti-PD1). However, the combination of BiTEDs with anti-VEGF was superior to combination with anti-PD1, based on findings of decreased peritoneal tumor burden and ascites with the former. This study shows the feasibility and efficacy of MUC16ecto- specific BiTEDs and provides a basis for the combination with anti-VEGF therapy for ovarian cancer.

IFNα Augments Clinical Efficacy of Regulatory T-cell Depletion with Denileukin Diftitox in Ovarian Cancer

AbstractPurpose:Immunotherapy treats some cancers, but not ovarian cancer. Regulatory T cells (Tregs) impede anti-ovarian cancer immunity but effective human Treg-directed treatments are lacking. We tested Treg depletion with denileukin diftitox (DD) ± IFNα as ovarian cancer immunotherapy.Patients and Methods:Mice with syngeneic ID8 ovarian cancer challenge were treated with DD, IFNα, or both. The phase 0/I trial tested one dose-escalated DD infusion for functional Treg reduction, safety, and tolerability. The phase II trial added IFNα2a to DD if DD alone failed clinically.Results:DD depleted Tregs, and improved antitumor immunity and survival in mice. IFNα significantly improved antitumor immunity and survival with DD. IFNα did not alter Treg numbers or function but boosted tumor-specific immunity and reduced tumor Treg function with DD by inducing dendritic cell IL6. DD alone was well tolerated, depleted functional blood Tregs and improved immunity in patients with various malignancies in phase 0/I. A patient with ovarian cancer in phase 0/I experienced partial clinical response prompting a phase II ovarian cancer trial, but DD alone failed phase II. Another phase II trial added pegylated IFNα2a to failed DD, producing immunologic and clinical benefit in two of two patients before a DD shortage halt. DD alone was well tolerated. Adding IFNα increased toxicities but was tolerable, and reduced human Treg numbers in blood, and function through dendritic cell–induced IL6 in vitro.Conclusions:Treg depletion is clinically useful but unlikely alone to cure ovarian cancer. Rational treatment agent combinations can salvage clinical failure of Treg depletion alone, even when neither single agent provides meaningful clinical benefit.

Tumour microenvironment-based molecular profiling reveals ideal candidates for high-grade serous ovarian cancer immunotherapy.

ObjectiveDue to limited immunological profiles of high‐grade serous ovarian cancer (HGSOC), we aimed to characterize its molecular features to determine whether a specific subset that can respond to immunotherapy exists.Materials and MethodsA training cohort of 418 HGSOC samples from TCGA was analysed by consensus non‐negative matrix factorization. We correlated the expression patterns with the presence of immune cell infiltrates, immune regulatory molecules and other genomic or epigenetic features. Two independent cohorts containing 482 HGSOCs and in vitro experiments were used for validation.ResultsWe identified immune and non‐immune groups where the former was enriched in signatures that reflect immune cells, infiltration and PD‐1 signalling (all, P < 0.001), and presented with a lower chromosomal aberrations but increased neoantigens, tumour mutation burden, and microsatellite instability (all, P < 0.05); this group was further refined into two microenvironment‐based subtypes characterized by either immunoactivation or carcinoma‐associated fibroblasts (CAFs) and distinct prognosis. CAFs‐immune subtype was enriched for factors that mediate immunosuppression and promote tumour progression, including highly expressed stromal signature, TGF‐β signalling, epithelial‐mesenchymal transition and tumour‐associated M2‐polarized macrophages (all, P < 0.001). Robustness of these immune‐specific subtypes was verified in validation cohorts, and in vitro experiments indicated that activated‐immune subtype may benefit from anti‐PD1 antibody therapy (P < 0.05).ConclusionOur findings revealed two immune subtypes with different responses to immunotherapy and indicated that some HGSOCs may be susceptible to immunotherapies or combination therapies.

Stromal infiltrating mast cells identify immunoevasive subtype high-grade serous ovarian cancer with poor prognosis and inferior immunotherapeutic response

ABSTRACT Tumor infiltrating mast cells (TIMs), with pro- or anti-tumorigenic role in different types of malignancies, have been implicated in resistance to anti-PD1 therapy. Here, we aimed to identify the relevance of TIMs with the prognosis, immune contexture, and immunotherapy in high-grade serous ovarian cancer (HGSOC). Tissue microarrays containing 197 HGSOC patients were assessed by immunohistochemistry (IHC) for detecting the expression of mast cell tryptase and other immune markers. Kaplan-Meier curve, log-rank test, and Cox regression model were applied to perform survival analysis. Single-cell RNA-seq analysis and flow cytometric analysis were selected to characterize TIMs. Furthermore, short-term HGSOC organoids were employed to validate the effect of TIMs on anti-PD1 therapy. Abundance of stromal TIMs (sTIMs) predicted dismal prognosis and linked to immunoevasive subtype of HGSOC, characterized by increased infiltration of pro-tumor cells (Treg cells, M2-polarized macrophages, and neutrophils) and impaired anti-tumor immune functions. Intensive inter-cell interactions between TIMs and other immune cells were identified, suggesting potential cross-talks to foster an immunosuppressive microenvironment. Organoids derived from sTIMs-low patients were associated with increased response to anti-PD-1 treatment other than the presence of high sTIMs infiltration. A nomogram, constructed by combining FIGO stage, sTIMs, and PD-L1, with an area under the curve (AUC) for predicting 5-year overall survival of 0.771 was better than that of FIGO staging system of 0.619. sTIMs/PD-L1-based classifier has potential clinical application in predicting prognosis of patients with HGSOC. sTIMs-high tumors correlate with immunosuppressive tumor microenvironment (TME) and possess potential insensitivity to immunotherapy.

Current and future immunotherapy approaches in ovarian cancer.

Ovarian cancer (OC) is the major cause of gynecologic cancer deaths and relapse is common despite advances in surgery and systemic chemotherapy. Therefore, novel treatments are required to improve long-term outcomes of the disease. Efficacy of immunotherapy was demonstrated in many tumors and it has been since incorporated into clinical practice for them. Although early data form preclinical studies imply that OC has an immunogenic microenvironment, immune checkpoint inhibitors (ICIs) did not produce favorable results in clinical trials to date. This review will highlight data from clinical studies regarding immunotherapy in OC and its combination with other agents as well as immunologic prospects which could strengthen the therapeutic armament against the disease in the future.

Impact of tumor heterogeneity and microenvironment in identifying neoantigens in a patient with ovarian cancer.

Identification of neoepitopes as tumor-specific targets remains challenging, especially for cancers with low mutational burden, such as ovarian cancer. To identify mutated human leukocyte antigen (HLA) ligands as potential targets for immunotherapy in ovarian cancer, we combined mass spectrometry analysis of the major histocompatibility complex (MHC) class I peptidomes of ovarian cancer cells with parallel sequencing of whole exome and RNA in a patient with high-grade serous ovarian cancer. Four of six predicted mutated epitopes capable of binding to HLA-A*02:01 induced peptide-specific T cell responses in blood from healthy donors. In contrast, all six peptides failed to induce autologous peptide-specific response by T cells in peripheral blood or tumor-infiltrating lymphocytes from ascites of the patient. Surprisingly, T cell responses against a low-affinity p53-mutant Y220C epitope were consistently detected in the patient with either unprimed or in vitro peptide-stimulated T cells even though the patient's primary tumor did not bear this mutation. Our results demonstrated that tumor heterogeneity and distinct immune microenvironments within a patient should be taken into consideration for identification of immunogenic neoantigens. T cell responses to a driver gene-derived p53 Y220C mutation in ovarian cancer warrant further study.

Immunotherapy for Ovarian Cancer: Adjuvant, Combination, and Neoadjuvant.

Ovarian cancer is the most lethal gynecologic malignancy. Surgery and chemotherapy are the primary treatments for ovarian cancer; however, patients often succumb to recurrence with chemotherapeutic resistance within several years after the initial treatment. In the past two decades, immunotherapy has rapidly developed, and has revolutionized the treatment of various types of cancer. Despite the fact that immunotherapy response rates among ovarian cancer patients remain modest, treatment with immune checkpoint inhibitors (ICIs), chimeric antigen receptor (CAR)- and TCR-engineered T cells is rapidly developing. Therapeutic efficiency could be improved significantly if immunotherapy is included as an adjuvant therapy, in combination with chemotherapy, radiation therapy, and the use of anti-angiogenesis drugs, and poly ADP ribose polymerase inhibitors (PARPi). Newly developed technologies that identify therapeutic targets, predict treatment efficacy, rapidly screen potential immunotherapy drugs, provide neoadjuvant immunotherapy, and utilize nanomedicine technology provide new opportunities for the treatment of ovarian cancer, and have the potential to prolong patient survival. However, important issues that may hinder the efficacy of such approaches, including hyperprogressive disease (HPD), immunotherapy-resistance, and toxicity of the treatments, including neurotoxicity, must be taken into account and addressed for these therapies to be effective.

Ovarian Cancer Cells Promote Glycolysis Metabolism and TLR8-Mediated Metabolic Control of Human CD4+ T Cells.

An immunosuppressive microenvironment is a major obstacle for successful tumor immunotherapy. Elucidating the regulatory mechanisms of energy metabolism and functionality in CD4+ T cells will provide insights for the development of novel immunotherapies for ovarian cancer (OC). An Agilent microarray was used to detect differences in gene expression between peripheral CD4+ T cells from five OC patients and those from five healthy controls. Functional pathway analysis was performed for differentially expressed genes. Gene expression profiles revealed significant differences in expression levels of 5,175 genes in peripheral CD4+ T cells from five patients with OC. Functional analysis indicated that the most significantly enriched pathways were metabolic pathways. Furthermore, eight glycolysis-related genes all showed significantly increased expression in peripheral CD4+ T cells of OC patients. Moreover, we established a coculture system of human CD4+ T cells with the OC cell line SKOV3, and then treated them with toll-like receptor 8 (TLR8) ligand ssRNA 40. Coculturing with SKOV3 cells could increase the expression of the eight glycolysis-related genes, promote glucose uptake and glycolysis in CD4+ T cells, induce the differentiation of CD4+ CD25+ Foxp3+ T cells, and enhance the suppression of naïve CD4+ T cells. Additionally, activated TLR8 signaling could mediate the reprogramming of glycolysis metabolism and function in CD4+ T cells. Overall, our study indicates that the SKOV3 coculture environment could regulate the glycolysis metabolism and function of CD4+ T cells, and also that TLR8 mediated the metabolic control of glycolysis in CD4+ T cells cocultured with SKOV3 cells. This provides a new direction for immunotherapy investigations in OC.

Generated SecPen_NY-ESO-1_ubiquitin-pulsed dendritic cell cancer vaccine elicits stronger and specific T cell immune responses

Dendritic cell-based cancer vaccines (DC vaccines) have been proved efficient and safe in immunotherapy of various cancers, including melanoma, ovarian and prostate cancer. However, the clinical responses were not always satisfied. Here we proposed a novel strategy to prepare DC vaccines. In the present study, a fusion protein SNU containing a secretin-penetratin (SecPen) peptide, NY-ESO-1 and ubiquitin was designed and expressed. To establish the DC vaccine (DC-SNU), the mouse bone marrow-derived DCs (BMDCs) were isolated, pulsed with SNU and maturated with cytokine cocktail. Then peripheral blood mononuclear cells (PBMCs) from C57BL/6 mice inoculated intraperitoneally with DC-SNU were separated and cocultured with MC38/MC38NY-ESO-1 tumor cells or DC vaccines. The results show that SNU was successfully expressed. This strategy made NY-ESO-1 entering cytoplasm of BMDCs more efficiently and degraded mainly by proteasome. As we expected, mature BMDCs expressed higher CD40, CD80 and CD86 than immature BMDCs. Thus, the PBMCs released more IFN-γ and TNF-α when stimulated with DC-SNU in vitro again. What's more, the PBMCs induced stronger and specific cytotoxicity towards MC38NY-ESO-1 tumor cells. Given the above, it demonstrated that DC-SNU loaded with SecPen and ubiquitin-fused NY-ESO-1 could elicit stronger and specific T cell immune responses. This strategy can be used as a platform for DC vaccine preparation and applied to various cancers treatment.

Enhancing delivery of small molecule and cell-based therapies for ovarian cancer using advanced delivery strategies.

Ovarian cancer is the most lethal gynecological malignancy with a global five-year survival rate of 30-50%. First-line treatment involves cytoreductive surgery and administration of platinum-based small molecules and paclitaxel. These therapies were traditionally administered via intravenous infusion, although intraperitoneal delivery has also been investigated. Initial clinical trials of intraperitoneal administration for ovarian cancer indicated significant improvements in overall survival compared to intravenous delivery, but this result is not consistent across all studies performed. Recently cell-based immunotherapy has been of interest for ovarian cancer. Direct intraperitoneal delivery of cell-based immunotherapies might prompt local immunoregulatory mechanisms to act synergistically with the delivered immunotherapy. Based on this theory, pre-clinical in vivo studies have delivered these cell-based immunotherapies via the intraperitoneal route, with promising results. However, successful intraperitoneal delivery of cell-based immunotherapy and clinical adoption of this technique will depend on overcoming challenges of intraperitoneal delivery and finding the optimal combinations of dose, therapeutic and delivery route. We review the potential advantages and disadvantages of intraperitoneal delivery of cell-based immunotherapy for ovarian cancer and the pre-clinical and clinical work performed so far. Potential advanced delivery strategies, which might improve the efficacy and adoption of intraperitoneal delivery of therapy for ovarian cancer, are also outlined.

Abstract 3309: Cardioprotective paraoxonase 2 enhances the tumoricidal activity of natural killer cells in a paracrine manner

Abstract Background: Immune system can recognize the neoplastic cells and target them for destruction through tumoricidal process. Further, immune cell dysfunction leads to escape of cancer cells from immune recognition, lead to the initiation and progression of tumor formation. Clinical shreds of evidence suggest that the presence of intratumoral immune cells correlates with improved progression free and overall survival among patients with ovarian carcinoma and provide evidence of activation of antitumor mechanisms. Hence identification of drugs that target the cancer cells as well as activate the immune cells is a sound strategy for cancer therapy. Paraoxonase 2 (PON2) is a membrane-associated lactonase with lipid transport, anti-oxidant and phagocytosis/efferocytosis properties. PON2 deficiency aggravates systemic inflammation, and causes the metabolic associated diseases. Recently we have reported that overexpression of PON2 reduces the ovarian cells proliferation and tumor formation by reducing IGF-1 production and its signaling pathway. Further, using microarray based experiment, we have identified several pathways that modulate the immune cells function such as activation of Natural Killer (NK) cells, dendritic cell maturation, B cells activation, and communication between innate and an adaptive cells signaling pathway were regulated by PON2. Hypothesis: PON2 enhances tumoricidal activity in a paracrine fashion. Methods and Results: NK cells were isolated from control C57BL/6 and purity was assessed with flow cytometry using FITC anti mouse CD49B and results revealed that 95 % of NK cells population. NK cells treated with condition medium from ID8hPON2 (human PON2 overexpressed in mouse ovarian cancer cell line) show an increase in tumoricidal activity compared to condition medium from ID8EV. Neither ID8hPON2 nor ID8EV condition medium affects the viability of NK cells. Moreover, there is no difference in CD49B positive staining in the tumor microenvironment between ID8EV and ID8hPON2 cells receiving mice. Condition medium obtained from ID8hPON2 ID8EV group shows increases the interleukin 18 level and neutralizing antibody against IL18 fail to induce the tumoricidal activity indicating mechanistically PON2 enhances the tumoricidal activity via IL18 dependent manner. Furthermore, IL18 levels are increased in tumor micro environment. PON2 increase the IL18 through transactivation of c-Myc gene that are independent of mitochondrial function. Conclusion: Taken together, our study for the first time showed that PON2 enhances the tumerocidal activity via paracrine manner by upregulating IL18 and PON2 could be used as conventional as well as immunotherapy for ovarian cancer. Citation Format: Asokan Devarajan, Dawoud Sulaiman, Feng Su, Ekambaram Ganapathy, Robin Farias-Eisner, Srinivasa Reddy. Cardioprotective paraoxonase 2 enhances the tumoricidal activity of natural killer cells in a paracrine manner [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3309.

Abstract 976: The systematic identification of novel immune checkpoint averting the metastasis of ovarian cancer: The role of IL20/IL24-IL20RA axis

Abstract Ovarian cancer has high mortality among all gynecological malignancies due to the lack of clear symptoms at early stage and the metastasis of cancer cells when diagnosed with advanced stage. The disseminated ovarian cancer cells have to overcome the immune surveillance in abdominal environment, which are rich in many types of immune cells, to survive and proliferate. However, the key genes and mechanisms preventing the metastasis of ovarian cancer cells into abdominal sites keep largely unexplored. Here, through an in vivo, genome-scale CRISPR/Cas9 knockout screening, we identified IL-20/IL24-IL20RA as a key checkpoint preventing the abdominal metastasis of ovarian cancer cells. In ovarian cancer patients, the level of IL20RA is dramatically decreased in metastatic cancer cells when compared with that in primary sites. Overexpression of IL20RA in highly metastatic ovarian cancer cells greatly suppressed the abdominal metastasis of ovarian cancer and the formation of ascites. In contrast, silencing IL20RA could increase the metastasis of ovarian cancer cells. We further examined IL20RA mediated crosstalk between the disseminated cancer cells and the immuno-microenvironment in mouse in vivo ovarian cancer models and in vitro co-culture experiments. Silencing IL20RA in ovarian cancer cells resulted in a dramatic decrease of M1 subtype of macrophages and increase of the M2 subtype of macrophages without affecting the population of T and B lymphocytes in ascites. Further mechanistic studies revealed that the ovarian cancer cells, when invaded into the abdominal sites, greatly induced the mesothelial cells in peritoneum to produce IL20RA ligands, i.e., IL-20 and IL-24, which in turn activated the IL20RA downstream signaling in ovarian cancer cells to trigger the production and maturation of IL-18. IL-18, when secreted into the microenvironment, was able to promote the polarization of macrophages into M1-subtype to clear the cancer cells. Thus, we identified a novel immune checkpoint IL-20/IL24-IL20RA that was always downregulated in metastatic ovarian cancer cells to prevent the formation of an anti-tumor immune microenvironment, which could be utilized to develop new strategies for the immunotherapy of ovarian cancers. Citation Format: Jia Li, Qin Xuan, JIE SHI, Longlong Wang, Yi Shi, Rong Xiang. The systematic identification of novel immune checkpoint averting the metastasis of ovarian cancer: The role of IL20/IL24-IL20RA axis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 976.

Immunotherapies in ovarian cancer

Ovarian cancer is the leading cause of death for gynaecological cancer, and new therapies are urgently awaited. Although the presence of tumour-infiltrating lymphocytes has been confirmed to be associated to a better prognosis, immunotherapy is not yet incorporated to the armamentarium in ovarian cancer. This review briefly summarises the strategies that have been tested or are under study for the three different groups of tumours: immune desert, inflamed and immune-excluded ovarian tumours. Finally, a better knowledge of the biology and immune microenvironment is needed for successfully developing new immunotherapy strategies.

Specific targeting of ovarian tumor-associated macrophages by large, anionic nanoparticles

Immunotherapy is emerging as one of the most effective methods for treating many cancers. However, immunotherapy can still introduce significant off-target toxicity, and methods are sought to enable targeted immunotherapy at tumor sites. Here, we show that relatively large (>100-nm) anionic nanoparticles administered intraperitoneally (i.p.) selectively accumulate in tumor-associated macrophages (TAMs). In a mouse model of metastatic ovarian cancer, fluorescently labeled silica, poly(lactic-co-glycolic acid), and polystyrene nanoparticles administered i.p. were all found to selectively accumulate in TAMs. Quantifying silica particle uptake indicated that >80% of the injected dose was in TAMs. Particles that were smaller than 100 nm or cationic or administered intravenously (i.v.) showed no TAM targeting. Moreover, this phenomenon is likely to occur in humans because when freshly excised human surgical samples were treated with the fluorescent silica nanoparticles no interaction with healthy tissue was seen but selective uptake by TAMs was seen in 13 different patient samples. Ovarian cancer is a deadly disease that afflicts ∼22,000 women per year in the United States, and the presence of immunosuppressive TAMs at tumors is correlated with decreased survival. The ability to selectively target TAMs opens the door to targeted immunotherapy for ovarian cancer.