Epithelial Ovarian Cancer: Microenvironment and Immunecheck Point Inhibitors

With poor prognosis for patients with advanced disease, ovarian cancer continues to be the most lethal gynecologic malignancy. Despite optimal treatment, the disease frequently recurs and develops chemoresistance. The tumor immune microenvironment has been predicted to play a special role in tumor development and treatment, by modifying immunosuppressive and cytotoxic responses in close proximity to tumor cells via metabolic reprogramming. A better understanding of the tumor microenvironment’s critical roles allows researchers to develop new therapeutic strategies, such as immunotherapy, to combat epithelial ovarian cancer chemoresistance. Unfortunately, the results of many clinical trials examining immune check-point blockade (ICB) have shown very low levels of efficacy for single-agent immune checkpoint inhibitors, and research on combination treatments has not yet identified any combinations with robust activity in a large number of epithelial ovarian cancer patients. The present research will begin with a review of the main concept of immune response inside the ovarian cancer microenvironment. Then, we’ll try to figure out what role various immune checkpoint inhibitors have in the ovarian cancer microenvironment. Finally, we’ll look into several exciting treatment options, such as immune checkpoint inhibition and the body’s anti-ovarian cancer immunological response.


Introduction
Ovarian cancer is a major contributor of cancer death among women [1].According to Casablanca's national registry from 2013 to 2017, 585 cases of ovarian cancer were registered, accounting for 4% of female malignancies and 2.3% of all recorded tumors in both sexes.It had a crude incidence of 5.4 per 100,000 women and a normalized incidence rate of 5.0 per 100,000 women in the global population [2].High-grade serous ovarian cancer is one of the most typical and aggressive types of ovarian cancer, as well as the major cause of ovarian cancer fatalities [3].Although high-grade serous ovarian cancer responds well to the first treatment including debulking resection & platinum-based chemotherapy, recurrence is widespread and severe in the majority of patients [4].First-line maintenance therapy innovations have sought to boost overall and recurrence-free survival in ovarian cancer patients.Bevacizumab or PARP inhibitor maintenance therapy has been demonstrated to increase progression-free survival (PFS) but not overall survival (OS), implying that more efficient maintenance therapy is required [5].
Immunotherapy, in particular, has aroused researchers' interest due to an improved understanding of the molecular underpinnings of immune identification and neutralization of cancer cells.Immune checkpoint inhibition, cancer vaccinations, and adoptive cell therapy are examples of such therapies.In fact, the Food and Drug Administration (FDA) has authorized numerous immune checkpoint inhibitors for a variety of cancers.Regrettably, no immunological treatments for ovarian cancer are currently available [6].This review will describe the immunological response inside the ovarian cancer microenvironment and to explore the role of several immune checkpoint inhibitor compounds used by ovarian cancer cells to avoid the immunological response.It will also report on certain

Immune Response to Ovarian Cancer
Submit Immunoediting of cancer is T cell identification of tumor antigens leading to immunological elimination or shaping of developing cancer.
It's a process that includes three states: elimination, equilibrium and escape [7]: Elimination involves the immunological surveillance system that detects, inhibits carcinogenesis and maintains cellular homeostasis by innate and adaptive immune responses.
Equilibrium arises when the body's defense system notices the tumor but mutations permit the tumor to grow.
Escape acquires the ability to avoide the immune system and proliferate unhindered, by different mechanisms including: reduced immune recognition or development of an immunosuppressive tumor microenvironment [8].
There's research supports for the involvement of immunosurveillance in the prevention of ovarian cancer.The identification of intraepithélial infiltrating lymphocyte CD 3+ (TILS), is associated with increased survival [9].The activity levels of immune effector cells such as CD3+, Natural-killer (NK) and Vγ 9Vδ2T has an improved prognosis [10].
T helper (Th)-17 cells, a type of T cells, were found in proportionally higher numbers in the epithelial ovarian cancer microenvironment compared to other immune cells, [11] their role is indeterminate, they can either act against the tumor or promote its development [12].
Different pathways are involved in immune escape in ovarian cancer: -Lowering of surface molecules that influence Ag presentation, like 2-microglobulin and the major histocompatibility complex [13].
-MHC class I related chain A expression is reduced, rendering tumor cells invisible by cytotoxic effector lymphocytes [14].
-Tumor amplification of chemicals that suppress the cytotoxic action of immune cells and safeguard tumor cells from lysis: CA125 binds to the inhibitory receptor of NK cells [15].
-Endothelin-B receptor upregulation is linked to TIL depletion in addition to a limited survival duration in ovarian cancer patients [19].
The article will provide an summary concerning the way the defense system operates in the malignant ovarian tumor microenvironment as well as the role of several various immune checkpoint inhibiting molecules used by cancer cells in order to elude immune reaction.It also presents some potential therapeutic approaches involving immune checkpoint blockade, exclusively or in conjunction with other therapies.

Immune Response Regulation by Conventional
Treatments against EOC Conventional epithelial ovarian cancer treatment uses debulking surgery and systemic chemotherapy.Cytotoxic therapies destroy any remaining tumour cells when surgery removes poorly vascularized tissue and reduces the tumour burden [22], [23].Major surgery would lead to immunosuppression due to down-regulation of the T helper (Th)-1 response [24], [25] despite this, the tumor debridement decreases the immuno-suppression induced by the tumor in cases of EOC [18], [22].Treg cells and naive CD4+ T cells proportion are reduced via surgery, while CD8+/Treg T cells and effector T cells are enhanced.Aside from that, surgery boosts IFN-production by peripheral CD8+ T cells while lowering the count of effector T cells in the circulation [22].As a result, tumor immunodeficiency is somewhat changeable, and gained immunity can be improved in the ovarian cancer by tumor debulking surgery [22].
Chemotherapy may have immunosuppressive effects due to the frequent lymphopenia it produces.However, current study suggests that the therapeutic mechanisms attributed to chemotherapy include immunity in a significant way [26], [27].For example, optimum tumor debulking outcome was more common among advanced ovarian cancer patients who were given platinum-based chemotherapy when CD3+ TILs were detected [28].Furthermore, paclitaxel or cisplatin administration in ovarian cancers increases Mannose-6-phosphate receptor expression in mouse cancer cells.It increases the vulnerability of tumor cells to the protease granzyme-B, which is secreted by cytotoxic T cells [29].Additionally, successful chemotherapy in advanced EOC is linked with greater efficiency and higher levels of CD8+ effector T cells [22], [30].Moreover, neoadjuvant chemotherapy minimizes immunodeficience via diminishing quantity of circulating Treg in ovarian tumors [22].Certain anticancer drugs may also cause immunogenic demise in malignant cells, triggering exposure or secretion of immunogenic signals from cancer cells that activate an anticancer immune response.Notably, all forms of chemotherapy except oxaliplatin and 3/25 of the anthracyclines tested do not cause immunogenic cell death.[31], [32].Overall, these results demonstrate that through direct and indirect effects, debulking surgery and chemotherapy may reestablish the equilibrium or elimination process in malignancies that have avoided immune control.
Despite optimal treatment, the disease recurs frequently and develops chemoresistance, causing a high fatality rate.As a consequence,a greater knowledge about the tumor microenvironment's critical researchers will be ready to propose new drugs, such as immunotherapy, to fight the chemoresistance of EOC [33].

Ovarian Cancer Immune Microenvironment
The ovarian cancer microenvironment is exceedingly intricate and varied consisting primarily T cells, NK cells, macrophages, MDSCs, and other immune cells.Such immune cells are classified into two types based on their specific functions: active immune cells which offer antibody surveillance and block tumor growth, like CD8+ T cells & NK cells, and suppressing immune cells that aid in tumor expansion and progression, which include mainly Tregs, M2 macrophages, MDSCs, and Fig. 1 [34].
The enigma of epithelial ovarian cancer immunity is that dual TME may exist in the same patients and tumor location, showing a high dynamic and variance in immune cell infiltration and, as a result, a different developing profile from patient to patient [35].

Dendritic Cells (DCs)
Dendritic cell infiltration is high in ovarian cancer lesions, but antigen presentation effectiveness is low due to dendritic cell acceptance, as evidenced via the decreased levels of costimulatory substance expression on the surface of dendritic cells [36].Dendritic cells dysfunction could be owed to metabolic variables like indoleamine 2, 3dioxygenase (IDO) and TGF-β [37].PD-1 inhibitors may be able to restore DC function, thereby promoting the antitumor effect in ovarian cancer [38].IDO-expressing dendritic cells drop tryptophan levels in the vicinity of Tregs while preserving Treg immunosuppression via mTORC-Akt signaling [39].In the ovarian cancer microenvironment, XBP1 is activated, inducing the accumulation of lipid bodies in tumor-infiltrating DCs & driving them into a tolerant position [40].
Overexpression of IGF1R in OC are linked with Dcs maturation into conventionnal DCs [41].
In order to trigger an antitumor response, DC vaccines may be able to restore the tumor's ability to present antigens.

T Lymphocytes
Ovarian tumor's microenvironment primary component, T lymphocytes, are essential to adaptive immunity.The primary immune cells that are activated are CD8+ cytotoxic T cells (CTLs).Cytolytic and inflammatory cytokines are generated when the T cell receptor interacts with the MHC-I molecule on tumor cells [42].Adequate T-cell infiltration, which are affected by CXCL9 released by antigen-expressing cells (APCs) with CCL5 generated in tumor cells [43], is essential for PD-L1 blocking in OC patients.The primary causes of T-cell abnormalities are mitochondrial T-cell metabolic suppression and a failure to create adequate energy precursors.

Natural Killer Cells (NK)
NK cells have a vital part in cancer early detection and treatment, and they have been related to tumor growth and management [44], [45].They have a significant impact on patient survival [46].When present in ovarian cancer, they exhibit minimized expansion, lower cytolytic activity & lower inflammatory cytokine generation [47], [48].To enhance their infiltration, function, and response potential in the tumor microenvironment, it is necessary to have a greater grasp of the techniques used to count, evaluate, and control NK cells.The products of ovarian tumors, many kinds of immunosuppressive cytokines released from Treg and myeloid derived suppressor cells (MDSC) affect the phenotypic & functional properties of NK cells [49].
Due to their variety of roles, NK cells are a prime choice for more individualized and efficient immunotherapy of HGSC.However, combination medicines or techniques are required to concurrently target various, co-existing aspects of the tumor.Researching NK cell interactions will aid in predicting the best outcomes [51].

T Regulatory Cells
A kind of CD4+ T cells is the T reg cell who encodes transcription factors including FoxP3, CTLA-4, & CD25 [52].They cause immunosuppressive effects in vivo and maintain immunological self-tolerance.The percent of Treg with CD8+ T cells is inversely associated to survival in cancer [53].IL-10 production is increased by accumulated Tregs, which also encourage tumor angiogenesis and immunological tolerance [54].Tregs formed by ovarian cancer include multiple TCR-involved receptors, involving PD-1, ICOS, & 4-1BB, which boost their sensitivity to anti-CD3/anti-CD28 stimulus as well as their potential to operate as powerful inhibitors [55].
Tregs may evolve to CD4+ effector T cells after activation of the GITR [56].In ovarian cancer malignant ascites, high levels of proinflammatory cytokines encouraged increased TNFR2 expression, which improved the expression of immunosuppressive molecules [57].TME can be reduced by focusing on the IL6/JAK/STAT3 signaling pathway [58].

Tumor Associated Macrophage (TAM)
M1 and M2 are the two primary phenotypes of macrophages.M2 macrophages, primarily support tumor growth by secreting inhibitory cytokines [59], [60], whereas M1 macrophages produce proinflammatory cytokines and chemokines, participate in immunological surveillance, that help the body's immune system destroy cancer.By controlling macrophage polarization, several proteins may impact the biochemical activities of OC cells [61].
Ovarian cancer cells can polarize M2 macrophages, leading to stimulate tumor development, by secreting miR-NAs and cytokines.
MDSCs perform a key part in immune suppression & neovascularization,in the TME.L-selectin levels on T cells can be reduced by ADAM17, which also limits lymph node T cell recruitment [64].
Oxidative stress correlates with ROS and RNS impacts on the receptors of T cells including the reactivity to antigen-MHC complexes [68].Hypoxia in malignancies stimulates MDSCs that generate VEGF, FGF2, and MMP9 [69].By generating MMPs, these elements promote invasion and metastasis.

Cancer-Associated Fibroblasts CAF
CAFs are a prominent ovarian cancer cell subpopulation it plays an important part in the disease's initiate [70].An extracellular coating of dense extracellular matrix (ECM) rich in fibronectin protects tumor cells from immune attack, however this line of defense can be compromised via ECM modification and excessive fibroblast deposition [71].CAFs recruit ascites for generating heterogeneous spheroids identified Mus and secrete EGF in order to retain MUs.CAFs, in cooperation with OC cells, can alter the ECM through emitting multiple kinds of cytokines and creating multiple paracrine signals [34].
CAFs are a type of growth factor derived from HGF and FGF-1.
HGF promotes OC tumor development and drug resistance [72], FGF-1, on the other hand, manages neoplasm growth through FGF-4, elevating Snail1 & MMP3 expression, whit engaging the pathway of MAPK/ERK [73].Cancer-associated fibroblasts also aid in ovarian neoplasm dissemination via releasing VEGF-A and tenascin-c [74], as well as recruiting immune cells and interacting with a variety of immune components.Interleukin (IL)-1 is an important immunosuppressor in the TME and has been linked to CAF-expressed PS1 [34].PS1 inhibition boosts CTL and DC proliferation and migration [75].
The pathogenesis of OC is significantly influenced by CAFs, and numerous therapeutic strategies have been suggested [76].

Immune Checkpoint in Ovarian Cancer
After obtaining adequate treatment with cytoreduction surgery and chemotherapy, a substantial proportion of women with ovarian cancer acquire secondary resistance.
Therefore, because of an imbalance between immunological tolerance and immune response, the majority of ovarian tumors manage to avoid the host immune system [78], and it's mainly due to the immunosuppressive microenvironment (TME) [79].
Blocking immunological checkpoints is a very promising way to boost antitumor immunity among patients with ovarian neoplasm, which is a great interest to the field of cancer research.Immune checkpoints are essential for preserving tolerance to tumors as well as defending tissues after a pathogen reply.Nevertheless, they significantly decreased the intensity, caliber, and timing of immune responses in the TME.A range of molecules, notably PD-1, CTLA-4, lymphocyte activation gene-3 (LAG-3), T-cell immunoglobulin, and mucin protein-3, are hypothesized to act as immune checkpoints [81].
Most research was centered on the CTLA-4 and PD-1 receptors, and various therapeutic trials have looked into monoclonal antibodies that target either PD-1/PD-L1 or CTLA-4 (binding to CD80 and CD86).

PD-1/PDL-1
A cell surface protein called PD-1 engages to the PD-L1 protein made by malignant cells and triggers the depletion of peripheral effector T cells and the change of effector T cells (Teff) into regulatory T cells [82].The final phase of immune response to cancer process is focused on anti-PD-1/PD-L1 agents functions [83].Immune checkpoint inhibitors, particularly PD-1/PD-L1 inhibitors, have changed the course of treatment in some malignancies.Latest research has demonstrated that PD-L1 levels, tumor mutational burden, microsatellite instability, and/or mismatch repair deficit are all potent indicators for anti-PD1/PD-L1 therapy.Although a multiple types of immune checkpoint inhibitors (ICPIs) are currently approved by FDA, research into ICPIs in gynecologic cancers lags behind that of other disease sites [84].Numerous trials in epithelial ovarian cancer (EOC), initially in recurring tumors, have been described.These trials contained assessments of PD-L1 expression as well as alternative measures of tumor immune phenotypes [85].For exemple, KEYNOTE-028 and KEYNOTE-100 studies demonstrated that PD-1 inhibitors monotherapy had a low ORR in ovarian cancer patients [86].Taken together, these research findings indicate that immunecheckpoint blockade possesses the ability to trigger long-lasting reactions in a tiny percentage of ovarian cancer patients.Without further delineation of accurate biomarkers to define the relevant subpopulations, clinical circumstances, and/or combinations that allow enhanced action, immune checkpoint inhibitors should not be utilized as monotherapy in ovarian neoplasm.Considering the disappointing outcome of anti-PDL1 monotherapy, certain trials have shown encouraging results when coupled with other therapies such as chemotherapy, targeted therapy (anti-VEGF), PARP inhibitors, or anti-CTLA4 antibodies [87].

Cytotoxic T-Lymphocyte Antigen-4 (CTLA-4)
CTLA-4 is a T-cell inhibitory molecule who has a significant impact in carcinogenesis and progression [88].CTLA-4 reduces the antitumor immune response mediated by T lymphocytes by inhibiting cytokine receptor signals, protein translation, triggering ubiquitin ligases, & mobilizing phosphatases [89].CTLA-4 strongly interacts to elements of the B7 family in extrinsic cell pathways and conveys inhibitory signals, lowering T cell activation [90].It also sends a reverse signal via B7 and promotes IDO production, resulting in tryptophan breakdown and T cell growth suppression [91].Anti-CTLA-4 antibodiy depress the body's defenses through restricting CTLA-4's ability to connect to B7 family elements, hence decreasing antigenspecific T cell recognition.A trial conducted in people with advanced ovarian cancer in which one patient maintained a steady CA-125 level with antibody injection, exhibited a 43% decrease in CA-125 levels during the first two months [92].Clinical research is in underway to investigate the anticancer efficacy of ipilimumab among patients with recurrent platinum-sensitive ovarian carcinoma [93].

Lymphocyte Activation Gene-3 (LAG-3)
The most promising immunological checkpoint is LAG3, along with PD-1 and CTLA-4.By dampening the immunological microenvironment, high levels of LAG3 and FGL1 expression encourage the growth of tumor [94].Variable amounts of LAG-3-positive tumorassociated lymphocytes, cytotoxic (CD8+) and regulatory (FOXP3+) T-cell infiltrates, and tumoral and immunological PD-L1 levels were found in OC.According to tumor cell expression with PD-L1 CPS ≥ 1, LAG-3-positive tumor-associated lymphocytes are linked with PD-L1 expression [95].Immunotherapies suppressing LAG-3 could profit certain patients with ovarian neoplasm, especially if combined to anti-PD-1/PD-L1 therapies [95].Nonetheless, generally modest amounts of LAG-3positive lymphocyte infiltration and PD-L1 levels indicate the potential for meaningful responses in this tumor type may be restricted.

T Cell Immunoglobulin and Mucin-Domain Containing-3 (TIM-3)
TIM3 adversely affects CD4 & CD8 T cells, causing their depletion and the formation of immunological cells that inhibit the immune system [96].As shown in other investigations on a variety of cancers it may function as a significant biomarker and possible therapeutic target [97].TIM-3 is the most prevalent co-inhibitory molecule in ovarian cancer, it inhibits immunity, diminishes the effectiveness of chemotherapy through preventing the transfer of nucleic acids to ovarian carcinoma cells' intracellular cysts [98].TIM-3 inhibition effectively reverses Treg-mediated reduction of CD8+T cell inflammation [99].Early clinical research proved that TIM-3-targeted treatment, singly as well as in conjunction with ICB therapies, might be an appropriate therapy for ovarian cancer patients [98].

T Cell Immunoglobulin and ITIM Domain (TIGIT)
TIGIT, a co-inhibitory receptor located upon the surface of lymphoid cells that primarily modulates immunological responses via Tregs [100].In ovarian cancer, DNAM-1 activity decreased among natural killers cells.Furthermore, DNAM-1 serves as an essential NK cell stimulating receptor in ovarian neoplasm, lower DNAM-1 level in patients with OC corresponds with higher CD155 level in ovarian cancer cells.According to the literature, patients with OC with poorer DNAM-1 activity on NK cells from peritoneal experience an inferior survival rate versus those who have greater DNAM-1 activity.Cytotoxicity of NK cells patients with OC coincides with increased CD155 activity [101].TIGIT, along with PD-1, are thought to be a hallmark of CD8+ T cell depletion [102].By a synergistic action, their dual blockage improves the effector role of CD8+ T cells.
According to ClinicalTrials [103], there will be just four recruiting clinical trials in 2022 using anti-TIGIT or anti-CD112R (COM701) mAbs in ovarian cancer therapy, primarily in addition to anti-PD-1 therapies.These are all in the initial phases of execution (phase 1 or 2).Nevertheless, no results have yet been published.

Exosome's as Immunotherapy Strategy for OC
The complex miRNAs, proteins, lipids, sugars, and nucleic acids found in small vesicles released via several kinds of cells in the extracellular microenvironment [104].They are extensively dispersed throughout the tumor microenvironment [105], and their primary functions are deletion and cancer promotion.From one side, exosomes may boost the innate and adaptive, through turning on Dcs cells, NK cells and T cells to, permitting these immune cells to have an anti-tumor impact.On the other hand, exosomes from ovarian neoplasm interact with immunosuppressive effectors, causing immune escape, one of the characteristics of cancer [106].
Exosomes are crucial for tumor initiation and development in ovarian cancer [107] as shown in the adjacent Fig. 2 by inducing T-cell arrest, allowing cancer cells to escape the immune system [108], [109].
According to numerous studies, exosomes offer a lot of potential for identifying and treating early-stage malignant cancers.
Exosomes might be employed as a possible diagnostic biomarker and disease target to eradicated cancer-derived exosomes and as a result reduce immunosuppression and increase the benefit of immunotherapy in ovarian neoplasm [110].

Conclusion
In this review, we had the opportunity to extract additional evidence suggesting how the body's defense system plays a role in ovarian neoplasm physiopathology and explain an overview of immune response inside the ovarian cancer microenvironment.
Immune cells are extensively repressed inside of the ovarian cancer microenvironment by a variety of mechanisms, including immune checkpoint inhibitors.Immune-checkpoint blockers, chiefly anti-PD-1/PD-L1 and anti-CTLA-4, have transformed cancer therapy over the last decade.
Thus far, it should be noted that ovarian neoplasm is one of the only neoplasms for which immune checkpoint inhibitors have yet to be approved by the FDA.
Significant heterogeneity was discovered across ovarian cancer patients at the genomic, proteomic, glycoproteomic, and immunologic levels, which we believe should be examined further to increase ICI efficacy.We also expect that combining ICIs with drugs with distinct modes of action will improve ICI efficacy in OC.Yet, in order to maintain tolerability, the schedule and timing must be optimized.Combinations with other medications should be investigated further to enhance efficacy while minimizing toxicity.
Apart from PD-L1, biomarkers having a predictive role for ICIs should be studied.Combining such biomarkers with genetic and immunologic profiling will provide an overall understanding of OC, leading clinical trials toward sensible medication combinations and sequencin.

Fig. 2 .
Fig.2.Bi-directional impact of exosomes from a variety of sources in the tumor microenvironment[110]: Exosomes have a significant role in the onset and progression of ovarian cancer.