Abstract: Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy, with poor survival rates among patients who have advanced disease despite recent significant advances in therapy, including therapy targeting the homologous recombination pathway. Evidence that cell-mediated antitumor immunity, as well as documented programmed death ligand 1 expression, is correlated with improved survival in EOC garnered early optimism regarding the utility of immune checkpoint blockade (ICB) in ovarian cancer. However, the results of multiple clinical trials investigating ICB have revealed very low levels of activity of single-agent immune checkpoint inhibitors, and the testing of combination therapies has not yet identified any combinations with robust activity in a significant proportion of patients who have EOC. In this review, we summarize the results of the major studies of ICB monotherapy and combinations; review novel combinations under investigation, including ICB with cellular therapies; and discuss potential candidate biomarkers for improving the selection of patients who may respond to ICB.
Epithelial Ovarian Cancer
Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy. Although surgery and platinum-based chemotherapy effectively induce remission,1 most women ultimately succumb to recurrent and therapy-resistant disease. Women with platinum-resistant ovarian cancer (PROC) have a median overall survival (OS) of less than 16 months, even with chemotherapy and bevacizumab,2 so that novel therapeutic strategies are needed. A great deal of interest has been shown in utilizing immunotherapy approaches in EOC, given the discovery nearly 2 decades ago that tumor-infiltrating lymphocytes (TILs) are detected in approximately 50% of these tumors, and their presence is associated with longer survival.3,4 Tumors with TILs have higher levels of intratumoral lymphocyte-activating cytokines and interferon gamma (IFN-γ), further supporting the importance of antitumor immunity in this disease.3,5 EOC TILs exhibit cytotoxic activity against autologous tumor-associated antigens in vitro6; however, it is well appreciated that a plethora of other cellular and noncellular factors in the tumor microenvironment (TME) interact to determine the overall tumor immune response.
In recent years, immune checkpoint blockade (ICB) has shifted the treatment paradigm in certain solid tumors and hematologic malignancies.7,8 Pembrolizumab (Keytruda, Merck) is now approved for patients with metastatic or unresectable cancers that have progressed following other treatment, who have no satisfactory alternative treatment options, and who have tumors that exhibit (1) mismatch repair deficiency (MMRd) or high microsatellite instability (MSI-H) or (2) a high tumor mutational burden (TMB). However, outside these disease-agnostic indications, which are unfortunately rare in EOC and even rarer in certain histologic subtypes, such as low-grade endometrioid, mucinous, and clear cell carcinomas, ovarian cancer remains a disease with no ICB-specific approvals. To date, the results of multiple studies of ICB therapy in newly diagnosed and recurrent EOC have been disappointing. Here, we review the results from ICB clinical trials in EOC to date, barriers limiting the success of single-agent ICB, the strategies currently under investigation to overcome these challenges, and potential biomarkers to guide the clinical development of ICB in ovarian cancer.
Single-Agent Immune Checkpoint Blockade Experience in EOC
Several trials testing the efficacy of single-agent ICB have been reported in EOC, initially in recurrent cancers. All of these trials demonstrated very modest activity (Table 1). Several of these ICB monotherapy and combination trials have included evaluations of programmed death ligand 1 (PD-L1) expression—measured in either tumor and immune cells with the combined positive score (CPS)9 or in tumor cells alone with the tumor proportion score (TPS)—or alternative assessments of the immune phenotype of tumors, such as the inflamed gene expression profile (GEP)10; these are discussed in more detail within the section on biomarkers below. In the ovarian cancer cohort of the phase 1b KEYNOTE-028 study, which included 26 patients with PD-L1–expressing recurrent ovarian cancer, the objective response rate (ORR) with pembrolizumab was 11.5% even though this was a biomarker-selected population.11 The phase 2 KEYNOTE-10012 trial examined the efficacy of pembrolizumab in women with recurrent EOC, who were evaluated in 2 separate cohorts according to number of prior lines of therapy. The ORR with pembrolizumab across the 2 cohorts was 8.0%, with a median progression-free survival (PFS) of 2.1 months.12,13 Similarly, in the phase 1b ovarian expansion cohort of the JAVELIN14 trial, which tested the efficacy of avelumab (Bavencio, EMD Serono/Pfizer) in recurrent PROC, the ORR was 9.6%, with a 1-year PFS rate of 10%.14 Single-agent nivolumab (Opdivo, Bristol Myers Squibb) has also been studied; initially promising activity was observed in a small phase 2 study of women with PROC, who had an ORR of 15%, including 2 durable complete responses.15 More recently, the phase 3 NINJA trial compared the efficacy and safety of nivolumab monotherapy vs chemotherapy in women with PROC.16 The trial attempted to answer definitively whether single-agent ICB should be used for women with PROC, especially given that these patients typically have high-grade serous tumors that are microsatellite stable (MSS) with a low TMB. Nivolumab ICB was inferior to nonplatinum chemotherapy, with significantly worse PFS and a nonsignificantly lower ORR (7.6% vs 13.2%; P=.191). Notably, in the patients who did respond to treatment, the median duration of response was 18.7 months (95% CI, 2.5-not evaluable) in those who received chemotherapy vs 7.4 months (95% CI, 3.0-10.3) in those who received nivolumab. Importantly, ICB trials in other tumor types have reported unbalanced effects on PFS and OS, with modest improvements in PFS but more significant extensions of OS,17 suggesting a potential effect on the immune microenvironment that translates to delayed but longer-term responses. These observations underscore the importance of both long-term follow-up in studies and continued efforts to identify populations of patients with EOC who may be likely to benefit from ICB (further discussed below). In addition to programmed death 1 (PD-1)/PD-L1–directed therapies, targeting of cytotoxic T-lymphocyte–associated antigen 4 (CTLA-4) has also been investigated in EOC; a phase 2 trial testing ipilimumab (Yervoy, Bristol Myers Squibb) monotherapy in women with recurrent platinum-sensitive ovarian cancer (PSOC) reported an ORR of 10.3%, but with poor tolerability (NCT01611558). As discussed below, anti–CTLA-4 therapy has more recently been investigated in combination with anti–PD-1 therapy in EOC. Taken together, these studies demonstrate that ICB has the potential to induce durable responses in only a very limited subset of patients with EOC, and that ICB should not be used as monotherapy in EOC without a further definition of reliable biomarkers to delineate the appropriate subpopulations, clinical settings, and/or combinations that allow improved activity.
Combination Immune Checkpoint Blockade
Anti–PD-1/PD-L1 agents exert their effects by targeting the final step of the well-described cancer immunity cycle.18 Although these agents have shown great potential to revolutionize therapy in some cancer types, their success is still limited by both primary and acquired resistance,19 and it is now clear that this approach is insufficient to induce effective antitumor immunity in EOC. A detailed understanding of the mechanisms driving the low response rates to ICB in EOC is rapidly being unveiled, including both tumor cell–intrinsic and –extrinsic characteristics that can promote cancer progression and limit the efficacy of anti–PD-1/PD-L1 therapy. Cell autonomous mechanisms limiting responses in EOC include a relatively low TMB, lack of an inflamed GEP,10 suppressed major histocompatibility complex (MHC) protein expression, and/or antigenic loss.20 Specific somatic alterations such as activating mutations in the phosphatidylinositol 3-kinase (PI3K)/AKT pathway and phosphatase and tensin homolog (PTEN) loss, as well as those increasing Wnt signaling, have also been associated with immune cell exclusion, suppression of T-cell cytotoxic function, lack of antitumor immunity, and poor prognosis in EOC.21-25 In addition, the unique TME within the peritoneal cavity permits interactions between tumor and immune cells, as well as fibroblasts26 and adipocytes,27 and dampens the response to ICB in EOC via many mechanisms.28 Innate and adaptive immune cells, including regulatory T cells (Tregs),29 M2-polarized tumor-associated macrophages (TAMs),30 tumor-associated neutrophils (TANs), and myeloid-derived suppressive cells (MDSCs),31 all contribute by elaborating cytokines and inflammatory factors that limit T-cell homing and suppress T-cell effector function, upregulate inhibitory receptors on tumor and immune cells, and alter dendritic cell function and maturation,32,33 overall shifting the balance to a tumor-permissive environment.34 Given the multiple co-occurring mechanisms of immune suppression in EOC tumors, combinatorial strategies to target various stages of the cancer immunity cycle are likely necessary for robust and durable antitumor immune responses. Approaches to enhance T-cell trafficking to the TME, increase T-cell priming and activation, and stimulate neoantigen generation/presentation are being tested by combining immune checkpoint inhibitors with various agents and are discussed below.
Immune Checkpoint Blockade
Combined With Chemotherapy
Preclinical work has demonstrated the ability of chemotherapy to potentiate the effect of immunotherapy in EOC by inducing local immune activation in the TME.35-39 These data, combined with the modest activity of single-agent ICB in ovarian cancer, have prompted investigation of combinations of anti–PD-1/PD-L1 therapy with chemotherapy (Table 1). The phase 3 JAVELIN-100 trial evaluated avelumab with and/or following carboplatin and paclitaxel in the upfront treatment of EOC. This study was negative and terminated early when the planned interim analysis demonstrated futility regarding the primary endpoint of PFS with the addition of avelumab. The median PFS (mPFS) was not reached in the chemotherapy-alone arm and was 16.8 months (hazard ratio [HR], 1.43; 95% CI, 1.05-1.95) and 18.1 months (HR, 1.14; 95% CI, 0.83-1.56) in the maintenance avelumab and concurrent avelumab plus maintenance avelumab arms, respectively.40 More recently, the IMagyn050 trial tested the safety and efficacy of first-line bevacizumab, carboplatin, and paclitaxel with or without the anti–PD-L1 agent atezolizumab (Tecentriq, Genentech).41 This trial was also designed to also test whether the addition of ICB to chemotherapy and antiangiogenic agents42,43 improved outcomes. IMagyn050 did not meet its co-primary endpoint of significantly improving PFS with the addition of atezolizumab, either in the intent-to-treat (mPFS, 19.5 vs 18.4 months; HR, 0.92; 95% CI, 0.79-1.07) or the PD-L1–positive population (mPFS, 20.8 vs 18.5 months; HR, 0.80; 95% CI, 0.65-0.99).41 It also did not meet its co-primary endpoint of OS at the first interim analysis, although these data remain immature. This trial is another example of a study in which longer-term follow-up may be necessary to unveil potential benefit in a subset of patients.44 In addition to being tested as first-line therapy, the combination of ICB and chemotherapy has also been tested in recurrent PROC in JAVELIN-200, a phase 3 trial randomly assigning women to avelumab combined with pegylated liposomal doxorubicin (PLD), avelumab alone, or PLD alone. This trial also did not demonstrate significant improvement in PFS or OS across arms with avelumab alone or in addition to PLD chemotherapy in women with PROC.45 The mPFS was 3.7 months, 3.5 months, and 1.9 months in the combination therapy, PLD-alone, and avelumab-alone groups, respectively, whereas the HR was 1.68 (93% CI, 0.59-1.24) for avelumab vs PLD and 0.78 (93% CI, 1.32-2.60) for the combination vs PLD. Although this was a negative study, a nonsignificant trend toward improved PFS was noted at the 12-month mark in the combination arm, again suggesting the need for the better identification of specific populations of patients who may benefit and experience a long duration of response.
Immune Checkpoint Blockade Combined With Antiangiogenic Therapies
The interplay between angiogenic signaling and immune suppression in the TME has been demonstrated preclinically, with vascular endothelial growth factor (VEGF) and other angiogenic factors contributing to immunosuppression by inducing vascular abnormalities, inhibiting antigen presentation, suppressing immune effector cells, and augmenting the activity of immunosuppressive Tregs, MDSCs, and TAMs.42,46 The resulting balance of immunosuppressive to effector cells in the TME further stimulates angiogenesis and perpetuates this cycle; thus, combinations targeting both angiogenesis and the PD-1/PD-L1 pathway are being investigated as a strategy to overcome resistance to ICB.47 A combination of nivolumab and bevacizumab was evaluated in a single-arm phase 2 study of 38 patients with recurrent EOC, demonstrating an ORR of 28.9.48 Notably, the response rate was 40% in PSOC vs 17% in PROC, suggesting some promising clinical activity in the platinum-sensitive setting. In recurrent PSOC, the ongoing ATALANTE/ENGOT-ov29 study is testing the combination of carboplatin-based chemotherapy, bevacizumab, and atezolizumab (Table 2; NCT02891824). In PROC, the NRG-GY009 phase 2/3 trial is testing PLD and atezolizumab with and without bevacizumab in recurrent PROC (NCT02839707), and the AGO-OVAR-2.29 study is evaluating the addition of atezolizumab to platinum-based chemotherapy plus bevacizumab (NCT03353831). Beyond bevacizumab, the combination of pembrolizumab with the multiple kinase inhibitor lenvatinib (Lenvima, Eisai)—which has shown impressive activity in MSS/MMR-proficient endometrial cancer49 (for which it now has an FDA approval)—resulted in an ORR of 29% in 31 patients with recurrent ovarian cancer at interim analysis of the LEAP-005 phase 2 basket study.50
Immune Checkpoint Blockade Combined With PARP Inhibitors
Poly(ADP-ribose) polymerase (PARP) inhibitors have become a major component of therapy for many women with EOC because approximately half of high-grade EOCs exhibit defects in the homologous recombination DNA damage repair pathway. Deleterious somatic and germline alterations in BRCA1 or BRCA2 account for up to 22% of high-grade serous carcinomas; thus, other mechanisms are leading to homologous recombination deficiency (HRD) in this disease.51,52 It has been demonstrated that BRCA-mutant (BRCAm) tumors possess more mutations,53 indels,54 and CD8+ TILs than do BRCA wild-type (BRCAwt) tumors. They also have a higher level of PD-L1 expression and a larger predicted neoantigen load55 and exhibit IFN-γ immune signatures,56 raising the theoretical yet still unproven possibility of increased sensitivity to ICB in EOC tumors with HRD. In addition, work in preclinical models has demonstrated that DNA damage induced by PARP inhibitors in BRCAm tumors induces an innate immune response via activation of the stimulator of interferon genes (STING) pathway, resulting in an improved response to anti–PD-L1 therapy.57,58 Thus, clinical trials have been developed to test the combination of PARP inhibition and ICB (Table 1). In the phase 2 MEDIOLA trial, which evaluated the combination of durvalumab (Imfinzi, AstraZeneca) and the PARP inhibitor olaparib (Lynparza, AstraZeneca) in BRCAm59 vs BRCAwt60 PSOC, the ORR for the combination was 72% in the BRCAm patients (notably, a population of patients expected to have a high response rate to PARP inhibition alone) vs 31.3% in the BRCAwt cohort.59 The ongoing ANITA trial is testing the addition of atezolizumab to the combination of carboplatin-based chemotherapy and niraparib (Zejula, GSK) maintenance in recurrent PSOC (NCT03598270). Also in PROC, the combination of pembrolizumab and niraparib was tested in the TOPACIO trial and demonstrated an ORR of 18%, with similar rates across BRCA and HRD subgroups.61 Even in patients without a RECIST (Response Evaluation Criteria in Solid Tumors) response, prolonged stable disease was often observed, the combination demonstrating promising clinical activity especially in the BRCAwt and homologous recombination–proficient populations with limited treatment options. In addition, immunogenomic profiling and single-cell imaging of tumor samples from TOPACIO participants identified mutational signature 3 (reflecting HRD) and a positive immune score (indicating exhausted CD8+ T cells) as 2 biomarkers of improved response to the combination.62 The phase 2 MOONSTONE trial (NCT03955471) testing the efficacy of niraparib in combination with dostarlimab (Jemperli, GSK) in women with BRCAwt PROC was closed to enrollment early on the basis of interim futility analysis. The combination of a PARP inhibitor with ICB is currently being compared with chemotherapy in PROC in the ongoing MITO 33 trial, which is comparing dostarlimab plus niraparib vs chemotherapy (NCT04679064).
In addition, several ongoing trials are comparing the combination of an anti–PD-1/PD-L1 agent and a PARP inhibitor added to and/or following first-line chemotherapy with or without bevacizumab in newly diagnosed EOC (Table 2). For example, the ATHENA trial is evaluating the combination of nivolumab plus rucaparib (Rubraca, Clovis Oncology) as maintenance therapy following response to first-line platinum-based chemotherapy in advanced EOC (NCT03522246). Several additional trials with similar strategies are also underway in the setting of first-line treatment of advanced disease. The phase 3 AGO/DUO/ENGOT trial is evaluating platinum-based chemotherapy and bevacizumab with or without concurrent durvalumab and maintenance durvalumab with or without maintenance olaparib in the first-line treatment of advanced EOC (NCT03737643). ENGOT-ov43 is testing pembrolizumab added to chemotherapy with or without bevacizumab and to maintenance with and without olaparib (NCT03740165). The FIRST trial is testing the benefit of adding dostarlimab to chemotherapy with or without bevacizumab and to niraparib maintenance (NCT03602859), and ENGOT-ov39 is evaluating the addition of atezolizumab to both front-line chemotherapy plus bevacizumab and maintenance with bevacizumab (NCT03038100).
Immune Checkpoint Blockade Combined With Anti–CTLA-4 Blockade
It is known that CTLA-4 is an inhibitory immune checkpoint molecule that promotes additional effector T-cell dysfunction beyond that of PD-1.63 Up to 50% of EOC TILs were found to express both PD-1 and CTLA-4, and combined blockade of both resulted in rescue of TIL function and led to tumor regression in murine models.64 It was recently demonstrated that CTLA-4 attenuates CD28 costimulatory signals by antigen-presenting cells in EOC, and that augmentation of CD28 costimulation by anti–CTLA-4 therapy enhances TIL activation in response to anti–PD-1.65 Clinically, the combination of anti–PD-1 and anti–CTLA-4 therapy was tested in NRG-GY003, a phase 2 study comparing nivolumab plus ipilimumab vs nivolumab alone in recurrent EOC. This study reported a superior ORR with the combination, 31.4% vs 12.2% (odds ratio, 3.28; P=.034), as well as longer mPFS (3.9 vs 2.0 months).66 However, related grade 3 or higher adverse effects were reported in 49% of patients receiving the combination vs 33% of those receiving nivolumab alone, findings consistent with those of prior studies of this combination. The Fc-enhanced anti–CTLA-4 antibody AGEN1181 is being tested in combination with the anti–PD-1 agent balstilimab in a phase 1 study of advanced solid tumors (NCT03860272). There were 2 confirmed PRs and 2 cases of stable disease in the 9 evaluable patients with ovarian cancer included in this study,67 and randomized phase 2/3 trials are being initiated in PROC, endometrial cancer, and MSS colorectal cancers.
Other ongoing studies are testing the combination of anti–PD-1 and anti–CTLA-4 therapy in EOC, in both the first-line and the recurrent setting (Table 2). The phase 2 TRU-D trial is testing the addition of durvalumab with or without the anti–CTLA-4 antibody tremelimumab to neoadjuvant carboplatin and paclitaxel in women with newly diagnosed advanced EOC (NCT03899610). A similar combination of nivolumab with or without ipilimumab added to neoadjuvant or adjuvant chemotherapy in advanced EOC is also being tested (NCT03245892). An ongoing phase 2 trial, MEDI4736, is also testing the combination of durvalumab and tremelimumab in recurrent EOC (NCT03026062).
Triplet and Other Targeted Therapy Combinations
Given the apparent additive activity of ICB combined with chemotherapy, PARP inhibitors, antiangiogenic agents, or anti–CTLA-4 drugs, triplet combinations are now being testing in EOC with the goal of rendering these tumors more vulnerable to ICB treatment and increasing the likelihood of potentially durable responses. In recurrent EOC, the previously mentioned MEDIOLA trial also includes a cohort of patients with non-germline BRCAm PSOC undergoing treatment with the combination of durvalumab, bevacizumab, and olaparib. Preliminary results for this combination are promising, with a reported disease control rate and ORR of approximately 77%.59 The phase 2 OPAL trial (NCT03574779) examined the combination of dostarlimab, niraparib, and bevacizumab in patients with recurrent EOC. In the PROC cohort, which comprised mostly BRCAwt patients, the ORR was 17.9%, with 7 partial responses and 23 of 39 evaluable patients with stable disease,68 suggesting clinical activity in a population predicted to have poor responses to systemic therapies. A single-arm phase 2 cohort study also evaluated the combination of pembrolizumab with oral metronomic cyclophosphamide and bevacizumab in patients with recurrent EOC, 75% of whom had PROC.69 There were 3 complete and 14 partial responses in the 40 patients treated with this combination, with an ORR of 47.5% and a clinical benefit rate of 95.0%. Moreover, durable responses of longer than 12 months were observed in 25%. A phase 1 study of the combination of durvalumab, olaparib, and the VEGF receptor 1-3 inhibitor cediranib in recurrent gynecologic cancers reported a clinical benefit rate of 67%, with 4 of 9 patients exhibiting a partial response.70 On the basis of these findings, the ongoing phase 2 NRG-GY023 trial is comparing this triplet combination with durvalumab and cediranib or physician’s choice of chemotherapy in women with PROC (NCT04739800).
Novel Investigational Strategies
Targeting PI3K, Wnt, and Notch Pathways. Several oncogenic pathways have been shown to facilitate the ability of tumor cells to evade antitumor immunity71,72; the PI3K/AKT and Wnt pathways are particularly relevant in EOC. Genomic alterations activating the PI3K/AKT pathway, loss of PTEN function, or both are frequently observed in EOC and are associated with reduced TIL numbers and cytotoxic function.22,73 Accordingly, PI3K and AKT inhibitors have been shown to increase immune infiltrating cells,74 activate CD8+ TILs,75 and selectively inhibit the proliferation of Tregs,73 thereby increasing sensitivity to anti–PD-1 therapy in preclinical models.74 Trials based on this rationale are underway (Table 2), testing the combination of PI3K and AKT inhibitors with ICB, including a phase 1 study of the AKT inhibitor capivasertib combined with durvalumab and olaparib in advanced solid tumors. An expansion cohort in gynecologic malignancies is ongoing (NCT03772561).
Wnt signaling in cancers, including EOC, has effects on immune cells, including effector T cells and dendritic cells, that can promote immune exclusion and resistance to ICB.36,76,77 Thus, trials are now underway (Table 2) testing the effect of suppressing Wnt signaling with porcupine acetyltransferase (PORCN) inhibitors (NCT01351103, NCT02521844). In addition, the combination of the PORCN inhibitor ETC-1922159 and pembrolizumab is being evaluated in a phase 1 basket study of solid tumors (NCT02521844).
Notch signaling plays critical roles in vascular homeostasis, and crosstalk between this pathway and VEGF regulates cancer angiogenesis. In EOC, Notch signaling is also implicated in stem cell maintenance, epithelial-mesenchymal transition, chemoresistance, and poor outcomes.78 Thus, novel therapies targeting gamma secretase and delta-like ligand 4 (DLL4) are being tested therapeutically in EOC. Navicixizumab, a novel anti-DLL4/VEGF bispecific antibody designed to target Notch and VEGF signaling simultaneously, has received an FDA fast track designation for the treatment of patients with recurrent EOC. A phase 1b study of navicixizumab combined with paclitaxel in 44 heavily-pretreated patients with PROC reported 1 complete response, 18 partial responses, and 15 patients with stable disease, with a manageable safety profile.79 A randomized, multicenter phase 3 study comparing navicixizumab with or without paclitaxel vs paclitaxel alone in patients with PROC expressing certain RNA markers is planned (NCT05043402).
Selective Stimulation of Cytotoxic T Cells. Strategies to activate cytotoxic T cells preferentially and avoid the stimulation of immunosuppressive T-cell populations are currently under active investigation. For example, nemvaleukin, an engineered protein comprising a modified interleukin 2 (IL-2) and high-affinity IL-2 alpha receptor chain, is hypothesized to selectively activate the intermediate-affinity IL-2 receptor complex. Nemvaleukin was recently granted FDA fast track designation for use in combination with pembrolizumab in advanced PROC. The phase 1/2 ARTISTRY-1 trial, which is evaluating nemvaleukin alone and in combination with pembrolizumab in advanced solid tumors, reported 1 complete response and 1 partial response in 13 evaluable patients with heavily pretreated PROC, with 9 experiencing disease control.80 Building upon this signal of activity, the phase 3 ARTISTRY-7 trial, which is a 4-arm study evaluating nemvaleukin alone or in combination with pembrolizumab vs pembrolizumab alone or physician’s choice of chemotherapy, is ongoing (NCT05092360).
Simultaneous Targeting of the Innate Immune System. Although the actions of anti–PD-1/PD-L1 therapy were historically attributed to activation of the adaptive immune system, it is now clear that stimulation of the innate immune system is also required to induce T-cell responses.81,82 Direct or indirect activation of the innate immune system via targeting of immune cells (eg, natural killer cells and dendritic cells) or activation of pattern recognition receptors, such as toll-like receptors (TLRs) and the cGAS/STING pathway, has demonstrated preclinical activity in various cancers.83,84 Proof of concept for the combination of innate and adaptive immune targeting has been demonstrated in EOC murine models.85 Currently, different combinations targeting both innate and adaptive immunity are under clinical investigation (Table 2). For example, ongoing phase 1/2 studies are testing the combination of doxorubicin, durvalumab, and the TLR8 agonist motolimod in PROC (NCT02431559) and the combination of the anti-CD27 antibody varlilumab and nivolumab in solid tumors, including ovarian cancer (NCT02335918).
Cancer Vaccines Combined With ICB. Cancer vaccines are now being investigated in combination with ICB in EOC, with several combinations exhibiting promising preliminary activity and acceptable safety profiles. For example, clinical trials testing the combination of WT-1 or NY-ESO-1 vaccine with nivolumab in recurrent EOC (NCT02737787; Table 3) and the combination of a multi-epitope antifolate receptor (anti-FR) vaccine with durvalumab (NCT02764333) in PROC have reported the combinations to be safe and tolerable.86,87 In addition, personalized neoantigen vaccines are being combined with ICB in EOC (NCT04024878) and other cancers88 in an attempt to “steer” the immune response while simultaneously “releasing the brakes” on the immune system, thereby inducing a more robust and specific antitumor immune response. Another novel approach being studied is the autologous tumor cell vaccine gemogenovatucel-T, also known as Vigil, which is engineered to express granulocyte-macrophage colony–stimulating factor (GM-CSF) and bi-shRNA-furin to transforming growth factor beta (TGF-β); the vaccine is being administered as maintenance therapy to women with newly diagnosed advanced EOC following response to upfront surgery and chemotherapy in the VITAL trial (NCT02346747).89 Initial efficacy results demonstrated longer median relapse-free survival (HR, 0.39; 90% CI, 0.20-0.75) and OS (HR, 0.34; 90% CI, 0.14-0.83) in the vaccine arm than in the placebo arm within the homologous recombination–proficient cohort.90 Additional trials are ongoing testing this agent in combination with atezolizumab (NCT03073525) and durvalumab (NCT02725489) in advanced gynecologic cancers.
Dendritic cell vaccines (DCVACs) have also been investigated in EOC, with the aim of enhancing antigen presentation in the TME. The production of DCVACs typically involves apheresis to obtain autologous immature dendritic cells, in vitro stimulation and maturation, loading of stimulated dendritic cells with tumor-associated antigens, and then administration of the vaccine into the patient, often in combination with chemotherapy or other therapies (reviewed in Zhang and colleagues91). For example, a randomized phase 2 trial comparing DCVAC administration following or during first-line adjuvant chemotherapy with chemotherapy alone in patients with newly diagnosed advanced EOC reported significantly longer PFS and a nonsignificant trend toward extended OS in those who received DCVAC following chemotherapy vs those who received chemotherapy alone.92 Furthermore, in an exploratory biomarker analysis, outcomes were significantly better in the patients with low levels of CD8+ TILs who received vaccine than in those with low levels of TILs in the chemotherapy-alone arm, suggesting a benefit of DCVAC in immunologically “cold” tumors. Ongoing clinical trials are planned or underway further investigating the safety and efficacy of DCVAC alone or in combination in both newly diagnosed and recurrent EOC91; a few of these are included in Table 3.
Adoptive Cellular Therapy Combined With ICB. Another active area of research in EOC is the use of adoptive cell therapies (ACTs), in which autologous immune effector cells are isolated from the patient, cultured and often genetically modified ex vivo, and then reinfused to enhance the antitumor response. These approaches are being combined with ICB in EOC, in part on the basis of initial observations of high levels of PD-1 expression on infused TILs in ACT trials.93 To date, trials testing TIL ACT alone in EOC have demonstrated feasibility, but modest efficacy (reviewed by Sarivalasis and colleagues94). Among other strategies, TIL ACT therapy combined with ICB has shown safety and feasibility, along with preliminary evidence of clinical and immunologic activity, in small, early-phase trials, improving TIL expansion and enhancing CD8+ T-cell tumor reactivity in comparison with TIL ACT alone.95 Current clinical trials (Table 3) are studying the combination of TIL infusion with aldesleukin (Proleukin, Clinigen Group) and either pembrolizumab (NCT01174121, NCT03158935) or ipilimumab and nivolumab (NCT03287674).
Beyond TILs, other ACT approaches, including T-cell receptor (TCR)–engineered T cells96 and chimeric antigen receptor (CAR) T cells, are being investigated in EOC. These strategies differ from TILs in that T cells are genetically altered to be directed to specific peptides, typically tumor-associated antigens (TAAs). To date, the most common CAR T-cell targets tested clinically include folate receptor alpha (FRα), mesothelin, and MUC16; several ongoing clinical trials are also testing third-generation CAR T cells directed against additional known TAAs in EOC (reviewed by Benard and colleagues97). TCRs engineered to recognize specific epitopes from TAAs, including WT1, p53, NY-ESO-1, and MAGE-A4, have been developed, and NY-ESO-1 has been tested in phase 1/2 clinical trials,98 with additional trials ongoing (Table 3). As of now, several barriers still limit the success of TCR-based and CAR T-cell therapies in EOC and other solid tumors, including the limited number and heterogeneous expression of membrane antigen targets, inadequate tracking of T cells to tumor sites, and limited fitness and survival in the TME. Novel strategies capitalizing on viral vector–based gene editing to overcome immunosuppressive and/or inhibitory signals are showing promising preclinical and early-phase clinical activity.99,100 For example, mesothelin-directed CAR T cells with CRISPR-Cas9–mediated knockout of PD-1 are being tested in mesothelin-positive solid tumors, including EOC (NCT03747965).
Bispecific antibodies are antibody constructs that recognize both a specific TAA expressed on tumor cells and the CD3 complex expressed on T cells.101 They therefore redirect endogenous polyclonal T cells to the tumor to induce tumor cell–specific lysis without the need for ex vivo expansion and genetic manipulation.102,103 An MUC16 bispecific antibody has shown potent preclinical activity and is currently being tested clinically alone and in combination with the anti–PD-1 agent cemiplimab (Libtayo, Sanofi-Aventis/Regeneron; NCT03564340).
Biomarkers of Response to ICB in Ovarian Cancer
Regardless of the response rates across populations of different cancer types, a common observation is that responses to ICB are often more durable than responses to other systemic therapies.104 Thus, the identification of biomarkers to select patients likely to respond to ICB or combinations containing immune checkpoint inhibitors are necessary, and this is an active area of investigation.
Tumor Cell–Intrinsic Biomarkers
PD-L1 expression in the TME is a well-studied potential biomarker of response to ICB, given its correlation with therapeutic responses across diverse cancer types.105,106 This correlation has been observed in EOC, although not uniformly across all studies. KEYNOTE-100 reported a positive correlation between response to pembrolizumab and CPS, with a CPS of 1 or higher corresponding to an ORR of 5% and a CPS of 10 or higher corresponding to an ORR of 17.1%.12 The incidence of cancers demonstrating a CPS of 10 or higher was very low, however. Notably, all 7 complete responses seen on study were in the subgroup with a CPS of 10 or higher. In IMagyn050, stratification by PD-L1 staining in immune cells with a cutoff of 1% did not identify a population that derived benefit from the addition of atezolizumab.41 However, a potential benefit from the addition of atezolizumab was seen in prespecified exploratory analyses of 2 populations, one with immune cell PD-L1 expression of 5% or higher (HR, 0.66; 95% CI, 0.44-0.98) and another with tumor cell PD-L1 expression of 1% or higher (HR, 0.45; 95% CI, 0.19-1.02). No differences were seen in OS according to PD-L1 expression in the NINJA study, although the PD-L1 analysis was restricted to tumor cells only by using a TPS of 1% or higher for stratification. Several challenges limit the clinical utility of PD-L1 as a biomarker,107-109 so that further optimization of PD-L1 testing is required, and its use in combination with other methods of assessing the immune-inflamed phenotype of a tumor is likely necessary.110
TMB is another leading biomarker candidate because of the assumption that a high proportion of mutations enhances immunogenicity.111,112 Pembrolizumab is now approved for use in all unresectable or metastatic tumors with a high TMB, determined with the FoundationOne CDx assay (Foundation Medicine). In EOC, the TMB is consistently low,12,113,114 and a prespecified exploratory analysis assessing TMB in KEYNOTE-100 revealed no association with rates of response to pembrolizumab.113 In IMagyn050, most tumors had a low TMB regardless of BRCA1/2 mutation/HRD status, and neither was associated with response to atezolizumab,114 confirming the KEYNOTE 100 data12,41 and underscoring the likely low utility of TMB assessment in EOC. BRCA mutations have also not been shown to be predictive of cancer responsiveness to ICB.12,14,59
Additional transcriptomic analyses have been performed to identify tumors with a T cell–inflamed phenotype115 or an immunoreactive or mesenchymal molecular subtype.51 In KEYNOTE-100, a prespecified exploratory analysis comparing expanded GEP signatures (eg, angiogenesis, hypoxia, granulocytic MDSCs, epithelial-mesenchymal transition (EMT)/TGF-β signaling, Wnt signaling) did not reveal any significant associations with clinical outcomes with pembrolizumab,116 and thus further work is needed to translate the findings of these studies into clinically relevant biomarkers.
Tumor Cell–Extrinsic Biomarkers
The importance of identifying biomarkers of response in the context of the TME is increasingly clear. Early work by Zhang and colleagues3 established the link between the presence of infiltrating T cells and improved outcomes in ovarian cancer; this was further supported by Li and colleagues,4 providing a rationale for pursuing immunotherapy-based therapeutic strategies in EOC. The ratio of immune-reactive to immune-tolerant cellular subpopulations in a tumor is correlated with clinical outcomes39 and may be an important consideration in terms of predicting response to ICB. Further, single-cell tissue-based cyclic immunofluorescence approaches paired with immunogenic profiling of EOC tumors from the TOPACIO trial identified specific tumor cell mutational signatures and CD8+ T-cell molecular states, as well as spatial interactions between them in the TME, that were associated with response to niraparib and pembrolizumab.62 These works suggest that single-cell spatially resolved data from clinical samples may be key to developing predictive biomarkers for determining response to therapy and aiding patient stratification.
The quest for more precise biomarkers to predict ICB response in EOC has extended beyond the TME to liquid biopsy–based approaches,117,118 including circulating tumor DNA,119 noncoding RNA,120 circulating tumor cells, and immune cells (CD8+ T cells, MDSCs, neutrophils),121-123 as well as soluble factors such as PD-L1,124 cytokines, and chemokines.125 Although associations with clinical benefit and survival are observed, these observations still require validation in prospective studies. In addition, the microbiome has recently been found to contribute to varied responses to ICB and may represent another source of predictive biomarkers.126-129 A recent review130 comprehensively summarizes the growing body of translational work that supports a key role of the gut microbiome in modulating the response to ICB, most convincingly to date in patients with melanoma.128,129,131 Furthermore, specific microbial phyla have been associated with response to ICB across solid tumors,132 and these have been identified as potential novel and modifiable biomarkers. As with liquid biopsy–based biomarker candidates, an improved understanding of the interactions between the host and the intestinal microbiome is necessary to identify microbiome-based biomarkers of ICB response. Nonetheless, these studies are exciting and raise the possibility of using the vaginal microbiome as an alternative source of candidate biomarkers in gynecologic cancers.
Conclusions
The low rates of response to ICB in EOC demonstrated thus far provide no justification for the use of single-agent ICB in unselected populations with recurrent EOC unless the cancer is found to be TMB-high or MSI-H/MMRd, and thus qualified for pembrolizumab or dostarlimab treatment on the basis of cancer site–agnostic approval. However, the clinical trials performed to date demonstrate the capability of ICB to produce durable responses in a very small subset of patients. These findings provide opportunities and pose challenges to the medical and scientific community to further uncover the specific barriers limiting the activity of ICB, to employ novel technologies to discover predictive biomarkers, and to develop immunotherapies that extend beyond ICB to help these agents work better. The identification of transformative biomarkers for predicting response to immunotherapy—which ultimately requires a method to test if a patient has a tumor-reactive T-cell repertoire that can access the TME and effectively eliminate cancer cells once there—remains challenging and will likely require the incorporation of multiple parameters. Combining patient data from across ICB trials with low numbers of responders may prove useful for the discovery of new candidates, which could then be validated in prospective trials. Building on what has already been learned and applying emerging technologies that can characterize tumors and their microenvironment at the single-cell level will undoubtedly lead to improved biomarkers; these not only will optimize the selection of patients for ICB therapies but also will also improve our ability to test novel ICB combinations and sequences in the most appropriate populations.
Disclosures
Dr Porter has no disclosures. Dr Matulonis has done consulting for AstraZeneca, Merck, Novartis, NextCure, Blueprint Medicines, and Trillium Therapeutics.
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