The implementation of immune checkpoint inhibitor therapy across solid tumors in recent years has been heralded by much enthusiasm given the promise of deep, durable therapy responses previously unseen in the treatment of advanced-stage malignancies. Although the therapy landscape of metastatic prostate cancer has changed considerably in the past decade – driven by success with novel agents and combinatorial approaches – enthusiasm for checkpoint inhibition (CPI) in prostate cancer has been tempered by limited efficacy demonstrated in clinical trials. Accordingly, attention has turned to better understanding the unique tumor-immune microenvironment of prostate cancer, its dynamics in conferring treatment resistance, and novel mechanisms that may be exploited alone or in combination with existing immune-targeting strategies. In parallel, there have been significant efforts to characterize molecular markers of immunotherapy response and create a new predictive taxonomy for clinical translation. Herein, we review key studies that have driven recent progress in immunotherapy and immune biomarker development in prostate cancer and further highlight important knowledge gaps and promising treatment approaches.
MULTIMODAL IMMUNOTHERAPY STRATEGIES
Prostate cancer is marked by a suppressive ‘cold’ tumor immune microenvironment and poor immunogenicity compared with other tumors, resulting in attenuated immune recognition and T-cell-mediated cytotoxicity . However, prostate cancer overexpresses antigens specific to prostate tissues, such as prostatic acid phosphatase (PAP), prostate-specific membrane antigen (PSMA) and prostate-specific antigen (PSA), which could potentially serve as targets for vaccine-based immunotherapeutics (Fig. 1). Indeed, the IMPACT trial of sipuleucel-T (Provenge), an autologous dendritic cell-based vaccine against PAP, demonstrated an overall survival benefit in men with metastatic castration-resistant prostate cancer (CRPC) and resulted in the first Food and Drug Administration (FDA) approval of a therapeutic cancer vaccine . Despite prolongation of survival, objective tumor responses and PSA declines are infrequent with sipuleucel-T. More recently, other antigen-presenting cell (APC) vaccines have been developed (GVAX, DCVAC/Pa, BPX-101) with the goal of inducing more potent and sustained APC activation. A trial of GVAX in combination with docetaxel chemotherapy (NCT00133224) was terminated early because of lack of efficacy. Results of a phase III study of DCVAC/PCa in the same setting (NCT02111577) are not yet reported, and there are no ongoing studies of BPX101 . Nonautologous therapeutic vaccines are appealing given that the process for ex-vivo manipulation of APCs can be cumbersome. Unfortunately, the Poxviral-based vaccine ProstVac-V/F (targeting PSA) did not demonstrate an overall survival benefit in a phase III study , and an mRNA-based vaccine CV9104 (targeting PSA, PSCA, PSMA, STEAP1, PAP, MUC1) did not show a survival benefit over placebo in a phase I/IIB study . Other nonautologous vaccine-based approaches being tested as monotherapy or in combination with other immunostimulatory approaches include strategies to target PSA (Proscavax, NCT03579654; ADXS31-142, NCT02325557; Adenovirus/PSA, NCT00583024), PAP (pTVG-HP, NCT02499835), telomerase (UV1/hTERT2012P, NCT01784913; GX301, NCT02293707), 5T4 (ChAdOx1.5T4, NCT03815942), brachyury (MVA-BN-Brachyury, NCT03493945), Bcl-xl (Bcl-xl_42-CAF09b, NCT03412786); or combinations of PSA, PSMA, PSCA (PF-06753512, NCT02616185) or PSA, MUC1 and brachyury (ETBX-071, ETBX-061, and ETBX-051, NCT03481816). Another strategy to promote an immune response is intratumoral injection of immunostimulants, such as poly-ICLC (NCT03262103), the TLR9 agonist SD-101 (NCT03007732), or the oncolytic virus ProstAtak(AdV-tk) (NCT01436968). However, a more personalized approach to vaccine therapy may show more promise in larger trials. For example, a neoantigen DNA vaccine based on DNA sequencing from a patient's tumor is being tested in combination with checkpoint immunotherapeutics in metastatic hormone sensitive prostate cancer (NCT03532217). A strategy taking into account preexisting host-specific immune features has been tested in a randomized controlled phase II trial, where a custom vaccine based on host immunoreactivity to a panel of HLA-type specific tumor peptides (such as PSA) in patients with chemotherapy-naïve CRPC resulted in significant improvements in PSA progression-free survival and overall survival .
Tasquinimod is a first-in-class oral immunomodulatory agent inhibiting S100A9, a calcium-binding protein that promotes myeloid-derived suppressor cell (MDSC) activity. In addition, tasquinimod has antiangiogenic, antiproliferative and antimetastatic properties. Having demonstrated efficacy as monotherapy in CRPC , recent studies have explored combination therapy with cabazitaxel  and a maintenance strategy after response or stability after first-line docetaxel chemotherapy for CRPC . In the latter trial, tasquinimod led to a significant improvement in radiographic progression-free survival (hazard ratio = 0.6); however, maintenance therapy was associated with excessive severe treatment-related adverse events (50 versus 27%) and deterioration in quality-of-life scores. Further development of tasquinimod in prostate cancer has since ceased.
CPI, the model of immunotherapy that changed the therapeutic landscape of many cancer types, has been met with more modest enthusiasm in prostate cancer owing to underwhelming efficacy. Ipilimumab, an inhibitor of cytotoxic T-lymphocyte antigen 4 (CTLA-4), failed to demonstrate a survival benefit compared with placebo in both chemotherapy-resistant and minimally symptomatic, chemotherapy-naïve CRPC cohorts [10,11]. Blockade of programmed death 1 (PD-1) receptor with pembrolizumab in the refractory prostate cancer cohort of the nonrandomized KEYNOTE-028 trial (selected for patients with PD-L1 expression in ≥1% of tumor or stromal cells) demonstrated an overall response rate of 17.4%; whereas the response rate in this biomarker-selected population was modest, most of these responses were over 1 year in duration . A more modest response rate (<10%) was recently reported in the KEYNOTE-199 study of docetaxel-refractory CRPC [13▪]; however, the response rate was reported to be higher in combination with enzalutamide in chemotherapy-naïve patients progressing on enzalutamide . What remains clear is that CPI monotherapy for unselected patients is a suboptimal strategy in advanced prostate cancer. In turn, approaches combining complementary or synergistic immune mechanisms (CPI with therapeutic vaccine; CPI with MDSC-targeted therapy ) or combining classes of checkpoint inhibitors (CTLA-4 inhibitor with PD-1 inhibitor) are rational strategies to improve the efficacy and are currently in clinical testing. Indeed, a recent phase II study of the combination of ipilimumab with the anti-PD1 agent nivolumab in mCRPC demonstrated promising overall response rates of 26% in cohort 1 (chemotherapy-naïve patients) and 10% in cohort 2 (patients who progressed after taxane-based chemotherapy); however, significant toxicity was seen leading to only 33% of patients in cohort 1 and 24% of patients in cohort 2 completing the planned four cycles of combination therapy . Furthermore, the biological underpinnings of resistance to immunotherapy in prostate cancer continues to be elucidated and will serve as a rationale for therapeutic targeting. For example, the inhibitory immune checkpoint VISTA appears to serve as a mechanism of dynamic, compensatory immunosuppression after exposure to ipilimumab in prostate cancer , and thus represents a potential second therapeutic target for combination therapy in future clinical trials.
The presence of prostate lineage-specific antigens has been exploited in the study of novel targeted therapies such Lutetium-177 PSMA for metastatic CRPC . In the immunotherapy space, the rationale for greater tumor specificity has led to the development of bispecific T-cell engager (BiTE) antibodies, which constitute an efficient system synapsing T cells and cancer cells to promote T-cell-mediated tumor cytotoxicity. MOR209/ES414, a BiTE targeting CD3ε and PSMA, induced robust T-cell activation and tumor kill in vitro and in vivo; however, clinical translation is limited by a short serum elimination half-life . To abrogate this limitation, depot-injectable controlled delivery of a BiTE with the same antigenic targets demonstrated feasibility in preclinical prostate cancer models . BiTEs have rapidly moved to early phase clinical studies of refractory prostate cancer and are distinguished by different targets including PSMA (BAY2010112, NCT01723475; AMG160, NCT03792841; ES414, NCT02262910), Her2 (NCT03406858) and EpCAM (MT110-101, NCT00635596).
Improving the efficacy of chimeric antigen receptor (CAR) T-cell therapy by overcoming a suppressive tumor-immune microenvironment remains a challenge in refractory solid tumors including prostate cancer. The global experience of CAR T-cell therapy for prostate cancer remains limited to early reports from PSMA CAR T-cell trials demonstrating some PSA declines and stable disease as best response in a small number of treated patients [21,22]. Recently, PSMA-directed CAR T cells with co-expression of a dominant-negative TGF-bRII (thereby inhibiting immunosuppressive TGF-b signaling) showed enhanced lymphocyte proliferation and tumor eradication in prostate cancer mouse models compared with wild-type PSMA CAR T cells . These promising findings have led to a phase I trial in CRPC (NCT03089203). Additionally, trials employing PSMA, PSCA and EpCAM-targeted CAR cellular therapy are ongoing, including a planned trial in China of anti-PSMA CAR Natural Killer (NK) cells for CRPC, in line with increased efforts to develop non-T CAR cellular therapies in hematological and solid cancers.
BIOLOGICAL CORRELATES OF IMMUNOTHERAPY RESPONSE
In light of the observation that a subset of patients with prostate cancer respond and durably benefit from immunotherapy, there has been intense focus on better understanding the determinants of immunotherapy response (and resistance) with the goal of improving the precision of patient care. In recent years, large-scale, systematic analyses of the prostate cancer genome have revealed a so-called ‘long tail’ of mutational drivers [24▪] as well as distinct molecular and phenotypic disease subtypes associated with response to immunotherapy.
The best characterized prostate cancer subtype linked to CPI response constitute tumors with deficient DNA mismatch repair (dMMR) and/or microsatellite instability (MSI). Following a proof-of-concept study in colorectal cancer, Le et al.  demonstrated the efficacy of PD-1 blockade across numerous mismatch repair-deficient cancers, including prostate cancer, with functional studies in responders confirming proliferation of neoantigen-specific T cells driven by the exceptionally high somatic mutation and mutant peptide load present in these tumors. Consequently, in May 2017, the FDA provided tumor-agnostic approval of pembrolizumab for dMMR or MSI-high advanced, resistant solid tumors.
What is the prevalence, clinical features and therapy response to CPI in dMMR/MSI-high prostate cancer? The largest case series to date examined tumors from 1346 patients with prostate cancer by targeted sequencing using a clinical assay [26▪▪], where MSI was quantified using a computational tool called MSIsensor. Additionally, immunohistochemistry (IHC) for MMR protein expression was performed in a subset of tumors. In total, 32/1033 (3.1%) of patients with evaluable tumor for MSIsensor analysis were MSI-high/dMMR of whom 22% had evidence of germline mutations in Lynch syndrome-associated genes. Prior studies indicate that the frequency of dMMR/MSI-high prostate cancer does not appear to differ between primary versus metastatic/resistant tumors [24▪,27]. Uniquely, 11 patients in the cohort with MSI-high/dMMR CRPC received CPI therapy, of whom more than half achieved a PSA decline of 50% or greater on therapy. Both radiographic and durable clinical responses were observed. These findings were recapitulated in an independent cohort of 13 patients with metastatic prostate cancer and deleterious tumor MMR mutations . Prolonged responses to hormonal therapy were observed despite aggressive clinicopathologic features, and two of four patients who received CPI therapy achieved a PSA response, with a median progression-free survival of 9 months among the CPI-treated.
Although increased neoantigen burden related to mismatch repair deficiency may explain sensitivity to CPI, Rodrigues et al. [29▪▪] sought to dissect the immunogenomic complexity of dMMR/MSI-high prostate cancer using an integrated, multiomic approach to refine biomarker-guided selection for immunotherapy. In total, 8.1% of advanced prostate cancer patients exhibited dMMR/MSI (by IHC and PCR, respectively) and these tumors were enriched for tumor-infiltrating T lymphocytes, PD-L1 protein expression and poorer clinical outcomes. Importantly, this study highlighted the significant degree of discordance between conventional and orthogonal approaches of detecting dMMR/MSI in prostate cancer (e.g. IHC; PCR; MMR gene mutation by sequencing) and suggests that a dMMR-associated DNA signature as the phenotypic readout of these alterations may perform better as a predictive biomarker.
Beyond the small subset of patients with dMMR/MSI-high prostate cancers (typically associated with a high rate of somatic mutation), it is currently unclear whether the larger set of patients with mutations in other genes involved in the DNA damage response (DDR) are enriched for responders to CPI, and studies of CPI with or without poly (ADP-ribose) polymerase inhibition are in progress [30,31]. Recent reports have described other somatic genetic alterations that may correlate with CPI responsiveness. Biallalic CDK12 loss is enriched in metastatic CRPC and is associated with a distinct mutational signature from MMR-deficient and homologous recombination-deficient tumors, and such tumors are marked by genomic instability, increased gene fusion-induced neoantigen burden and high inferred immune infiltration [32▪]. Two of four CDK12-mutant patients treated with a PD-1 inhibitor monotherapy in this study exhibited exceptional PSA responses. Recently, a case report of a patient with metastatic CRPC and an isolated somatic POLE V411L mutation, which is associated with extreme hypermutation despite microsatellite stability, detailed exceptional response to pembrolizumab without evidence of disease progression after 24 cycles of therapy . Collectively, these data suggest that in prostate cancer, DNA-based biomarkers may have an important role in defining discrete molecular/mutational subtypes likely to respond to immunotherapy.
Beyond single gene mutations, global tumor mutational burden (TMB) is also promising as a predictive biomarker of response to CPI therapy. There is a significant correlation between TMB and response to anti-PD-1 or anti-PD-L1 therapy across tumor types  as well as improved survival among CPI-treated patients . Responses to the combination of ipilimumab with nivolumab in mCRPC appeared to be enriched in patients with high TMB , but these findings will require validation in a larger cohort. Furthermore, non-DNA-based biomarkers are being studied and will provide additional layers of biological information for integration, in order to improve our understanding of the complexity of tumor-immune interactions and to develop robust, multimodal biomarkers. For example, TMB and a T-cell-inflamed gene expression profile (GEP) exhibited joint predictive utility in identifying responders and nonresponders to the PD-1 antibody pembrolizumab across tumor types . The role of tumor immune infiltration, tumor immune-related transcriptomic signatures (both activating and repressive) [29▪▪,37], PD-L1 expression and host immune factors (including the dynamic and modulatory effect of the microbiome) are all active areas of investigation across cancer types.
Continued efforts to develop novel therapeutic immune-targeting agents and test rational combinations in the clinic, hold the promise of improving the efficacy of immunotherapy across the clinical spectrum of prostate cancer. In parallel, there is a clear unmet need to better understand the complexity of the prostate cancer immune microenvironment as well as train and validate predictive biomarkers to improve the precision of patient selection for existing and novel therapies. Both efforts will necessarily inform each other in the pursuit of effective and biomarker-driven immunotherapy for prostate cancer.
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Conflicts of interest
There are no conflicts of interest.
REFERENCES AND RECOMMENDED READING
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