Abstract
Background: Prostate cancer (PCa) is a malignancy with significant immunosuppressive properties and limited immune activation. This immunosuppression is linked to reduced cytotoxic T cell activity, impaired antigen presentation, and elevated levels of immunosuppressive cytokines and immune checkpoint molecules. Studies demonstrate that cytotoxic CD8+ T cell infiltration correlates with improved survival, while increased regulatory T cells (Tregs) and tumor-associated macrophages (TAMs) are associated with worse outcomes and therapeutic resistance. Th1 cells are beneficial, whereas Th17 cells, producing interleukin-17 (IL-17), contribute to tumor progression. Tumor-associated neutrophils (TANs) and immune checkpoint molecules, such as PD-1/PD-L1 and T cell immunoglobulin-3 (TIM-3) are also linked to advanced stages of PCa. Chemotherapy holds promise in converting the “cold” tumor microenvironment (TME) to a “hot” one by depleting immunosuppressive cells and enhancing tumor immunogenicity. Summary: This comprehensive review examines the immune microenvironment in PCa, focusing on the intricate interactions between immune and tumor cells in the TME. It highlights how TAMs, Tregs, cytotoxic T cells, and other immune cell types contribute to tumor progression or suppression and how PCa’s low immunogenicity complicates immunotherapy. Key Messages: The infiltration of cytotoxic CD8+ T cells and Th1 cells correlates with better outcomes, while elevated T regs and TAMs promote tumor growth, metastasis, and resistance. TANs and natural killer (NK) cells exhibit dual roles, with higher NK cell levels linked to better prognoses. Immune checkpoint molecules like PD-1, PD-L1, and TIM-3 are associated with advanced disease. Chemotherapy can improve tumor immunogenicity by depleting T regs and myeloid-derived suppressor cells, offering therapeutic promise.
Introduction
Prostate cancer (PCa) is considered a malignancy with a high degree of immunosuppression and restricted capacity for immune activation. This is primarily attributed to the low levels of activated cytotoxic T cells, hampered antigen-presenting processes, and elevated concentrations of immunomodulatory molecules, including immunosuppressive cytokines and immune checkpoint molecules [1]. Furthermore, these factors are supported by a significant recruitment of immunosuppressive cell populations, further exacerbating the inhibitory tumor microenvironment (TME) [2].
As shown in Figure 1, various types of immune cells make up the unique tumor-immune microenvironment (TIME) in PCa and the infiltration with tumor-infiltrating lymphocytes (TILs) is usually accompanied by an infiltration with tumor-associated macrophages (TAMs) [3]. The repertoire of tumor-specific antigens (TSAs) that demonstrate higher specificity for PCa remains relatively modest [4]. Biomarkers such as prostate-specific antigen (PSA), prostate-specific membrane antigen (PSMA), and prostate stem cell antigen (PSCA) are recognized for their diagnostic utility in PCa screening [5, 6]. In contrast, a broader range of pan-carcinoma TSAs, such as Globo H, GM2, B7-H3, and the human six-transmembrane epithelial antigen of the prostate (STEAP) protein family, is commonly expressed across multiple malignancies, including PCa [7‒10].
PCa diagnosis is primarily based on microscopic evaluation. Although various diagnostic and supportive criteria have been suggested, only a few are truly specific to PCa. The most dependable indicators include an infiltrative growth pattern, prominent nucleoli, and the absence of basal cells, which are best identified at high magnification [11]. Figure 2 presents histological images of PCa from patients with Gleason scores 3 and 4.
Upon closer examination of the immunosuppressive immune populations that infiltrate PCa, it becomes evident that these populations predominantly comprise regulatory T cells (Tregs), myeloid-derived suppressor cells (MDSCs), and TAMs that exhibit the M2-like polarization phenotype [12]. The activity of cytotoxic T cells, crucial for tumor cell destruction, is suppressed by each of these cell populations.
PCa is often considered immunologically “cold,” primarily due to its low mutational burden, minimal T-cell infiltration, reduced inflammatory signaling, and the presence of immune checkpoint molecules [1]. In contrast, “hot” tumors typically exhibit high levels of TILs, an abundance of cytokines and chemokines that attract immune cells, and a high mutational load, which results in the generation of numerous abnormal proteins that can be targeted by the immune system [13]. Nonetheless, despite these characteristic features of PCa, the TME is far from inert and possesses a degree of complexity that could affect its immunogenic potential. Several critical components are involved, including TAMs, MDSCs, and locally synthesized and released cytokines and chemokines (Fig. 1), which collectively shape the interaction between the tumor and the immune system. Moreover, emerging evidence has shown the existence of tertiary lymphoid structures (TLSs) within the PCa tumors. These structures, in conjunction with the identification of novel TSAs, hold substantial promise in furthering our understanding of patient prognoses and allowing the development of novel immunotherapeutic strategies in the management of PCa. In this comprehensive review, we aim to elucidate the delicate balance between immune cells and tumor cells within the TME of PCa.
Immune Cell Subpopulations of PCa
Both adaptive and innate immune responses operate concurrently in PCa. Nonetheless, owing to the low immunogenic profile and relatively slow progression of PCa, there are only a few antigens present in the tissue. As a result, the activation and recruitment of T cells are limited [14]. Genetic analyses of the tumor cells demonstrated that approximately 90% of prostate PCa manifest a state of immunological ignorance. This implies that genes associated with the cancer immunity cycle exhibit a lack of upregulation within the tumor tissue [15]. The cancer immunity cycle can be explained in seven pivotal steps, starting with the release of antigens from the cancer cell and culminating in the targeted elimination of cancer cells (Fig. 3). Intermediate stages of this cycle include antigen presentation, priming and activation of T cells, their subsequent trafficking to tumor sites, infiltration into tumor tissue, and specific recognition of tumor antigens by T cells. To effectively eliminate cancer cells, it is critical to upregulate this cycle within the TME [14].
Mirroring the mechanisms observed in inflammatory states, tumors exhibit infiltration by a broad spectrum of immune cell populations [16]. The immune response against PCa cells is initiated by the detection of PCa antigens by antigen-presenting cells (APCs), such as dendritic cells (DCs) [2]. Mature APCs are characterized by elevated expression levels of major histocompatibility complex (MHC) class II, alongside with the upregulation of surface molecules CD86 and CD80, which serve as costimulatory molecules [17, 18]. The priming of both CD4+ and CD8+ naive T cells predominantly falls under the purview of professional APCs with DCs playing a crucial role in this process. The activation of T cells mediated by DCs necessitates direct physical interaction between these cellular entities, wherein DCs present the antigen within the framework of MHC, augmented by the synergistic influence of cytokines and additional costimulatory surface molecules [19]. To facilitate the activation of CD4+ T cells, DCs internalize exogenous peptides and subsequently present them via MHC-II molecules [20]. While CD4+ T cells activate other cellular components of the immune system, such as CD8+ T cells and macrophages, they also assist B cells in isotype switching and affinity maturation. Especially important is the interaction between CD40 on B cells and CD40 ligand on follicular helper T cells, enhancing antibody diversity and specificity [21]. CD8+ T cells, once activated, release cytotoxic granules, express death receptor ligands and produce cytotoxic cytokines [22].
So far, T cells have received the most extensive exploration in terms of their presence within the prostate TME [23, 24]. The infiltration of PCa with cytotoxic CD8+ T cells has consistently demonstrated associations with favorable survival outcomes across diverse studies. Similarly, to other solid tumors, both overall survival (OS) and recurrence-free survival were found to be related to high CD8+ T cell infiltration [25, 26]. Further research has also underscored the correlation between high loads of CD8+ T cells and the improvement in progression-free survival among PCa patients [26]. A study by Petitprez et al. [27] demonstrated a reduced risk of metastases in PCa cases exhibiting robust infiltration by CD8+ T cells. Nevertheless, the phenotypic profiles of CD8+ TILs, with a particular focus on the expression of immune checkpoint molecules, is of major importance, as elaborated in the next sections of this review.
The CD4+ T cell subset is remarkably diverse. Th1 cells have been shown to be particularly beneficial for PCa patients and associated with positive outcomes [28]. On the other hand, individuals with higher loads of CD4+ T regs in the TME face a higher risk of PCa-related mortality. In fact, the risk of death is nearly twice as high in patients with higher proportions of CD4+ T regs [23].
T regs exhibit multifaceted mechanisms to suppress immune responses. They achieve this through various pathways: secreting immunosuppressive cytokines, such as TGF-β, IL-10, and IL-35; inducing cytotoxic effects via perforin/granzyme B, Fas/FasL, and galactin-1 in target cells; and metabolizing ATP into adenosine through the enzymatic actions of CD39/CD73. Moreover, Tregs impact DCs by employing direct suppression, releasing soluble factors, and interfering with costimulatory pathways. These actions involve molecules like cytotoxic T lymphocyte antigen-4 (CTLA-4), upregulation of indoleamine 2,3-dioxygenase, and heightening CD25 affinity for IL-2 [29, 30]. According to Karpisheh et al. [29] inhibiting the functions of T regs or reducing their numbers may be a viable strategy for immunotherapy in PCa. This could be achieved through nonspecific targeting using cyclophosphamide or tyrosine kinase inhibitors, as well as blocking chemokines or their receptors. T regs can be depleted using approaches such as anti-CD25 antibodies or fusion protein denileukin diftitox. Furthermore, therapeutic interventions targeting anti-CTLA-4, anti-PD-1, or inhibitors of glucocorticoid-induced TNFR-related and T-cell immunoglobulin and mucin domain 3 (TIM-3) exhibit notable promise in the context of T reg modulation. Currently, several primary subtypes of Tregs are recognized, such as effector Tregs (eTregs) and naive Tregs, each distinguished by their specific function and phenotype [31]. The master gene regulating the function of T regs is the transcription factor FOXP3. Elevated infiltration of FOXP3+ cells of tumors has been associated with unfavorable clinical outcome across various cancer types [32]. Flamminger et al. [33] discovered a significant association between elevated counts of intratumoral FOXP3+ T regs and a more advanced stages of PCa. C-C chemokine receptor 4 (CCR4) is predominantly expressed on eTregs and plays a crucial role in the chemotactic migration of eTregs into tumor tissues. Watanabe et al. [31] revealed that PCa patients with high levels of CCR4+ T regs infiltrating the prostate tissue had poor prognostic outcomes and the degree of infiltration was directly associated with the patient’s prognosis. Furthermore, these patients showed an increased likelihood of disease progression to castration-resistant PCa (CRPCa).
Research has also revealed that in patients with PCa, there is a tendency toward the differentiation of T cells into Th17 phenotype, known for its inflammatory nature and production of interleukin-17 (IL-17) [24]. The role of Th17 cells appears to be cancer-type dependent [34]. Research indicates a dual role of IL-17 in tumorigenesis. Some studies identify its tumor-suppressive function, while others highlight a positive correlation between IL-17 levels and the malignancy’s aggressiveness [35]. In a murine study on PCa, aged mice exhibited enlarged prostate glands, accompanied by heightened inflammatory cell infiltration and increased levels of Th17 cytokines within prostate tissue compared to younger mice. Additionally, factors released by aged CD4+ T cells, particularly those derived from ex vivo differentiated Th17 cells, notably enhanced PCa cell viability, migration, and invasion [36]. Another studies have demonstrated that IL-17 blockade halts disease progression [34, 37].
In contrast to T cells, which serve as a relatively minor component of the PCa TME, TAMs constitute the predominant nonneoplastic cell population within prostate tumors [38, 39]. TAMs have the capacity to constitute up to 50% of the infiltrative cell population within the tumor mass, highlighting their significant role in shaping the TME, as shown in Figure 4 [40]. Elevated levels of TAMs are mostly associated with unfavorable prognostic outcomes [41‒43]. However, TAMs possess the capability to exhibit both antitumor and pro-tumor activity depending on the conditions within the microenvironment [44]. This is mostly dependent on tissue-specific regulatory factors and the stage of tumor progression [45]. However, an increased load of TAMs was shown to facilitate metastatic spread and tumor growth in PCa [40]. Moreover, TAMs contribute to immunosuppression within the TME and therapeutic resistance [40, 46]. Several studies have indicated that elevated levels of TAMs served as markers for metastatic progression and accelerated tumor growth and increased number of TAMs compromised the efficacy of antitumor therapies [40]. A substantial rise in density of CD68+ macrophages was observed in cases associated with elevated Gleason scores [47]. Interestingly, another study demonstrated that the quantity of CD68+ TAMs across total tumor tissue exhibited a negative correlation with the TNM clinical stage, while within the cancer cell area, TAM density showed a direct positive correlation with Gleason score [48].
A study conducted by Erlandsson et al. [49] revealed that patients exhibiting elevated levels of M2 macrophages faced nearly double of the mortality risk compared to those with diminished levels. This adverse effect was attributed to the immunosuppressive functions of M2 macrophages, which predominantly produced the anti-inflammatory cytokine, TGF-β [50]. Several studies have also observed a correlation between M2 macrophages and Treg activation [49, 50]. This intriguing interrelationship was shown to be mediated through diverse biochemical pathways. One of these pathways involves T regs suppressing the secretion of IFNγ by CD8+ T cells, subsequently enhancing macrophage activity [51].
While the majority of TAMs in PCa exhibit the M2-like phenotype, it is crucial to acknowledge the presence of M1-like macrophages, which manifest pro-inflammatory and antitumor properties [52]. Those are facilitated through the recruitment of Th1 cells via the secretion of IFN-γ and TNF-alpha [53]. The role of M1-like macrophages is critical when discussing TAMs due to the plastic nature of macrophages; these cells exist on a functional spectrum and can transition from an M1 to M2 phenotype under specific conditions, such as hypoxia, thereby exerting different functional roles [44].
In a murine model, Di Mitri et al. [38] evaluated a therapeutic strategy aimed at targeting TAMs through the use of a CXCR2 selective antagonist. This approach seeks to harness the anti-tumorigenic potential of macrophages specifically in Pten-null PCa. Masetti et al. [39] focused on macrophage lipid metabolism pathways in PCa. The subset of TAMs exhibiting dysregulated transcriptional pathways associated with lipid metabolism demonstrates a positive correlation with the progression of PCa and shorter disease-free survival. This subset is characterized by an accumulation of lipids dependent on macrophage receptor with collagenous structure (MARCO). The authors revealed that inhibiting lipid accumulation through MARCO blockade not only hinders tumor growth and invasiveness but also improves the effectiveness of chemotherapy in mice models. Interestingly, the introduction of a fatty diet to tumor-bearing mice results in an increased number of lipid-loaded TAMs [39]. Similar to TAMs, tumor-associated neutrophils (TANs) exhibit dual functionality within the TME. This is ascribed to their inherent cellular plasticity [54]. Depending on the cytokines present, TANs can differentiate into either antitumoral (TAN1) or pro-tumoral (TAN2) phenotypes. Specifically, granulocyte colony-stimulating factor and TGFβ are involved in the formation of TAN1 and TAN2 subtypes, respectively [55]. The antitumoral actions of TANs are mediated through multiple pathways. First, TANs release reactive oxygen species, which exert a direct cytotoxic effect on tumor cells. Second, they participate in antibody-dependent cellular cytotoxicity by binding to opsonized tumor cells. Finally, TANs activate various components of both the adaptive and innate immune system, thereby contributing to an integrated anti-tumoral response [56, 57]. Trogoptosis represents another important mechanism through which TANs are capable of eliminating antibody-opsonized tumor cells (Fig. 5) [58]. According to Costanzo-Garvey et al. [59], bone-resident neutrophils exert an inhibitory effect on PCa, while metastasis to the bone can advance by avoiding neutrophil-mediated killing. Thus, enhancing neutrophil cytotoxicity within the bone might offer a new therapeutic strategy. Alsamraae et al. [60] reported that second-generation androgen-deprivation therapy suppresses the cytotoxicity of peripheral-blood neutrophils through increased transforming growth factor beta receptor I (TGFβ receptor I). The administration of high-dose testosterone and the inhibition of TGFβ receptor I was shown to reverse androgen receptor (AR)-mediated neutrophil suppression, thereby reinstating the anti-tumor immune response of neutrophils.
TANs may also facilitate pro-tumoral activities through an array of mechanisms that contribute to tumor progression, angiogenesis, and immunosuppression [56]. Moreover, through trogocytosis, TANs may limit the efficacy of therapeutic monoclonal antibodies [61].
Given the plasticity of TANs, their roles within the TME require further investigation for potential therapeutic applications [62]. When exploring peripheral blood for prognostic or predictive neutrophil-associated markers in PCa, the neutrophil count emerged as an independent prognostic factor, distinctly linked to overall mortality in PCa [63]. While the role of natural killer (NK) cells in the context of PCa remains underexplored, extant literature does indicate a positive correlation between elevated levels of CD56+ NK cells in peripheral-blood samples, robust NK cell infiltration within PCa tumors, and favorable prognostic outcomes [64‒66]. Tumor-associated NK cells may act as effector cells capable of supporting angiogenesis in PCa through direct interactions with endothelial cells and by influencing macrophage polarization [67]. A study by Pasero et al. [64] corroborated these observations, indicating an improved prognostic outlook for patients with elevated levels of NKp30 and NKp46 cytotoxic activity. A high-dose androgen, acting through the AR, can hinder NK cell cytotoxicity toward enzalutamide-influenced cell lines via the circFKBP5/miR-513a-5p/PD-L1 pathway. Consequently, targeting PD-L1 with an anti-PD-L1 antibody or reducing PD-L1 expression could amplify the inhibitory impact of high-dose androgen treatment, leading to enhanced suppression of CRPC cell growth by boosting NK cell cytotoxicity [68].
MDSCs represent a distinct population of myeloid cells with immunosuppressive functions that can be derived from both granulocytic and monocytic lineages [69]. In the context of human physiology, two primary subclasses of MDSCs are recognized: monocytic (M-MDSC) and polymorphonuclear or granulocytic (PMN-MDSC). These cells can be immunophenotypically distinguished through specific marker expressions. Both subsets express the myeloid lineage marker CD11b, while CD14 is exclusively expressed in M-MDSCs. PMN-MDSCs are uniquely characterized by the expression of CD15 and CD66b markers. The involvement of these MDSCs in the TIME largely attenuates the anti-tumor efficacy of cytotoxic T cells [70]. In patients with PCa, a notable increase in various MDSC subtypes in the peripheral blood was identified compared to healthy individuals. Their baseline absolute counts, along with their alterations during treatment, are being investigated as potential predictive and prognostic biomarkers [71]. In a study of 23 PCa patients, the CD14+HLA-DR− cell subset was higher than in healthy individuals and significantly decreased after prostatectomy [72]. Studies have demonstrated that the monocytic fraction of MDSCs can suppress the proliferation of autologous T cells [73, 74]. There is also an observed correlation between MDSC levels and circulating PSA levels [74]. Recent evidence suggests that the monocytic subset of immunosuppressive MDSCs correlates with increased disease severity and poorer clinical outcomes in PCa patients. Another study highlights the granulocytic subset as the predominant determinant of prognostic implications [71]. However, the debate about which MDSC subpopulations have the most significant clinical relevance continues [71].
B cells exhibit versatility, engaging in immune responses through various mechanisms. Intratumoral PCa regions were shown to contain higher B cell proportions compared to extratumoral benign prostate tissue in sections stained for the CD20 marker [75]. Weiner et al. [76] concluded that prostate tumors in Black men or men of African ancestry exhibit heightened plasma cell infiltrate, enhanced markers of NK cell activity, and increased IgG expression. These observations correlated with improved recurrence-free survival post-surgery, suggesting that plasma cells play a crucial role in driving immune responsiveness in PCa. Ammirante et al. [77] concluded that the development of CRPCa in mice is accelerated by inflammatory processes involving B cells recruited into androgen-deprived PCa through the chemoattractant C-X-C motif chemokine 13 (CXCL13). Furthermore, interventions that inhibit CXCL13 expression, such as immunodepletion of myofibroblasts, blockade of TGF-β signaling, and the use of phosphodiesterase-5 inhibitors, can effectively hamper the recruitment of B cells into androgen-deprived prostate tumors. Additionally, these treatments mitigate the development of a more aggressive cancer phenotype [78]. Shalapour et al. [79] showed that B cells in mice play a role in modulating the response to low-dose oxaliplatin. In three distinct mouse models of PCa, oxaliplatin showed limited efficacy unless B cells were genetically or pharmacologically depleted. The removal of IgA-producing plasmocytes commonly found in therapy-resistant PCa in humans, supports the CD8+ T cell-dependent eradication of tumors treated with oxaliplatin. Distinguishing between immunosuppressive and immune-supportive B-cell phenotypes is crucial, as this differentiation is not clear and may contribute to the negative perception of B cells in the context of PCa [80].
Mechanisms Underlying Immunosuppression in PCa
Tumor cells can escape the immune system through various mechanisms of immunosuppression, including the expression of immune checkpoint molecules, restriction of antigen presentation, secretion of immunosuppressive molecules, recruitment of immunosuppressive cells, or downregulation of tumor antigens [81].
Expression of Checkpoint Molecules and Therapeutic Implications
Prostate tumor cells have the ability to attenuate anti-tumor immune responses through the upregulation of specific immune checkpoint molecules (Fig. 6). Among these proteins are programmed cell death protein 1 (PD-1) and programmed death-ligand 1 (PD-L1). The receptor for PD-L1, programmed cell death protein 1 (PD-1), is not only located on the surface of activated T cells, but is also detected on various subtypes of leukocytes [82]. Recent research findings indicate that the majority of CD4+ and CD8+ T cells found in PCa express PD-1 [83]. Once PD-L1 binds to its corresponding receptor, the activation signals through T cell specific receptors are consequently decreased [82]. Notably, expression of this protein is often observed in various human malignancies and has been established as a powerful prognostic marker [84]. Gevensleben et al. [85] provided first evidence that PD-L1 is not just highly prevalent in PCa but also serves as an independent prognostic marker for biochemical recurrence. Another study demonstrated that PD-1 promoter hypermethylation was inversely related to PD-1 mRNA expression and positively associated with higher preoperative PSA score, Gleason grade and was linked to biochemical reoccurrence [86]. A study by Petitprez et al. [27] established that higher load of PD-L1+ correlated with more advanced stages of PCa. Complementing these findings, another study has also found a correlation between the progression of PCa and increased presence of PD-L1+ tumor cells [79]. These findings cumulatively underscore the significance of the PD-1/PD-L1 axis, emphasizing its profound impact on disease progression in patients with PCa.
Another example of immune checkpoint molecules is CTLA-4, which is also localized on the surface of T cells. Upon engagement with its ligands, CD80 and CD86, CTLA-4 induces the suppression of T cell activation by opposing the action of CD28-mediated co-stimulation which is necessary for effective T-cell activation, proliferation, differentiation, and effector function [87]. High expression of CTLA-4 has been observed in cases of prostate adenocarcinoma. Moreover, the expression and dynamics of CTLA-4 molecule was significantly different when comparing tumor tissue to normal tissue [88, 89]. Although CTLA-4 is generally considered a cell surface receptor, studies have shown that it can also be produced as naturally secreted soluble molecules by certain cells [90]. In a cohort study conducted by Wang et al. [89] it was concluded that serum levels of CTLA-4 exhibited a correlation with both the risk of disease progression and the incidence of biochemical recurrence in patients with PCa. Anti-CTLA-4 has been observed to promote the increased expression of the immune checkpoints PD-1 and PD-L1, which suggests a potential synergy in employing a dual blockade strategy [91].
T-cell immunoglobulin-3 (TIM-3), also known as Hepatitis A virus cellular receptor 2 (HAVCR2), along with lymphocyte activation gene-3 (LAG-3) have been identified as critical regulators of the immune response [92]. TIM-3 serves as expression inhibitor of cytokines such as IFN-γ and TNF, which simultaneously attenuate type 1 immune responses [93]. TIM-3 has been identified as a negative regulator of CD8+ cytotoxic cells and CD4+ T helper cells. Increased expression of TIM-3 has been documented across various malignancies, primarily due to its effect on cancer stem cells and functionally exhausted T cells [92]. Piao and Jin [94] demonstrated that positive staining of TIM-3 was observed in PCa. Furthermore, little or no staining of TIM-3 was observed in benign prostate hyperplasia epithelium. This study further revealed that TIM-3 expression in CD4+ and CD8+ T cells could potentially signify disease progression in PCa [94].
Increased expression of LAG-3 is necessary to mitigate excessive lymphocyte activation and to prevent the onset of autoimmunity. When LAG-3 engages with its ligands, typically MHC class II molecules, it delivers inhibitory signals that dampen T-cell activation and function [92, 95]. In PCa, LAG-3 expression levels were shown to be relatively high, especially when exposed to the pro-inflammatory cytokine IL-6.
The V-domain immunoglobulin suppressor of T-cell activation (VISTA) has been well-documented as a pivotal immune regulatory receptor. Current preclinical research suggests that its impact on cancer immunity is more complex than earlier recognized [96]. The use of ipilimumab (anti-CTLA-4) or nivolumab (anti-PD-1) as monotherapy has not shown significant clinical benefits in PCa patients. Gao et al. [91] suggested that VISTA could serve as a compensatory inhibitory pathway in prostate tumors after ipilimumab therapy [97]. In the CT26 colorectal cancer model, anti-VISTA increased stimulated antigen presentation and reduced myeloid-mediated suppression. Single-cell RNA sequencing of CD8+ T cells revealed distinct pathways induced by anti-VISTA, different from anti-PD-1. Unlike anti-CTLA-4/PD-1, anti-VISTA promoted costimulatory genes and reduced T-cell quiescence regulators [98].
The B7 homolog 3 protein (also known as B7-H3 or CD276) belongs to the B7 family and holds promise as a target for cancer immunotherapy. Its considerable expression within tumor tissues, minimal occurrence in healthy tissues, and its influential role in shaping the TME highlights its potential as a target for immunotherapeutic interventions [99, 100]. Zang et al. [101] demonstrated elevated expression of B7-H3, along with another member of the B7 family of molecules, B7x, in PCa. This heightened expression was associated with extracapsular spread, seminal vesicle invasion, metastasis, hormone resistance, and increased mortality. Another study by Shi et al. [102] showed that inhibition of B7-H3 combined with blockade of PD-L1 or CTLA-4 resulted in durable anti-tumor effects and demonstrated curative potential in PTEN/TP53-deficient preclinical CRPCa model.
Nevertheless, immune checkpoint inhibitors (ICIs) are not yet part of standard therapeutic portfolio for PCa. Ipilimumab, a human monoclonal antibody against CTLA-4, has been studied in a phase III randomized trial enrolling 799 patients with metastatic castration refractory PCa (mCRPC) progressing on or after docetaxel chemotherapy. Prior to initiating treatment with ipilimumab, radiotherapy was administered to a bone metastasis (8 Gy). Although the median OS was similar in the ipilimumab arm and the placebo arm, subgroup analysis indicated that patients with an indolent course of the disease may derive some clinical benefit from the treatment [103]. Several strategies involving ICI have failed in PCa, including the combination of ipilimumab and nivolumab in a large, randomized phase II study CheckMate 650, and the phase III studies with pembrolizumab + docetaxel (Keynote-921), nivolumab + docetaxel (CheckMate-DX), enzalutamide + atezolizumab (IMbassador-250) and enzalutamide + pembrolizumab (Keynote-641) in the mCRPC setting [104‒107].
Impairment of Antigen Presentation
Major histocompatibility complex class I (MHC-I) plays an important role in the presentation of antigens to T cells. Downregulation of MHC-I represents another mechanism employed by solid tumor cells to evade immune surveillance and counteraction [108]. A low MHC-1 phenotype has been observed across several human malignancies, including colorectal, breast, and prostate tumors [109, 110]. The expression of MHC-I within these solid tumors may differ significantly, indicating variable levels among distinct tumor cells. Furthermore, MHC-I expression may change as the tumor undergoes progression and metastasizes to secondary locations. MHC-low tumors, encompassing PCa, typically contain fewer TILs. Downregulation of MHC-I may thus be associated with a more adverse clinical trajectory [109, 110]. According to Rodems et al. [111], significant decrease in the expression of MHC-I was found to be correlated with a shorter interval to biochemical recurrence after radical prostatectomy. MHC-I may be transcriptionally downregulated in a particular subset of primary prostate tumors as well as in the majority of metastatic PCa. This downregulation appears to be a risk factor for biochemical recurrence [111].
The diminished capacity for antigen presentation in MHC-I-deficient tumor cells has been linked to heightened resistance against certain immunotherapeutic strategies, especially antibodies targeting the T-cell inhibitory receptors CTLA-4 and PD-1 [108]. Mutations in genes integral to the MHC-I antigen-processing pathway have emerged as a frequent cause of resistance to ICIs [112, 113].
The Immunosuppressive Nature of Cytokines and Chemokines in PCa
Interleukins (ILs) are specialized cytokines that play essential roles in communication between various immune cells [114]. These molecules also contribute significantly to the pathophysiology of tumor progression and carcinogenesis. Several IL superfamilies have been associated with cancer development and progression. These include the IL-6 superfamily, the IL-17 superfamily, the IL-10 superfamily, the IL-1 superfamily, the TNF superfamily, the IL-8/CXCL superfamily, and the IL-2 (common γc) cytokine family [115, 116].
IL-1 plays an important role in both the tumor initiation and progression. Specifically, IL-1 facilitates key mechanisms, such as tumor angiogenesis, the accumulation of MDSCs, and the induction of chronic inflammation. Consequently, these activities contribute to tumor invasion and metastasis, highlighting its multifaceted impact on various stages of cancer progression [115, 117]. Lindmark et al. [118] described association between a common haplotype in the IL1RN gene, encoding IL-1 receptor antagonist (IL-1RA), and PCa in a Swedish population. Furthermore, IL-1β has been identified as an important factor in transforming bone stroma to create a niche favorable for metastasis in PCa, as well as in promoting the colonization of metastatic tumor cells in bone [119, 120].
IL-6 is commonly overexpressed across a variety of malignancies and serves as a protective factor for cancer cells. Specifically, IL-6 assists in avoiding apoptosis and mitigates DNA damage inflicted by therapeutic interventions, such as radiotherapy and chemotherapy. Through its involvement in specific intracellular signaling pathways, IL-6 creates resistance to radiotherapy, a phenomenon particularly observed in the treatment of PCa. This is mediated by the enhanced upregulation of key DNA repair-associated signaling molecules, including breast cancer susceptibility genes BRCA 1 and 2, ataxia-telangiectasia and Rad3 related (ATR) gene, and ataxia-telangiectasia mutated (ATM) gene [115, 121]. Increased serum levels of IL-6 have been observed in patients with metastatic PCa [122]. In both human and murine models of PCa, IL-6 has exhibited multifaceted effects on the proliferation of tumor cells, demonstrating a spectrum of influences on their growth dynamics [123]. Given these findings, the IL-6-mediated pathways in PCa need further investigation as potential avenues for addressing treatment resistance.
IL-8 expression may be modulated by factors like IL-1β, chemical and environmental stress, or steroid hormones, such as androgens [124‒126]. High levels of IL-8 in the serum of men diagnosed with PCa were previously correlated with unfavorable prognostic outcomes [127]. In parallel with the effects of IL-6, IL-8 serves to facilitate tumor cell proliferation while simultaneously inhibiting apoptosis in these malignant cells [128].
In contrast, IL-15 stands out for its distinct capacity to trigger cytotoxic activities of NK cells, particularly in the presence of PCa cells within the tissue [115, 121, 129, 130]. IL-15, a member of gamma chain family, holds promise as a therapeutic agent for both solid tumors and hematological malignancies [131, 132]. Sakellariou et al. [130] have demonstrated that IL-15 is capable of stimulating both the activity and proliferation of NK cells as well as CD8+ T cells in vitro upon exposure to PCa cells. For that reason, the therapeutic potential of soluble recombinant IL-15 has been under investigation across a range of malignancies [131, 133].
It has been demonstrated that AR signaling is pivotal in the development and progression of PCa. IL-23 serves as a crucial cytokine in activating the AR pathway, thereby facilitating the survival and proliferation of PCa cells even under androgen-deprived conditions. Elevated concentrations of IL-23, which originates from MDSCs, have been consistently identified in both tumor biopsies and peripheral-blood samples of individuals afflicted with CRPCa [115, 134].
Chemokines serve as immunomodulatory molecules that are secreted by both malignant cells and leukocytes, regulating the recruitment of various cell types including TAMs and Tregs into the TME. Dysregulated expression of chemokines and imbalances in homeostatic mechanisms associated with these molecules is linked to the development and progression of PCa [135]. Chemokines are also involved in establishing tumor-host interactions and help cancer cells to avoid the immune system by creating immunosuppressive TIME. This is accomplished via the recruitment of M2-like TAMs and Tregs. Both TAMs and Tregs contribute to the promotion of tumor angiogenesis and growth [115, 135].
Chemokines, such as CXCL12 (also known as stromal cell derived factor-1), CXCL8, CCL2, and CCL5, have been linked to the onset and progression of PCa. These chemokines are frequently upregulated in prostate tumor tissues. For example, CXCL12 contributes to the onset of PCa via interaction with it is cognate receptor, CXCR4, which is upregulated by progenitor cells and tumorigenic tumor associated fibroblasts in PCa [136]. CXCR4 enhances cancer cell survival and facilitates chemotactic migration of mesenchymal stem cells to the tumor site. In addition, CXCR4/CXCL12 signaling contributes to metastasis as CXCR4+ tumor cells migrate toward CXCL12 expressed constitutively by hematopoietic stem cells and stromal cells in the bone marrow [137‒139]. The significance of CXCR4/CXCL12 signaling in PCa is highlighted as in humans the CXCL12 G801A polymorphism is a risk factor for sporadic PCa [140]. Thus, both CXCL12 and CXCR4 have been proposed as potential prognostic markers/therapeutic targets [141].
CCL2 in PCa recruits CCR2+ monocytes from the circulation to the primary tumor site. These monocytes differentiate into TAMs and CCL2 further exacerbates the favoring of pro-angiogenic M2-like macrophage differentiation. This directly enhances the immunosuppressive nature of the TME. In addition, these M2 macrophages secrete CXCL12, promoting tumor survival via interaction with CXCR4 on tumor cells [141]. Furthermore, monocytes can differentiate into osteoclasts and CCL2 is therefore associated with bone metastasis in PCa. CCL2 exacerbates metastasis by activating the small GTPase Rac, which promotes the transendothelial migration of PCa cells into the bone stroma [142, 143].
In addition, CXCL8 expression is regulated by TGF-β and secreted by the primary tumor in PCa. CXCL8 has been reported to support angiogenesis during PCa via modulation of vascular endothelial growth factor (VEGF) expression by tumor cells [144]. This is highlighted as TGF-β inhibition can reduce tumor size [145, 146]. Ultimately, chemokines establish tumor-host interactions the support the development of an immunosuppressive TME in PCa.
The international phase 1 trial (NCT03177187) explored myeloid cell involvement in resistance to AR signaling inhibitors. Using a combination of CXCR2 inhibitor and enzalutamide, the trial targeted metastatic castration-resistant prostate adenocarcinoma resistant to AR signaling inhibitors. The treatment reduced circulating neutrophil levels and intratumor CD11b+HLA-DRloCD15+CD14− myeloid cell infiltration and provided durable clinical benefits. These results support the notion that senescence-associated myeloid inflammation may drive metastatic CRPC progression and resistance to AR blockade [147].
Additionally, key pro-inflammatory cytokines including interferon-gamma, TNF-alpha, and TGF-β play important roles in modulating tumor growth and metastatic potential. Nevertheless, the specific mechanism by which these cytokines and chemokines contribute to cancer pathophysiology remain incompletely understood [115, 148].
Tumor-Specific and Tumor-Associated Antigens in PCa
TSAs and tumor-associated antigens (TAAs) represent two main groups of antigens expressed by PCa cells [149, 150]. TSAs are primarily presented via MHC class I molecules on the surface of cancer cells, distinguishing them from healthy cells where such antigens are generally not expressed. TAAs, on the other hand, are expressed on both neoplastic and normal cells. However, their expression in normal tissues is rather limited [151]. Both categories of antigens significantly contribute to eliciting antitumor immune responses. PCa is characterized by a high prevalence of targetable TSAs and TAAs, making these antigens suitable targets for cancer vaccine therapies [149].
Antigens originating from prostate tissue function as activators for APCs, which subsequently initiate the activation of circulating CD8+ T cells. PSA serves as a typical example of organ-specific antigen. In patients with PCa, PSA elicits a series of events, leading to the activation of both helper and cytotoxic T cells [152]. In healthy individuals, most men exhibit functional PSA-specific CD8+ T cells, whereas in cancer patients, more than half display an impairment of CD8+ T-cell response [152]. PSA has been crucial in early detection of PCa; however, the expression of PSA in normal cells compromises its diagnostic specificity [5, 153]. Additionally, elevated PSA levels are not confined solely to PCa and can also be observed in conditions, such as benign prostatic hyperplasia [154]. To enhance the accuracy of PCa diagnosis, several new metrics have been developed that utilize derivatives of PSA. These include PSA doubling time, the percentage ratio of free to total PSA (%fPSA), free PSA fraction (fPSA), and prostate health index [5, 153].
PSMA serves as an important TAA that is notably overexpressed in PCa. PSMA is characterized as a type II integral membrane glycoprotein that possesses enzymatic activities such as carboxypeptidase and folate hydrolase, along with internalization capabilities. While its primary expression site is the prostate tissue, PSMA can also be found in other tissues such as the small intestine, proximal renal tubules, salivary glands, and neuroglia [5, 155]. Studies have demonstrated that elevated levels of PSMA are associated with increased serum PSA levels, as well as with prostate differentiation and growth. Owing to the regulatory influence of the AR on PSMA expression, PSMA levels markedly elevate during the course of androgen-deprivation therapy in patients [156‒158]. Upon expression of PSMA, there is a consequential increase in cellular folate content, which in turn facilitates the migration, proliferation and survival of PCa cells [5]. PSMA also plays a pivotal role in the metastatic processes of PCa. In a study conducted by Xu et al. [159], they found that the expression of certain genes linked to PSMA – namely MMP3, CDH6, and MTSS1 – had a negative correlation with both the stage of PCa and the levels of PSMA. This negative correlation suggests that PSMA may exert a regulatory influence on the mechanisms underlying PCa metastasis [159].
In addition to PSA and PSMA, several other antigens merit attention for their diagnostic potential. Prostatic acid phosphatase (PAP) and Prostate Stem Cell Antigen (PSCA) have been identified as viable biomarkers for the screening of PCa [160]. Employing multiple biomarkers for PCa detection can enhance diagnostic accuracy and reduce the incidence of false-negative biopsies. PSCA is a prostate-specific TAA encoded by a gene located on chromosome 8. PSCA has been found to positively correlate with advanced clinical stages and the presence of metastasis in PCa. Given that PSCA, similarly to PSMA, is present on the cell surface, it stands as a potential candidate for targeted immunotherapeutic approaches. PAP serves as a prostate tumor antigen and plays a role in regulating epithelial growth of the prostate. In healthy tissue, PAP levels are expected to be low. However, they rise in patients with PCa in direct correlation with the stage of the tumor [161]. While PAP is not exclusive to prostate tissue, its expression is elevated in prostate tissue compared to other anatomical sites.
Importantly, PAP serves as a target for Sipuleucel-T, an autologous active cellular immunotherapy. This therapy involves activating autologous peripheral-blood mononuclear cells, including antigen-presenting cells (APCs), ex vivo with a recombinant fusion protein (PA2024) [162, 163]. Leukapheresis is performed to collect the patient’s peripheral-blood mononuclear cells, and monocytes are enriched via density-gradient centrifugation. These monocytes are then incubated with a fusion protein combining granulocyte–macrophage colony-stimulating factor and PAP before intravenous administration. Once infused, they mature into functional APCs that activate PAP-specific CD4+ and CD8+ T cells, which then traffic to tumor lesions and mediate an antitumor response [164, 165]. Sipuleucel-T was shown to extend the OS of men diagnosed with CRPCa [166]. Furthermore, Fong et al. [167] demonstrated that administering Sipuleucel-T in the preoperative setting induces a local immune response. The authors observed more than a threefold increase in infiltrating CD3+, CD4+ FOXP3−, and CD8+ T cells in radical prostatectomy tissues compared to pretreatment biopsy samples [167]. Since Sipuleucel-T facilitates the trafficking of T cells to tumors, it may serve as an ideal combination partner with other immunotherapies, such as ICIrs, or with radiation therapy [168].
McNeel et al. [169] showed that a DNA vaccine encoding PAP, pTVG-HP, is safe and may be associated with an increased PSA doubling time. Phase II trial did not show improvement in survival in patients with castration-sensitive metastatic PCa, except possibly in those with rapidly progressing disease. However, imaging suggests the vaccine may have detectable effects on micrometastatic bone disease [170]. A recent study from the same research group has demonstrated that combining nivolumab with pTVG-HP vaccination in castration-sensitive, nonmetastatic PCa is safe, elicits an immune response, and prolongs the time to disease progression [171]. Another TAAs-based therapeutic vaccine, PROSTVAC, has been evaluated in phase III clinical trial. PROSTVAC is a PCa vaccine composed of a recombinant vaccinia vector with an inserted gene for human PSA and costimulatory or adhesion molecules for T lymphocytes (CD80, ICAM-1, LFA-3). PROSTVAC, following subcutaneous injection, enters DCs, triggering a potent immune response against its viral proteins and encoded tumor antigen PSA. This prompts anti-PSA cytotoxic T cells to target and destroy PCa cells, unveiling other TAAs, activating additional T cells, a process called antigen spreading, broadening the anticancer immune effect. In a phase III study, PROSTVAC was well tolerated in patients with mCRPC but failed to improve outcomes [172]. Evidence suggests that PCa vaccines could be used to activate and expand tumor-specific T cells in combination therapies with agents that counteract the tumor’s mechanisms of immune resistance [173, 174].
A broader range of pan-carcinoma TSAs, which are expressed across various malignancies including PCa, is more prevalent. According to Roth et al. [9] the cell surface protein B7-H3 is abnormally expressed in all PCas and serves as an independent predictor of cancer progression post-surgery. Liu et al. [175] found that elevated B7-H3 expression in PCa correlates with increased levels of the proliferation marker Ki-67. High Ki-67 expression was also linked to clinical progression in multivariate analyses [175]. Globo H, a glycosphingolipid antigen, is reported to be overexpressed in various epithelial cancers, including PCa [7]. Cancer cells with high levels of Globo H exhibit increased tumorigenicity and angiogenicity compared to those with low Globo H expression [176]. Ganglioside GM2 is preferentially expressed on the surface of various tumor cells and has been identified as a potential target antigen for specific immunotherapy of PCa [177, 178]. Subsequent research, however, did not yield satisfactory results, and this strategy was not pursued further [174]. Burnell et al. [8] stated that STEAP2 expression levels significantly correlated with Gleason score, while STEAP4 was a strong predictor of relapse. These findings suggest STEAP2 as a potential prognostic marker, and STEAP4 as an indicator of relapse [8]. STEAP proteins, due to their membrane-bound localization and elevated expression in cancers, including PCa, have emerged as promising targets for immunotherapy [179, 180].
The Presence and Significance of Tertiary Lymphoid Organs in PCa
The Development of Tertiary Lymphoid Organs within TME
Tertiary lymphoid organs (TLOs), also known as tertiary lymphoid structures (TLSs), are ectopic lymphoid formations that arise in regions marked by chronic inflammation or neoplastic activity (Fig. 7), [181, 182]. TLOs are unencapsulated structures, exhibiting heterogeneity of their cellular composition and maturation, which are dependent on the specific microenvironment conditions and stimuli that initiate their formation [183].
In the context of the TME, tumor-associated TLOs (TA-TLOs) display ontogenetic patterns similar to those observed in secondary lymphoid organs. TA-TLOs originate with the influence of lymphoid tissue inducer cells (LTi), but their continuous development follows a different path than that of traditional secondary lymphoid organs, removing the need for ongoing LTi intervention. Chronic inflammatory states also serve as a potent catalyst for de novo formation of TLOs [184]. Various cell types are involved in the neogenesis of TLOs. Specifically, DCs, M1-like macrophages, B cells, T follicular helper cells (Tfh), and Th17 cells are crucial contributors to this process [185]. The initiation of TLO formation is regulated by the binding interaction between the cytokine complex lymphotoxin α1β2 (LTα1β2) and its cognate LTβ receptor (LTβR). This interaction activates a signaling cascade that results in the production of numerous adhesion molecules and chemokines [185]. Prominent among these chemokines are CCL19 and CCL21, which play a crucial role in recruiting CCR7+ T cells, integral for the constitution of the T-cell zone in the TLO. Additional chemokines, such as CXCL12 and CXCL13, specifically attract B cells expressing CXCR4 and CXCR5 through the CXCL12-CXCR4 and CXCL13-CXCR5 axes [186, 187].
The signaling cascade initiated by the binding of LTα1β2 to LTβR also results in the upregulation of specific adhesion molecules, such as MADCAM1, VCAM1, and ICAM-1. These molecules are integral to the recruitment and retention of immune cells within the TLOs [185, 188]. Additionally, this molecular pathway stimulates the production of VEGF-C, which, in turn, facilitates the formation of high endothelial venules [188]. High endothelial venules have a crucial role in initiating and sustaining immune reactions. They are responsible for transporting both naive and memory lymphocytes from the circulatory system to APCs within lymph nodes, irrespective of antigen receptor specificity, under homeostatic conditions [189]. They also enable lymphocyte migration and extravasation into TLSs [188].
The Multifaceted Structure of TLOs
TLOs can be stratified based on their spatial relationship to the tumor, being either intratumoral or peritumoral [194]. Further classification of TA-TLOs can be made according to their structural characteristics. Two principal types can be delineated: the nonclassical type, which lacks distinct cellular compartments for B and T lymphocytes, and the classical type, featuring separate compartments for B lymphocytes, in association with follicular dendritic cells, and for T lymphocytes, colocalized with DCs [185]. Beyond these structural classifications, a complex array of chemokines orchestrates the recruitment of T and B cell populations. The chemokines CCL19 and CCL21 act as crucial mediators in the recruitment and homing of T-cell subsets to TLOs. Conversely, CXCL12 and CXCL13 serve as key chemokines responsible for recruitment and spacial organization of B cell populations [184]. Fibroblastic reticular cells serve as stromal elements in the T-cell compartment that maintain the structure of the TLO [195]. Within the T-cell compartment of TLOs, mature DCs expressing lysosomal associated membrane protein (DC-LAMP) are present [196]. Additionally, a specialized subset of macrophages, known as “tingible body macrophages” was identified within the TLOs. These macrophages play an important role in the efficient phagocytic removal of apoptotic cells within the microenvironment of the TLOs.
The Functional and Clinical Significance of TLO within PCa
Due to the cellular and architectural heterogeneity of TLOs, these structures can lead to both positive and negative prognostic outcomes [197]. The density and specific cellular components within the TLOs are crucial factors in determining their association with survival outcomes [198‒200]. The immunosuppressive characteristics of TLOs are attributed to the secretion of IL-10, as well as the presence of T regs [185, 201]. T regs exert a prognostically adverse influence by suppressing the antitumor immune responsiveness [202]. In a number of studies, particularly those focused on pancreatic and lung carcinomas, the TNM classification system was not associated with the predictive role of TLOs [188, 190, 203].
As previously mentioned, the prognostic significance of TLOs is influenced by a multitude of factors. In some malignancies, such as non-small cell lung cancer, evidence substantiate a positive correlation between the presence of TLOs and favorable clinical outcome [204]. Corroborating this notion, research conducted on melanomas demonstrated the correlation of the presence of CD8+ T cells and CD20+ B cells with positive prognosis [205]. In another investigation focused on colorectal carcinoma, it was revealed that the maturity of TLOs was positively associated with an improved prognosis. Importantly, this study further emphasized that the level of maturation of TLOs serves as a superior prognostic marker than their mere presence [206].
Conversely, in certain malignancies, the presence of TLOs correlates with adverse clinical outcomes. For instance, a study conducted on breast cancer patients concluded that the presence of TLOs is associated with more aggressive and invasive tumor phenotypes, thereby resulting in unfavorable prognoses [207]. Several studies focusing on pancreatic cancer and hepatocellular carcinomas have yielded consistent evidence demonstrating that intratumoral TLOs confer a significantly more favorable prognosis in comparison to their peritumoral analogs [190, 208]. Such data highlight the pivotal role of the spatial distribution of TLOs within neoplastic sites as a potential prognostic indicator for some malignancies.
While TLOs have been extensively studied across various malignancies, their specific role and significance in PCa are gaining attention. In a study by García-Hernández et al. [209], TLOs were observed across various stages of PCa, including the incipient stages of intraepithelial neoplasia. Interestingly, the presence of TLOs was also detected in patients who experienced spontaneous remission. In a murine study examining PCa with Pten-null tumorous tissue, authors found that the presence of the classical form of intratumoral TLOs potentiated the efficacy of combined therapy involving BAY1082439 and ICI immunotherapy. BAY1082439 serves as an anti-PI3K inhibitor that elicits several immunostimulatory effects. Specifically, it stimulates CXCL10/CCL5 secretion, activates IFNα/IFNγ signaling pathways, and induces the expression of β2-microglubin. This suggests potential for leveraging TLOs to enhance chemotherapy strategies [210].
The Shifting Dynamics of Tumor-Immune Interactions
It has been well-established that the TME can undergo temporal changes induced by a variety of factors. These dynamic alterations complicate our understanding of the TME, as both cell populations and cytokines are not static but constantly evolving [211]. While this presents challenges in targeting specific cells within the TME, it also offers opportunities to explore therapeutic strategies aimed at converting nonresponders to responders by inducing specific alterations in the TME.
Chemotherapy Induced Changes of the TME
Chemotherapy was shown to stimulate the immune system by directly targeting cancer or immune cells, as well as by inducing changes in overall body physiology. The “on-target” effects of chemotherapy include increasing the antigenicity or adjuvanticity of malignant cells. In contrast, the “off-target” effects may lead to an increase in effector cells, such as DCs, CD4+ Th1 cells, CD8+ T cells, NK cells, and M1 macrophages. Additionally, chemotherapy can result in a reduction of suppressor cells, including T regs, M2-like macrophages, and MDSCs [212]. Hence, the immunostimulatory effect of chemotherapeutic agents, in particular, has led to the emerge of selected chemotherapies as promising partners for combination therapies with ICIs [213].
In certain cancer types, it has been observed that despite the lack of tumor regression following chemotherapy, notable alterations in immune cell signatures occur within paratumoral tissue compartments, including adjacent lymph nodes. These alterations predominantly enhance CD8+ T-cell responses and modulate death receptor expression, thereby presenting new opportunities for optimizing chemotherapy to enhance immune-mediated tumor control [214].
In PCa, the use of chemotherapy for immunomodulation has been extensively studied as a strategy to convert a “cold” tumor into a “hot” one. A study by Vicari et al. [215] in a murine PCa model demonstrated that treatment with paclitaxel resulted in a shift of macrophage populations toward the M1-like phenotype, accompanied by a reduction in T regs. Similarly, a study by Garnett et al. [216] showed that in a murine PCa model treated with docetaxel, there was an enhanced cytotoxic T cell response and a simultaneous decrease in the frequency of MDSCs.
In humans, a study by Rozkova et al. [217] demonstrated that combining chemotherapy with immunotherapy, specifically alternating courses of chemotherapy and vaccination using mature DCs loaded with the LNCaP cell line, resulted in significant improvements in both clinical and laboratory outcomes. This approach also led to a reduction in PSA levels by over 90%.
In PCa, the observed benefits from immunomodulation by chemotherapy appear to stem largely from the depletion of T regs and MDSCs, which reduces immunosuppression, as well as the generation of additional TAAs, thereby increasing tumor immunogenicity [1]. Other studies, primarily based on case series, have shown that approved chemotherapies for the treatment of mCRPC can be safely combined with checkpoint inhibitor therapy to enhance the antitumor immune response [1].
Shaping the Tumor Microenvironment through Radiation
The immunomodulatory effects of radiation vary depending on factors such as the dose administered, the extent of fractionation, and potentially the location of metastasis [218, 219]. Prolonged or fractionated radiation can be harmful to lymphocytes, potentially reducing the ability of the immune system to affect the treatment outcome. On the other hand, findings from animal models suggest that newer approaches like hypofractionated RT offer a more favorable balance, presenting opportunities to enhance the cancer-killing effects of radiation while simultaneously boosting immune activation for improved therapeutic outcomes [220].
In a phase I/II clinical trial investigating the combination of ipilimumab and radiation, patients with mCRPC showed improved outcomes when radiation was added to checkpoint inhibitor monotherapy. Importantly, this enhancement in therapeutic benefit occurred without a significant increase in adverse events [221].
Primary versus Metastatic Tumors
The bone is the most common site for distant metastasis in patients with advanced PCa [222, 223]. The distinct bone-forming phenotype of PCa bone metastasis contributes to both tumor progression in the bone and resistance to therapy [224]. Several studies have demonstrated that patients with PCa bone metastasis exhibit elevated markers of osteoclast activity [225, 226]. Even in osteoblastic bone metastases, osteolytic events may occur. The degradation of the extracellular matrix releases bone matrix-bound growth factors. Once activated, these growth factors can be crucial for promoting the proliferation of metastatic cancer cells [227].
PAP may be a crucial factor in driving the osteoblastic phenotype and abnormal bone mineralization in PCa bone metastasis. Targeting PAP could offer a therapeutic strategy to reduce morbidity and mortality associated with osteoblastic bone metastases in PCa [228].
The overexpressed transcription factor BHLHE22, related to the transformation of advanced PCa cells, recruits protein arginine methyltransferase 5, leading to colony-stimulating factor 2 (CSF2) upregulation [229, 230]. This, in turn, drives an influx of neutrophils and monocytes, and suppression of CD4+ and CD8+ T-cell activity, fostering an immunosuppressive bone microenvironment [231]. CSF2 or protein arginine methyltransferase 5 antagonists were shown to improve ICT response by reducing the immunosuppressive bone microenvironment in BHLHE22-expressing PCa, offering a strategy against ICT resistance in advanced cases [223]. VEGF plays a crucial role in cancer dissemination to the skeleton by contributing to the development of the bone premetastatic niche, facilitating tumor cell recognition of bone, and influencing bone remodeling [232].
Conclusion
In conclusion, the immunological landscape of PCa is profoundly shaped by the dynamic interplay between the tumor and the immune system within the TME. Various components, including the proportions, phenotypes, spatial distribution of tumor-infiltrating immune cells, expression of TSAs and TAAs, cytokine profiles, and immune checkpoint molecules, dictate largely disease progression and treatment outcomes. In the PCa, most studies have centered on CD4+ and CD8+ T cells due to their remarkable anticancer properties. Nevertheless, the efficacy of these cells is often hampered within the TME because of the evasion mechanisms of tumor cells. As a result, immunosuppressive immune cell populations, including M2-like macrophages, MDSCs and Tregs, tend to dominate the PCa TME, shaping the tumor’s biological behavior. Local cytokines and chemokines play a dual role, fueling tumor progression while orchestrating the recruitment, trafficking, and activation of various immune cell components such as CD8+ T cells, NK cells, and M1-like macrophages. Furthermore, chemokines facilitate the development of TLOs within the TME of PCa and the maturation status of TLOs was shown additionally to shape both disease progression and the treatment response. The presence or absence of TSAs represents the last area with a significant impact on the tumor-immune system interplay. While the infiltration with tumor-infiltrating immune cells and their phenotype dictate the response to ICIs, the development of novel cellular-based treatments, such as adoptive cell transfer or oncolytic viruses, on the other hand, critically depend on the identification of novel TSAs. Extensive preclinical research and ongoing clinical trials aim to address these challenges, seeking breakthroughs in improving therapeutic outcomes for PCa.
This study has several limitations. First, the review discusses mechanisms observed in the TME of different cancer types to provide broader context. Mechanisms identified in other cancers may not be directly applicable to PCa, as biological differences among cancers can be disease-specific. Second, the review does not fully address the histopathological heterogeneity within PCa, as this type of cancer can exhibit a wide range of molecular subtypes, which may impact treatment responses and disease progression differently. Lastly, while the review discusses the latest data on the established elements of the TME and several emerging therapeutic approaches, a comprehensive overview of ongoing research and potential future directions for immunotherapy in PCa is still needed.
Acknowledgments
We would like to extend our sincere gratitude to Jan Balko, MD, PhD, for providing the histological images of PCa and for sharing his invaluable expertise in their interpretation and description.
Conflict of Interest Statement
The Rene Novysedlak, Robin Bartolini, Lily Koumbas Foley, Miray Güney, Majd Al Khouri, Iva Benesova, Andrej Ozaniak, Vojtech Novak, Stepan Vesely, Pavel Pacas, Tomas Buchler, and Zuzana Ozaniak Strizova declare no conflict of interest. Tomas Buchler declares following research support: AstraZeneca, Roche, Bristol Myers Squibb, Exelixis, Merck KGaA, MSD, and Novartis; consulting fees from Bristol Myers Squibb, Astellas, Janssen, and Sanofi/Aventis; payment or honoraria for lectures, presentations, speakers’ bureaus, manuscript writing, or educational events from Ipsen, Bristol-Myers Squibb, AstraZeneca, Roche, Servier, Accord, MSD, and Pfizer. All unrelated to the present paper.
Funding Sources
The study has been supported by funding from the Ministry of Health, Czech Republic – projects AZV NU23J-08-00031.
Author Contributions
Conceptualization: Rene Novysedlak, Zuzana Ozaniak Strizova, Robin Bartolini, and Tomas Buchler. Formal analysis and investigation: Zuzana Ozaniak Strizova, Rene Novysedlak, Miray Guney, Majd Al Khouri, Lily Koumbas Foley, Robin Bartolini, Andrej Ozaniak, Iva Benesova, and Tomas Buchler; writing – original draft preparation: Zuzana Ozaniak Strizova, Rene Novysedlak, Miray Guney, Majd Al Khouri, and Tomas Buchler; writing – review and editing: Rene Novysedlak, Miray Guney, Majd Al Khouri, Robin Bartolini, Lily Koumbas Foley, Iva Benesova, Andrej Ozaniak, Vojtech Novak, Stepan Vesely, Pavel Pacas, Tomas Buchler, and Zuzana Ozaniak Strizova; funding acquisition: Zuzana Ozaniak Strizova and Andrej Ozaniak; supervision: Zuzana Ozaniak Strizova.
Declaration of Generative AI in Scientific Writing
During the preparation of this work, the English language has been reviewed and improved using an AI language model, specifically ChatGPT. This tool was used solely for linguistic assistance and did not contribute to the scientific content, analysis, or interpretation of the study. After using this tool, the authors reviewed and edited the content as needed and take full responsibility for the content of the publication.