Virus against cancer: paradigm-shifting biological concepts

Abstract

Purpose of Review:

Virotherapy has emerged as a promising approach to cancer treatment. Over the past two decades, early-phase clinical trials have demonstrated the safety and promising efficacy of virotherapy in subsets of cancer patients. However, a significant knowledge gap needs to be filled to propel the field further and achieve substantial anti-cancer benefits for more than the current 10–20% of treated patients. This article reviews the most relevant current challenges in cancer virotherapy.

Recent Findings:

Recent clinical observations suggest that patients who respond to virotherapy experience a shift in their immune response from an initial or concomitant response against the virus toward the tumor.

Summary:

Strategies aimed at facilitating the temporary escape of the virus from the immune response and ultimately redirecting the immune response from the virus to the tumor may propel the development of cancer viroimmunotherapy as a potent and versatile approach to cancer treatment. Here, we examine this issue and other current challenges in cancer virotherapy.

Introduction

The concept of oncolytic virotherapy has experienced at least one critical paradigm shift since its modern inception (1). Oncolytic viruses (OVs) were discovered or developed with the intention to possess inherent or engineered tumor-selective properties, making them attractive candidates for cancer therapy. Various OVs achieve cancer selectivity by exploiting different aberrant cellular pathways that are preferentially activated in malignant cells, such as uncontrolled cellular division, disrupted apoptosis, or altered interferon response (1). Results from preclinical studies and clinical trials have later revealed that in addition to induce direct cell killing, oncolytic viruses elicit potent anti-tumor immune responses (2–7). Therefore, the success of virotherapy relies on a delicate balance between the initial viral replication within cancer cells, resulting in direct oncolysis, and the subsequent activation of the immune response, leading to tumor debulking and, in some cases, total tumor eradication. Due to this bimodal mechanism of action, OVs may represent an effective therapeutic alternative for certain solid tumors, including brain and other “cold” tumors, for which immune checkpoint blockade and other types of immunotherapies have failed to be efficacious (8). Consequently, the success of virotherapy requires a thorough understanding of the cellular interplays that lead to the development of immune responses upon tumor infection.

The anti-viral immune response has long been recognized as both a necessity and an obstacle to virotherapy. On one side, response against the virus induces inflammation, promotes antigen spreading, and triggers anti-tumor immunity; and on the other side, activation of the immune system also limits viral replication, oncolysis and virus spread. Possibly in agreement with the dual action of the immune system, reports from preclinical and clinical studies provide mixed interpretations on the role of anti-viral antibody responses during virotherapy. Some studies have suggested that virus-specific neutralizing antibodies limit the efficacy of OVs (6, 9), while others have indicated that seropositivity is associated with a better prognosis (4). Still others found no significant link between the two (10). On the other hand, we are only beginning to explore the role of anti-viral cytotoxic T cells. Clinical data have demonstrated that the intratumoral injection of OVs results in the infection of a small proportion of cancer cells, even after several rounds of replication, leaving the majority of the tumor bulk uninfected (11). Interestingly, most patients develop strong antiviral cellular immunity that eliminates virus-infected cancer cells within a few weeks of virus administration. Still, only a subset of patients develops robust and sustained anti-tumor cellular immunity sufficient to eradicate the uninfected tumor mass.

Collectively, all this evidence underscores the unique challenge in the cancer viroimmunotherapy field that arises from our currently limited understanding of the kinetics and the mechanism of the key transition from immunity against OVs to immunity against tumors, and why this shifting occurs only in a small subset of patients. Therefore, solving the “immune redirection problem” appears to be a categorical imperative to propel the next vertical advance in the field of viroimmunotherapy of cancer. In this review, we discuss the results from clinical trials that have renewed our understanding of OVs as an immunotherapeutic agent, with a particular focus on recent trials of Delta-24-RGD for brain tumors. We also describe existing challenges and potential approaches to harness anti-tumor immune responses to enhance the therapeutic efficacy of OVs.

Oncolytic Viruses as Immunotherapeutic Bioagents

In the mid-19th century, investigators already collected anecdotal evidence that natural infections coincided, in certain cases, with tumor regressions (Figure 1). Also in the 19th and early 20th centuries, the discovery of viruses ushered the initial attempts to investigate their applications in oncology (12). Recently, during the coronavirus pandemic (COVID-19), reports emerged indicating that a small subset of cancer patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) experienced a paradoxical benefit from the coronavirus infection. These patients, predominantly with liquid tumors, exhibited partial or complete remissions of their cancers without the need for chemotherapy or radiation therapy (13–15). While these cases are rare, they underscore the potential role of viral infections in triggering spontaneous tumor remissions in cancer patients, thereby reinforcing the viability of oncolytic viruses as a therapeutic strategy for cancer treatment.

Figure 1. Historical milestones in virotherapy.

Anecdotal cases of spontaneous, yet transient, tumor regressions were observed even before the discovery of viruses. Early clinical trials using non-engineered viruses for cancer therapy blossomed for a few decades following the 1950s. Still, limited efficacy and safety concerns halted the field until the advent of molecular cloning. Engineered viruses with enhanced tumor tropism were investigated starting in the 1990s, leading to the global approval of four oncolytic viruses today. * Discontinued in Latvia in 2019. [Created with BioRender.com]

In 1949, a pioneer clinical trial was conducted to test the efficacy of the Hepatitis B virus as a treatment for the lymphoma of Hodgkin (16). Four out of these 22 patients treated with human sera or tissue extract containing the hepatitis virus experienced tumor regressions. Subsequent trials explored the use of other viruses. Thus, in 1952, the Egypt 101 isolate of the West Nile virus was administered to 34 patients with advanced, treatment-unresponsive tumors, resulting in transient tumor regression in four patients (17). In 1956, adenovirus was administered to 30 patients with cervical carcinoma, causing hemorrhage and necrosis, at the tumor site in 26 patients (18). However, the immune system quickly eliminated the virus without conferring long-term survival benefits. In one of the most intriguing clinical trials conducted in 1974, the mumps virus was administered to 90 patients with 18 types of terminal cancers, including gastric, pulmonary, and uterine tumors (19). Notably, 37 patients experienced complete or at least a 50% reduction in tumor size. With strengthened regulations for injecting pathogens into patients, combined with a limited understanding of the mechanism of action of the therapies, the virotherapy field faced a temporary halt in its progress until the advent of molecular and genetic engineering.

In the 1990s, advances in recombinant technology enabled the development of cancer-selective engineered viruses (20, 21). In 2000, the first clinical trial of a genetically modified herpes simplex virus to treat human gliomas was reported (22). Over the last three decades, preclinical and clinical investigations have culminated in the approval of four OVs worldwide, as described in detail elsewhere (10): Rigvir, an oncolytic ECHO-7 virus, was approved in Latvia in 2004 for the treatment of melanoma (10); Oncolytic adenovirus H101 was approved in China in 2005 for the treatment of nasopharyngeal carcinoma (23); oncolytic herpes simplex virus (HSV) T-VEC was approved in the US in 2015 and subsequently in Europe, Australia, and Israel to treat unresectable melanoma (24); and most recently, oncolytic HSV G47Δ was granted a time-limited approval in Japan in 2021 for the treatment of recurrent glioblastoma (25).

Clinical data have repeatedly demonstrated the involvement of the immune system, including immune cell tumor infiltrations and tumor pseudo-progressions followed by complete tumor regressions. These and other data have fueled a paradigm shift in our understanding of the anti-cancer effects of OVs. Whereas OVs were initially thought to act solely by direct oncolysis, it is now accepted that the long-term anti-tumor effects of the virus are likely due to the elicitation of a cytotoxic T-cell anti-tumor immune response (1) (Figure 2).

Figure 2. Schematic representation of the historical evolution of the mechanisms of anti-cancer effect of virotherapy.

A) The initial model proposed that oncolytic viruses infect tumors (1), leading to selective virus replication and amplification of the input dose (2), with eventual viral spreading within the tumor, inducing regression (3). B) Recent preclinical and clinical data reveal a different sequence of events. Upon tumor infection (1), the virus initiates direct oncolysis, destroying tumor cells (2). This process, in turn, results in the release of tumor antigens into the tumor microenvironment, triggering inflammation and an anti-tumor immune response (3). While direct oncolysis is essential, the immune response ultimately ensures the destruction of the tumor mass. Consequently, oncolytic virotherapy is now recognized as a form of cancer immunotherapy. [Created with BioRender.com]

Our experiences similarly reflect the findings of other groups and highlight the importance of the immune system. Over the last two decades, we have developed a platform of oncolytic viruses for the treatment of brain tumors based on the backbone of an Rb-targeted virus, termed Delta-24 (26). The second generation of this adenovirus, Delta-24-RGD, developed in collaboration with the University of Alabama at Birmingham, has already been translated into the clinical setting (5–7). Thus, we completed our first-in-human Phase I clinical trial with biological endpoints of Delta-24-RGD in patients with recurrent malignant gliomas (NCT00805376) (5). In this trial, we observed durable responses (> 3 years) in 20% of patients. Subsequent animal studies evaluating the effects of Delta-24-RGD in tumor-bearing, immunocompetent mice confirmed the potential of Delta-24-RGD to induce cytotoxic CD8+ T-cell responses against tumor-associated antigens (TAA) (27). We recently completed a dose-escalation Phase 1 study of Delta-24-RGD in patients with newly diagnosed diffuse intrinsic pontine glioma (DIPG (NCT03178032) (6).) The patients received a single virus infusion through a catheter placed in the cerebellar peduncle, followed by radiotherapy. While the overall survival of patients with DIPG is limited to a year after the initiation of conventional treatment, the median survival of the 12 patients involved in the study was 17.8 months, with two patients surviving more than four years, and one patient is still alive at the moment of writing this manuscript. Importantly, we showed that the combined treatment of Delta-24-RGD and radiation therapy resulted in changes in the tumor microenvironment, including T-cell activity in these immune-desert tumors. In a third study, a multicenter Phase 1/2 clinical trial, we evaluated the combination of intratumoral delivery of Delta-24-RGD with anti-PD-1 (Programmed cell death protein 1) antibody in 49 patients with recurrent malignant gliomas (NCT02798406) (7). 56.2% of patients experienced a clinical benefit, and three patients completed treatment with durable responses, remaining alive at 45, 48, and 60 months at the time of publication. Interestingly, gene expression and immunophenotypic analyses revealed that the balance between immune cell infiltration and the expression of checkpoint inhibitors may potentially inform on response to treatment and mechanisms of resistance.

The radical shift in the perspective of the mechanism of action of virotherapy, from an exclusively direct oncolytic effect to a combination of oncolysis and anti-tumor immune response, suggested that the most relevant way to improve oncolytic virus therapy would be to enhance the “immune arm” of the anti-cancer effect. In the following section, we will dissect and discuss the main obstacles to enhancing the immune arm of oncolytic viruses and propose some potential solutions that are or should be tested in the clinical setting (Table 1).

Table 1.

Effective activation of the immune system in virotherapy: Challenges and potential solutions.

Virotherapy hurdles	Potential solutions
Immune evasion mechanisms	Combination with immune checkpoint inhibitors Cytokine-expressing oncolytic viruses Inhibition of myeloid cells and/or Tregs Combination with natural killer cells Mitigate tumor immunosuppression Combination of oncolytic viruses with tumor-antigen-based vaccines including mRNA-designed vaccines
Viral clearance and neutralization	Modification of virus to reduce immunogenicity Tolerogenic dendritic cells Chimeric virus Combination with immunosuppressant drugs Local/intratumoral administration Regional/endovascular delivery Cell vehicles
Immunogenicity and antigen presentation	Viruses expressing immune costimulatory molecules Toll-like receptors agonists Targeting dendritic cells Viruses expressing tumor antigens Mitigation of tumor immunosuppression Avoidance of combination with immunosuppressant drugs (corticoids)
Immunodominance	Use of rare viral serotypes to avoid pre-existing anti-viral immune response (avoiding the “boost” of a vaccine effect) Modification of viral dominant epitopes to reduce recognition (using chimeric viruses) Tolerogenicity of viral antigens Viruses expressing patient tumor-specific antigens Redirect the anti-tumor immune response to drivers of the neoplastic phenotype

Immune Evasion Mechanisms

Accumulating evidence has demonstrated that tumors have evolved sophisticated immune evasion mechanisms, creating a hostile microenvironment that impedes the action of oncolytic viruses. Understanding and overcoming these immune evasion mechanisms are crucial for enhancing the therapeutic outcomes of oncolytic viruses. Tumor cells often upregulate immune checkpoint molecules, such as PD-L1 (programmed death-ligand 1), to inhibit the activity of cytotoxic T cells (28). To overcome this problem, we and others have proposed combining oncolytic virotherapy with immune checkpoint inhibitors to block the PD-L1/PD-1 interaction, thereby unleashing the anti-tumor immune response (3, 7). Furthermore, tumors release immunosuppressive cytokines (e.g., TGF-β, IL-10) that create an inhibitory microenvironment, thereby impairing the function of effector immune cells (28). To counter this mechanism, investigators have generated cytokine-expressing oncolytic viruses that produce pro-inflammatory cytokines, such as IL-12 (29). These viruses can shift the balance toward an anti-tumor immune response, counteracting the immunosuppressive effects of tumor-derived cytokines.

Another main mechanism of tumor immune evasion is the recruitment (or differentiation of CD4+ cells into) of regulatory T cells (Tregs). Tregs suppress the activity of cytotoxic T cells and dampen the overall immune response (28). Although there are few strategies to counteract the effect of Tregs, it would be extremely interesting to develop oncolytic viruses that selectively deplete or inhibit Tregs within the tumor microenvironment. In addition to Tregs, tumors are characterized by the accumulation of myeloid-derived stem cells (MDSCs), which also suppress immune responses by inhibiting T-cell activation (28). Therefore, inhibiting myeloid cells or shifting their phenotype from pro-tumoral to pro-inflammatory should enhance the immune-mediated destruction of tumor cells. This combined approach may be more relevant in “cold tumors,” such as gliomas, where myeloid cells are the predominant immune population.

Another common mechanism of immune evasion is the downregulation of major histocompatibility complex (MHC) molecules in cancer cells, thereby preventing the efficient presentation of antigens to cytotoxic T cells (28). The inherent downregulating effect of some viral infections on MHC expression can further enhance this immune-evasive mechanism (30). Since NK cells recognize and kill cells that do not express HLA, the downregulation of MHC expression in cancer cells upon virus infection can be utilized as a rationale for combining oncolytic viruses with NK cells (31).

The clonal evolution of tumors and the process of tumor editing also play a significant role in tumor evasion. Thus, different clones of tumor cells characteristically exhibit antigenic heterogeneity, resulting in a diverse array of tumor antigens that also undergo changes under treatment pressure (32). These dynamic changes in antigen expression by the different clones significantly contribute to incomplete immune recognition of the tumor. As such, clonal evolution and antigen presentation evolution have become two of the major engines driving tumor recurrence mediated by immune evasion (32). To overcome this significant hurdle inherently linked to tumor evolution, some investigators have proposed developing oncolytic viruses that can target a broad spectrum of tumor antigens to eliminate as many tumor clones as possible (33). This strategy can be more effective if the targeted antigens represent proteins that function as drivers of the neoplastic phenotype. Others have proposed combining therapies that simultaneously address multiple antigens, including immune checkpoint antibodies (3, 29). This strategy aims to ensure a more comprehensive and effective immune response against heterogeneous populations of tumor cells. We should remember that overcoming the hurdles posed by antigen heterogeneity is a significant challenge for oncolytic viruses and other immunotherapies, such as monoclonal antibodies and CAR-T cells, in solid tumors.

Viral Clearance and Neutralization

The host immune system has evolved to recognize and respond to foreign and invasive pathogens at several levels. Due to these immune responses, oncolytic viruses may be dispelled before they can effectively reach and replicate in enough tumor cells to trigger a significant therapeutic effect. Furthermore, pre-existing immunity resulting from prior exposure to similar viruses may directly neutralize the therapeutic potential of oncolytic agents during the early stages of the infection cycle. For these reasons, strategies designed to evade host immune surveillance while maintaining the virus potency and specificity for tumor cells are ideal for the success of virotherapy. Understanding the mechanisms of viral clearance and neutralization is crucial for developing those strategies.

Upon the administration of oncolytic viruses, the innate immunity of the host recognizes viral nucleic acids and proteins as foreign, eventually eliciting an adaptive immune response (34). Viruses can be modified to incorporate stealth approaches that evade immune detection, including the cloning of genes encoding immune-evading proteins on the viral genome backbone (35). Riskier, yet perhaps more effective in the long term, is to attempt to induce virus tolerance. This tolerance can be achieved, for instance, by loading the host with viral proteins to numb the dendritic cells before injecting the therapeutic agent. Although never complete or perfect, inducing viral tolerance should allow the virus to persist in, replicate within, and remain in the tumor cells for extended periods before the inevitable clearance, generating a window of opportunity for eliciting an anti-tumor immune response.

Because patients recruited in clinical trials using oncolytic viruses may have pre-existing immunity to the oncolytic virus or closely related viruses, investigators have proposed utilizing chimeric serotype-switching viruses that express antigens with a low prevalence in the population (36–38). This strategy can help avoid the “vaccine effect.” Viruses can also be engineered to eliminate the dominant epitope to mitigate cross-reactivity to pre-existing immunity (39). Alternatively, to prevent the elimination of the virus in the bloodstream, oncolytic viruses can be administered intratumorally or locally, coated with liposomes or other methods, or used as cargo of cell vehicles such as neural or mesenchymal stem cells (40, 41).

In addition to antibodies, cytotoxic T cells can recognize and eliminate virus-infected cells, contributing to the clearance of oncolytic viruses. To counteract these effects, investigators have combined the virus with immunosuppressant drugs, such as corticosteroids or cyclophosphamide (42), to temporarily suppress the antiviral T-cell response during the initial stages of oncolytic virotherapy, thus allowing for effective, albeit transient, viral replication to progress. This immunosuppressant strategy may, in turn, carry some additional risks, particularly when using aggressive, highly pathogenic viruses whose effects cannot be halted with anti-viral treatments. Therefore, it requires adjusting doses and regimens to maintain a timed balance between virus propagation and immune evasion.

In the case of systemic delivery of the therapeutic agent, components of the serum can target viruses, inducing their clearance from the bloodstream. The complement system can be activated in response to oncolytic viruses, leading to their opsonization and subsequent clearance (43). This challenge might be approached by modifying the viral capsid or membranes or by using complement inhibitors (43). Local or regional delivery, as mentioned before, can also be a partial solution to this problem.

Immunogenicity and Antigen Presentation

Immunogenicity refers to the ability of a substance, such as an oncolytic virus, to induce an immune response. In oncolytic virotherapy, viruses induce immunogenic cell death within tumors, thereby triggering a robust immune response that will encounter cells expressing tumor-associated antigens and cancer neoantigens (1). Effectively presenting the cancer-related antigens to immune cells is crucial for eliciting a potent anti-tumor immune response. For that reason, incorporating co-stimulatory molecules into oncolytic viruses can enhance antigen presentation by providing a second signal that activates immune cells against the infected tumor (44). The co-stimulatory molecules, such as OX40, GITR, and 4–1BB ligands, ensure the proper activation of T cells and further promote the proliferation of intratumoral lymphocyte clones that have already interacted with cancer antigens, thereby amplifying and perpetuating the anti-tumor immune response (44).

Another possibility to increase tumor antigenicity is designing a treatment that involves vaccination of the patients with tumor antigens before virotherapy (45). In this strategy, investigators aim to educate the immune system to elicit more robust immune responses against cancer antigens, rather than viral antigens. An ideal approach would be to personalize oncolytic virotherapy based on the unique antigenic profile of individual tumors to optimize specific antigen presentation. This approach involves tailoring the oncolytic virus to express antigens specific to the tumor of the patient, increasing the likelihood of a precise immune response. The selection of antigens should be based not only on their presence in the tumor but also on their role as driver proteins for the neoplastic phenotype. When possible, the selected antigen should represent a protein required for the viability of the cancer.

In the realm of virus and vaccines, it is possible to engineer oncolytic viruses to selectively infect and activate dendritic cells against cancer cells (31). This strategy can indeed enhance the efficiency of antigen presentation and, if successful, would ensure optimal priming of T cells and promote a robust and sustained anti-tumor immune response. Another approach combines oncolytic virotherapy with immune modulators, such as checkpoint inhibitors, cytokines, or toll-like receptor agonists, which can synergistically enhance T-cell activation, immunogenicity, and antigen presentation (3, 46).

In conclusion, maximizing cancer immunogenicity and optimizing the presentation of cancer antigens are pivotal for the success of oncolytic virotherapy. Strategic engineering of oncolytic viruses and integrating complementary immunomodulatory strategies should ensure and enhance the anti-tumor immune response triggered during tumor infection, resulting in a robust and sustained anti-tumor immune response.

Immunodominance

Immunodominance refers to the selective recognition and response of the immune system to a few specific antigens, often resulting in the dominance of certain epitopes over others. In the presence of several immunogenic targets, the immune system does not react against each part of the antigenic proteins but focuses the response on a few antigens (47). These antigens are referred to as immunodominant epitopes. Thus, the immune response against one or several targets is modulated by the phenomenon of immunodominance.

In the context of oncolytic viruses, immunodominance may prevent the trigger of an immune response against the tumor, therefore affecting the overall therapeutic outcome (48). Ample evidence from clinical trials has shown that oncolytic viruses face immediate and effective immune surveillance, leading to rapid clearance even within the immunosuppressive environment of tumors (5–7). While the anti-tumor response is effective in a small percentage of patients, the oncolytic virus is consistently eliminated in most cases. Thus, the immunodominant epitopes in the case of a tumor infection are the viral antigens and the dominance of virus antigens versus tumor antigens may dramatically reduce the overall efficacy of oncolytic viruses.

For all these reasons, overcoming immunodominance is essential for unlocking the full potential of oncolytic viruses as a robust and effective modality in cancer treatment (Figure 3). Addressing this challenge requires a nuanced understanding of the immune response dynamics and the development of innovative strategies to optimize therapeutic outcomes. It requires strategies to broaden the immune response, enhance viral persistence, and optimize anti-tumor effects. Engineering oncolytic viruses to express or modify viral antigens can alter the immunodominance landscape. This may involve incorporating novel epitopes or modifying existing ones to reduce their recognition by the immune system (49). Introducing a repertoire of tumor-associated antigens within the oncolytic virus to ensure that a broader spectrum of antigens is presented to the immune system, hoping that some of them will be included in the immune response, can be achieved by encoding multiple tumor-specific antigens or using strategies to induce the release of various cellular antigens during viral replication (33).

Figure 3. Central role of immunodominance in viroimmunotherapy.

Immunodominance is defined by a hierarchical response to competing antigens. In the context of tumor infection, T-cell activity predominantly targets viral antigens (dominant clones) and, secondarily, tumor antigens (subdominant clones). This viral antigen immunodominance poses a challenge, hindering the induction of an effective anti-tumoral immune response in virotherapy. To overcome this challenge, which is considered the hardest problem in virotherapy, investigators should devise strategies aimed at redirecting the immune response of the patient from the virus toward cancer cells. If successful, this therapeutic redirection of immune dominance will increase the number of patients that respond to virotherapy and prevent tumor recurrence after treatment. [Created with BioRender.com]

As we mentioned earlier, administering pro-inflammatory cytokines or incorporating co-stimulatory molecules into oncolytic viruses may result in the augmentation of an anti-tumor immune response (44). We have already discussed incorporating co-stimulatory signals into the backbone of the virus. Concerning cytokines, co-administration of interferons or engineering oncolytic viruses to produce interferons or IL-12 may be helpful, as interferons e interleukins play a crucial role in orchestrating both innate and adaptive immune responses (34). Another strategy would involve incorporating toll-like receptor agonists into oncolytic viruses to stimulate innate immune responses (46). This combination creates a more inflammatory microenvironment, promoting increased antigen presentation and a diversified anti-tumor immune response. One caveat is that these strategies may reinforce the anti-viral immune response, as the canonical immune response against tumors is similar to the immune response against viruses. However, cytokines, co-stimulatory molecules, or even immune checkpoint antibodies can stimulate a more robust and diversified immune reaction against both viral and tumor antigens.

Another step in the right direction would be to develop patient-specific (tumor-specific) antigen targeting by tailoring oncolytic viruses to express antigens specific to the individual tumor profile of a patient. This personalized approach aims to overcome immunodominance by focusing on antigens uniquely expressed by a particular tumor, thereby potentially evading pre-existing immune responses against viruses. Conducting comprehensive immune profiling of patients to understand their immune landscape can guide the customization of oncolytic viruses and combination therapies to address immunodominance.

Vectorologists and virologists will attempt to address the problem of the immunodominance of viral antigens by modifying viral proteins. For instance, modifying the viral capsid or envelope may significantly reduce virus recognition by neutralizing antibodies (36, 43). This lack of recognition, and the resulting clearance, can extend the circulation time of oncolytic viruses, increasing the likelihood of the virus reaching and infecting tumor cells before being cleared by the immune system.

In addition, immunodominance is not confined to the viral components alone. It may also involve the tumor-associated antigens released during oncolytic virus-mediated cell lysis. The immune system may prioritize specific tumor antigens over others, potentially leading to suboptimal recognition and elimination of tumor cells (47, 50). For instance, recognizing antigens that are not drivers of the neoplastic phenotype may have little or no effect on the leading and more aggressive clones of the tumor. Thus, this selective immune response can undermine the desired anti-tumor effects of oncolytic virotherapy.

In summary, overcoming immunodominance in oncolytic virotherapy requires a comprehensive and adaptable approach that addresses both viral and tumor-associated antigens. The development of personalized, combination therapies that modulate the immune response and enhance viral persistence holds promise for unlocking the full potential of oncolytic viruses in cancer treatment. Continued research efforts and clinical trials are crucial for validating and refining these strategies for practical clinical application.

Conclusions

The paradigm-shifting concept that constitutes the basis for cancer viroimmunotherapy proposes that this cancer treatment modality is effective if, after the first wave of oncolysis, appears a second therapeutic wave propelled by the development of an anti-tumor immune response. These two-wave phenomenon is only observed in a minority of patients treated with oncolytic viruses. The evolution in the line of thinking about virotherapy as viroimmunotherapy implies that increasing the percentage of survivors requires the development of several strategies aimed at facilitating the temporary escape of the virus from the immune response elicited after the administration of the therapeutic virus and ultimately to redirect the immune response from the virus to the tumor. More specifically, the foreseen challenges to maximizing viroimmunotherapy efficacy include tumor immune evasion mechanisms, viral clearance and neutralization within and outside the tumor, high viral immunogenicity and low tumor immunogenicity with limited antigen presentation, and immunodominance.

The immunosuppressive tumor microenvironment constitutes a barrier characterized by an intricate network of immune-suppressive cells, cytokines, and other factors that hinder the immune system from mounting an effective response against tumor cells. Several approaches are being developed to overcome tumor immune evasion mechanisms, including the combination of oncolytic viruses with immune modulators, the generation of viruses that express immune checkpoints or cytokines, and the combination of the virus with strategies to inhibit the immunosuppression provided by myeloid cells and T regulatory cells.

Furthermore, viruses administered for therapeutic purposes are often swiftly recognized and eliminated by the immune system, thereby limiting their efficacy. Addressing viral clearance and neutralization involves modifying viruses to reduce immunogenicity and employing techniques such as the use of tolerogenic dendritic cells and chimeric viruses. Local or intratumoral administration, as well as regional or endovascular delivery, can also help bypass systemic neutralization and enhance viral persistence within the tumor microenvironment. Additionally, utilizing cell vehicles for viral transport can enhance targeted delivery and reduce off-target effects. Notably, the use of cell vehicles can be combined with locoregional delivery for better results.

The low immunogenicity and compromised antigen presentation of the tumor can be overcome by enhancing antigen presentation using viral vectors engineered to express tumor-specific antigens, or by co-administration of the immune costimulatory molecule, which can potentiate the immune response and bolster the efficacy of oncolytic virotherapy. Targeting dendritic cells and Toll-like receptor agonists can further increase the anti-tumor immune response while mitigating tumor immunosuppression. Furthermore, combining oncolytic viruses with natural killer cells and tumor-antigen-based vaccines, including mRNA-designed vaccines, can enhance the capability of the immune system to recognize and attack cancer cells.

Although the virus infection may neutralize immunosuppressive factors, stimulate the infiltration and activation of cytotoxic immune cells within the tumor milieu, and potentiate the anti-tumor immune response, the awakening of this immune response will primarily be directed against the immunodominant viral antigen. Strategies to overcome immunodominance, such as utilizing rare viral serotypes, modifying dominant epitopes, avoiding pre-existing humoral immune responses against adenoviruses, and expressing patient-specific tumor antigens simultaneously with viral antigens, may lead to the redirection of the immune response toward the tumor. Directing the immune response from the virus towards subdominant clones or neoantigens expressed by the tumor cells may broaden and sustain a robust anti-tumor immune response. Another strategy may involve inducing systemic tolerogenicity against viral antigens during tumor infection.

In conclusion, cancer viroimmunotherapy stands out as a potent and versatile approach to cancer treatment and future developments will propel new clinical trial and the final legitimization of viromimmunotherapy as a standard treatment for tumors including malignant brain tumors.

Data availability.

No datasets were generated or analyzed during the current study.