TNFa and IL-2 armed adenoviruses enable complete responses by anti-PD-1 checkpoint blockade

Releasing the patient's immune system against their own malignancy by the use of checkpoint inhibitors is delivering promising results. However, only a subset of patients currently benefit from them. One major limitation of these therapies relates to the inability of T cells to detect or penetrate into the tumor resulting in unresponsiveness to checkpoint inhibition. Virotherapy is an attractive tool for enabling checkpoint inhibitors as viruses are naturally recognized by innate defense elements which draws the attention of the immune system. Besides their intrinsic immune stimulating properties, the adenoviruses used here are armed to express tumor necrosis factor alpha (TNFa) and interleukin-2 (IL-2). These cytokines result in immunological danger signaling and multiple appealing T-cell effects, including trafficking, activation and propagation. When these viruses were injected into B16.OVA melanoma tumors in animals concomitantly receiving programmed cell-death protein 1 (PD-1) blocking antibodies both tumor growth control (p <0.0001) and overall survival (p <0.01) were improved. In this set-up, the addition of adoptive cell therapy with OT-I lymphocytes did not increase efficacy further. When virus injections were initiated before antibody treatment in a prime-boost approach, 100% of tumors regressed completely and all mice survived. Viral expression of IL2 and TNFa altered the cytokine balance in the tumor microenvironment towards Th1 and increased the intratumoral proportion of CD8+ and conventional CD4+ T cells. These preclinical studies provide the rationale and schedule for a clinical trial where oncolytic adenovirus coding for TNFa and IL-2 (TILT-123) is used in melanoma patients receiving an anti-PD-1 antibody.Peer reviewe

Cytokine-Coding Oncolytic Adenovirus TILT-123 Is Safe, Selective, and Effective as a Single Agent and in Combination with Immune Checkpoint Inhibitor Anti-PD-1

Oncolytic viruses provide a biologically multi-faceted treatment option for patients who cannot be cured with currently available treatment options. We constructed an oncolytic adenovirus, TILT-123, to support T-cell therapies and immune checkpoint inhibitors in solid tumors. Adenoviruses are immunogenic by nature, are easy to produce in large quantities, and can carry relatively large transgenes. They are the most commonly used gene therapy vectors and are well tolerated in patients. TILT-123 expresses two potent cytokines, tumor necrosis factor alpha and interleukin-2, to stimulate especially the T-cell compartment in the tumor microenvironment. Before entering clinical studies, the safety and biodistribution of TILT-123 was studied in Syrian hamsters and in mice. The results show that TILT-123 is safe in animals as monotherapy and in combination with an immune checkpoint inhibitor anti-PD-1. The virus treatment induces acute changes in circulating immune cell compartments, but the levels return to normal by the middle of the treatment period. The virus is rapidly cleared from healthy tissues, and it does not cause damage to vital organs. The results support the initiation of a phase 1 dose-escalation trial, where melanoma patients receiving a tumor-infiltrating lymphocyte therapy are treated with TILT-123 (NCT04217473).Peer reviewe

Abscopal Effect in Non-injected Tumors Achieved with Cytokine-Armed Oncolytic Adenovirus

Cancer treatment with local administration of armed oncolytic viruses could potentially induce systemic antitumor effects, or the abscopal effect, as they self-amplify in tumors, induce danger signaling, and promote tumor-associated antigen presentation. In this study, oncolytic adenovirus coding for human tumor necrosis factor alpha (TNF-alpha) and interleukin-2 (IL-2) Ad5/3-E2F-d24-hTNF-alpha-IRES-hIL-2 (also known as [a.k.a.] TILT-123) provoked antitumor efficacy in tumors that were injected with Ad5/3-E2F-d24-hTNF-alpha-IRES-hIL-2 and those that were left non-injected in the same animal. Importantly, the virus was able to travel to distant tumors. To dissect the effects of oncolysis and cytokines, we studied replication-incompetent viruses in mice. Systemic antitumor effects were similar in both models, highlighting the importance of the arming device. The cytokines induced positive changes in immune cell infiltrates and induced the expression of several immune-reaction-related genes in tumors. In addition, Ad5/3-E2F-d24-hTNF-alpha-IRES-hIL-2 was able to increase homing of adoptively transferred tumor-infiltrating lymphocytes into both injected and non-injected tumors, possibly mediated through chemokine expression. In summary, local treatment with Ad5/3-E2F-d24-hTNF-alpha-IRES-hIL-2 resulted in systemic antitumor efficacy by inducing immune cell infiltration and trafficking into both treated and untreated tumors. Moreover, the oncolytic adenovirus platform had superior systemic effects over replication-deficient vector through spreading into distant tumors.Peer reviewe

Oncolytic Adenoviruses Armed with Tumor Necrosis Factor Alpha and Interleukin-2 Enable Successful Adoptive Cell Therapy

Adoptive cell therapy holds much promise in the treatment of cancer but results in solid tumors have been modest. The notable exception is tumor-infiltrating lymphocyte (TIL) therapy of melanoma, but this approach only works with high-dose preconditioning chemotherapy and systemic interleukin (IL)-2 postconditioning, both of which are associated with toxicities. To improve and broaden the applicability of adoptive cell transfer, we constructed oncolytic adenoviruses coding for human IL-2 (hIL2), tumor necrosis factor alpha (TNF-alpha), or both. The viruses showed potent antitumor efficacy against human tumors in immunocompromised severe combined immunodeficiency (SCID) mice. In immunocompetent Syrian hamsters, we combined the viruses with TIL transfer and were able to cure 100% of the animals. Cured animals were protected against tumor re-challenge, indicating a memory response. Arming with IL-2 and TNF-alpha increased the frequency of both CD4(+) and CD8(+) TILs in vivo and augmented splenocyte proliferation ex vivo, suggesting that the cytokines were important for T cell persistence and proliferation. Cytokine expression was limited to tumors and treatment-related signs of systemic toxicity were absent, suggesting safety. To conclude, cytokine-armed oncolytic adenoviruses enhanced adoptive cell therapy by favorable alteration of the tumor microenvironment. A clinical trial is in progress to study the utility of Ad5/3-E2F-d24-hTNFa-IRES-hIL2 (TILT-123) in human patients with cancer.Peer reviewe

Comparison of Clinically Relevant Oncolytic Virus Platforms for Enhancing T Cell Therapy of Solid Tumors

Despite some promising results, the majority of patients do not benefit from T cell therapies, as tumors prevent T cells from entering the tumor, shut down their activity, or downregulate key antigens. Due to their nature and mechanism of action, oncolytic viruses have features that can help overcome many of the barriers currently facing T cell therapies of solid tumors. This study aims to understand how four different oncolytic viruses (adenovirus, vaccinia virus, herpes simplex virus, and reovirus) perform in that task. For that purpose, an immunocompetent in vivo tumor model featuring adoptive tumor-infiltrating lymphocyte (TIL) therapy was used. Tumor growth control (p < 0.001) and survival analyses suggest that adenovirus was most effective in enabling T cell therapy. The complete response rate was 62% for TILs + adenovirus versus 17.5% for TILs + PBS. Of note, TIL biodistribution did not explain efficacy differences between viruses. Instead, immunostimulatory shifts in the tumor microenvironment mirrored efficacy results. Overall, the use of oncolytic viruses can improve the utility of T cell therapies, and additional virus engineering by arming with transgenes can provide further antitumor effects. This phenomenon was seen when an unarmed oncolytic adenovirus was compared to Ad5/3-E2F-d24-hTNFa-IRES-hIL2 (TILT-123). A clinical trial is ongoing, where patients receiving TIL treatment also receive TILT-123 (ClinicalTrials.gov: NCT04217473).</p

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival