Rational combination of checkpoint blockade and gene therapy with ARF and IFN-b in melanoma models

Author(s)

Ana Carolina Martins Domingues

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina (FM/SBD)
• Defense date: 2024-06-21

Examining board members

Bryan Eric Strauss; Luciana Nogueira de Sousa Andrade; Adriana Bastos Carvalho

• Advisor: Bryan Eric Strauss

Abstract

In the last decades, the therapeutic options for oncologic patients improved in unprecedented ways, mainly after the advent of checkpoint inhibitors, a class of antibodies that block inhibitory pathways in immune cells thus restoring the patients own immune activity against the tumor. However, many patients do not respond to this therapy, indicating the need for new approaches to treat these tumors. Our group has previously developed a gene therapy using non-replicating adenoviral vectors coding for ARF and interferon beta (IFNb). In previous publications our team showed that this combination promotes immunogenic cell death in different melanoma models. Moreover, both p53 (restored by ARF reestablishment) and IFNb pathways have been described as inducers of PD-L1 upregulation, a molecule that binds to PD-1, an inhibitory T cell receptor. The upregulation of this pathway is known to increase treatment resistance and to cooperate with tumor progression, in such a way that blocking the PD-L1/PD-1 axis (known as checkpoint blockade) concomitantly to delivering the ARF and IFNb vectors could work synergistically. To test this hypothesis, we first assessed PD-L1 expression on checkpoint blockade-resistant mouse cell lines. As we detected an upregulation of this molecule upon gene transfer to B16 cells, we rationally combined the checkpoint blockade and gene therapy in vivo, also comparing these effects in the MC38 checkpoint blockade-sensitive mouse model. We observed, however, that there is no robust synergy between the treatments in either model. When immunophenotyping the B16-treated tumors, we observed that the gene therapy alone increases innate immune responses. To further explore the immune context related to our proposed treatment, nineteen patient-derived tumors were cultured ex vivo and treated with the vectors. As in this model the immune cells present in the patients original tumor are preserved, we could correlated that, among the more than 50% responders to the gene therapy alone, the majority was cold tumors with a major myeloid infiltration, which also had the biggest cytokine increased expression upon treatment, even when comparing to anti-PD-1. Importantly, the mutational status of the locus that encodes ARF were not important when the spheroids were treated with a constitutive promotor, while it was important when using a p53-responsive promoter. Lastly, from the treated tumors, the gastrointestinal and liposarcoma samples had the best responses to the gene therapy. These results indicate that, even though there is a rationale for combining the two therapies, these assays revealed no benefit. However, these analyses point to as yet unexplored directions, mainly the use of the gene therapy for cold tumors that cannot benefit from checkpoint inhibitors, importantly MDM2-amplified liposarcomas (AU)

• FAPESP's process: 18/25555-9 - Rational combination between checkpoint blockade and gene therapy with p19Arf and IFN-beta in a mouse model of melanoma
• Grantee: Ana Carolina Martins Domingues
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)