Highly stable mini-circRNA vaccine with maximally activated and long-lasting T cells for HPV-related cancer combination immunotherapy

Human papillomavirus (HPV) is responsible for a large proportion of global cancer cases, including head and neck, cervical and anal cancers, among others. HPV-related cancers are caused by certain high-risk strains of the virus, which can cause long-lasting infections that eventually lead to the development of cancerous changes. The mainstay for addressing HPV-related cancer is prevention, particularly through prophylactic vaccination and screening, and definitive treatment of early pre-cancerous lesions. Despite the success of prevention efforts, the prevalence of early and late-stage HPV-related cancers is still staggering, with over 18,000 deaths annually in the US. One promising new treatment approach is immunotherapy, in particular immune checkpoint inhibition (ICI). However, only 15-30% of patients respond to ICI treatment, highlighting the need for new techniques to boost efficacy. Although ex vivo adoptive T cell immunotherapies, in which human T cells are genetically modified outside the body to target cancer cells, are promising, they have significant limitations due to complexity, cost, and the need for myeloablative lymphodepletion. Effective therapeutic cancer vaccines have long been the holy grail as potentially safe and cost-effective in vivo-elicited T cell therapies that could be used in all stages of disease to boost cancer immunotherapy without the challenges and risks of ex vivo adoptive T cell therapies. While conventional vaccines largely have not been effective, newer highly immunogenic mRNA cancer vaccines have demonstrated clear evidence of efficacy with a reduction in recurrence and death in late-stage melanoma and strong anti-tumor responses in pancreatic cancer. Mini-circular RNA (mcRNA) is a highly biostable therapeutic cancer vaccine platform that can elicit robust activation and expansion of long-lasting antigen-specific T cells, achieving breakthrough levels up to 8-fold higher than state-of-the-art mRNA vaccines with, surprisingly, improved safety and tolerability. Our goal is to develop a highly immunogenic HPV E6 and E7 mcRNA vaccine that could be used as an off-the-shelf HLA-agnostic product or as part of a personalized HLA-specific cancer vaccine. In this proposal, we plan to build on a significant body of highly encouraging in vitro and in vivo data to facilitate rapid development of the mcRNA therapeutic vaccine platform with a highly immunogenic vaccine for HPV-related cancers. Key results from the proposed investigational studies will demonstrate the superior efficacy of a LNP-encapsulated multivalent mcRNA HPV16 E6/E7 vaccine compared to a N1-methylΨ mRNA vaccine. In Aim 1, we will show that a multivalent HPV mcRNA vaccine can sustain the expression of long concatemers of encoded HPV E6/E7 epitopes that are processed and presented to elicit highly potent and durable T cell responses that can yield a robust immunotherapy in combination with ICI for HPV-related cancer in mice. In Aim 2, we will characterize the immunogenicity of 5 epitope clusters encoding all of the top human MHC-binding epitopes of HPV16 E6 and E7 in PBMCs collected from human subjects, and assess the immunogenicity of a multivalent HPV16 epitope cluster vaccine compared to a benchmark HPV16 mRNA vaccine.