P-06.02 Enhancing cancer immunotherapy through synthetic biology: a novel heterologous prime-boost strategy

<h3>Background</h3> Effective cancer immunotherapy is driven by robust T cell responses against neoantigens. A sufficient number of neoantigens is crucial to stimulate at least 2–3 cancer-specific T cell responses that can home into the tumor microenvironment and exert therapeutic activity. Current clinical trials support these principles, showing promising results with combinations of approved immunotherapies, including anti-PD1 antibodies. However, most existing delivery systems rely on single-platform approaches, such as long peptides, plasmid DNA or mRNA-LNPs, which often result in limited and short-lived immune responses. <h3>Materials and Methods</h3> To address the limitations of conventional delivery platforms, we investigated a synthetic approach combining the Hermes™ lipopolyplex nanoparticle platform encapsulating mRNA in combination with delivery of synthetic linear hairpin (hp)DNA™. Synthesis of hpDNA™ is fully enzymatic, facilitating fast production of DNA vaccine devoid of bacterial backbone with improved quality over traditional plasmid fermentation processes. Meanwhile, Hermes™ nanoparticles exploit a cationic ligand, combined with traditional lipids, to drive payload condensation and extend shelf life at refrigerator temperature. <h3>Results</h3> To explore the efficacy of these technologies, vaccines targeting 20 neoantigens of the MC38 tumor model were generated and combined in an innovative heterologous prime-boost protocol. This strategy partners the potent but transient nature of mRNA with synthetic DNA to enhance immunogenicity and durability of the anti-tumor response. While hpDNA encoding a large number of neoantigens is effective in inducing significant immune responses and antitumor activity, its combination with the new mRNA-Hermes™ formulation provide a faster T cell induction and improved tumor responses. This new therapeutic protocol represents the first example of a completely synthetic heterologous prime-boost vaccination protocol for cancer therapy. The results from our study suggest that the heterologous prime-boost regimen induces a polyfunctional and polyspecific T cell response against neoantigens, offering several advantages in the clinical setting. Responses in the MC38 tumor model are characterized by long-lasting activity and significant anti-tumor effects. The heterologous synthetic platform demonstrates a potent ability to elicit sustained immune responses, surpassing the performance of individual delivery systems including plasmid DNA or mRNA-LNP-based systems. <h3>Conclusions</h3> In conclusion, our findings provide compelling evidence that a synthetic biology-based heterologous prime-boost strategy is feasible and may offer an alternative valid immunotherapy platform. This approach holds significant potential for developing more effective and durable cancer immunotherapies, offering new hope for patients with challenging malignancies. I. Esposito: None. C. Tonini: None. A. Bianchi: None. E. Young: None. A. Dhir: None. M. Esposito: None. L. Aurisicchio: None. A. Walker: None. F. Palombo: None.