Preclinical Toxicology: Safety Evaluation of pNGVL4a-CRT-E6E7L2 (detox) Recombinant DNA Vaccine in Mice

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pNGVL4a-hCRT/E6E7L2, is a naked DNA vaccine directed at HPV E6, E7, and L2 proteins, with the goal to develop a therapeutic vaccine capable of inducing cellular immunity to eliminate HPV infection and HPV-associated lesions. The newly licensed HPV vaccines, Gardasil® and Cervarix®, both based upon L1 virus-like particles (VLPs), are effective only for prevention of new genital infections by the HPV genotypes included in the vaccines. Unfortunately, the L1 VLP vaccines do not alter the course of established HPV infections, and thus will not reduce rates of HPV-related cancer for several decades; additionally, they are not effective against all oncogenic HPV types. There is a clear unmet need for a broadly protective HPV vaccine that is also able to trigger the clearance of established HPV infections and associated neoplasia, particularly in immunocompromised persons such as human immunodeficiency virus (HIV)+ patients who have higher rates of HPV-related cancers. pNGVL4a-hCRT/E6E7L2 was generated by fusing human calreticulin (CRT) to HPV16 E6 and E7 genes, each containing multiple inactivating mutations and codons optimized for high level expression, and capsid L2 protein amino acid residues 11−200. In proof-of-concept (PoC) studies in mice, vaccination with the pNGVL4a-hCRT/E6E7L2 DNA vaccine was shown to be capable of generating a strong E6/E7-specific CD8+ T cell immune response that resulted in a potent therapeutic effect against E6/E7-expressing TC-1 tumor cells. Furthermore, significant L2-specific antibodies were generated that were able to neutralize HPV 16 virions. In another PoC study in mice, vaccination with hCRTE6E7L2 DNA elicited a potent antitumor effect against E7-expressing tumors even in mice depleted for CD4+ T cells, suggesting that this vaccine may be active even in immune-compromised hosts. A head-to-head comparison of various administration methods for delivery of a DNA vaccine encoding CRT linked to HPV16 E7 (pNGVL4a-CRT/E7) was performed in a preclinical model, and tested with conventional intramuscular (IM) needle administration, electroporation (EP)-mediated IM delivery, and particle-mediated epidermal delivery via gene gun. Results showed that EP-mediated IM delivery generated higher levels of circulating protein and higher numbers of E7-specific CD8+ T cells among splenocytes compared to conventional IM injection. The enhanced CD8+ T cell immune response detected in EP-vaccinated mice translated into superior antitumor effects in E7-expressing tumors in tumor-bearing mice suggesting that EP-mediated delivery may provide an advantage compared to conventional IM delivery of the vaccine. Based on this promising preclinical data, a contract manufacturer has generated a GMP Master Cell Bank and manufactured clinical grade pNGVL4a-hCRT/E6E7L2 DNA. A toxicology study to assess the safety of pNGVL4a-hCRT/E6E7L2 DNA vaccine in mice when administered using the TriGrid Delivery System (TDS)-IM EP device (Ichor Medical Systems, Inc.) is required to support a planned clinical trial evaluating the safety and feasibility of DNA vaccination of patients with HPV16-associated cervical intraepithelial neoplasia (CIN) 2/3, as part of the Cervical Cancer SPORE program.