Preclinical Development of a Balanced, Tetravalent, Live-Attenuated Vaccine Against the Four Dengue Virus Serotypes

Background: This project is a critical step in the development of a tetravalent live attenuated vaccine (LAV) to prevent dengue fever. It falls under the Military Infectious Diseases Research Program (MIDRP) Preventative Measures for Infectious Diseases objective of developing a broadly protective vaccine to prevent dengue. The mosquito-borne dengue virus (DENV) is a major international health concern, and in 2019, a military prioritization panel ranked DENV as the top viral threat to the U.S. military and the third most significant infectious disease threat (behind malaria and bacterial diarrhea) overall. The only prevention strategy is to avoid mosquito bites; there are no vaccines indicated for adults or specific therapies. CodaVax-DENV, an LAV DENV vaccine, was generated using Codagenix's Synthetic Attenuated Viral Engineering (SAVE) technology, which uses synthetic biology, genome "deoptimization," and de novo DNA synthesis to generate vaccine strains. SAVE-attenuated viruses have identical amino acid sequences to that of the wild-type (wt) virus, but are translated more slowly. We developed a tetravalent vaccine, CodaVax-DENV, that (1) can be grown to production scale, (2) is attenuated in vitro and in vivo, (3) is identical to currently circulating wt strains at the amino acid level for each serotype, (4) induces a balanced neutralizing antibody response in two DENV susceptible mouse strains and rhesus monkeys, and (5) protects mice and monkeys against wt DENV challenge. Because it uses only a two-dose regimen spaced 30 days apart, the Codagenix vaccine would be far more practical and potentially safer, especially for use in the military, than current vaccines that require widely spaced dose regimens. Hypothesis/Objective: Our objective is to develop a safe tetravalent DENV vaccine that when administered to non-human primates reduces viremia in response to wt DENV challenge and generates a balanced neutralizing antibody response that persists at least 24 weeks. Our ultimate goal is to develop a DENV vaccine that is safe for use in humans that can protect individuals against all four DENV serotypes and not predispose recipients to enhanced disease irrespective of pre-existing serostatus. Specific Aims: Aim 1: Determine the attenuation, neutralizing antibody response and balance, and extent and durability of protection in rhesus monkeys vaccinated with CodaVax-DENV. Aim 2: Manufacture at least 300 vials of Good Manufacturing Practices (GMP) CodaVax-DENV product. Aim 3: Complete the Good Laboratory Practices (GLP) repeat dose CodaVax-DENV toxicology studies in rhesus monkeys that are needed to open a U.S. Investigational New Drug (IND). Aim 4: Prepare and submit an IND application to the U.S. Food and Drug Administration (FDA). Study Design: Aim 1: A total of 25 animals will be vaccinated subcutaneously (SC) with 106 FFU (i.e., 106/serotype) CodaVax-DENV, a mixture of the four serotypes of wt virus, or vehicle and boosted on day 28. Animals will be bled to assess post-vaccination viremia, development of neutralizing antibodies, and cellular immunity. One-half of the animals will be challenged with wt virus that elicits the lowest immunogenicity on day 70 and the other half on day 196 (24 weeks) post-vaccination to assess durability of protection over time. Aim 2: Process for CodaVax-DENV production and formulation will be developed at Codagenix. Manufacture of CodaVax-DENV at Charles River Laboratories (CRL) under current GMP will occur following a technology transfer from Codagenix. CRL will perform a test run with the viruses and then a full GMP run. The four resulting bulk drug substances will be purified and formulated. They will then be pooled at calculated volumes to achieve a titer of 106 FFU/ml for each serotype in the formulated bulk drug product. The formulated bulk drug product will be vialed (at least 300 vials) and final release testing performed. Aim 3: We will conduct a toxicology study under current GLP in non-human primates, as is appropriate for a live virus vaccine against dengue. A total of 24 rhesus macaques will be vaccinated with CodaVax-DENV, wt DENV, or vehicle on days 0, 21, and 42. Animals will be bled at regular intervals to assess post-vaccination viremia, blood chemistry, and safety labs. Animals will be sacrificed on day 46 (n=4 from each group) and 56 for a gross examination of vital organs and tissue histopathology. Aim 4: Regulatory activities will run for the duration of this project, starting with a pre-IND to present our GMP manufacturing results and plan for GLP toxicology study to the FDA and culminating in an IND submission at the end of the project. Impact: This work will enable Codagenix to complete the remaining preclinical studies, process development, and manufacturing needed to prepare and submit an IND package to the FDA. Once the IND is opened, Codagenix will be able to begin first-in-human clinical trials. Successful development of CodaVax-DENV would address a critical unmet need in protecting active-duty and retired Service Members and the general public against DENV. It would also be a tremendous boon to the billions of people living in DENV endemic regions. Less