ABSTRACT Genital tract infection with Neisseria gonorrhoeae does not induce a state of specific protective immunity and it can be acquired repeatedly. Despite public health measures, the disease persists at an unacceptably high frequency; there is no vaccine against it, and resistance even to the latest generations of antibiotics continues to emerge. Our SBIR Phase I work established proof-of-principle that intra-vaginal immunization with a sustained-release interleukin-12 microsphere formulation plus proprietary gonococcal (GC) outer membrane vesicles (NGoXIM, previously GvaX12) induces durable Th1-driven adaptive immune responses that protect mice against genital tract infection with N. gonorrhoeae. Those studies demonstrated: (i) induction of GC-specific Th1 cells, (ii) generation of anti-GC antibodies in serum and genital tract secretions, (iii) protection against diverse strains of N. gonorrhoeae, (iv) recall of specific antibodies and reactivation of T cells after challenge infection, and (v) duration of protection against infection for at least 6 months. Protection depended upon both antibodies and IFN? production. In SBIR Phase II, we established efficacy of intranasal (i.n.) immunization in mice and, importantly, in non-human primates. We confirmed similar IFN? production and generation of GC- specific antibodies and by this route, demonstrated activity against additional antigenically diverse strains of N. gonorrhoeae, including clinical isolates, and showed functional antibody activity against GC in bactericidal assays. Finally, we concluded a type C pre-pre-IND meeting with the FDA, garnering Agency agreement with our single species primate toxicology plans. The specific aims of this Phase IIb application are partially based on Agency recommendations communicated during that meeting and the need to optimize NGoXIM for human application. In Aim 1 NGoXIM will be further optimized in non-human primates using the i.n. route in preparation for human trials. Aim 2 studies will conclude FDA-compliant pharmaceutical quality and development activity for the vaccine components. Upon successful completion of Aims 1 and 2, Therapyx will request a type B meeting with the FDA to confirm that the completed non-clinical IND-enabling program, including an NHP toxicology study plan and preliminary clinical study design, is sufficient to support an IND (Aim 3). In Aim 4, following FDA guidance and the results of our type B meeting, we will use validated products generated in Aim 2 to evaluate initial serum pharmacokinetics (PK) and toxicity profiles of NGoXIM in cynomolgus monkeys. The overall goal of this project is to produce the world's first prophylactic vaccine against genital tract infection with Neisseria gonorrhoeae.