The failure to elicit high (>80%) protective immunity in recent human and animal trials may be due to the inclusion of vaccine epitopes that enhance lentiviral infection. Therefore, 3 unique approaches were utilized in the selection and delivery of T-cell based vaccine epitopes. First, cytotoxic T lymphocyte (CTL)-associated epitopes conserved among lentiviruses: FIV, HIV-1 and SIV were analyzed for T-cell proliferation, cytokine and CTL-associated cytotoxin production, and viral enhancement. Second, selected epitopes from p24, MA, RT and Env were evaluated for reactivity to human (HLA) and feline (FLA) leukocyte antigen supertypes common in the general population of both humans and cats. Evaluation of FLA binding pockets in the cat model have demonstrated considerable similarity to those of HLA supertypes A3, B27, B44, A24, B7, A2, and A1 (EpiVax Inc). Third, lentivirally-conserved (LC) epitopes identified from p24 and RT peptides were constructed into multi-antigenic peptides (MAPs), and tested in the FIV-cat model. In a pilot vaccine study, MAP constructs of 4 FIV epitopes (2-p24, 2-RT) conferred complete protection in 1 cat with delayed infection in 2 cats (vaccine group, n = 4), whereas all 4 challenge-control cats were unprotected. Notably, the non-LC epitope and its MAP construct enhanced FIV infection, whereas the MAP consisting of overlapping epitopes had an overall inhibitory activity. Selected LC epitopes from p24, MA, RT and Env induced robust CTL and polyfunctional T-cell activity in humans and cats. An expanded study including 9 LC epitopes is ongoing (n = 18). Current results underscore the need for careful selection of protective epitopes and targeted exclusion of enhancing epitopes for any vaccine design.