436 Immunization Against SIV Gag Using Recombinant SV40 Vectors Elicits Strong, Durable Cell-mediated Immune Responses H. J. McKee*, D. S. Strayer Jefferson Med Coll, Philadelphia, PA
Background: A promising approach to anti-HIV vaccine development is the use of live virus vectors to deliver HIV antigens in vivo. The parameters of protective immunity against HIV infection remain unclear, but strong cell-mediated immune responses are likely to be very important. Since recombinant SV40 vectors (rSV40s) do not elicit immune responses against themselves, they can be used to deliver multiple inoculations in order to boost immune responses against even relatively weak antigens. We investigated whether multiple immunizations with a rSV40 encoding SIVmac239 gag elicited antigen-specific immune responses.
Methods: A recombinant SV40 encoding SIVmac239 gag (SV[gag239]) was generated by cloning a PCR fragment which encodes gag from the cloned SIVmac239 proviral genome into a plasmid carrying a large T-antigen deleted SV40 genome. SV(gag239), then was used to inoculate BALB/cJ mice monthly. Cloned P815 cells stably transfected with a gag-expressing plasmid (pT7A5-gag) were used as target cells for both cell-based ELISAs and 51Cr-release assays to measure gag-specific antibody and cytotoxic lymphocyte responses, respectively, in immunized mice.
Results: Flow cytometry analysis was used to screen gag-expressing P815 stable-transfectants, and high gag-expressing clones were used as targets in all subsequent experiments. Significant binding antibody responses against gag were detectable after multiple inoculations (> 3) with SV(gag239). Very strong gag-specific CTL responses (= 70% lysis) were observed in direct CTL assays, with unselected effector cells from mice receiving multiple immunizations. Low E:T ratios (10:1 and 20:1) were used. In these studies, the final inoculation was administered 6 months after the penultimate inoculation. Thus, high specific CTL activity suggests that a durable, memory cytotoxic response was elicited by immunization with SV(gag239).
Conclusions: Recombinant SV40 vectors can be used successfully to deliver SIV antigens multiple times, boosting immune responses to them. Very powerful cytotoxic responses are seen in immunized mice, and there is evidence that cytolytic memory is elicited. These results demonstrate that rSV40s may be useful vaccine delivery vehicles.
