Safety and immunogenicity of a polyvalent peptide C4-V3 HIV vaccine in conjunction with IL-12

We examined the safety and immunogenicity of a human leukocyte antigen-based HIV envelope polyvalent synthetic peptide vaccine, C4-V3, alone and in combination with subcutaneous IL-12 in nine HIV-infected patients. Lymphocyte proliferative responses increased threefold or more to all four peptides at two consecutive post-immunization timepoints for four individuals. Three responders had received IL-12, suggesting a possible adjuvant effect of IL-12. Transient mild injection site reactions (7/9) and systemic symptoms (3/9) occurred. An ideal HIV vaccine would induce both cytotoxic T-cell and neutralizing antibody responses. In a previous study, a human leukocyte antigen (HLA)-based polyvalent HIV envelope synthetic peptide vaccine, C4-V3, elicited neutralizing antibodies (four out of eight patients) and cellular immune responses, as measured by lymphocyte proliferative assays (five out of eight patients) [1]. IL-12 is under study as a vaccine adjuvant because it enhances the activity of CD8 T lymphocytes and natural killer cells and the production of certain antibody isotypes [2,3]. Two groups have shown in mice that IL-12 as an adjuvant to HIV vaccination may be a feasible approach [4,5]. The AIDS Clinical Trials Group 5049 Study was a phase I, limited center, sequential cohort trial in HIV-infected patients on stable antiretroviral therapy for 90 days or more with CD4 cell counts greater than 400 cells/mm3 and two screening HIV-RNA assays of less than 50 copies/ml designed to evaluate the safety of and cellular immune responses to C4-V3 administered alone and in combination with low and intermediate doses of subcutaneous IL-12. Study subjects were required to be HLA-B7 positive and naive to HIV vaccination and IL-12. The study was designed to enroll four patients into each of five cohorts. All subjects were to receive four injections of 0.25 ml C4-V3 vaccine intramuscularly (each deltoid and quadriceps muscle) at entry and weeks 4, 8, and 16. At the same timepoints, cohort I received no IL-12, cohort II received 0.25 μg IL-12 subcutaneously within 1 cm of one quadricep vaccine injection site, and cohort III received 0.25 μg IL-12 subcutaneously within 1 cm of two vaccine injection sites. Cohorts IV and V were to receive 0.5 μg and 1 μg IL-12 subcutaneously within 1 cm of two vaccine injection sites, respectively. IL-12 was administered in proximity to the vaccine site based on mouse data indicating improved cytotoxic T-lymphocyte (CTL) responses with the co-delivery of antigen and IL-12 [6]. Clinical assessments, measurements of lymphocyte subsets and plasma HIV-RNA levels, and immunology assays were performed pre and post-immunization at entry and weeks 4, 8, 12, 20, 24, and 48. C4-V3 vaccine (Wyeth-Lederle Vaccines, Pearl River, NY, USA) is an HIV polyvalent immunogen consisting of linearly synthesized hybrid peptides representing T helper determinants from the fourth constant (C4; amino acids 428–443) and third variable regions (V3, HLA-B7-restricted CTL epitope) formulated with incomplete Freund's adjuvant (IFA, Montanide ISA-51, Seppic Company, Paris, France). Primary amino acid sequences from four clade B HIV isolates (MN, RF, EV91, and CanOA) are represented [1]. The vaccine was emulsified using a mechanical device, the EmulsiFlex 1000 (Kirkland Products, El Cajon, CA, USA). Each injection series consisted of 1 mg total peptide (0.25 mg of each individual peptide) administered to each subject in four divided doses. Lymphocyte proliferation assays (LPA) were performed using the AIDS Clinical Trials Group consensus method utilizing the four immunogen components (C4-V3MN, C4-V3RF, C4-V3EV91, and C4-V3CanOA, each at 10 μg/ml) with a non-sense peptide control (Duke University Medical Center, Durham, NC, USA). LPA responses to HIV-1 antigen and HIV-1 native p24 antigen (0.5 μg/ml; Immune Response Corporation, Carlsbad, CA, USA) and Candida albicans skin test antigen (10 μg/ml; Greer Laboratory, Lenoir, NC, USA) were measured and used unstimulated medium as the control [7]. At the same timepoints CTL assays utilized enzyme-linked immunospot assays to measure IFN-γ-producing CD8 T cells after stimulation with the four immunogen components. Because of the small sample size, the analyses are descriptive and exploratory. For the LPA analyses, the stimulation index (SI), defined as the median counts per minute (CPM) in the stimulated wells divided by the median CPM in the appropriate control wells, was used. If the observed SI was less than 1, then the value of 1 was assigned. A positive LPA response was at least a threefold increase in SI over baseline on two consecutive post-immunization timepoints. This examination of threefold increases represents a post-hoc exploratory analysis after first examining fivefold increases [1]. The number of C4-V3 antigens (0 to 4) to which a subject developed a response was compared between subjects who received IL-12 versus those who did not using the Wilcoxon rank-sum test. Delta-CPM was defined as the difference in median CPM between the stimulated wells and the appropriate control wells. Among the responders to each antigen, the absolute increase in delta-CPM from baseline to the average of the two threefold increase timepoints was summarized. Because of an unanticipated expiration date of 31 July 2001 for the C4-V3 vaccine, the study accrued only nine (seven men, two women) of the desired 20 subjects. Four subjects were enrolled into cohort I, four into cohort II, and one into cohort III. Each subject received vaccine at entry and weeks 4, 8, and 16, except for subject 9 in cohort III, who only received three series of injections because the vaccine expired between his week 8 and 16 visits. The median CD4 cell count was 761 cells/mm3 (30%). No significant changes in CD4/CD8 cell counts or in plasma HIV-RNA levels occurred post-immunization. Only two of the nine subjects had a transient increase in plasma HIV-RNA levels to a maximum of 561 copies/ml before returning to less than 50 copies/ml. Increased lymphocyte proliferative responses were seen in six out of nine subjects (Fig. 1). Four subjects developed stimulation indices threefold or more above baseline on two consecutive visits against all four vaccine peptides. Although three of these four had been co-administered IL-12, the IL-12 effect was not significant (P = 0.33). The median delta-CPM increase from baseline among the responders was 5700 for the C4-V3MN antigen (range 1200–8700), 4400 for C4-V3RF (range 1800–7100), 6200 for C4-V3EV91 (range 2200–10 900), and 5800 for C4-V3CanOA (range 3900–8800). All subjects showed an SI greater than 3 to Candida (six at baseline and the remaining three at weeks 4 or 8). Only one subject, in cohort II, had a positive LPA response to either HIV-specific antigen. CTL responses to targets coated with peptide vaccine components were not observed in any of the subjects; however, the peripheral blood mononuclear cell yield was suboptimal.Fig. 1. Longitudinal lymphocyte proliferation assay responses to the four C4-V3 vaccine peptides, in terms of stimulation index.: Bold lines represent subjects with confirmed threefold increases in the stimulation index (SI) for two or more consecutive timepoints. The top panels display the four cohort I subjects who received vaccine only. The bottom panels display the five subjects from cohorts II and III who received IL-12 in addition to the C4-V3 vaccine. The line containing filled circles represents the one subject in cohort III who received the higher dose of IL-12 but only three series of vaccinations.Injection site reactions were common but mild. Seven subjects experienced grade 1 injection site reactions (erythema, edema, or induration < 25 cm2) of which two (both in cohort II) also experienced grade 2 injection site reactions (erythema, edema, or induration 25–100 cm3). Grade 2 injection site reactions in the two subjects occurred after only one of the four injection series for each subject. In each individual subject, injection site reactions did not occur with each series of injections. The one subject in cohort III who received a higher dose of IL-12 developed grade 1 injection site symptoms and body aches with the first injection series, grade 1 fever and grade 2 body aches and chills with the second injection series, but only grade 1 body aches with the third injection series. No ulcerations, necrosis, or sterile abscesses were seen in any subject. Grade 1 or 2 systemic symptoms consisting of fever, chills, headache or body aches occurred in three subjects, two in cohort I and one in cohort III. Grade 3 bilirubin and alanine/aspartate aminotransferase elevations occurred in one subject with chronic active hepatitis C, who had grade 2 elevations at baseline. Previously, DATRI 010 examined the safety and immunogenicity of the C4-V3 vaccine (2.0 mg total peptide) in 10 HIV-infected subjects [1]. Five out of eight C4-V3 recipients had LPA responses that increased fivefold or more over the baseline SI to at least one of the immunogen peptides in two consecutive post-immunization timepoints compared with none of the two controls who received IFA alone. Three out of eight vaccine recipients had fivefold or greater increases to all four peptides. In our study, increased lymphocyte proliferation responses were seen in six out of nine subjects, of which four subjects developed stimulation indices threefold or greater above baseline on two consecutive visits against all four peptides. Given our small numbers, no statistically significant effect of IL-12 could be demonstrated; however, there appeared to be a trend towards increasing LPA responses with the receipt of IL-12. The C4-V3 vaccine was well tolerated with low doses of subcutaneous IL-12. The tolerability was comparable to that reported in DATRI 010, in which four out of eight vaccine recipients and both of the two IFA-alone recipients experienced grade 1 site reactions [1]. AVEG 020 used the same emulsification technique but larger volumes of the C4-V3 vaccine in HIV-uninfected subjects (0.5 ml injected into each deltoid; 1 or 4 mg total peptide) and was halted as a result of severe injection site reactions [8]. Presumably the larger volume injected or the immunocompetence of the HIV-uninfected subjects led to severe injection site reactions that were not seen in our study. This study suggests that low doses of IL-12 can be given safely in conjunction with an HLA-based HIV envelope polyvalent synthetic peptide vaccine, and that IL-12 may have an adjuvant effect. Future studies should study next-generation peptide vaccines with higher doses of IL-12. Acknowledgements The authors would like to acknowledge the contributions of the following staff at sites where subjects were enrolled: University of Texas at Galveston, TX (Gerianne Casey RN); Duke University, Durham, NC (Martha Silberman RN, and Kenneth Shipp RPh); and Rush University Medical Center, Chicago, IL (Ruth M. Davis RN, and Gail Ruderman BS, RPh). They would also like to thank the following individuals who contributed to the conduct of the study: Nicole Grosskopf BS, Scharla G. Estep RPh, Vicki Stocker RN, MS, John Spritzler ScD, Miriam Chernoff PhD, and Richard Ginsberg MD. Finally, the authors thank the subjects who participated in this study.

Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.