Safety and immunogenicity of novel recombinant BCG and modified vaccinia virus Ankara vaccines in neonate rhesus macaques

Abstract

Although major inroads into making antiretroviral therapy available in resource-poor countries have been made, there is an urgent need for an effective vaccine administered shortly after birth, which would protect infants from acquiring human immunodeficiency virus type 1 (HIV-1) through breast-feeding. Bacillus Calmette-Guérin (BCG) is given to most infants at birth, and its recombinant form could be used to prime HIV-1-specific responses for a later boost by heterologous vectors delivering the same HIV-1-derived immunogen. Here, two groups of neonate Indian rhesus macaques were immunized with either novel candidate vaccine BCG.HIVA(401) or its parental strain AERAS-401, followed by two doses of recombinant modified vaccinia virus Ankara MVA.HIVA. The HIVA immunogen is derived from African clade A HIV-1. All vaccines were safe, giving local reactions consistent with the expected response at the injection site. No systemic adverse events or gross abnormality was seen at necropsy. Both AERAS-401 and BCG.HIVA(401) induced high frequencies of BCG-specific IFN-gamma-secreting lymphocytes that declined over 23 weeks, but the latter failed to induce detectable HIV-1-specific IFN-gamma responses. MVA.HIVA elicited HIV-1-specific IFN-gamma responses in all eight animals, but, except for one animal, these responses were weak. The HIV-1-specific responses induced in infants were lower compared to historic data generated by the two HIVA vaccines in adult animals but similar to other recombinant poxviruses tested in this model. This is the first time these vaccines were tested in newborn monkeys. These results inform further infant vaccine development and provide comparative data for two human infant vaccine trials of MVA.HIVA.

FIG. 1.

HIVA immunogen and experimental design. (A) Schematic representation of the HIVA expression cassettes in BCG.HIVA⁴⁰¹ and MVA.HIVA. The immunogen HIVA consists of consensus HIV-1 clade A Gag p24-p17 domains and a string of partially overlapping epitopes (Ep). The polyepitope region contains epitopes derived from Gag, which are not present in the p24-p17 domains, Pol, Nef, and Env (14). To facilitate the preclinical development of the HIVA vaccines, the polyepitope region also includes immunodominant Mamu-A*01-restricted epitope CTPYDINQM derived from SIV Gag (CM9; residues 181 to 189) (red) (29). In BCG.HIVA⁴⁰¹ (top), the HIVA transcription is driven by the mycobacterial 85B antigen promoter, and nucleotides coding for the 19-kDa protein signal peptide (SP) are attached to the HIVA open reading frame. In MVA.HIVA (bottom), the gene transcription is controlled by vaccinia virus promoter P7.5 (14). (B) Experimental design. *, The specific ages of neonates at the time of AERAS-401/BCG.HIVA⁴⁰¹ vaccination were as follows: N3 and N8, 5 days; N1 and N2, 6 days; N4 and N7, 9 days; N6, 20 days; and N5, 22 days.

FIG. 2.

Local reaction observed on infant macaques at the site of AERAS-401/BCG.HIVA⁴⁰¹ administration. Neonatal rhesus macaques were given 10⁷ CFU i.d. of either AERAS-401 or BCG.HIVA⁴⁰¹, i.e., 10× the human infant dose. Pictures of the injection sites were taken at 1 and 4 weeks after the vaccine injection.

FIG. 3.

BCG-specific T-cell responses elicited by AERAS-401/BCG.HIVA⁴⁰¹ administration to infant macaques. Two groups of four infant rhesus macaques were immunized with 10⁷ CFU i.d. of either empty parental AERAS-401 or BCG.HIVA⁴⁰¹, and the responses elicited to BCG were determined in an ex vivo IFN-γ ELISPOT assay using PPD as the antigen. The times of vaccinations are shown below in weeks. Bp, parental AERAS-401; B, BCG.HIVA⁴⁰¹; M, MVA.HIVA. The panel shows median responses for AERAS-401 (gray bars) or BCG.HIVA⁴⁰¹ (black bars); the mock-stimulated background yielded a vast majority of wells with no spots. *, Statistically significant difference (P = 0.015) in a two-tail Student t test, which was not reached at any other time points. nd, not done.

FIG. 4.

HIV-1-specific T-cell responses elicited by HIVA. Two groups of four infant rhesus macaques were immunized with 10⁷ CFU i.d. of either empty parental AERAS-401 (Bp; panel A) or BCG.HIVA⁴⁰¹ (B; panel B) at week 0 and boosted by 5 × 10⁷ PFU of MVA.HIVA i.m. at weeks 11 and 14 (M11 and M14). HIV-1-specific T-cell responses were measured in an ex vivo IFN-γ ELISPOT assay using HIVA-derived peptide pools 1 to 5 (from light gray to black bars). Note that Pool 5 contains Mamu-A*01-restricted epitope CM9. For the ELISPOT data analysis, control counts from each neonate were pooled to form the control group. For each peptide pair, a Wilcoxon test was performed, and P values were corrected within each animal using Benjamini-Hochberg correction. Any P value that was <0.05 postcorrection was deemed significantly different from the control pool. Only significant values were represented in the graph, with the exception of bars marked by “x”. Software was used for the statistical analysis (http://www.r-project.org). *, Mamu-A*01⁺ animals.