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While DNA vaccines have been shown to prime cellular immune responses, levels are often low in nonhuman primates or humans. Hence, efforts have been directed toward boosting responses by combining DNA with different vaccination modalities. To this end, a polyvalent DNA prime/protein boost vaccine, consisting of codon optimized HIV-1 env (A, B, C, E) and gag (C) and homologous gp120 proteins in QS-21, was evaluated in rhesus macaques and BALB/c mice. Humoral and cellular responses, detected following DNA immunization, were increased following protein boost in macaques and mice. In dissecting cellular immune responses in mice, protein-enhanced responses were found to be mediated by CD4+ and CD8+ T cells with a Th1 cytokine bias. Our study reveals that, in addition to augmenting humoral responses, protein boosting of DNA-primed animals augments cellular immune responses mediated by CD8+ CTL, CD4+ T-helper cells and Th1 cytokines; thus, offering much promise in controlling HIV-1 in vaccinees.

HV MNE is an Epstein–Barr virus (EBV)-like lymphocryptovirus (LCV) originally isolated from a Macaca nemestrina with CD8 + T cell mycosis fungoides/cutaneous T cell lymphoma (Blood 98 (2001), 2193). HV MNE transforms rabbit T cells in vitro and causes T cell lymphoma in New Zealand white rabbits. Here we demonstrate that HV MNE also immortalizes T cells from mustached tamarins but not those from owl monkeys, common marmosets, squirrel monkeys, black-capped capuchins, and humans. Cytogenetic and FACS analysis revealed the true origin and T cell lineage of the transformed tamarin T cell lines. Tamarin T cells contained HV MNE DNA sequence and displayed a decreased requirement for the IL-2 cytokine for growth. Thus, this EBV-like virus from M. nemestrina differs from the other EBV-like viruses found in nonhuman primates inasmuch as it appears to preferentially transform T cells.

Rhesus macaques were immunized with a replication-deficient vaccinia virus (MVA) expressing human immunodeficiency virus type 1 89.6 envelope (env) and SIV gagpol (MVA/SHIV89.6) with or without a protein boost consisting of soluble 89.6 env (gp140). Immunization with MVA/SHIV89.6 alone elicited binding antibodies in all animals and neutralizing antibodies in 5 of 15 animals. Both types of antibodies were enhanced by protein boosting. In addition, CD8 cells exhibiting CM9 tetramer binding were detected in the subset of animals that were Mamu-A*01 positive. Animals were challenged intravenously with either SHIV-89.6 (Study 1) or the more pathogenic derivative SHIV-89.6P (Study 2). In Study 1, all control and vaccinated animals except one became infected. However, the levels of viremia were as follows: controls > rMVA alone > rMVA + protein. The differences were statistically significant between immunized and control groups but not between the two immunized groups. In Study 2, all animals became infected; however, the vaccinated group exhibited a 5-fold reduction in peak viremia and a 10-fold reduction in the postacute phase viremia in comparison to the controls. All of the controls required euthanasia by 10 months after challenge. A relationship between vaccine-induced antibody titers and reduction in virus burden was observed in both studies. Thus, immunization with MVA/SHIV89.6 alone or with a protein boost stimulated both arms of the immune system and resulted in significant control of viremia and delayed progression to disease after challenge with SHIV-89.6P.

The immunogenicity of a polyvalent HIV-1 vaccine comprised of Env antigens from primary R5 isolates was evaluated in rhesus macaques. DNA vaccines encoding four Env antigens from multiple HIV-1 subtypes and HIV-1 Gag antigen from a single subtype elicited a persistent level of binding antibodies to gp120 from multiple HIV-1 isolates that were markedly enhanced following boosting with homologous gp120 proteins in QS-21 adjuvant irrespective of the route of DNA immunization. These sera neutralized homologous and, to a lesser degree, heterologous HIV-1 isolates. Four of the six immunized animals were completely protected following rectal challenge with a SHIV encoding Env from HIV-1Ba-L, whereas the virus load was reduced in the remaining animals compared to naïve controls. Hence priming with DNA encoding Env antigens from multiple HIV-1 clades followed by boosting with homologous Env proteins elicits anti-HIV-1 immune responses capable of protecting macaques against mucosal transmission of R5 tropic SHIV isolate.

A seven-year-old male gibbon ape (Hylobates lar) with malignant lymphoma and leukemia was obtained from a colony in Bermuda. Tissues obtained at necropsy were infiltrated with tumor cells to varying degrees, the most extensively involved being the kidneys, ureters, liver, lymph nodes, heart, salivary glands, and mesentery. These cells showed typical lymphoblast morphology. Type-C virus particles were abundant in the plasma and were detectable in all tissues tested. Intracytoplasmic virus was seen in some nonleukemic parenchymal cells. However, budding virus particles were observed only from the leukemic peripheral blood lymphoblasts. Tissue extracts contained viral p30, reverse transcriptase, viral RNA, and proviral DNA in amounts which generally correlated with the degree of leukemic cell infiltration. Viral markers were particularly abundant in tissue from the region of the oral cavity. Proviral DNA was found in all tissues except muscle and brain, which contained little or no leukemic cell infiltration. Replicating virus was transmitted to heterologous cells in tissue culture from many tissues, including muscle and brain. This virus, called GaLVH was classified as a member of the gibbon ape leukemia virus-simian sarcoma (simian sarcoma-associated) virus group by comparative tests of the antigenic relatedness of p30 and reverse transcriptase, and by nucleotide sequence homology of RNA from this virus and from other members of this group. These results: (a) highlight the extraordinary replication of this type of virus in gibbons; (b) further implicate this virus group in the etiology of spontaneous gibbon ape leukemia; (c) suggest that the oral route is a likely mode of virus transmission; and (d) fail to provide evidence for a genetic reservoir of virus outside of the tumor cells.

Topical DNA vaccination (DermaVir) facilitates antigen presentation to naive T cells. DermaVir immunization in mice, using HIV-1 Env and Gag, elicited cellular immune responses. Boosting with HIV-1 gp120 Env and p41 Gag augmented Th1 cytokine levels. Intramuscular DNA administration was less efficient in priming antigen-specific cytokine production and memory T cells. In rhesus macaques, DermaVir immunization induced Gag- and Env-specific Th1 and Th2 cytokines and generation of memory T cells. Boosting of DermaVir-primed serum antibody levels was noted following gp140(SHIV89.6P)/p27(SIV) immunization. Rectal challenge with pathogenic R5-tropic SHIV162P3 resulted in control of plasma viremia (4/5 animals) that was reflected in jejunum, colon and mesenteric lymph nodes. An inverse correlation was found between Gag- and Env-specific central memory T cell responses on the day of challenge and plasma viremia at set point. Overall, the topical DermaVir/protein vaccination yields central memory T cell responses and facilitates control of pathogenic SHIV infection.

The productive infection and transformation of fresh human lymphocytes by several HTLV isolates have recently been reported. We extend these observations here with the description of multiple immortalized, non-producer, human umbilical cord blood lymphocyte cultures developed by cocultivation or fusion of fresh cells with T cells cultured from leukemia-lymphoma patients. These transformed neonatal leukocytes exhibit morphological, cytochemical, and other phenotypic characteristics similar to those of other HTLV-infected cells but, in contrast to the usual productive infection seen, these cells contain only low amounts of viral proteins and do not release virus particles. These cells contain at least one copy of HTLV proviral DNA/cell and transcribe viral RNA similar in size to virus-producing cells. Virus expression in these cultures was not enhanced by IUdR treatment. These cell cultures should be useful in studies of the regulation of viral expression in human cells and of the viral proteins and nucleic acids involved in T-cell immortalization and growth.