David R. Kaufman

Trafficking of Antigen-Specific CD8+ T-Lymphocytes to Mucosal Surfaces Following Intramuscular Vaccination

A critical goal of vaccine development for a wide variety of pathogens is the induction of potent and durable mucosal immunity. However, it has been assumed that this goal would be difficult to achieve by systemic vaccination due to the anatomic and functional distinctness of the systemic and mucosal immune systems and the resultant compartmentalization of immune responses. In this study, we show that Ag-specific CD8+ T lymphocytes traffic efficiently to mucosal surfaces following systemic vaccination. Intramuscular immunization with recombinant adenovirus (rAd) vector-based vaccines expressing SIV Gag resulted in potent, durable, and functional CD8+ T lymphocyte responses at multiple mucosal effector sites in both mice and rhesus monkeys. In adoptive transfer studies in mice, vaccine-elicited systemic CD8+ T lymphocytes exhibited phenotypic plasticity, up-regulated mucosal homing integrins and chemokine receptors, and trafficked rapidly to mucosal surfaces. Moreover, the migration of systemic CD8+ T lymphocytes to mucosal compartments accounted for the vast majority of Ag-specific mucosal CD8+ T lymphocytes induced by systemic vaccination. Thus, i.m. vaccination can overcome immune compartmentalization and generate robust mucosal CD8+ T lymphocyte memory. These data demonstrate that the systemic and mucosal immune systems are highly coordinated following vaccination.

Route of Adenovirus-Based HIV-1 Vaccine Delivery Impacts the Phenotype and Trafficking of Vaccine-Elicited CD8+ T Lymphocytes

ABSTRACT Candidate HIV-1 vaccine regimens utilizing intramuscularly (i.m.) administered recombinant adenovirus (rAd)-based vectors can induce potent mucosal cellular immunity. However, the degree to which mucosal rAd vaccine routing might alter the quality and anatomic distribution of vaccine-elicited CD8+ T lymphocytes remains unclear. We show that the route of vaccination critically impacts not only the magnitude but also the phenotype and trafficking of antigen-specific CD8+ T lymphocytes in mice. I.m. rAd immunization induced robust local transgene expression and elicited high-frequency, polyfunctional CD8+ T lymphocytes that trafficked broadly to both systemic and mucosal compartments. In contrast, intranasal (i.n.) rAd immunization led to similarly robust local transgene expression but generated low-frequency, monofunctional CD8+ T lymphocytes with restricted anatomic trafficking patterns. Respiratory rAd immunization elicited systemic and mucosal CD8+ T lymphocytes with phenotypes and trafficking properties distinct from those elicited by i.m. or i.n. rAd immunization. Our findings indicate that the anatomic microenvironment of antigen expression critically impacts the phenotype and trafficking of antigen-specific CD8+ T lymphocytes.

Multiple Innate Immune Pathways Contribute to the Immunogenicity of Recombinant Adenovirus Vaccine Vectors

ABSTRACT The innate immune pathways that contribute to the potent immunogenicity of recombinant adenovirus (rAd) vaccine vectors remain largely undefined. Previous studies assessing innate immunity triggered by vaccine vectors have largely focused on in vitro studies involving antigen-presenting cells and on early in vivo inflammatory responses. Here, we systematically explore the Toll-like receptor (TLR) signaling requirements for the generation of cellular immune responses by intramuscular immunization with common and alternative serotype rAd vectors in mice. Antigen-specific CD8+ T-lymphocyte responses elicited by these rAd vectors were significantly diminished in MyD88−/− mice but not in TRIF−/− or TLR3−/− mice, suggesting the importance of MyD88-dependent TLR signaling. However, the absence of each individual TLR resulted in minimal to no effect on vaccine-elicited cellular immune responses. Moreover, responses were not diminished in IL-1R−/− or IL-18R−/− mice. These data suggest that rAd vectors engage multiple MyD88-dependent signaling pathways, none of which are individually critical; rather, they are integrated to contribute to the potent immunogenicity of rAd vectors. Stimulation of multiple innate immune mechanisms may prove a generalizable property of potent vaccines, and this strategy could be harnessed in the development of next-generation vaccine vectors and adjuvants.

Vitamin A Deficiency Impairs Vaccine-Elicited Gastrointestinal Immunity

Vitamin A deficiency is highly prevalent in much of the developing world, where vaccination programs are of paramount importance to public health. However, the impact of vitamin A deficiency on the immunogenicity and protective efficacy of vaccines has not been defined previously. In this article, we show that the vitamin A metabolite retinoic acid is critical for trafficking of vaccine-elicited T lymphocytes to the gastrointestinal mucosa and for vaccine protective efficacy in mice. Moderate vitamin A deficiency abrogated Ag-specific T lymphocyte trafficking to the gastrointestinal tract, gastrointestinal cellular immune responses, and protection against a mucosal challenge following immunization with a recombinant adenovirus vaccine vector. Oral vitamin A supplementation as well as retinoic acid administration fully restored the mucosal immune responses and vaccine protective efficacy. These data suggest that oral vitamin A supplementation may be important for optimizing the success of vaccines against HIV-1 and other mucosal pathogens in the developing world, highlighting a critical relationship between host nutritional status and vaccine efficacy.

Differential antigen requirements for protection against systemic and intranasal vaccinia virus challenges in mice

ABSTRACT The development of a subunit vaccine for smallpox represents a potential strategy to avoid the safety concerns associated with replication-competent vaccinia virus. Preclinical studies to date with subunit smallpox vaccine candidates, however, have been limited by incomplete information regarding protective antigens and the requirement for multiple boost immunizations to afford protective immunity. Here we explore the protective efficacy of replication-incompetent, recombinant adenovirus serotype 35 (rAd35) vectors expressing the vaccinia virus intracellular mature virion (IMV) antigens A27L and L1R and extracellular enveloped virion (EEV) antigens A33R and B5R in a murine vaccinia virus challenge model. A single immunization with the rAd35-L1R vector effectively protected mice against a lethal systemic vaccinia virus challenge. The rAd35-L1R vector also proved more efficacious than the combination of four rAd35 vectors expressing A27L, L1R, A33R, and B5R. Moreover, serum containing L1R-specific neutralizing antibodies afforded postexposure prophylaxis after systemic vaccinia virus infection. In contrast, the combination of rAd35-L1R and rAd35-B5R vectors was required to protect mice against a lethal intranasal vaccinia virus challenge, suggesting that both IMV- and EEV-specific immune responses are important following intranasal infection. Taken together, these data demonstrate that different protective antigens are required based on the route of vaccinia virus challenge. These studies also suggest that rAd vectors warrant further assessment as candidate subunit smallpox vaccines.

Longitudinal Requirement for CD4+ T Cell Help for Adenovirus Vector–Elicited CD8+ T Cell Responses

Despite the widespread use of replication-incompetent recombinant adenovirus (Ad) vectors as candidate vaccine platforms, the mechanism by which these vectors elicit CD8+ T cell responses remains poorly understood. Our data demonstrate that induction and maintenance of CD8+ T cell responses by Ad vector immunization is longitudinally dependent on CD4+ T cell help for a prolonged period. Depletion of CD4+ T cells in wild type mice within the first 8 d following Ad immunization resulted in dramatically reduced induction of Ag-specific CD8+ T cells, decreased T-bet and eomesodermin expression, impaired KLRG1+ effector differentiation, and atypical expression of the memory markers CD127, CD27, and CD62L. Moreover, these CD8+ T cells failed to protect against a lethal recombinant Listeria monocytogenes challenge. Depletion of CD4+ T cells between weeks 1 and 4 following immunization resulted in increased contraction of memory CD8+ T cells. These data demonstrate a prolonged temporal requirement for CD4+ T cell help for vaccine-elicited CD8+ T cell responses in mice. These findings have important implications in the design of vaccines aimed at eliciting CD8+ T cell responses and may provide insight into the impaired immunogenicity of vaccines in the context of AIDS and other CD4+ T cell immune deficiencies.

Translational Mini-Review Series on Vaccines for HIV: T lymphocyte trafficking and vaccine-elicited mucosal immunity

Many pathogens use mucosal surfaces to enter and propagate within the host, making particularly desirable vaccines that target immune responses specifically to mucosal compartments. The majority of mucosal vaccine design strategies to date have been empirical in nature. However, an emerging body of basic immunological knowledge is providing new insights into the regulation of tissue‐specific lymphocyte trafficking and differentiation. These insights afford the opportunity for the rational design of vaccines that focus immune responses at mucosal surfaces. Mucosal cellular immunity may prove critical for protection in the context of HIV infection, and thus there has been considerable interest in developing vaccines that target HIV‐specific cellular immune responses to the gastrointestinal and vaginal mucosa. However, the optimal strategies for eliciting mucosal cellular immune responses through vaccination remain to be determined. Here, we review both recent vaccine studies and emerging paradigms from the basic immunological literature that are relevant to the elicitation of potent and protective mucosal cellular immune memory. Increasing the synergy between these avenues of research may afford new opportunities for mucosal vaccine design.

Focus and breadth of cellular immune responses elicited by a heterologous insert prime-boost vaccine regimen in rhesus monkeys

The global sequence diversity of HIV-1 presents a daunting challenge for vaccine development. We investigated whether a heterologous insert prime-boost regimen could expand global coverage by selectively boosting cellular immune responses to conserved epitopes. Rhesus monkeys were primed and boosted with recombinant adenovirus vectors expressing homologous or heterologous HIV-1 Gag sequences that were optimized to focus responses on highly conserved epitopes. We observed comparable responses directed to specific regions of the Gag protein in all experimental groups without evidence of improved coverage or expanded breadth in the heterologous insert group. These data suggest that antigen-independent factors contribute to the immunodominance patterns of vaccine-elicited cellular immune responses.

Efficient Generation of Mucosal and Systemic Antigen-Specific CD8+ T-Cell Responses following Pulmonary DNA Immunization

ABSTRACT Although mucosal CD8+ T-cell responses are important in combating mucosal infections, the generation of such immune responses by vaccination remains problematic. In the present study, we evaluated the ability of plasmid DNA to induce local and systemic antigen-specific CD8+ T-cell responses after pulmonary administration. We show that the pulmonary delivery of plasmid DNA formulated with polyethyleneimine (PEI-DNA) induced robust systemic CD8+ T-cell responses that were comparable in magnitude to those generated by intramuscular (i.m.) immunization. Most importantly, we observed that the pulmonary delivery of PEI-DNA elicited a 10-fold-greater antigen-specific CD8+ T-cell response in lungs and draining lymph nodes of mice than that of i.m. immunization. The functional evaluation of these pulmonary CD8+ T cells revealed that they produced type I cytokines, and pulmonary immunization with PEI-DNA induced lung-associated antigen-specific CD4+ T cells that produced higher levels of interleukin-2 than those induced by i.m. immunization. Pulmonary PEI-DNA immunization also induced CD8+ T-cell responses in the gut and vaginal mucosa. Finally, pulmonary, but not i.m., plasmid DNA vaccination protected mice from a lethal recombinant vaccinia virus challenge. These findings suggest that pulmonary PEI-DNA immunization might be a useful approach for immunizing against pulmonary pathogens and might also protect against infections initiated at other mucosal sites.

Response to Comment on “Trafficking of Antigen-Specific CD8+ T Lymphocytes to Mucosal Surfaces following Intramuscular Vaccination”

A critical goal of vaccine development for a wide variety of pathogens is the induction of potent and durable mucosal immunity. However, it has been assumed that this goal would be difficult to achieve by systemic vaccination due to the anatomic and functional distinctness of the systemic and mucosal immune systems and the resultant compartmentalization of immune responses. In this study, we show that Ag-specific CD8(+) T lymphocytes traffic efficiently to mucosal surfaces following systemic vaccination. Intramuscular immunization with recombinant adenovirus (rAd) vector-based vaccines expressing SIV Gag resulted in potent, durable, and functional CD8(+) T lymphocyte responses at multiple mucosal effector sites in both mice and rhesus monkeys. In adoptive transfer studies in mice, vaccine-elicited systemic CD8(+) T lymphocytes exhibited phenotypic plasticity, up-regulated mucosal homing integrins and chemokine receptors, and trafficked rapidly to mucosal surfaces. Moreover, the migration of systemic CD8(+) T lymphocytes to mucosal compartments accounted for the vast majority of Ag-specific mucosal CD8(+) T lymphocytes induced by systemic vaccination. Thus, i.m. vaccination can overcome immune compartmentalization and generate robust mucosal CD8(+) T lymphocyte memory. These data demonstrate that the systemic and mucosal immune systems are highly coordinated following vaccination.