Adenovirus vector vaccination induces expansion of memory CD4 T cells with a mucosal homing phenotype that are readily susceptible to HIV-1

A recently conducted phase 2B HIV vaccine trial in high risk HIV-negative volunteers (the STEP study) using an adenovirus vaccine (Ad5 vector expressing Gag, Pol and Nef) was ineffective. In that study, individuals were divided into two groups on the basis of their Ad5 antibody titers. Besides being ineffective, individuals who received this vaccine were more susceptible to HIV-1 infection in comparison to the placebo. This susceptibility was more significant in the group of vaccinees with high Ad5 antibody titers. The reason for this is not clear.

Natural Ad5 infection occurs via gut or nasopharynx and the virus replicates in the epithelial cells of mucosal tissues, inducing mucosal immunity. Thus, the authors of this study hypothesized that vaccination of individuals immune to Ad5 with adenovirus vector would activate and expand T cells expressing a mucosal homing phenotype, and these cells would migrate to the gut mucosa, increasing the number of permissive HIV-1 target cells and the risk of infection.

Citation: Benlahrech A, Harris J, Meiser A, Papagatsias T, Hornig J, Hayes P,

Lieber A, Athanasopoulos T, Bachy V, Csomor E, Daniels R, Fisher K, Gotch F, 

Seymour L, Logan K, Barbagallo R, Klavinskis L, Dickson G, Patterson S. 

Adenovirus vector vaccination induces expansion of memory CD4 T cells with 

a mucosal homing phenotype that are readily susceptible to HIV-1. 

Proc Natl Acad Sci U S A. 2009 Nov 24;106(47):19940-5.

 

RESULTS:

1. Adenovirus specific cytokine responses do not correlate with Ad5 antibody titers: Ad5 and Ad11 antibody titers were measured in 20 healthy donors. IFN-gamma ELSIPOT was performed on PBMCs stimulated with Ad5 or Ad11 from 15 of these donors. 73% and 66.7% of individuals showed responses against Ad5 and Ad11, respectively. When Ad5 and Ad11 IFN-gamma responses were plotted against Ad5 antibody titers, no significant correlation was observed in between the Ad5 or Ad11 IFN-gamma ELISPOT responses and Ad5 antibody titer. Furthermore, the investigators pulsed dendritic cells from the donor with Ad5, Ad11, tetanus toxoid, heat inactivated influenza or Staphylococcus enterotoxin B (SEB) and measured cytokines in lymphocytes. It was observed that majority of IFN-gamma responses against Ad5 and Ad11 were mediated by CD8 T cells. CD8 cells produced more TNF-alpha in response to stimulation with Ad5 or Ad11 in comparison to CD4 T cells. There was no or very little IL-12 production in response to either Ad5 or Ad11 by any cell. The authors then assessed the functional quality of the responding Ad5 and Ad11-specific CD4 T cells. There was no difference between Ad5 and Ad11 responding CD4 cells in terms of cytokine profile as both groups of cells predominantly produced TNF-alpha alone or in combination with IFN-gamma or IL-2.
   
2. Adenovirus induced T cell proliferation correlates with Ad5 antibody titers: The investigators next sought to determine the ability of Ad vectors to induce T cell expansion ex vivo. For this, DCs were pulsed with Ad5, Ad11, tetanus toxoid, heat inactivated influenza or Staphylococcus enterotoxin B (SEB) and co-cultured with autologous CFSE-stained T cells. It was observed that majority of proliferationg T cells (CSFR low) in response to Ad5 or Ad11 stimulation were CD4 T cells. No significant difference in CD4 or CD8 cells was observed in response to tetanus toxoid or influenza. The authors also observed a positive correlation between Ad5- or Ad11- induced CD4 T cell proliferation and Ad5 serostatus.
   
3. Alpha4beta7 expression by expanded adenovirus specific memory CD4 T cells: Since, migration of lymphocytes to gastrointestinal tissues is dependent on the expression of a heterodimer composed of alpha4 and beta7 integrins, the authors measured expression of this molecule on expanded CSFE-low (dividing) CD4 T cells co-cultured with Ad5-, Ad11-, tetanus toxoid-, heat inactivated influenza- or Staphylococcus enterotoxin B (SEB)-pulsed DCs for five days. Majority of the expanded Ad5-specific CD4 T cells expressed high levels of alpha4beta7 in comparison to CSFE high (undivided) CD4 cells.
   
4. Adenovirus induced alpha4beta7 increases correlate with Ad5 titers: The authors next determined the fold increase in the alpha4beta7 expression by stimulation of T cells with Ad5, Ad11, tetanus toxoid, influenza or SEB-pulsed DCs in relation to the background expression levels of CD4 T cells cultured with unpulsed DCs. A positive correlation between the fold increase in total CD4 T cells expressing alpha4beta7 in response to Ad5 or Ad11 stimulation was observed. The authors also found a positive correlation between the fold increase in total CD4 T cells expressing alpha4beta7 and Ad5 antibody titers.
   
5. CCR9 and CCR5 up-regulation in response to Ad5 and Ad11 challenge: As chemokine receptor CCR9 also helps in the migration of lymphocytes to mucosal tissues, the authors next studied the level of CCR9 expression by activated adenoviral-specific memory CD4 T cells in four donors. They observed significantly increased levels of surface CCR9 in cells expanded by stimulation with first and second generation Ad5 and Ad11 vectors in comparison to CSFE high cells. Similar to CCR9, CCR5, which functions as a co-receptor for HIV-1 entry is up-regulated by T cells with effector or memory phenotype and Th1 cells. The authors thus, investigated the expression of CCR5 on expanded adenovirus specific memory T cells. CCR5 was found to be up-regulated on CSFE low CD4 T cells when co-cultured with Ad5 or Ad11 pulsed DCs in comparison to CSFE high undivided T cells.
   
6. Re-stimulated adenovirus specific CD4 T cells are more permissive to HIV-1 infection: To determine the significance of increased expression of CCR5 on expanded adenovirus specific memory CD4 T cells, the authors stimulated DCs from these donors with first or second generation Ad5 or Ad11 and co-cultured them with lymphocytes. They were then infected with the R5 virus, HIV-1. The authors observed that intracellular staining for HIV-1 p24 was significantly higher in the proliferating CFSE-low CD4 T cells in comparison to un-dividing CFSE-high cells.