Post Vaccination Testing

Many preventive vaccines function by stimulating the development ofneutralizing antibodies to the immunogen(s), these antibodies being sufficient to protect against a subsequent infection by the pathogen. These vaccines are designed to induce a "humoral" or B-cell response that translates into an enduring production of protective, high-affinity, neutralizing antibodies against the pathogen [23]. The measurement of these specific antibodies in serum can be directly correlated with the efficacy of a vaccine to provide immunity against distinct pathogens [24]. Traditionally, such measurements are made by immunoassays, with no requirements for cell-based assays. However, "B-cell vaccination strategies" do not provide effective immunity against all pathogens, especially against rapidly mutating viruses, bacteria, and parasites. The control and elimination of these pathogens is subject to the effectiveness of the infected host's T-cell responses and cellular immune mechanisms.

For all vaccines, the development of antigen-specific T cells - as helper T cells, CTL and memory T cells - is essential for initiating and maintaining humoral and cellular immunity. The classic view is that peptides derived from the invading pathogen are recognized in combination with major histocompatibility complex (MHC) class II molecules by CD4+ T lymphocytes. These help B-cells in the induction of antibody responses and are crucial for the development of CD8+ T-cell-mediated protective immunity, while cytotoxic CD8+ T lymphocytes (CTL) - which kill infected cells and vanquish infective pathogens, especially viruses - recognize peptides in combination with MHC class I molecules. Indeed, in some infections (e.g., by HIV-1), cellular immunity due to CTL has been considered of greater importance than the presence of neutralizing antibodies (see below). It is evident from many studies of HIV-1-infected individuals that, although they develop neutralizing antibodies to the primary infecting HIV-1 strain, these rapidly become ineffective as the virus mutates [25, 26]. Moreover, the only HIV-1 antigen - recombinant envelope glycoprotein gp120 - to have been fully assessed in clinical trials induced neutralizing antibodies, but completely failed to provide a protective effect [27, 28, 118].

In continuing to use HIV-1 infection for illustration purposes, there is evidence that CD8+ T cells are involved in providing some resistance to the infective process by the virus. For instance, following primary infection with HIV-1, a strong CD8+ CTL response develops during the acute viremia phase, which generally persists into the chronic phase [29-31]. In HIV-1-infected individuals that do not progress to AIDS, strong HIV-1-specific CD8+ CTL activity, as well as CD4+ T-cell proliferative responses, against multiple epitopes have been measured [32, 33]. Furthermore, individuals that are multiply-exposed to HIV-1 (e.g., sex workers in the Gambia and Kenya) but do not contract the infection, have been shown to have HIV-1-specific CD8+ CTL [34, 35]. This observation, in addition to other supporting evidence from HIV-exposed seronegatives [36, 37] and from the SIV/rhesus-macaque model system [38-41], strongly suggest that CTL have a protective role against HIV-1 infection. However, more recent data on the Kenyan sex workers indicate that, while CTL were independently associated with age and recent HIV-1 exposure, they were not prospectively associated with protection [42]. Furthermore, CTL appear to kill HIV-1-infected cells only slowly in vivo [43], which has implications for vaccines designed to induce a lytic CTL response. For example, unless the vaccine-induced CTL response is several-fold greater than the natural response to HIV-1 infection, it is unlikely it will prevent infection, or mediate complete viral clearance in the case of a "therapeutic" vaccine (one that is administered to HIV-1 seropositives). However, a CTL response elicited by a "therapeutic" vaccine might reduce viral load and lengthen the asymptomatic period, and thus have some beneficial effect.

Despite the uncertainty over the effectiveness of CTL in conferring protection against HIV-1 infection [118], research is continuing into modern vector technologies with the potential for generating a cellular immune response to HIV-1. The hope is that a vaccine will emerge that induces a strong cellular immune response (especially a CTL response) which, while not being able to provide total protection from HIV infection, should enable "vaccines" to limit viral replication, reduce virus load, and slow the progression towards disease.

Based on the assumption that CTL do play some protective role against HIV-1 infection, and because of the ineffectiveness of neutralizing antibody inducing vaccines, much recent research has focused on developing HIV-1 vaccines that induce cellular immunity in the form of HIV-1-specific CTL [95, 118]. Starting with SIV in the rhesus-macaque model, potent CTL responses and protection against the pathogenic SIV-HIV hybrid virus, SHIV-89.6P, have been achieved with plasmid DNA [44, 45] or recombinant (MVA) poxvirus [46] vaccines expressing SIVmac239 Gag and HIV-1 89.6P Env. In addition, an effective CTL response and pronounced attenuation of subsequent SHIV challenge infection was demonstrated using either a replication-incompetent Ad5 viral vector expressing SIV gag [47] or an attenuated vesicular stomatitis virus (VSV) viral vector expressing env and gag genes and boosting with vectors having viral envelope glycoproteins from different VSV serotypes [48]. Although protection against SHIV 89.6P challenge has been repeatedly shown, it has been found to be more difficult to protect against other immunodeficiency viruses (e.g., SIVmac239) with these types of vaccines [49]. Numerous plasmid DNA and recombinant viral vector vaccines have been developed for immunizing against HIV-1; details of these T-cell vaccines are summarized in Table 6.1. An accurate monitoring ofT-cell activation is required in order to evaluate their effectiveness in stimulating T-cell responses, for correlating vaccine-induced T-cell status with overall protection, and for making future improvements. However, a number of specific tests are applicable to this monitoring, and these are described in the following section.

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