Variable viral genes as genetic immunogens

Abstract

Influenza and HIV-1 are two highly variable viruses that cause extensive morbidity and mortality in many countries. The most cost-effective medical intervention against influenza is vaccination but this has to be repeated annually because of the low antigenicity of the vaccine. Moreover, the antigenic determinants of the viral envelope proteins hemagglutinin (HA) and neuraminidase (NA) undergo continuous immunoselection during viral spread in the population (antigenic drift).

In the case of HIV-1, medical treatment is lifelong, expensive and has severe side effects. The lack of an effective vaccine may be due in part to the intricate life cycle of the virus and its antigenic variability. The genetic variability of viruses, leading to antigenic variation, has important immunological and pathogenic consequences as well as epidemiological and evolutionary implications. In addition, the genetic variability is especially troublesome when identifying vaccine targets. DNA vaccines may provide solutions to some of these problems. In genetic immunization, one or several genes from a pathogen are cloned into an expression vector and subsequently introduced to a host tissue to elicit an immune response. This process ingeniously mimics the events during the natural infection of an intracellular parasite, and may consequently induce cellular as well as humoral immune responses.

In this thesis, we have selected highly variable envelope genes from influenza A and HIV-1, and investigated their ability to mediate protective humoral and cell mediated immune responses against influenza and primary HIV-1 infection.

Specifically, we have developed a methodology to rapidly adapt envelope-based genetic influenza vaccines to the genetic drift. The method was based on the construction of chimeric HA genes encoding the antigen determinant region from one virus and the less variable parts of the HA protein from another virus. We have subsequently demonstrated the immunogenicity of the chimeric HA genes in mice and errets. Furthermore, we established that the antibody specificity was restricted to the antigen determinant region. The methodology allows for rapid adaptation of an influenza vaccine to the genetic drift of circulating influenza strains.

To provide background information for a prospective HIV-1 vaccine trial, we determined the viral subtype by sequence analysis in samples from HIV-1 infected individuals from six different regions in Bangladesh. We found unexpected subtype heterogeneity in the samples, subtype C being the predominant subtype. Subtypes A and G as well as recombinant forms were also found. The subtype heterogeneity reflects a recent epidemic in the Bangladeshi population.

We have investigated the potential of using the HIV-1 envelope gene from multiple subtypes for vaccination. Gene combinations containing the HIV-1 env gene originating from subtypes A, B and C were shown to induce stronger antibody responses in particular as well as recall immune responses, after experimental challenge of mice with syngeneic splenocytes infected with an HIV-1/MuLV pseudovirus. In a follow-up study, recombinant GM-CSF was shown to be a potent adjuvant to the envelope genes. Again, the multi-subtype immunized animals had the most prominent antibody responses. Moreover, the antibodies had strong neutralizing capacities against several HIV-1 strains. A boost with rgp160 raised the antibody titers, but did not further enhance neutralization. When CpG-ODN was added to the boost, antibody profiles and IFN-gamma production indicated a shift from a Th2 type response towards a more balanced Th1/Th2 profile. The use of several envelope genes in a prime boost scheme may thus constitute a potent strategy for HIV-1 vaccination.