This article discusses the production process of the major influenza antigen, hemagglutinin (HA), by rDNA methods in E. coli. Fusing the gene for HA to the gene for flagellin, a Toll-like receptor (TLR) ligand, yields a bi-functional protein. The HA moiety contains the structures recognized by the immune system as it generates neutralizing antibody, and the flagellin targets the HA antigen into the appropriate compartment of an antigen presenting cell. Having the antigen and the TLR ligand physically connected drives a robust antibody and cellular immune response. Producing this protein in E. coli provides the additional benefit of high yield per culture volume and global portability. These vaccines could help immunize the global population rapidly during an influenza pandemic.
Despite these successes, the fact is that the vast majority of viral antigens do not self assemble into regular arrays and thus are not presented efficiently by APCs. For the past two decades, researchers have been cloning, expressing, and purifying proteins from pathogens and testing them as vaccines. Overall, these monomeric proteins have been found to be poorly immunogenic. The collective experience has shown that while natural infection generally results in a robust and durable immune response to a variety of components of the pathogen, vaccination with the same components as purified proteins does not. Clearly, something was missing from vaccine candidates based solely on purified recombinant viral proteins.
The Role of Adjuvants
For years, researchers have known that one can improve the immunogenicity of an antigen by adding an adjuvant. Freund's Complete Adjuvant (FCA), has been the benchmark for laboratory work. FCA, an oily emulsion containing killed mycobacteria, can raise both robust antibody and cytotoxic T-cell immune responses in vaccinated animals. But this comes at the expense of a high frequency of sterile abcesses, making FCA unsuitable for human use. The adjuvant that is extensively used in man is alum, a generic term for a range of particulate forms of aluminum sulfate mixed with sodium hydroxide and varying amounts of phosphate. Aluminum-containing adjuvants work both as depots—most antigens can be made to stick to it—and as irritants, drawing cells of the immune system to the site of injection. Antigens formulated on aluminum-containing adjuvants tend to elicit good antibody responses, but not cytotoxic T-cell responses. However, for most naturally caused infectious diseases, both antibody and cytotoxic T-cell responses are required to control the disease and protect against future infection. This suggests that naturally infecting pathogens must carry some kind of FCA-like adjuvant activity.