Overcoming Challenges of HIV Vaccine Antigen Production
- Author(s): Byrne, Gabriel Patrick
- Advisor(s): Berman, Phil P
- et al.
Progress on the development of a vaccine to HIV-1 has been slowed by the inability to efficiently produce antigens at the quantities needed for clinical trials. Adding further difficulties, there is not currently a cellular substrate capable of large scale production that imparts the correct glycoforms required of an HIV-1 vaccine. As the biopharmaceutical industry matured over the last thirty years, the production of glycoproteins has increasingly focused on incorporating as much complex, sialic acid-containing N-linked glycans as possible onto recombinant proteins; seeking the increased circulatory half-life and immune tolerability these structures impart. The field of HIV-1 vaccine research followed suit, with material produced for clinical trials manufactured using the same paradigms applied to therapeutics such as monoclonal antibodies, recombinant enzymes (e.g. tissue plasminogen activator), chimeric receptors (e.g. TNFR-IgG), and cytokines (e.g. erythropoietin). In the case of those therapeutic examples, complex glycosylation helps to prolong the half-life, lowering cost, and avoid an immune response. Recent studies have demonstrated that this strategy has been counter-productive for an HIV-1 vaccine, where high mannose forms of N-linked glycosylation are required for stimulating broadly neutralizing antibodies required for inducing a protective immune response. This presents a challenge as there is currently no available expression system suitable for the biopharmaceutical production of recombinant glycoproteins possessing high-mannose glycans. This work describes the genetic engineering of a Chinese hamster ovary cell line to limit glycosylation to high-mannose and its applications for HIV-1 antigen production. This cell line was then used as the substrate to establish two permanent clonal lines expressing recombinant HIV-1 envelope protein antigens of clades C (TZ97008) and CRF01_AE (A244). Generating in excess of 1g/L, with glycosylation mimicking that of native virions, these lines solve two road blocks of HIV-1 vaccine development: inability to produce sufficient quantities of antigen and incorporation of high-mannose glycans.