Progress in HIV Vaccines
With 2.53.5 million estimated deaths worldwide from HIV/AIDS, the vast majority of those occurring in sub-Saharan Africa, the AIDS epidemic and HIV infection represent an obvious and urgent target for vaccine development.
John Henessey, Jr., Ph.D., senior director of bioprocess and bioanalytical research at Merck Research Laboratories (Whitehouse Station, NJ) identified the three major challenges for HIV vaccine R&D: the diversity of the target population; the diversity in amino acid sequence between different clades of the HIV virus; and issues related to vaccine supply, demand, and delivery.
The Merck HIV vaccine is a replication-incompetent adenovirus type 5 construct that is E1 deficient and propagated in an E1-complemented PER.C6 cell line. It incorporates transgenes for gag, pol, and nef, which produce the corresponding proteins when taken up by host cells, inducing an immune response to the virus.
Merck employs a variety of assays to measure the in vitro biological activity of a series of genetic constructs, including end-point dilution assays and standard curve assays to assess in vitro viral replication and infectivity, in vitro antigen expression assays intended to mimic in vivo activity, and an adenovirus genome quantitation assay based on quantitative PCR to detect full versus empty virus particles. Dr. Hennessey emphasized the need to define standardized assays for adenovirus-based vaccines
Bryan Butman, Ph.D., vp of quality at GenVec (Gaithersburg, MD) described his company's work to develop a multiclade adenoviral vector-based vaccine against HIV. GenVec is working with the Vaccine Research Center of the NIH to create novel adenoviral vectors that encode for multiclade and multigene HIV antigens.
The company combined four different adenoviral vectors to make one tetravalent vaccine; it includes the gag and pol genes representing clade B and pg140 genes from clades A, B, and C. The expression cassette lacks E1 and E2, and E3 has been partially deleted. The complementing 293-ORF6 cell line leads to the production of replication-deficient vectors that encode for a fusion protein representing the multigene products.
GenVec is developing a scalable bioreactor system for vector production in serum-free suspension cultures with column chromatographic purification. Dr. Butman described a series of potency and identity assays used to characterize the vectors and the viral particle concentration. These include immunofluorescent assays, a transgene expression Western blot, PCR identity studies, and full-length sequencing.
Quality studies included reverse-phase HPLC of the blended product and are supported by safety and stability cell testing, aging studies, and tumorigenicity and oncogenicity studies. The HIV adenoviral vector induces a cellular immune response in cynomolgus macaques.
The Vaccine Research Center submitted an IND for clinical testing to assess the efficacy of a single administration of the vaccine at three dosing levels. GenVec has produced four bulk vaccine lots, which were quality-tested and released, and a Phase I dose-escalation clinical trial in humans began in July 2004.
The long-term goal is to assess the immunogenicity of the blended adenoviral HIV vector alone and in combination with a DNA prime-boost strategy. Challenges include the need for an efficacy trial, a more readily scalable manufacturing process, correlates of protection, quantitative potency assays for the transgene, and formulations that would allow storage of the vaccine at temperatures above -20C.
Boro Dropulic, Ph.D., founder and CSO of VIRxSYS (Gaithersburg, MD), presented the company's work using lentiviral-based vectors for HIV gene therapy. VRX496 is the firm's lentiviral vector expressing a 937-base antisense insert that targets the HIV envelope protein.
The goal of this approach is to transduce T lymphocytes in HIV-infected patients with the antisense payload, which then binds and destroys HIV RNA in T cells, thereby minimizing HIV replication to levels not associated with AIDS.
The company reported greater than 94% transduction of T lymphocytes in preclinical studies and greater than 100-fold inhibition of HIV replication and is currently completing a Phase I safety trial in HIV-infected patients who have failed at least two regimens of highly active anti-retroviral therapy (HAART).
Preliminary findings indicate no serious adverse events, no evidence of replication competent lentivirus, and steady CD4 counts in all patients. Two of the five patients had viral loads below baseline levels at six months post dosing, two maintained baseline viral loads, and the fifth patient had not yet undergone monitoring.