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Clinical and Vaccine Immunology, February 2009, p. 233-240, Vol. 16, No. 2
1071-412X/09/$08.00+0     doi:10.1128/CVI.00066-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Production of a Dendritic Cell-Based Vaccine Containing Inactivated Autologous Virus for Therapy of Patients with Chronic Human Immunodeficiency Virus Type 1 Infection{triangledown}

Theresa L. Whiteside,1,4* Paolo Piazza,3 Amanda Reiter,4 Joanna Stanson,4 Nancy C. Connolly,2,3 Charles R. Rinaldo Jr.,1,3 and Sharon A. Riddler2,3

Departments of Pathology,1 Medicine, University of Pittsburgh School of Medicine,2 Departments of Infectious Diseases and Microbiology, Graduate School of Public Health, University of Pittsburgh,3 University of Pittsburgh Cancer Institute, Pittsburgh, Pennsylvania4

Received 19 February 2008/ Returned for modification 18 May 2008/ Accepted 17 November 2008

In preparation for a pilot clinical trial in patients with chronic human immunodeficiency virus type 1 (HIV-1) infection, a novel dendritic cell (DC)-based vaccine is being manufactured. The trial will test the hypothesis that isolated endogenous virus presented by DCs serves as a potent immunogen for activation of CD8+ and CD4+ T cells specific for a broad range of autologous HIV-1 antigens. Production of the vaccine under good manufacture practice conditions involves (i) autologous virus isolation; (ii) superinfection of CD4+ T cells with the virus; (iii) inactivation of the virus in CD4+ T cells, T-cell apoptosis, and coincubation of T cells with autologous DCs; and (iv) product testing and release. Endogenous virus was isolated from peripheral blood-derived CD4+ T cells of three HIV-1-positive subjects by coincubation with autologous OKT-3-stimulated CD4+ T cells. CD4+ T-cell supernatants were tested for p24 levels by enzyme-linked immunosorbent assay (>25 ng/ml) and for the 50% tissue culture infective doses (TCID50; which ranged from 4,642 to 46,416/ml on day 19 of culture). Autologous CD4+ T cells that were separated on immunobeads (>95% purity) and superinfected with virus-expressed p24 (28 to 54%) had TCID50 of >400/ml on days 5 to 10. Virus inactivation with psoralen (20 µg/ml) and UVB irradiation (312 nm) reduced the TCID50 of the supernatants from 199,986 to 11/ml (>99%). 7-Amino-actinomycin D-positive, annexin V-positive CD4+ T cells were fed to autologous DCs generated by using the Elutra cell separation system and the Aastrom system. Flow analysis showed that DC loading was complete in 24 h. On the basis of these translational results and experience with the generation of DCs from HIV-1-infected patients in a previous clinical trial, the Investigational New Drug application for clinical vaccination was submitted and approved by the FDA (application no. BB-IND-13137).

* Corresponding author. Mailing address: University of Pittsburgh Cancer Institute, Research Pavilion at the Hillman Cancer Center, 5117 Centre Avenue, Suite 1.27, Pittsburgh, PA 15213-1863. Phone: (412) 624-0096. Fax: (412) 624-0264. E-mail: whitesidetl{at}upmc.edu

{triangledown} Published ahead of print on 26 November 2008.

Clinical and Vaccine Immunology, February 2009, p. 233-240, Vol. 16, No. 2
1071-412X/09/$08.00+0     doi:10.1128/CVI.00066-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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