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Journal of Virology, September 2001, p. 7840-7847, Vol. 75, No. 17
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.17.7840-7847.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Ex Vivo Stimulation and Expansion of both CD4+ and CD8+ T Cells from Peripheral Blood Mononuclear Cells of Human Cytomegalovirus-Seropositive Blood Donors by Using a Soluble Recombinant Chimeric Protein, IE1-pp65

Jocelyn Vaz-Santiago,1 Jacqueline Lulé,2 Pierre Rohrlich,3 Céline Jacquier,1 Nicolas Gibert,2 Emmanuelle Le Roy,2 Didier Betbeder,1 Jean-Luc Davignon,2 and Christian Davrinche2,*

Inserm U395, IFR 30, UPS, CNRS, CHU, 31024 Toulouse Cédex,2 Biovector Therapeutics, 31676 Labège Cédex,1 and Unité d'Immunité Cellulaire Anti-Virale, Institut Pasteur, 75724 Paris,3 France

Received 6 March 2001/Accepted 29 May 2001

The transfer of anti-human cytomegalovirus (HCMV) effector T cells to allogeneic bone marrow recipients results in protection from HCMV disease associated with transplantation, suggesting the direct control of CMV replication by T cells. IE1 and pp65 proteins, both targets of CD4+ and CD8+ T cells, are considered the best candidates for immunotherapy and vaccine design against HCMV. In this report, we describe the purification of a 165-kDa chimeric protein, IE1-pp65, and its use for in vitro stimulation and expansion of anti-HCMV CD4+ and CD8+ T cells from peripheral blood mononuclear cells (PBMC) of HCMV-seropositive donors. We demonstrate that an important proportion of anti-HCMV CD4+ T cells was directed against IE1-pp65 in HCMV-seropositive donors and that the protein induced activation of HLA-DR3-restricted anti-IE1 CD4+ T-cell clones, as assessed by gamma interferon (IFN-gamma ) secretion and cytotoxicity. Moreover, soluble IE1-pp65 stimulated and expanded anti-pp65 CD8+ T cells from PBMC of HLA-A2, HLA-B35, and HLA-B7 HCMV-seropositive blood donors, as demonstrated by cytotoxicity, intracellular IFN-gamma labeling, and quantitation of peptide-specific CD8+ cells using an HLA-A2-peptide tetramer and staining of intracellular IFN-gamma . These results suggest that soluble IE1-pp65 may provide an alternative to infectious viruses used in current adoptive strategies of immunotherapy.

* Corresponding author. Mailing address: INSERM U395, IFR 30, UPS, CNRS, CHU, BP 3028, 31024 Toulouse Cédex, France. Phone: 33 5 62 74 83 85. Fax: 33 5 62 74 83 86. E-mail: davrinch{at}purpan.inserm.fr.

Journal of Virology, September 2001, p. 7840-7847, Vol. 75, No. 17
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.17.7840-7847.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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