ABSTRACT
The use of WT1-specific CTL is one potential strategy to treat leukemic relapse following allogeneic stem cell transplant (SCT). Previous studies have largely focused on generating WT1-CTL from adult donors by cloning. We demonstrate that WT1-CTL can be generated from healthy adult donors and from cord blood by stimulating with an overlapping pool of peptides derived from full length WT1 and selecting antigen-specific cells based on the expression of CD137. The rapid expansion with anti-CD3 and IL-2 resulted in a 100-200-fold expansion. These CTL lysed WT1 expressing targets, including leukemia lines, in a HLA restricted manner.
Subject(s)
Fetal Blood/cytology , Hematopoietic Stem Cells/cytology , Leukemia/immunology , Leukemia/therapy , T-Lymphocytes, Cytotoxic/immunology , Tissue Donors , WT1 Proteins/immunology , Adult , Cell Line, Tumor , Feasibility Studies , Fetal Blood/immunology , Flow Cytometry , HLA-A2 Antigen/immunology , Hematopoietic Stem Cell Transplantation , Hematopoietic Stem Cells/immunology , Humans , Immunophenotyping , Immunotherapy, Adoptive , Interferon-gamma , Leukemia/genetics , Lymphocyte Count , Lymphocytes/immunology , Peptide Fragments/immunology , Transplantation, Homologous , WT1 Proteins/geneticsABSTRACT
We report a stem cell transplant patient with a therapy-refractory cytomegalovirus (CMV) infection who received CMV-specific T cells from his sero-negative stem cell donor. This donor received the Towne strain CMV vaccine, and T cells were expanded using monocytes pulsed with pp65 overlapping peptides. CMV DNA decreased after the CTL infusion, and CMV-specific cytotoxicity increased. This strategy could be implemented in similar situations or with persistent viremia post-transplant.
Subject(s)
Cytomegalovirus Infections/therapy , Cytomegalovirus/immunology , Immunotherapy, Adoptive/methods , Phosphoproteins/immunology , T-Lymphocytes, Cytotoxic/immunology , Viral Matrix Proteins/immunology , Cell Culture Techniques , Cells, Cultured , Cytomegalovirus Infections/immunology , Cytomegalovirus Vaccines , Humans , Male , Stem Cell Transplantation/adverse effects , Young AdultABSTRACT
Adoptive immunotherapy with antigen-specific cytotoxic T lymphocytes (CTLs) has proven effective in restoring cellular immunity to cytomegalovirus (CMV) and preventing viral reactivation after allogeneic stem cell transplantation (SCT). In an effort to develop a cost-effective, relatively rapid method of CMV CTL expansion, we investigated the use of a pool of overlapping CMV peptides. Because the possibility exists of vaccinating CMV-seronegative donors, and these individuals may have T cell responses predominantly against IE-1, commercially available peptide mixes for pp65 as well as IE-1 were used to stimulate CTLs from 10 seropositive donors. Of these 10 donors, 4 responded to pp65 only, 1 did not respond to either pp65 or IE-1, 4 responded to both pp65 and IE-1, and 1 responded to IE-1 only. These CMV- specific T cells included a mixture of CD4(+) and CD8(+) effectors, and specific cytotoxicity correlated with interferon-gamma production. The costs associated with a 28-day maintenance course of intravenous ganciclovir, cidofovir, foscarnet, and valganciclovir, as well as the preparation and shipping a single dose of CTLs, were determined. The price of generating CMV CTLs using this method was comparable to or less expensive than a 28-day maintenance course for these agents, not including the costs associated with drug administration, supportive care, and the treatment of drug-related complications. Considering the relative ease, low cost, and the fact that CTL administration can result in CMV-specific immune reconstitution, this option should be considered for patients with CMV reactivation or for prophylaxis in patients at high risk for infection.
Subject(s)
Antigens, Viral/therapeutic use , Cytomegalovirus Infections/therapy , Cytomegalovirus/immunology , Immunotherapy/methods , T-Lymphocytes, Cytotoxic/immunology , Antigens, Viral/immunology , Cell Proliferation , Cells, Cultured , Humans , Immediate-Early Proteins/immunology , Lymphocyte Activation/immunology , Phosphoproteins/immunology , T-Cell Antigen Receptor Specificity/immunology , T-Lymphocytes, Cytotoxic/cytology , Viral Matrix Proteins/immunologyABSTRACT
The etiologic agent of adult T-cell leukemia (ATL) is human T cell lymphotropic virus type I (HTLV-I). The HTLV-I protein Tax alters gene expression, including those of cytokines and their receptors, which plays an important role in early stages of ATL. Here we demonstrate that expression of interleukin-9 (IL-9) is activated by Tax via an NF-kappaB motif in its proximal promoter, whereas IL-9 receptor-alpha (IL-9Ralpha) expression is not induced by Tax. However, supporting a role for IL-9/IL-9Ralpha in ATL, a neutralizing monoclonal antibody directed toward IL-9Ralpha inhibited ex vivo spontaneous proliferation of primary ATL cells from several patients. Fluorescence-activated cell sorter analysis of freshly isolated peripheral blood mononuclear cells from these patients revealed high level expression of IL-9Ralpha on their CD14-expressing monocytes. Furthermore, purified T cells or monocytes alone from these patients did not proliferate ex vivo, whereas mixtures of these cell types manifested significant proliferation through a contact-dependent manner. Taken together, our data suggest that primary ATL cells, via IL-9, support the action of IL-9Ralpha/CD14-expressing monocytes, which subsequently support the ex vivo spontaneous proliferation of malignant T cells. In summary, these data support a role for IL-9 and its receptor in ATL by a paracrine mechanism.