Cytomegalovirus (CMV) infections remain a serious problem in hematopoietic stem cell transplant patients. Following transplantation CMV infections can cause peumonititis, hepatitis, enteritis, and marrow failure. CMV seropositive transplant recipients can be treated with antiviral agents such as ganciclovir at the onset of infection or at the time of stem cell engraftment, but ganciclovir therapy is associated with renal toxicity and suppression of neutrophil counts. Preliminary studies have found that adoptive immune therapy using CMV-reactive cytotoxic T lymphocytes (CTL) is an effective and less toxic alternative to prevention of CMV infection in transplant recipeints. The purpose of this study is to develop new treatment strategies for producing CMV-reactive CTLs that can be used for adoptive immunetherapy and to better understand the cellular immune response to CMV. Current studies are focused on identifying the immune dominant peptides that can be used to stimulate CMV reactive CTLs. CMV contains over 200 proteins, but one protein, CMV matrix protein pp65, has been found to be immune dominant. We found that the peptide CMV pp65 91-100 was an immune dominant peptide restricted to HLA-A*33 and pp65 328-337 was an immune dominant peptide restricted to HLA-A*2402. It has been generally thought that there is only one virus- or tumor-specific immune dominant peptide for each HLA type, but our studies have found that there are often more than one immune dominant pp65 peptide for some HLA types. We found three different pp65 peptides to be immune dominant for HLA-A*03. In addition, some pp65 peptides are not restricted to a single HLA antigen. We found that peptide pp65 341-350 stimulated T cells expressing either HLA-A*0101 or HLA-A*2402. We have also begun collaborative studies with Drs. John Barrett and Scott Solomon of the NHLBI. They are studying a canary pox vaccine containing the gene for CMV pp65. The vaccine is being given to hematopoietic stem cell donors prior to the collection and transplantation of their stem cells. We are monitoring the donors' immune response to the pp65 vaccination. We will also be monitoring the recipients' immune response to pp65 following transplantation. In a pilot study we found that it is possible to monitor the immune response to CMV by stimulating lymphocytes with pools of peptides 15 amino acids in length that over lap at 4 amino acids and measuring intracellular production of interferon-gamma. This procedure will be used to monitor the pp65 vaccinated donor and the transplant recipient.
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