CMV pp65 (113-121) – VYALPLKML – Epitope peptide of Cytomegalovirus

CMV-specific T cells generated from naïve T cells recognize atypical epitopes and may be protective in vivo

Adoptive transfer of cytomegalovirus (CMV)-specific T cells derived from adult seropositive donors can effectively restore antiviral immunity after transplantation. However,CMV-seronegative donors lack CMV-specific memory T cells,which restricts the availability of virus-specific T cells for immunoprophylaxis. We demonstrate the feasibility of deriving CMV-specific T cells from naïve cells for T cell therapy. Naïve T cells primed to recognize CMV were restricted to different,atypical epitopes than T cells derived from CMV-seropositive individuals; however,these two cell populations had similar avidities. CMV-seropositive individuals also had T cells recognizing these atypical epitopes,but these cells had a lower avidity than those derived from the seronegative subjects,which suggests that high-avidity T cells to these epitopes may be lost over time. Indeed,recipients of cord blood (CB) grafts who did not develop CMV were found by clonotypic analysis to have T cells recognizing atypical CMVpp65 epitopes. Therefore,we examined unmanipulated CB units and found that T cells with T cell receptors restricted by atypical epitopes were the most common,which may explain why these T cells expanded. When infused to recipients,naïve donor-derived virus-specific T cells that recognized atypical epitopes were associated with prolonged periods of CMV-free survival and complete remission. These data suggest that naïve-derived T cells from seronegative patients may be an additional source of cells for CMV immunoprophylaxis.

Cytomegalovirus (CMV) phosphoprotein 65 makes a large contribution to shaping the T cell repertoire in CMV-exposed individuals

Antigen-specific,cytokine flow cytometry was used to analyze the prevalence and frequency of CD4 and CD8 memory T cells specific for the abundantly expressed cytomegalovirus (CMV) phosphoprotein 65 (pp65) in healthy CMV IgG-seropositive individuals. Stimulation of peripheral blood mononuclear cells with peptide pools and individual peptides derived from the pp65 amino acid sequence in 40 donors revealed that 63% of donors had a detectable CD4 T cell response and that 83% of donors had a detectable CD8 T cell response against this protein. The overall frequencies of T cells directed against pp65 were analyzed for 20 donors by stimulation with peptide pools covering the complete pp65 protein and were as high as 2 in 1000 and 9 in 1000 (median) peripheral blood CD4 and CD8 T cells,respectively. In addition,a comparison between CD4 responses to a CMV lysate containing various CMV proteins and pp65-specific responses in 9 donors indicated that pp65 was a dominant target of the CMV-specific CD4 T cell response in some,but not all,donors. Several new T cell epitopes were identified.

CMV pp65 antigen testing is of limited utility in the diagnosis of concomitant CMV disease in HIV-infected patients in the HAART era

BACKGROUND: There are limited data on the utility of the CMV pp65 antigen (Ag) test for the diagnosis of concomitant CMV end-organ disease (EOD) in HIV+ patients in the highly active antiretroviral therapy era.

OBJECTIVES: We sought to assess the predictive value of a single pp65 test for the diagnosis of concomitant CMV EOD in HIV-infected patients.

METHODS: A review of all pp65 Ag tests conducted at a large county teaching hospital from January 1998 through July 1999 was conducted. A diagnosis of CMV EOD required histopathologic evidence (except for retinitis). Concomitant disease was defined as CMV EOD within 30 days of Ag test. Results were reported as number of Ag positive cells/300000 cells counted.

RESULTS: Two-hundred and thirty patient charts (308 antigen tests) were reviewed. The median follow-up time was 334 days. Thirty-two patients had a prior diagnosis of CMV EOD. The most common reasons for testing were fever (45),pneumonia (10),and monitoring for recurrent retinitis (8). Ag tests were positive (range 1-1042 cells) in 51 patients. Twelve patients were diagnosed with concomitant CMV EOD. A diagnosis other than CMV was made in a significant majority of patients (154). The mean initial pp65 level was significantly higher in patients with concomitant CMV EOD versus those without concomitant CMV: 314 cells versus 13 cells,P<0.0001. The overall sensitivity,specificity,positive predictive value (PPV),and negative predictive value (NPV) were 67,81,16 and 98%,respectively. Using a 50-cell cutoff and including only patients with CMV symptoms and CD4<100,improved test accuracy: sensitivity,specificity,PPV,and NPV of 60,91,60,and 91%. The CD4 cell count but not the HIV viral load was predictive of CMV EOD.

CONCLUSION: The CMV pp65 Ag test is useful in excluding concomitant CMV disease,but has limited utility in the diagnosis of acute CMV disease. The highest test utility will be in patients with a high likelihood of CMV disease based on symptoms,and CD4 cell count.