Cytomegalovirus Immune Reconstitution Occurs in Recipients of Allogeneic Hematopoietic Cell Transplants Irrespective of Detectable Cytomegalovirus Infection

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Cytomegalovirus Immune Reconstitution Occurs in Recipients of Allogeneic Hematopoietic Cell Transplants Irrespective of Detectable Cytomegalovirus Infection Ghislaine Gallez-Hawkins, Lia Thao, Simon F. Lacey, Joybelle Martinez, Xiuli Li, Anne E. Franck, Norma A. Lomeli, Jeff Longmate, Don J. Diamond, Ricardo Spielberger, Stephen J. Forman, John A. Zaia Biology of Blood and Marrow Transplantation Volume 11, Issue 11, Pages 890-902 (November 2005) DOI: 10.1016/j.bbmt.2005.07.008 Copyright © 2005 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 1 ICC/IFN-γ–positive samples expressed as a percentage of all samples tested in patients with CMV reactivation or without CMV reactivation (no-CMV group). After overnight stimulation with individual CMV peptides (pp65-495, IE-1-256, IE-1-297, and IE-1-316), samples were analyzed for IFN-γ by using an intracellular cytokine (ICC) assay as described in “Patients and Methods.” The detection limit was 0.01% of CD8+/IFN-γ–positive cells or 1 × 105 cells per liter after subtraction of background data by using an HIV peptide. Biology of Blood and Marrow Transplantation 2005 11, 890-902DOI: (10.1016/j.bbmt.2005.07.008) Copyright © 2005 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 2 Levels of CMV-specific CD8+/IFN-γ–positive cells at various times after HCT. CMV peptide–specific CD8+ cells were stimulated in the CMV-reactivation group (A and B) and in the no-CMV group (C and D). A and C, Cells stimulated with pp65-495. B and D, Sum of cells stimulated with IE-1-256, IE-1-297, and IE-1-316. The number of IFN-γ–positive cells, expressed as x times 1 × 105 cells per liter, is shown after subtraction of background data by using HIV peptide stimulation. The median value was calculated at each time point and is shown on the graph as a continuous line. Biology of Blood and Marrow Transplantation 2005 11, 890-902DOI: (10.1016/j.bbmt.2005.07.008) Copyright © 2005 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 3 CMV immunity and CMV infection. CMV reactivation is shown for each individual (patients 54, 70, 93, 94, and 105) as a bold line at the top of the graph showing the time to PCR positivity after HCT. The number of pp65-495–specific /IFN-γ–positive cells is shown in (A), and IE-1–specific peptides consisting of IE-1-256, IE-1-297, and IE-1-316 stimulation are shown in (B). The number of IFN-γ–positive cells, expressed as x times 1 × 105 cells per liter (y-axis), is shown after subtraction of background data by using HIV peptide stimulation. The time after HCT for each subject is on the x-axis, and, when available, the number of cells is shown for the donor before and after granulocyte colony-stimulating factor treatment (PreG and PostG, respectively). Biology of Blood and Marrow Transplantation 2005 11, 890-902DOI: (10.1016/j.bbmt.2005.07.008) Copyright © 2005 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 4 Time course of tetramer-positive T cells after HCT. The number of tetramer-positive cells specific to pp65-495 (A and C) and the sum of tetramer-positive cells specific to IE-1-297 and IE-1-316 (B and D) are shown. The left panels (A and B) represent results for subjects with documented CMV reactivation, and the right panels (C and D) represent results in subjects with no documented CMV infection. The number of IFN-γ–positive cells is shown as x times 1 × 105 cells per liter, and the median value is calculated at each time point and shown on the graph as a continuous line. Biology of Blood and Marrow Transplantation 2005 11, 890-902DOI: (10.1016/j.bbmt.2005.07.008) Copyright © 2005 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 5 Levels of CMV-specific CD8+/IFN-γ–positive cells at various times after HCT. An ICC assay on fresh blood from 33 HLA-A*0201 subjects stimulated with CMV-pp65 peptide (A and B) or a mixture of IE-1–specific peptides (CMV IE-1-256, -297, and -316; C and D) is shown. A and C, Number of cells obtained in recipients with detectable CMV reactivation (23 subjects). B and D, Results from those with no detectable CMV infection (10 subjects). Biology of Blood and Marrow Transplantation 2005 11, 890-902DOI: (10.1016/j.bbmt.2005.07.008) Copyright © 2005 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 6 Percentage of HCT recipients with CMV-specific T-cell immune reconstitution. The cumulative incidence of IFN-γ–positive cells stimulated with pp65 (A) or a mixture of IE-1 peptides (B) is shown at the indicated times after HCT. The recipients with detectable CMV infection (CMV group) are shown as a continuous line, and those with no detectable CMV infection (no CMV) are shown as a hatched line. The log-rank test showed no significant differences between the 2 curves in (A) and (B). Biology of Blood and Marrow Transplantation 2005 11, 890-902DOI: (10.1016/j.bbmt.2005.07.008) Copyright © 2005 American Society for Blood and Marrow Transplantation Terms and Conditions