Expression of activating KIR2DS2 and KIR2DS4 genes after hematopoietic cell transplantation: relevance to cytomegalovirus infection.

The important role of activating killer immunoglobulin-like receptors (KIRs) in protecting against cytomegalovirus (CMV) reactivation has been described previously in patients undergoing hematopoietic cell transplantation (HCT). More specifically, the presence of multiple activating KIRs and the presence of at least KIR2DS2 and KIR2DS4 in the donor genotype identified a group of HCT patients at low risk for CMV reactivation. However, CMV infection still occurs in patients with the KIR protective genotype, and the question has been raised as to whether this is related to the lack of KIR expression.

Increased programmed death-1 molecule expression in cytomegalovirus disease and acute graft-versus-host disease after allogeneic hematopoietic cell transplantation.

To study the role of the programmed death-1 molecule (PD-1) in cytomegalovirus (CMV) infection and disease after allogeneic hematopoietic cell transplantation (HCT), 206 subjects were followed prospectively for immune response to CMV and assigned to 3 groups based on CMV outcome. The subjects were analyzed retrospectively for PD-1 expression in cryopreserved CD4+ and CD8+T cells collected at days 40, 90, 120, 150, 180, and 360 posttransplantation. HCT recipients with CMV disease (n=14) were compared with recipients with prolonged CMV infection, but no CMV disease (median duration of infection, 3 months; n=14) and with controls with no CMV infection who received similar transplants (n=22).

Biologic and immunologic effects of knockout of human cytomegalovirus pp65 nuclear localization signal.

The human cytomegalovirus (CMV) pp65 protein contains two bipartite nuclear localization signals (NLSs) at amino acids (aa) 415 to 438 and aa 537 to 561 near the carboxy terminus of CMV pp65 and a phosphate binding site related to kinase activity at lysine-436. A mutation of pp65 with K436N (CMV pp65mII) and further deletion of aa 537 to 561 resulted in a novel protein (pp65mIINLSKO, where NLSKO indicate NLS knockout) that is kinaseless and that has markedly reduced nuclear localization. The purpose of this study was to biologically characterize this protein and its immunogenicity compared to that of native pp65.

Impact of donor CMV status on viral infection and reconstitution of multifunction CMV-specific T cells in CMV-positive transplant recipients.

Reconstitution of cytomegalovirus (CMV)-specific CD8(+) T cells is essential to the control of CMV infection in CMV-positive recipients (R(+)) after allogeneic hematopoietic stem cell transplantation (HCT). Six-color flow cytometry was used to assess the functional profile of CMV-specific CD8(+) T cells in 62 of 178 R(+) HCT recipients followed virologically for CMV reactivation. R(+) recipients receiving grafts from CMV-negative donors (D(-); D(-)/R(+)) reconstituted fewer multifunctional CD8(+) T cells expressing tumor necrosis factor-alpha (TNF-alpha), macrophage inflammatory protein-1beta (MIP-1beta), and CD107 in addition to interferon-gamma (IFN-gamma), compared with D(+)/R(+) recipients.

The effect of single and combined activating killer immunoglobulin-like receptor genotypes on cytomegalovirus infection and immunity after hematopoietic cell transplantation.

It has been shown that activating killer Ig-like receptor (aKIR) genes are important for control of cytomegalovirus (CMV) reactivation after hematopoietic cell transplantation (HCT). To date, using the broad classification of KIR haplotypes A and B, the precise role of individual KIR genes in the control of infection cannot be discerned. To address this, a consecutive case series of 211 non-T cell-depleted HCT patients all at risk for CMV were monitored biweekly for CMV DNA in plasma by quantitative polymerase chain reaction (Q-PCR) and at intervals for CMV-specific T cell immunity.

Functional comparison of T cells recognizing cytomegalovirus pp65 and intermediate-early antigen polypeptides in hematopoietic stem-cell transplant and solid organ transplant recipients.

The functional status of cytotoxic T lymphocyte (CTL) populations recognizing cytomegalovirus intermediate-early antigen (IE1) and pp65 polypeptides was investigated in peripheral blood mononuclear cells from hematopoietic stem-cell transplant (HSCT) and solid organ transplant recipients. Combined flow-based CD107a/b degranulation/mobilization and intracellular cytokine (ICC) assays using peptide libraries as antigens indicated that a significantly higher proportion of pp65-specific CTLs were in a more mature functional state, compared with IE1-specific CTLs. Degranulation/multiple cytokine ICC assays also indicated that a significantly higher proportion of pp65-specific than IE1-specific CTLs secreted both interferon- gamma and tumor necrosis factor- alpha and possessed greater cytotoxic potential.

Cytomegalovirus immune reconstitution occurs in recipients of allogeneic hematopoietic cell transplants irrespective of detectable cytomegalovirus infection.

The question of when immune reconstitution of cytomegalovirus (CMV)-specific CD8 T cells occurs after hematopoietic cell transplantation and, more specifically, to which CMV targets this immunity is likely to be directed remains poorly understood. The dependence of immune reconstitution on CMV reactivation is even less clear. To better understand these events, 44 CMV-seropositive HLA-A*0201 subjects were followed up at approximately days 40, 90, 120, 150, 180, and 360 after hematopoietic cell transplantation for CMV immunity as measured by 2 types of assays: (1) an HLA-A*0201 tetramer-binding assay for both CMV pp65 (pp65) and immediate-early 1 (IE-1) or (2) intracellular cytokine interferon gamma responses induced by pp65 or IE-1-derived peptides.

Simultaneous reconstitution of multiple cytomegalovirus-specific CD8+ cell populations with divergent functionality in hematopoietic stem-cell transplant recipients.

A panel of 7 human cytomegalovirus (CMV) epitope peptides and corresponding major histocompatibility class 1 tetramers was used to evaluate cellular immunity in healthy seropositive donors and in hematopoietic stem-cell transplant recipients. Broad CMV-specific T cell responses to epitopes were found within several CMV polypeptides and were restricted by multiple human leukocyte antigen alleles. Their cytotoxic functionality was evaluated by use of an assay that measures transient surface levels of lysosomal membrane proteins LAMP-1 (CD107a) and LAMP-2 (CD107b) after peptide stimulation.

DNA and low titer, helper-free, recombinant AAV prime-boost vaccination for cytomegalovirus induces an immune response to CMV-pp65 and CMV-IE1 in transgenic HLA A*0201 mice.

A prime-boost immunization regimen allowed the use of low titer, helper-free rAAV-pp65mII and rAAV-IE1 virus to elicit specific humoral and cellular responses to two important cytomegalovirus (CMV) antigens: the immediate-early 1 (IE-1) and pp65 proteins. Simultaneous immunization of both CMV proteins, using DNA vaccine priming followed by rAAV boost, induced antibody (Ab) response, CD8 lymphocytes with cytotoxic function, and detectible binding of the cognate peptide epitopes for human HLA A*0201 restriction using tetramer technology.

Synthesis and biological evaluation of a new class of geldanamycin derivatives as potent inhibitors of Hsp90.

The heat shock protein Hsp90 has increasingly become an important therapeutic target especially for treatment of cancers. Inhibition of the ATPase activity of Hsp90 by natural products (e.g.

A high-affinity conformation of Hsp90 confers tumour selectivity on Hsp90 inhibitors.

Heat shock protein 90 (Hsp90) is a molecular chaperone that plays a key role in the conformational maturation of oncogenic signalling proteins, including HER-2/ErbB2, Akt, Raf-1, Bcr-Abl and mutated p53. Hsp90 inhibitors bind to Hsp90, and induce the proteasomal degradation of Hsp90 client proteins. Although Hsp90 is highly expressed in most cells, Hsp90 inhibitors selectively kill cancer cells compared to normal cells, and the Hsp90 inhibitor 17-allylaminogeldanamycin (17-AAG) is currently in phase I clinical trials.