Betaherpesvirus Reactivation and Toll-Like Receptor Expression After Allogeneic Stem Cell Transplantation

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4924-4924
Author(s):  
Cristina Skert ◽  
Manuela Fogli ◽  
Simone Perucca ◽  
Simona Fiorentini ◽  
Emirena Garrafa ◽  
...  
Keyword(s):  
Stem Cell ◽  
Immune System ◽  
Stem Cell Transplantation ◽  
Nucleic Acids ◽  
Cell Transplantation ◽  
Allogeneic Stem Cell Transplantation ◽  
Receptor Expression ◽  
Structural Proteins ◽  
Type I ◽  
Cmv Reactivation

Abstract Abstract 4924 Introduction. β-herpesviruses, such as CMV and HHV6, are important pathogen in transplanted patients. The morbidity because of CMV reactivation after allogeneic stem cell transplantation (SCT) has led to the monitoring of this virus and to introduction of preemptive therapy. However, CMV infection is still one of the most challenging complications, because CMV disease may occur as life-threatening pneumonitis, and may increase the risk of opportunistic infections. HHV6 reactivation has been demonstrated after SCT and this virus is recognized as important pathogen, either by direct infection or via interaction with CMV. Innate and adaptive immune response against these viruses involves the activation of Toll-like receptors (TLRs). TLRs belong to type I transmembrane glycoprotein receptor family and recognize pathogen-associated molecular patterns (PAMPs). Viral nucleic acids and viral structural proteins, such as glycoproteins, are considered as PAMPs. Endosomal TLRs (TLR3, 7, 8 and 9) recognize viral nucleic acids and some surface TLRs may be involved in the detection of structural proteins. Some clinical and experimental evidences indicate that CMV and HHV-6 can modulate the immune system and influence the immune reconstitution after SCT. However, the role of TLRs in this complex interplay remains unclear, especially in the setting of allogeneic SCT. The aim of this study was to evaluate the expression of TLRs on lymphocytes and monocytes in relation to CMV and HHV6 reactivation in the early period after allogeneic SCT. Methods. CMV and HHV6 reactivation was monitored weekly by quantitative real-time PCR until the second month after SCT. The expression of TLRs on lymphocytes and monocytes was analysed by flow cytometry as mean fluorescence intensity at day +30 and in any case before CMV or HHV6 reactivation. Functional data were obtained by ELISA assay after TLRs activation. The cell supernatants were collected and assayed for TNF-alpha, IFN-gamma and MCP-1. Relative induction of these cytokines was calculated in relation with unstimulated controls. Results. CMV reactivation within 2 months after transplantation was observed in 13 out of 33 patients. CMV pneumonitis was observed in 1 patient. HHV-6 reactivation was detected in 1 patient. Median age was 45 years (range, 22–64) and 21 patients were male. TLRs expression and function did not significantly differ in controls and patients without CMV. Lymphocytes of patients with CMV reactivation showed an increased expression of TLR5 (4,1±2,4 vs 2,0±1,7 p=0,008). TLR8 expression was lower on monocytes with CMV reactivation (0,8±0,9 vs 2,0±1,7 p=0,03). MCP-1 relative induction post-stimulation of TLR1 and 8 was significantly decreased in patients with CMV reactivation (p<0,04). Conclusions. Surface TLR2 and intracellular TLR3 and 9 are reported to recognize CMV by some authors. In our study, surface TLR5 and intracellular TLR8 seem to be involved in the interaction between CMV and the immune system of transplanted patients. In particular, TLR8 could play a protective role. MCP-1 production upon TLR1 and 8 activation negatively correlates with CMV reactivation. The defective immune system after SCT could explain these results, which could be confirmed by the assessment of a larger number of patients and the analysis of other possible interfering factors. Disclosures: No relevant conflicts of interest to declare.

Expression of Toll-Like Receptors on Peripheral Blood Cells After Allogeneic Stem Cell Transplantation: Results of a Prospective Study,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4071-4071
Author(s):  
Cristina Skert ◽  
Manuela Fogli ◽  
Simone Perucca ◽  
Simona Fiorentini ◽  
Emirena Garrafa ◽  
...  
Keyword(s):  
Stem Cell ◽  
Immune System ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Allogeneic Stem Cell Transplantation ◽  
Acute Gvhd ◽  
Adaptive Immune System ◽  
Type I ◽  
Ifn Gamma ◽  
Adaptive Immune

Abstract Abstract 4071 Introduction. Emerging trends emphasize the importance of both innate and adaptive immune system in the response against infections, and in the pathogenesis of autoimmune and graft-versus-host (GVHD) diseases. Pattern recognition receptors such as Toll-like receptors (TLRs) play a key role in the cross-talk between innate and adaptive immune system. TLRs belong to type I transmembrane glycoprotein receptor family and recognize pathogen-associated molecular patterns (PAMPs), such as common protein, carbohydrate or DNA/RNA pattern motifs. TLRs are also receptors for endogenous ligands and damaged tissue, suggesting that both pathogen-derived molecules and products of damaged tissue can trigger signals which are responsible for the regulation of innate and adaptive immune responses. Extracellular ligands are recognized by surface TLRs (TLR1,TLR2,TLR4,TLR5, and TLR6). Intracellular TLRs (TLR3,TLR7,TLR8 and TLR9) bind mainly to foreign nucleic acids and sometimes detect self DNA/RNA. Aim of the study. Very little is known about expression and function of TLRs in vivo in patients who underwent allogeneic stem cell transplantation (SCT). The aim of this study was to evaluate the expression of TLRs on lymphocytes and monocytes in relation to the onset of acute GVHD. Methods. The expression of TLRs on lymphocytes and monocytes was analysed by flow cytometry as mean fluorescence intensity at day +30 and at the onset of GVHD. Functional data were obtained by ELISA assay after TLRs activation. The cell supernatants were collected and assayed for TNF-alpha, IFN-gamma and MCP-1. Relative induction of these cytokines was calculated in relation with unstimulated controls. Results. We analyzed 17 healthy donors and 34 patients. Median age was 46 years (range, 22–64) and 22 patients were male. Acute GVHD developed in 19 patients (12 with grade >=2). Clinical and transplant characteristics did not differ in patients with and without GVHD. Lymphocytes and monocytes of patients with acute GVHD showed higher levels of TLR5 (3,5±2,3 vs1,9±1,6 p=0,03; 25,8±25,9 vs 9,0±5,0 p=0,02) and a decreased expression of TLR1 (2,5±2,8 vs 4,3±2,8 p=0,02; 21,4±21,9 vs 54,9±37,4 p=0,005) and TLR9 (63,8±30,4 vs 111,1±62,9 p=0,03; 85,3±73,9 vs 164,2±90,6 p=0,01). IFN-gamma relative induction post-stimulation of TLR2,3,4 and 9 was significantly decreased in patients with acute GVHD (p< 0,04). Conclusions. TLRs show a different profile of expression in patients with acute GVHD in comparison with patients without it. These results suggest that the innate immune response via TLRs activation could be involved in the development of GVHD. In particular, a decreased expression of TLR-9 (receptor of hypomethylated DNA) on lymphocytes and monocytes can promote TLR-7 activation, inducing type I interferons and other pro-inflammatory cytokines. TLR-1 and −5, which are ligands for bacterial cell wall, could also be involved in the pathogenesis of GVHD. Moreover, acute GVHD negatively correlates with IFN-gamma production upon TLR2,3,4 and 9 activation. The assessment of a larger number of patients could be useful to understand the complex interplay among pathogens, self or non-self DNA and RNA, and the immune system. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Are the Polyomaviruses BK and JC Associated with Opportunistic Infections, Graft-versus-Host Disease, or Worse Outcomes in Adult Patients Receiving Their First Allogeneic Stem Cell Transplantation with Low-Dose Alemtuzumab?

2017 ◽  
Vol 138 (1) ◽  
pp. 3-9 ◽  
Author(s):  
Laila Schneidewind ◽  
Thomas Neumann ◽  
Florian Knoll ◽  
Kathrin Zimmermann ◽  
Sigrun Smola ◽  
...  
Keyword(s):  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Graft Versus Host Disease ◽  
Allogeneic Stem Cell Transplantation ◽  
Opportunistic Infections ◽  
Hemorrhagic Cystitis ◽  
Adult Patients ◽  
Graft Versus Host ◽  
Cmv Reactivation

Background: The association of polyomaviruses BK and JC with other opportunistic infections and graft-versus-host disease (GvHD) in allogeneic stem cell transplantation is controversially discussed. Methods: We conducted a retrospective study of 64 adult patients who received their first allogeneic stem cell transplantation between March 2010 and December 2014; the follow-up time was 2 years. Results: Acute leukemia was the most frequent underlying disease (45.3%), and conditioning included myeloablative (67.2%) and nonmyeloablative protocols (32.8%). All patients received 10 mg of alemtuzumab on day -2 (20 mg in case of mismatch) as GvHD prophylaxis. Twenty-seven patients (41.5%) developed cytomegalovirus (CMV) reactivation. BKPyV-associated hemorrhagic cystitis was diagnosed in 10 patients (15.6%). Other opportunistic infections caused by viruses or protozoa occurred rarely (<10%). There was no association of BKPyV or JCPyV with CMV reactivation, Epstein-Barr virus reactivation, human herpes virus 6, or parvovirus B19 infection requiring treatment. There was a significant correlation of BKPyV-associated hemorrhagic cystitis with toxoplasmosis (p = 0.013). Additionally, there was a significant link of simultaneous BKPyV and JCPyV viruria with toxoplasmosis (p = 0.047). BKPyV and JCPyV were not associated with GvHD, relapse, or death. Conclusion: We found no association of BKPyV or JCPyV with viral infections or GvHD. Only the correlation of both polyomaviruses with toxoplasmosis was significant. This is a novel and interesting finding.

Valganciclovir in the Prevention of Cytomegalovirus Following Allogeneic Stem Cell Transplantation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2851-2851
Author(s):  
Margarida Magalhaes-Silverma ◽  
Karen Parrott ◽  
Thomas Carter ◽  
Christina Leopold ◽  
Hohl Raymond ◽  
...  
Keyword(s):  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Allogeneic Stem Cell Transplantation ◽  
Prospective Trial ◽  
Intravenous Ganciclovir ◽  
Shell Vial ◽  
Early Results ◽  
Antigenemia Assay ◽  
Cmv Reactivation

Abstract BACKGROUND: Cytomegalovirus (CMV) infection remains an important cause of morbidity and mortality after allogeneic stem cell transplantation. Valganciclovir, the valine ester of ganciclovir, has excellent oral bioavailability and has the potential to replace intravenous ganciclovir in may situations, namely CMV prophylaxis or pre-emptive therapy after allogeneic stem cell transplantation. METHODS: From August 2002 to December 2005, 58 patients (pts) who were either CMV positive or had a seropositive donor were enrolled in a prospective trial of intravenous (IV) ganciclovir at 5 mg/Kg twice a day for one week, followed by oral valganciclovir 900 mg once daily for a total of 180 days. IV ganciclovir was started at time of engraftment. Dose adjustments of valganciclovir were made according to renal function; growth factors were allowed in the event of neutropenia. Patients were monitored with weekly antigenemia. Study endpoints included incidence of CMV reactivation and disease during the first 180 days after transplantation. For the study, viremia was defined as a positive CMV blood culture by shell vial or conventional culture. A positive antigenemia assay in pts with severe (grade III–IV) GVHD was defined as 1 positive cell on either of 2 duplicate slides. For pts with no or mild GVHD, a positive antigenemia assay was defined as 2 positive cells/slide. RESULTS: Fifty eight pts were enrolled. The median age was 50 years (range 18–71). Twenty eight pts underwent a sibling transplant and 30 pts unrelated transplant. Twenty pts were transplanted using a reduced intensity regimen (12 received Campath-1H) and 38 pts received a standard ablative regimen with T cell depletion. There were 47 seropositive recipients and 33 seropositive donors. Fourteen pts were not able to proceed after consenting due to screening positive antigenemia (2 pts), financial reasons (2 pts), severe gut GVHD unabling oral intake (4 pts), death (4 pts), acyclovir resistant HSV infection treated with other antiviral agents (2 pts). Forty four pts received valganciclovir. Neutropenia was observed in 7 pts, resolving in all of them. Valganciclovir was discontinued temporarily in these pts. Four of them were receiving concomitant mycophenolate mofetil. No adverse effects were otherwise reported. Nineteen pts failed to complete the study due to diagnosis of severe gut GVHD (8 pts), financial reasons (1 pt), death due to disease progression (5 pts), relapse/progression of underlying disease (5pts). Twenty six pts have completed at least 3 months of therapy and 21 pts completed 180 days. One pt experienced CMV reactivation (not taking the precribed dose). No other pt experienced CMV reactivation or disease while on therapy. CONCLUSION: These early results suggest that valganciclovir has a role in the prevention of CMV after allogeneic stem cell transplantation. Valganciclovir is well tolerated and is an attractive oral alternative for prevention of CMV.

The role of Vδ2-negative γδ T cells during cytomegalovirus reactivation in recipients of allogeneic stem cell transplantation

Blood ◽  
2010 ◽  
Vol 116 (12) ◽  
pp. 2164-2172 ◽  
Author(s):  
Andrea Knight ◽  
Alejandro J. Madrigal ◽  
Sarah Grace ◽  
Janani Sivakumaran ◽  
Panagiotis Kottaridis ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Allogeneic Stem Cell Transplantation ◽  
Γδ T Cells ◽  
Cell Receptor ◽  
Barr Virus ◽  
Cmv Reactivation

Abstract Reactivation of cytomegalovirus (CMV) remains a serious complication after allogeneic stem cell transplantation, but the role of γδ T cells is undefined. We have studied the immune reconstitution of Vδ2negative (Vδ2neg) γδ T cells, including Vδ1 and Vδ3 subsets and Vδ2positive (Vδ2pos) γδ T cells in 40 patients during the first 24 months after stem cell transplantation. Significant long-term expansions of Vδ2neg but not Vδ2pos γδ T cells were observed during CMV reactivation early after transplantation, suggesting direct involvement of γδ T cells in anti-CMV immune responses. Similarly, significantly higher numbers of Vδ2neg γδ T cells were detected in CMV-seropositive healthy persons compared with seronegative donors; the absolute numbers of Vδ2pos cells were not significantly different. The expansion of Vδ2neg γδ T cells appeared to be CMV-related because it was absent in CMV-negative/Epstein-Barr virus-positive patients. T-cell receptor-δ chain determining region 3 spectratyping of Vδ2neg γδ T cells in healthy subjects and patients showed restricted clonality. Polyclonal Vδ2neg cell lines generated from CMV-seropositive healthy donors and from a recipient of a graft from a CMV-positive donor lysed CMV-infected targets in all cases. Our study shows new evidence for role of γδ T cells in the immune response to CMV reactivation in transplantation recipients.

scholarly journals Clonal Dynamics and Publicness of CMV-Specific TCR Repertoire after Allogeneic Stem Cell Transplantation

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4551-4551
Author(s):  
Takashi Toya ◽  
Ayumi Taguchi ◽  
Kazutaka Kitaura ◽  
Yuki Otsuka ◽  
Ryosuke Konuma ◽  
...  
Keyword(s):  
Stem Cell ◽  
Next Generation Sequencing ◽  
T Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Allogeneic Stem Cell Transplantation ◽  
Peripheral Blood ◽  
Tcr Repertoire ◽  
Cmv Reactivation ◽  
Generation Sequencing

[Background] Cytomegalovirus (CMV) disease is a major complication after allogeneic stem cell transplantation (SCT). However, mechanisms of adoptive immunity against CMV are not fully elucidated. Recently, high-throughput next-generation sequencing (NGS) technology made it possible to shed light on the detailed and comprehensive landscape of T-cell receptor (TCR) repertoire. In this study, we analyzed TCR repertoire of CMV-specific cytotoxic T-cells (CMV-CTLs) in patients who suffered from CMV reactivation after SCT to clarify the diversity and dynamics of CMV-specific T-cell immunity. [Methods] We sequentially collected peripheral blood mononuclear cells from patients with HLA-A*24:02 who received SCT in our institution. Samples were collected weekly or every two weeks from their neutrophil engraftment until approximately 100 days after SCT. CMV reactivation was evaluated weekly with CMV antigenemia test. CD8 and CMV pp65 tetramer positive cells were sorted and unbiased next-generation sequencing-based analyses of TRBV/TRBJ gene segments were performed at the timing of CMV reactivation in 16 patients, and TRA gene segments were also analyzed in 10 patients. In addition, TCR beta repertoires after 2-4 weeks were analyzed in 12 patients. In the 12 patients, the dynamics of TCR repertoire diversity and proportional changes of each clone were assessed. We evaluated the diversity by Shannon-Weber index, and we defined TCR beta clonotypes found in two or more patients using the same TRBV/TRBJ gene segments and CDR3 amino acid sequences as public. This study was approved by the ethics committee of Tokyo Metropolitan Komagome Hospital. [Results] Among 16 patients, 11 received bone marrow, 3 received peripheral blood stem cells, and 2 received cord blood transplant. Underlying diseases were acute myeloid leukemia in 7, acute lymphoblastic leukemia in 7, and myelodysplastic syndromes in 2 patients. Median age at SCT was 50 years old (range: 20-71). Median duration from SCT to first CMV reactivation was 39 days (range: 16-55) and 7 patients (43.8%) were administered systemic corticosteroid at the time of reactivation (prednisolone 10-30mg/day). Median peripheral blood CMV-CTLs count at that time was 29.47/uL (range: 4.65-229.6). In most patients TCR beta repertoire of CMV-specific CTLs when CMV reactivated was highly skewed (median Shannon-Weber index was 1.44 [range: 0.542-3.164]). TCR alpha and beta were analyzed together in 10 patients and their diversity correlated well (p<0.001). Interestingly, 12 of 16 patients (75.0%) had at least one public TCR and, in all 12 patients with public TCR, the most frequent TCRs were public. Diversity of TCR repertoire was significantly lower in patients with public TCR compared with those in patients without (p=0.039). There was no obvious association between presence of public TCR and multiple events of CMV reactivation (p=0.57). When we evaluated clonal transition in 12 patients whose TCR beta repertoires were analyzed sequentially, the public clonotypes were continuously detected in 10 of 11 patients with public TCR and remained dominant in 9, while in one patient frequency of the most frequent public clone declined massively (from 50.25% among assigned reads to 6.63%). In addition, there were two patterns of subsequent clonal behavior in TCR repertoire of CMV-CTLs. In 10 patients, prevailing clones persisted and TCR repertoires of CMV-CTLs remained oligoclonal (Figure A). However, in other two patients, TCR repertoires of CMV-CTLs became more diverse (Shannon-Weber index at the time of reactivation and a few weeks after was 0.558/2.958 in one patient and 1.471/3.86 in another), major clones lost, and new private clones appeared afterwards (Figure B). Polyclonal pattern (TCR repertoire of CMV-CTLs was diverse at the time of reactivation or after a few weeks) was detected in 3 patients out of 16 patients, and the pattern was exclusively seen in patients who were administered corticosteroid when CMV reactivated (42.9% vs 0.0%, p=0.063). [Conclusion] TCR repertoire of CMV-CTLs at the time of CMV reactivation after SCT is highly oligoclonal and frequently shared among different patients, but can dynamically change in a short period in some patients. Functional analyses of the dominant TCRs to understand their reactivity against CMV epitope and elucidation of the clinical significance and developmental mechanisms of clone shift are strongly warranted. Figure Disclosures Kitaura: Repertoire Genesis Inc.: Employment. Suzuki:Repertoire Genesis Inc.: Equity Ownership.

CMV Reactivation after Allogeneic Stem Cell Transplantation Is Associated with a Preponderance of Circulating Late-Stage CMV-Specific CD8+ T Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3250-3250
Author(s):  
Krishna V. Komanduri ◽  
Lisa S. St. John ◽  
Elizabeth J. Shpall ◽  
Richard E. Champlin ◽  
Jeffrey J. Molldrem ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Cd8 T Cells ◽  
Allogeneic Stem Cell Transplantation ◽  
Late Stage ◽  
Viral Reactivation ◽  
Tetramer Staining ◽  
Cmv Reactivation

Abstract CMV reactivation remains a major cause of morbidity after after allogeneic stem cell transplantation. Using HLA-peptide tetramer staining and cytokine flow cytometry, we previously demonstrated that higher CD4+ and CD8+ T cells specific for CMV were found in the peripheral circulation of individuals who experienced CMV reactivation, but that more of these cells were dysfunctional in subjects experiencing viral reactivation. Recent studies have examined the role of maturation status of virus-specific T cells in the setting of of cleared, persistent and progressive viral infections. CD57 is a marker of replicative senescence that is expressed on a subset of human CD8+ memory T cells that generally lacks CD28 expression, and that is characterized by downregulation of the naïve and central memory T cell markers CD45RA and CCR7. To examine whether CMV reactivation after SCT was associated with late-stage differentiation of CMV-specific CD8+ T cells specific for viral proteins, we first examined the fraction of CD57-expressing CD8+ T cells in patients stratified by the occurrence of post-SCT viral reactivation. In 87 patients examined with HLA-peptide tetramer staining at approximately 3 months after allogeneic SCT, we found that individuals experiencing CMV reactivation within the first 100 days after SCT had significantly higher absolute numbers of circulating CD57+ tetramer-stained CD8+ T cells specific for CMV than those who did not experience viral reactivation (mean 11.8 vs. 1.6 CD57+ tetramer-stained cells/μl, median 3.3 vs. 0.08 cells/μl, p&lt;0.0001). In individuals in whom &gt;75% of tetramer-stained CD8+ T cells were CD57+, the incidence of CMV reactivation was 83%, in contrast to 52% of others. In other experiments, we found that CD57+ tetramerstained CD8+ T cells failed to expand following co-culture with autologous monocyte-derived dendritic cells pulsed with CMV peptide pools, confirming that CD57+ CMV-specific CD8+ T cells may have impaired proliferative capacity. Taken together, these results suggest that late-stage differentiation of memory CD8+ T cells may be indicative of an exhausted antigen-specific immune response, and that optimal post-SCT immune restoration should consider the differentiation status and not simply the numbers of antigen-specific T cells targeted. Figure Figure

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