Integrative Analysis of Long Noncoding RNAs in Patients with Graft-versus-Host Disease

2020 ◽  
Vol 143 (6) ◽  
pp. 533-551 ◽  
Author(s):  
Feiyan Wang ◽  
Lan Luo ◽  
Zhenyang Gu ◽  
Nan Yang ◽  
Li Wang ◽  
...  
Keyword(s):  
B Cells ◽  
Signaling Pathway ◽  
Microarray Analysis ◽  
Graft Versus Host Disease ◽  
Differentially Expressed ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host ◽  
Qrt Pcr ◽  
Bcr Signaling

<b><i>Background:</i></b> Chronic graft-versus-host disease (cGVHD) remains a major cause of late non-recurrence mortality despite remarkable improvements in the field of allogeneic hematopoietic stem cell transplantation. Although recent studies have found that B-cell receptor (BCR)-activated B cells contribute to pathogenesis in cGVHD, the specific molecular mechanisms of B cells in this process remain unclear. <b><i>Methods:</i></b> In our study, human long noncoding RNA (lncRNA) microarrays and bioinformatic analysis were performed to identify different expressions of lncRNAs in peripheral blood B cells from cGVHD patients compared with healthy ones. The differential expression of lncRNA was confirmed in additional samples by quantitative real-time polymerase chain reaction (qRT-PCR). <b><i>Results:</i></b> The microarray analysis revealed that 106 of 198 differentially expressed lncRNAs were upregulated and 92 were downregulated in cGVHD patients compared with healthy controls. Intergenic lncRNAs accounted for the majority of differentially expressed lncRNAs. A KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis showed that the differentially expressed mRNAs, which were coexpressed with lncRNA, between the cGVHD group and the healthy group were significantly enriched in the BCR signaling pathway. Further analysis of the BCR signaling pathway and its coexpression network identified three lncRNAs with the strongest correlation with BCR signaling and cGVHD, as well as a series of protein-coding genes and transcription factors associated with them. The three candidate lncRNAs were further validated in another group of cGVHD patients by qRT-PCR. <b><i>Conclusions:</i></b> This is the first study on the correlation between lncRNA and cGVHD using lncRNA microarray analysis. Our study provides novel enlightenment in exploring the molecular pathogenesis of cGVHD.

Host Plasmacytoid or Conventional Dendritic Cells Alone Are Sufficient To Initiate Graft-Versus-Host Disease.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2164-2164
Author(s):  
Motoko Koyama ◽  
Daigo Hashimoto ◽  
Kazutoshi Aoyama ◽  
Ken-ichi Matsuoka ◽  
Kennosuke Karube ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
B Cells ◽  
Mhc Class Ii ◽  
Graft Versus Host Disease ◽  
Class Ii ◽  
Wild Type ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host

Abstract Graft-versus-host disease (GVHD) is a major complication after allogeneic hematopoietic stem cell transplantation. Alloantigen expression on host dendritic cells (DCs) is critical to initiate GVHD. DCs can be divided into two main subpopulations; conventional DCs (cDCs) and plasmacytoid DCs (pDCs), however, the contribution of each DC subset to elicit GVHD remains unclear. We examined the ability of cDCs and pDCs to initiate GVHD. pDCs, cDCs and B cells were isolated from C57BL/6 (B6: H–2b) mice treated with Flt3 ligand in order to expand DCs. pDCs were enriched from bone marrow by depleting CD3+, CD19+, CD11b+, and CD49b+ cells, followed by a FACS sorting of CD11cint B220+ cells. cDCs and B cells were sorted from splenocytes as CD11chi B220− cells and CD11c− B220+ cells, respectively. Isolated pDCs showed plasmacytoid morphology, produced IFN-α in response to CpG oligonucleotide. Although pDCs stimulated allogeneic T cells far less potently than cDCs, stimulation with CpG enhanced their allostimulatory capacity as potent as cDCs. We compared the ability of each DC subset to initiate GVHD by an add-back study of MHC class II-expressing DCs into MHC class II-deficient (II−/−) mice that were resistant to CD4-dependent GVHD. Lethally irradiated II−/− B6 mice were injected with 2 × 106 pDCs, cDCs or B cells from wild-type (II+/+) B6 mice on day -1 and injected with 2 × 106 CD4+ T cell from BALB/c (H–2d) mice on day 0. A flow cytometric analysis of the mesenteric lymph nodes on day +5 demonstrated significantly greater expansion of donor CD4+ T cells in recipients of pDCs or cDCs than those of B cells (Table). While injection of B cells did not cause any sign of GVHD, injection of pDCs or cDCs alone was sufficient to produce clinical and pathological GVHD (Table), thus breaking GVHD resistance of II−/− mice. We next examined the ability of pDCs to induce CD8-dependent GVHD in MHC-matched transplant using mice deficient in functional MHC class I expression (β2m−/−). Again, injection of pDCs or cDCs alone was sufficient to cause expansion of donor CD8+ T cells (p&lt;0.05). We next asked whether signaling through Toll-like receptors (TLRs) could be required for pDCs to initiate GVHD. However, injection of pDCs isolated from MyD88/TRIF-double deficient mice was able to initiate GVHD as potent as wild-type pDCs, thus demonstrating that pDCs initiate GVHD in a TLR signaling independent manner. These results provide important information for developing strategies aimed at inactivating host DCs to prevent GVHD. Impact of each APC subpopulation on GVHD APC Donor CD4 expansion (×103±SE) Clinical GVHD score (mean±SE) Pathological GVHD score (mean±SE) *p&lt;0.05 compared with B cells B cell 0.1 ± 0.0 2.1 ± 0.2 2.1 ± 0.2 pDC 5.3 ± 2.4* 4.3 ± 0.3* 7.4 ± 0.5* cDC 9.7 ± 3.8 * 3.8 ± 0.5 * 7.2 ± 0.7*

scholarly journals Pirfenidone Reverses Murine Chronic Graft Versus Host Disease

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1158-1158
Author(s):  
Jing Du ◽  
Ryan P Flynn ◽  
Katelyn Paz ◽  
Ante Vulic ◽  
Tara M. Robinson ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
B Cells ◽  
Graft Versus Host Disease ◽  
Flow Cytometry Analysis ◽  
T Follicular Helper Cells ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host ◽  
Helper Cells ◽  
Follicular Helper

Abstract Allogeneic hematopoietic stem cell transplantation (aHSCT) is hampered by chronic graft-versus-host disease (cGVHD), which results in multi-organ fibrosis and loss of function. In particular, bronchiolitis obliterans (BO) and scleroderma resulting from fibrotic bronchiolar and cutaneous response, respectively, are two devastating outcomes for cGVHD patients. Fibrotic manifestations often are considered irreversible and progressive. Therefore, new therapies targeting fibrosis are urgently needed. Pirfenidone (5-methyl-1-phenyl-2- (1H)-pyridone) exhibits a well-documented anti-inflammatory and anti-fibrosis function in multiple pre-clinical models and is the first and only FDA-approved drug for idiopathic pulmonary fibrosis. For this study, Pirfenidone was synthesized as a crystalline solid and found to be pure both by melting point and NMR spectroscopy. We evaluated Pirfenidone's anti-fibrosis function in 2 pathophysiologically distinct cGVHD murine models: 1. a major mismatched multi-organ system model (C57BL/6 to B10.BR) that induces BO as a result of a cGVHD-induced germinal center (GC) reaction, antibody deposition and fibrosis in the lung; and 2. a minor antigen mismatched model (B10.D2 to BALB/c) in which severe scleroderma is the major disease manifestation. In the BO model, pulmonary function loss in cGVHD mice (as reflected by increased resistance, elastance and decreased compliance of the lung) was restored by Pirfenidone treatment (400mg/kg) during both the early (day28-56) (Fig A, representative of 3 experiments with 5-8 mice per group) and late stages (day56-84) of the disease. Pathologic changes in the lung, such as collagen deposition and narrowing of bronchioles, were significantly reduced by Pirfenidone. The size and frequency of GCs in the spleen, and the frequency of GC B cells (Fig B, representative of 2 experiments with 5-8 mice per group) and T follicular helper cells were all significantly reduced in Pirfenidone- treated groups. To determine whether GCs were directly affected by Pirfenidone, we evaluated Pirfenidone in C57BL/6 mice immunized with sheep red blood cells (SRBC) to induce GCs. Interestingly, Pirfenidone did not reduce the SRBC-induced GC reaction (Fig C) (comparable frequencies of splenic GC B cells, T follicular helper cells and serum IgG levels were seen between Pirfenidone and vehicle-treated groups). These results suggested that Pirfenidone suppresses the GC reaction through a cGVHD-specific mechanism, rather than through immune regulation. Mechanistically, Pirfenidone administration attenuated the sequestration of pro-fibrogenic F4/80+ macrophages (Fig D, representative of 2 experiments) and TGF-β (Fig E, representative of 2 experiments) production within the lung. These results have led us to elucidate a potential mechanism of cGVHD: antibody deposition in the lung results in the activation of macrophages and TGF-β that drive fibrotic change and tissue damage, resulting in the exposure of auto- and allo- antigens to the immune system that support and sustain pathologic GC reactions. In the B10.D2 to BALB/c sclerodermatous cGVHD model, Pirfenidone treatment (400mg/kg, day21-55) improved clinical signs of scleroderma and reduced macrophage infiltration in the skin (Fig F). In summary, this is the first study evaluating a commercially available anti-fibrosis drug on pathologically distinct pre-clinical cGVHD models. Our data suggests Prifenidone reversed cGVHD in the BO model and, to a lesser extent, in the scleroderma model. Thus, Pirfenidone is a novel therapeutic agent for treating cGVHD patients with fibrosis that have been typically refractory to therapies. A. Resistance of lungs was measured on day56 of transplantation; Elastance and compliance correlated with resistance but were not shown here. B. Flow cytometry analysis of GC B cells of no cGVHD vs cGVHD mice treated with Pirfenidone or vehicle; C. Flow cytometry analysis of GC B cells from SRBC-immunized mice treated with Pirfenidone or vehicle; D and E. Macrophage F4/80 and TGF-β quantification of day56 lungs of no cGVHD vs cGVHD mice treated as indicated; F. Skin GVHD scores were recorded on indicated dates of irradiated BALB/c mice transplanted with B10.D2 donor BM alone or with T cells and treated as indicated. Unpaired student T test was used for statistical analysis. ****:P<0.0001; ***: P<0.001; **: P<0.01; *: P<0.05; ns: not significant. Figure Figure. Disclosures No relevant conflicts of interest to declare.

scholarly journals Altered B-cell homeostasis and excess BAFF in human chronic graft-versus-host disease

Blood ◽  
2009 ◽  
Vol 113 (16) ◽  
pp. 3865-3874 ◽  
Author(s):  
Stefanie Sarantopoulos ◽  
Kristen E. Stevenson ◽  
Haesook T. Kim ◽  
Corey S. Cutler ◽  
Nazmim S. Bhuiya ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Graft Versus Host Disease ◽  
High Plasma ◽  
Cell Homeostasis ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host ◽  
B Cell Homeostasis

Abstract Chronic graft-versus-host disease (cGVHD) causes significant morbidity and mortality in patients otherwise cured of malignancy after hematopoietic stem cell transplantation (HSCT). The presence of alloantibodies and high plasma B cell–activating factor (BAFF) levels in patients with cGVHD suggest that B cells play a role in disease pathogenesis. We performed detailed phenotypic and functional analyses of peripheral B cells in 82 patients after HSCT. Patients with cGVHD had significantly higher BAFF/B-cell ratios compared with patients without cGVHD or healthy donors. In cGVHD, increasing BAFF concentrations correlated with increased numbers of circulating pre–germinal center (GC) B cells and post-GC “plasmablast-like” cells, suggesting in vivo BAFF dependence of these 2 CD27+ B-cell subsets. Circulating CD27+ B cells in cGVHD comprised in vivo activated B cells capable of IgG production without requiring additional antigen stimulation. Serial studies revealed that patients who subsequently developed cGVHD had delayed reconstitution of naive B cells despite persistent BAFF elevation as well as proportional increase in CD27+ B cells in the first year after HSCT. These studies delineate specific abnormalities of B-cell homeostasis in patients with cGVHD and suggest that BAFF targeting agents may be useful in this disease.

scholarly journals Serum and Extracellular Vesicle Micrornas MiR-423, MiR-199 and MiR-93* As Biomarkers for Acute Graft Versus Host Disease

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4545-4545
Author(s):  
Rachel E Crossland ◽  
Jean Norden ◽  
Kim F Pearce ◽  
Mateja Kralj Juric ◽  
Clare Lendrem ◽  
...  
Keyword(s):  
Graft Versus Host Disease ◽  
Roc Analysis ◽  
Rna Extraction ◽  
Cell Transplant ◽  
Diagnostic Ability ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host ◽  
Qrt Pcr ◽  
Acute Graft

Abstract Introduction MicroRNAs are small, non-coding single-stranded RNAs and regulate approximately 50% of all genes by repressing translation. They are present in bodily fluids, where they are protected from RNase-mediated degradation by encapsulation into extracellular vesicles (EVs) and demonstrate a novel capacity to regulate the cellular differentiation of blood cells and immune function. Candidate microRNAs miR-377, miR-199, miR-93* and miR-423 have previously been associated with acute graft versus host disease (aGvHD) in post-hematopoietic stem cell transplant (HSCT) patient plasma. However, validation in independent cohorts is necessary, as well as further exploration to assess expression in the EV fraction of the blood. Methods MicroRNA expression was evaluated in early HSCT time point exploratory (n=34), validation (n=47) and diagnostic (n=65) serum cohorts by TaqMan qRT-PCR. Expression was also assessed in serum EVs (exploratory n=16 and validation n=47 cohorts) by EV isolation, RNA extraction and TaqMan qRT-PCR analysis. Results In sequential pre- and post-HSCT serum samples (n=34; pre-HSCT, Day0 (D0), D7, D14 & D28), miR-423 (p=0.03), miR-199 (p=0.06), miR-93* (p=0.04) and miR-377 (p=0.03) were upregulated at D14 in patients who developed aGvHD vs. no aGvHD. MiR-423 was also significantly upregulated at D0 (p=0.04), D7 (p=0.03) and D28 (p=0.03) in aGvHD patients. In relation to aGvHD severity, miR-423 (p=0.05), miR-199 (p=0.007) and miR-93* (p=0.09) were differentially expressed at D14 according to aGvHD grade. MiR-423 (p=0.02), miR-199 (p=0.07) and miR-93* (p=0.01) expression was validated at D14 in an independent cohort (n=47). When the exploratory and validation D14 samples were combined (n=81), miR-423 (p<0.001), miR-199 (p=0.04) and miR-93* (p<0.001) expression was upregulated in patients that developed aGvHD vs. no GvHD and ROC analysis identified miR-423 (p<0.001, AUC=0.75), miR-199 (p=0.09, AUC=0.62) and miR-93* (p<0.001, AUC=0.74) to have diagnostic ability. MiR-423 (p=0.001), miR-199 (p=0.01) and miR-93* (p<0.001) expression was higher in severe (III-IV) vs. no aGvHD, miR-423 (p=0.006) and miR-93* (p=0.01) were higher in mild (I-II) vs. no aGvHD and miR-199 was higher in severe (III-IV) vs. mild (I-II) aGvHD (p=0.002). All microRNAs demonstrated significant positive correlation (p<0.001), thus, principle component analysis (PCA) was performed. The PC1 composite score was used for ROC analysis and showed diagnostic ability for aGvHD incidence (p<0.001, AUC=0.73). In an independent diagnostic cohort (n=65), from a separate Institution, miR-423 (p=0.02), miR-199 (p=0.007) and miR-93* (p=0.004) expression was higher at aGvHD onset and showed diagnostic ability by ROC analysis (miR-423 p=0.03 AUC=0.66; miR-199 p=0.04 AUC=0.65; and miR-93* p=0.01 AUC=0.68). The three microRNAs had diagnostic ability with respect to aGvHD incidence based on composite ROC analysis of PC1 (p=0.019, AUC=0.68). MicroRNAs were also investigated within serum EVs (n=15). MiR-199 (p=0.008), miR-93* (p=0.001) and miR-423 (p=0.09) expression was lower at D14 in patients who developed aGvHD. Results were confirmed in a D14 validation cohort (n=47), with lower EV miR-423 (p=0.02) and miR-199 (p=0.04), but not miR-93* (p=0.15) expression in patients who developed aGvHD. MiR-423 and miR-199 had diagnostic ability based on composite PC1 ROC analysis (p=0.06, AUC=0.69). By D14, patients remaining aGvHD free had higher expression of miR-423 (p=0.03), miR-199 (p=0.05) and miR-93* (p<0.001) in the EV fraction compared to whole serum. Conclusions Results validate the capacity for circulating serum miR-423, miR-199 and miR-93* to act as diagnostic and prognostic biomarkers for aGvHD. Novel findings of differential expression between whole serum and the EV compartment prior to disease onset suggest a role for EV microRNAs in the biology of aGvHD, which warrants further investigation. Disclosures No relevant conflicts of interest to declare.

scholarly journals Bob.1 Blocks a SYK Protein Degradation Pathway in Mice That Develop Chronic Graft Versus Host Disease Manifestations

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3815-3815
Author(s):  
Wei Jia ◽  
Jonathan C Poe ◽  
Hsuan Su ◽  
Rachel A. DiCioccio ◽  
Sonali Bracken ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
B Cells ◽  
Western Blot ◽  
B Cell ◽  
Graft Versus Host Disease ◽  
Degradation Pathway ◽  
Host Disease ◽  
Graft Versus Host ◽  
Protein Levels ◽  
Bcr Signaling

Abstract After elucidating a role for aberrant B Cell Receptor (BCR) signaling in chronic graft versus host disease (cGVHD) (Allen et al, Blood. 2014;123:2108), we showed that the proximal BCR molecule SYK is a viable therapeutic target in mice and patients (Flynn et al, Blood 2015; 125:4085. Poe et al, JCI Insight. 2018;3:e122430. NCT02611063). Increased SYK protein levels associate with enhance phosphorylation of SYK activation sites, leading to immediate BCR responsiveness in cGVHD. In mice that develop cGVHD manifestations after allogeneic bone marrow transplantation (BMT), BCR responsiveness is promoted by B Cell Activating Factor (BAFF) (Jia et al, Blood. 2021;137:2544). Instead of SYK protein levels decreasing upon engagement with surrogate antigen to regulate excessive BCR-signaling, SYK protein was maintained when BAFF and alloantigen were present. In the present study, we address a potential molecular mechanism underpinning SYK protein maintenance in cGVHD B cells. In an unbiased scRNAseq study examining purified B cells from active cGVHD patients, we found that Pou2af1 was significantly altered within a BCR-activated B cell subset (Poe et al, Blood. 2019; 134; Suppl1:874-874). Pou2af1 encodes B cell Oct binding factor 1 (Bob.1), a prime candidate for further study. Bob.1 is a transcriptional coactivator that is also able to bind to SYK and regulate SYK protein stability (Siegel et al, Cell. 2006; 125:761). To determine whether BAFF promoted increased Bob.1 expression we examined B cells from BAFF transgenic (Tg) mice. Consistent with a role for this molecule in blocking SYK degradation, Bob.1 protein levels were notably higher in BAFF-Tg B cells. Intracellular flow cytometry analyses of B cells at day 40 after BMT (cGVHD n=9 vs control n=5 mice) confirmed that Bob.1 is significantly higher in cGVHD mouse B cells (p=0.0002, data not shown). We then investigated whether phosphorylation of a SYK negative regulatory site after BCR-activation was affected by BAFF. Since tyrosine 317 (SYK-Y317) phosphorylation is known to initiate ubiquitination and degradation of SYK, we stimulated purified B cells through the BCR before preparing cell lysates and performing Western blot analyses to measure SYK-Y317 phosphorylation, total SYK, LYN, Bob.1 and BLNK. As shown in a representative blot in Fig 1A, phosphorylation of SYK-Y317 (pSYK-Y317) was significantly decreased in B cells from BAFF transgenic (BAFF-Tg) mice compared to those from wild type (WT) mice. These data suggest that BAFF promotes SYK through Bob.1. Hypothesizing that Bob.1 is required to maintain SYK protein after allogeneic BMT, we employed Bob.1 knockout mice. After verifying that heterozygous mice (Bob.1 +/-) had lower Bob.1 protein levels and comparable B cell numbers to WT mice, we investigated the effect of Bob.1 reduction on SYK levels. Bob.1 +/- mice were crossed with BAFF-Tg mice and then SYK expression in B cells was determined using intracellular flow cytometry. We found that SYK protein levels were significantly increased in B cells exposed to a high BAFF environment and that SYK levels were significantly lower when Bob.1 protein was decreased (Fig 1B). These data demonstrate that Bob.1 is needed in vivo to maintain BAFF-mediated SYK protein maintenance after BCR engagement. Finally, we evaluated whether Bob.1 affected phosphorylation at SYK-Y317 in B cells from cGVHD mice. Splenic B cells were purified 41 days after BMT from recipients of B6 syngeneic (Syn), BM alone (BM only, control) or BM plus splenocytes (BM+Sp, cGVHD) and then stimulated with anti-IgM for 30 seconds before pSyk-Y317 and total SYK were detected by Western blot (Fig 1C). Using Image Studio Lite software, we normalized the intensity of pSyk-Y317 to actin in n=4 (Syn-BM+Sp), n=5 (BM only) and n=4 (BM+Sp) and found a significant decrease in pSyk-Y317 in cGVHD mice. We are currently investigating potential pathologic effects of Bob.1 after BMT in our mouse model and in patient cells. In summary, we have now demonstrated that, Bob.1 is required to maintain SYK protein in B cells after allogeneic BMT in mice that develop cGVHD. Our data suggest that Bob.1 maintains the stability of BCR proximal SYK by blocking a known SYK degradation pathway (Fig 1D). Improving our understanding of Bob.1 and SYK protein homeostasis after HCT will impact how we implement SYK inhibitors in patients. This work was supported the National Institutes of Health (NHLBI R01 HL 129061-06). Figure 1 Figure 1. Disclosures Sarantopoulos: Rigel: Other: Advisory Board.

Circulating B-Lymphocyte Subpopulations as Novel Biomarker for Measuring Activity of Chronic Graft-Versus-Host Disease.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2881-2881
Author(s):  
David Pohlreich ◽  
Hildegard T. Greinix ◽  
Karin Feldmann ◽  
Ulrike Koermoeczi ◽  
Imke Lohmann ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
B Lymphocytes ◽  
Graft Versus Host Disease ◽  
Memory B Cells ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host ◽  
Cell Counts ◽  
Severe Infections

Abstract Introduction: Chronic graft-versus-host-disease (cGvHD) is a major complication of allogeneic hematopoietic stem cell transplantation (HSCT) and a leading cause of non-relapse mortality due to profound immunodeficiency. The approach currently used to establish the diagnosis of cGvHD depends almost exclusively on the clinical history, physical examination, and histopathologic confirmation. Biology-based markers for confirmation of diagnosis or monitoring progression of cGvHD are urgently warranted. The pathogenesis of cGvHD is poorly understood. Mounting evidence implies a role of B-cells in this autoimmune-like disorder. In this study we analyzed whether changes in B-cell subpopulations in the peripheral blood (PB) of HSCT recipients could serve as a biomarker for immune-modulatory consequences of cGvHD. Methods: Within six months 70 consecutive patients (45 men, 25 women; median age 47.5 years) with complete donor cell engraftment a median of 45.5 (range, 5–149) months after allogeneic HSCT were enrolled including 21 never having cGvHD and 49 with cGvHD (35 active and/or progressive, 14 resolved at study entry). The latter received standard anti-infective prophylaxis and immunosuppressive therapy with a median duration of 36 (range, 3–104) months. A third of patients with active/progressive cGvHD had more than 2 organs involved. Evaluations in the study consisted of clinical parameters including cGvHD severity and treatment, serum immunoglobulin (Ig) levels, and infections. PB was analyzed by multiparameter flow cytometry to define B-lymphocytes (CD19+), which were further subdivided by staining for surface Ig (IgD+/M+) and the B-cell memory marker CD27+ and more immature B-lymphocytes (CD19+/CD21−). Results: No significant differences in absolute B, T, and NK-cell counts between the groups with and without cGvHD were seen. However, the relative number of both class-switched and non-class-switched memory B-cells was significantly lower (below 3%) in patients with active cGvHD compared to patients never having cGvHD (up to 7%) (p<0.001, c2 =22.0 for class-switched and p=0.002, c2 =9.2 for non-class-switched). Absolute cell counts confirmed these findings (10.3 vs. 27.2 cells/uL, p=0.02 for class-switched and 12.1 vs. 22.1 cells/ul, p=0.047 for non-class-switched). Significantly more patients with active cGvHD had elevated numbers of immature CD19+/CD21− B-cells (>15% cut-off) than the groups with resolved or never having cGvHD (37% vs 21% vs 9%, p=0.024). In patients with >15% immature CD19+/CD21− B-cells significantly more severe infections (according to NCI-CTC Infection module grade III–IV) were seen compared to patients with low CD19+/CD21− B-cell counts (p=0.002, c2 =9.5). In the group of active cGvHD 12/13 (92%) patients with >15% CD19+/CD21− B-cells compared to 4/22 (18%) patients with ≤ 15% CD19+/CD21− B-cells experienced severe infections (p<0.001, c2 =18.1). First results in an ongoing prospective study showed an increase of memory B-cell numbers in patients with decreasing cGvHD activity. Discussion: Monitoring of class-switched and non-class-switched memory B-cells may allow measuring of disease activity of chronic GvHD. In combination with assessment of immature CD19+/CD21− B-cells new insights into immune-modulatory consequences of cGvHD focusing on B-cell defects and activation status are possible.

Glutathione S-Transferase T1 Mismatch and Presence of Anti-GSTT1 Antibodies In Hepatic Gvhd

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2323-2323
Author(s):  
María José Martínez-Bravo ◽  
Inmaculada Tallón ◽  
Ildefonso Espigado ◽  
Alvaro Urbano-Ispizua ◽  
Antonio Núñez-Roldán ◽  
...  
Keyword(s):  
Stem Cell ◽  
B Cells ◽  
Graft Versus Host Disease ◽  
Glutathione S Transferase ◽  
Serum Samples ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host ◽  
Post Transplant ◽  
Gstt1 Gene

Abstract Abstract 2323 Graft-versus-host disease (GVHD) remains a major complication after allogenic hematopoietic stem-cell transplantation (alloHSCT) from HLA-identical donor. It is due to donor T-cell responses against minor histocompatibility antigens (mHags) of the recipient. Nevertheless, there is a complete lack of studies addressing the B-cell response to mHags in GVHD. Glutathione S-transferase T1 (GSTT1) is a drug metabolizing enzyme that is involved in detoxification processes. It is highly expressed in liver, kidney and erythrocytes. The GSTT1 gene is absent in 20% of the Caucasian population. In liver and renal transplantation, we have reported that some GSTT1-negative patients who received a GSTT1-positive graft produce anti-GSTT1 antibodies (abs) and present signs of chronic rejection. The aims of this study were to analyze the effects of GSTT1 donor/recipient mismatches in the development of hepatic GVHD and to detect production of anti-GSTT1 abs after alloHSCT. For that purpose, we have studied a group of 40 patients that received an alloHSCT from HLA-identical donors between January 2004 and July 2009 in HU Virgen del Rocío, whose DNA samples and their corresponding donor samples were available. All patients and their donors gave written informed consent. GSTT1 genotyping was performed by PCR and a commercially available ELISA test with the GSTT1 human recombinant protein was used to detect anti-GSTT1 abs. We studied abs in serum samples corresponding to different pre and post transplant phases. The distribution of the four possible GSTT1 genetic combinations was as follows: 25 donor+/recipient+, 6 donor+/recipient-, 5 donor-/recipient+ and 4 donor-/recipient-. Anti-GSTT1 antibodies were detected in 5 patients all of them with a donor that carries the null genotype. Four of the 5 patients included in the donor-/recipient+ group developed anti-GSTT1 abs (of the IgG class) after the infusion and were diagnosed with acute hepatic GVHD. In all of these cases the donor was a multiparous female. The presence of anti-GSTT1 abs is significantly associated with hepatic GVHD (p=0,01, Table 1). The fifth recipient within this group had a non-transfused male donor and did not produce abs. In all the cases, anti-GSTT1 abs were found in post-transplant serum samples of the recipient (never in pre-transplant samples) under a condition of 100% chimerism. Our hypothesis is that the immune system of the null donors has encountered the GSTT1 antigen during pregnancy of a positive-fetous and memory B-cells would be in the peripheral blood stem-cell infusion. These cells, once in a positive recipient, would be activated when contacted with the antigen present in the recipientxs liver. In the patients, the B-cells produced high levels of anti-GSTT1 abs that were associated with GVHD episodes. In conclusion, we describe that the presence of anti-GSTT1 abs is associated with the development of hepatic graft-versus-host-disease and that the null genotype of the donor is a necessary condition for antibody-production. These results confirm the GSTT1 gene as new minor histocompatibility antigen. With Hepatic GVHD Without Hepatic GVHD With 4 1 5 anti-GSTT1 abs Without anti-GSTT1 abs 6 29 35 10 30 n = 40 Disclosures: No relevant conflicts of interest to declare.

scholarly journals Effect of Early Post-Transplantation Tacrolimus Concentration on the Risk of Acute Graft-Versus-Host Disease in Allogenic Stem Cell Transplantation

Cancers ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 613
Author(s):  
Nidhi Sharma ◽  
Qiuhong Zhao ◽  
Bin Ni ◽  
Patrick Elder ◽  
Marcin Puto ◽  
...  
Keyword(s):  
Stem Cell ◽  
Graft Versus Host Disease ◽  
Tacrolimus Concentration ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host ◽  
The Impact ◽  
Risk Of Relapse ◽  
Acute Graft

Acute graft versus host disease (aGVHD) remains a leading cause of morbidity and mortality in allogeneic hematopoietic stem cell transplant (allo-HSCT). Tacrolimus (TAC), a calcineurin inhibitor that prevents T-cell activation, is commonly used as a GVHD prophylaxis. However, there is variability in the serum concentrations of TAC, and little is known on the impact of early TAC levels on aGVHD. We retrospectively analyzed 673 consecutive patients undergoing allo-HSCT at the Ohio State University between 2002 and 2016. Week 1 TAC was associated with a lower risk of aGVHD II–IV at TAC level ≥10.15 ng/mL (p = 0.03) compared to the lowest quartile. The cumulative incidence of relapse at 1, 3 and 5 years was 33%, 38% and 41%, respectively. TAC levels at week 2, ≥11.55 ng/mL, were associated with an increased risk of relapse (p = 0.01) compared to the lowest quartile. Subset analysis with acute myeloid leukemia and myelodysplastic syndrome patients showed significantly reduced aGVHD with TAC level ≥10.15 ng/mL at week 1 and a higher risk of relapse associated with week 2 TAC level ≥11.55 ng/mL (p = 0.02). Hence, achieving ≥10 ng/mL during the first week of HCT may mitigate the risk of aGVHD. However, levels (>11 ng/mL) beyond the first week may be associated with suppressed graft versus tumor effect and higher relapse.

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