ST2 and Reg3α: Can they predict aGvHD, steroid refractoriness and transplant-related mortality in pediatric patients after HSCT?

BACKGROUND/AIM: There are several complications of hematopoietic stem cell transplantation. Without any doubt, most important of these is aGvHD that increases transplant-related mortality. The aim of this study is to investigate whether ST-2 and Reg3α levels measured at an early stage in pediatric patients undergoing allogeneic hematopoietic stem cell transplantation can be individual biomarkers identifying future GvHD and predicting treatment response. MATERIALS AND METHODS: From January 2019 to January 2021, 27 patients undergoing hematopoietic stem cell transplantation for primary immunodeficiency or hematopoietic diseases formed the study group. During their follow-up, the patients were classified into two groups as those developing and those not developing aGvHD. Nineteen healthy volunteers from a similar age group who needed their blood samples drawn for other reasons and who did not have any history of chronic disease, infection or medication use formed the control group. Blood samples of patients scheduled to have allogeneic HSCT were obtained before the administration of the preparative regimen, on Day +7 post-transplant and on the day of diagnosis if they developed aGvHD. Serum samples were stored at -20ºC until the day of processing. ST2 and Reg3α levels were measured using the ELISA method. RESULTS: For patients who developed aGvHD (n = 13), ST2 levels obtained before the transplantation, on Day +7 post-transplant and on the day of aGvHD diagnosis (in patients developing GvHD) were significantly higher compared to the healthy Control Group (p-value <0.05). As regards to the samples obtained on the same days, ST2 levels did not differ significantly among patients who developed and those who did not develop GvHD (n = 14; p-value >0.05). ST2 levels of samples obtained on the days that acute skin and gastrointestinal tract GvHD developed did not differ significantly between these two groups (p-value >0.05). Reg3α levels of the pre-transplant samples, on Day +7 after the transplantation and on the day of aGvHD diagnosis did not show any difference between any of the groups (p-value >0.05). As only two patients died after transplantation, thus correlation of ST2 and Reg3α levels with transplant-related mortality could not be proven. CONCLUSION: The results of this study suggest that ST2 and Reg3α levels are neither diagnostic nor prognostic or predictive biomarkers of aGvHD, steroid resistance or transplant-related mortality in pediatric patients. This study can be regarded as a pilot study because of the small patient population; more research involving a larger patient population is required.

Assessing cell chimerism in an acute graft-versus-host disease model established in TLR4 knockout mice.

BACKGROUND: Graft-versus-host disease (GVHD) is a major complication after allogeneic hematopoietic stem cell transplantation. OBJECTIVE: To elucidate the role of Toll-like receptor 4 (TLR4), the major receptor for bacterial lipopolysaccharide, in the development of GVHD, we constructed a GVHD model in TLR4 knockout (TLR4-/-) mice and monitored the cell chimerism. METHODS: In this study, we used polymerase chain reaction to identify whether TLR4 knockout (TLR4-/-) mice were established. Before transplantation, we pretreated mice with irradiation so as to obtain an appropriate irradiation dose. Flow cytometry was applied to measure the chimerism status, the distributions of antigen-presenting cells (APCs), and T-cells in TLR4+/+ and TLR4-/- recipient mice. RESULTS: The general condition of TLR4-/- recipients was better than that of TLR4+/+ recipients, and the TLR4-/- recipient mice showed less severe GVHD manifestations than the TLR4+/+ recipient mice. Most of the APCs and T-cells in the host mouse spleen were derived from donor cells, and CD4+ T-cells, including memory T-cells, were in the majority in host mice. CONCLUSION: In general, our data show that TLR4 deletion attenuated GVHD development, which suggests that TLR4 could be used as a novel target and therapeutic paradigm in GVHD therapies.

ANTECEDENTES: La enfermedad de injerto contra huésped (EICH) es una complicación importante después del trasplante alogénico de células madre hematopoyéticas. OBJETIVOS: Para dilucidar el papel de TLR4, el principal receptor de LPS bacteriano, en el desarrollo de GVHD, construimos un modelo de GVHD en ratones knockout para TLR4 (TLR4-/-) y monitoreamos el quimerismo celular. MÉTODOS: En este estudio, usamos PCR para identificar si se establecieron ratones knockout para TLR4 (TLR4-/-). Antes del trasplante, pretratamos a los ratones con irradiación para obtener la dosis de irradiación adecuada. Se aplicó citometría de flujo para medir el estado de quimerismo, las distribuciones de APC y células T en ratones receptores TLR4+/+ y TLR4-/-. RESULTADOS: El estado general de los receptores de TLR4-/- fue mejor que el de los receptores de TLR4+/+, y los ratones receptores de TLR4-/- mostraron manifestaciones de GVHD menos graves que los ratones receptores de TLR4+/+. La mayoría de las APC y las células T en el bazo del ratón huésped se derivaron de las células del donante, y las células T CD4+, incluidas las células T de memoria, se encontraban en su mayoría en los ratones huéspedes. CONCLUSIÓN: En general, nuestros datos muestran que la eliminación de TLR4 atenuó el desarrollo de GVHD, lo que sugiere que TLR4 podría usarse como un nuevo objetivo y paradigma terapéutico en las terapias de GVHD.

Umbilical Cord Mesenchymal Stromal Cells for Steroid-Refractory Acute Graft-versus-Host Disease.

BACKGROUND: Steroid-refractory acute graft-vs.-host disease (SR-aGVHD) is a complication of allogeneic hematopoietic stem cell transplantation with a dismal prognosis and for which there is no consensus-based second-line therapy. Ruxolitinib is not easily accessible in many countries. A possible therapy is the administration of mesenchymal stromal cells (MSCs). METHODS: In this retrospective study, 52 patients with severe SR-aGVHD were treated with MSCs from umbilical cord (UC-MSCs) in nine institutions. RESULTS: The median (range) age was 12.5 (0.3-65) years and the mean ± SD dose (×106/kg) was 4.73 ± 1.3 per infusion (median of four infusions). Overall (OR) and complete response (CR) rates on day 28 were 63.5% and 36.6%, respectively. Children (n = 35) had better OR (71.5% vs. 47.1%, p = 0.12), CR (48.6% vs. 11.8%, p = 0.03), overall survival (p = 0.0006), and relapse-free survival (p = 0.0014) than adults (n = 17). Acute adverse events (all of them mild or moderate) were detected in 32.7% of patients, with no significant difference in children and adult groups (p = 1.0). CONCLUSIONS: UC-MSCs are a feasible alternative therapy for SR-aGVHD, especially in children. The safety profile is favorable.

The level of Tim-3+CD8+ T cells can serve as a potential marker for evaluating the severity of acute graft-versus-host disease after haplo-PBSCT

Early and accurate diagnosis of acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation is crucial for the prognosis of patients. This study identified a potential biomarker for the severity of aGVHD after human leukocyte antigen (HLA)-haploidentical peripheral blood hematopoietic stem cell transplantation (haplo-PBSCT). We included 20 healthy subjects and 57 patients who underwent haplo-PBSCT. Of these patients, 22 developed aGVHD after haplo-PBSCT. The results showed that patients with aGVHD had significantly increased levels of Tim-3+/Perforin+/Granzyme B+CD8+ T cells, but significantly decreased Galectin-9. The differences in Galectin-9 and Tim-3+/Granzyme B+CD8+ T cells between grade I-II aGVHD and III-IV aGVHD were also significant. In vitro, the apoptosis of CD8+ T cells from aGVHD patients was significantly increased after Tim-3/Galectin-9 pathway activation, which decreased Granzyme B secretion. As revealed by univariate analysis, the level of Tim-3+CD8+ T cells was a risk factor for severe aGVHD. ROC analysis demonstrated that high levels of Tim-3+CD8+ T cells had a significant diagnostic value for severe aGVHD, with an area under the curve of 0.854 and cut-off value of 14.155%. In conclusion, the binding of Tim-3 with exogenous Galectin-9 can promote apoptosis of CD8+ T cells and affect the secretion of Granzyme B. Tim-3+CD8+ T cells have the potential to serve as immunological markers for assessing the severity of aGVHD after haplo-PBSCT and identifying patients at a higher risk for severe aGVHD.

Broad-Spectrum Antibiotics and Risk of Graft-versus-Host Disease in Pediatric Patients Undergoing Transplantation for Acute Leukemia: Association of Carbapenem Use with the Risk of Acute Graft-versus-Host Disease

Variation in the gastrointestinal (GI) microbiota after hematopoietic cell transplantation (HCT) has been associated with acute graft-versus-host disease (aGVHD). Because antibiotics induce dysbiosis, we examined the association of broad-spectrum antibiotics with subsequent aGVHD risk in pediatric patients undergoing HCT for acute leukemia. We performed a retrospective analysis in a dataset merged from 2 sources: (1) the Center for International Blood and Marrow Transplant Research, an observational transplantation registry, and (2) the Pediatric Health Information Services, an administrative database from freestanding children's hospitals. We captured exposure to 3 classes of antibiotics used for empiric treatment of febrile neutropenia: (1) broad-spectrum cephalosporins, (2) antipseudomonal penicillins, and (3) carbapenems. The primary outcome was grade II-IV aGVHD; secondary outcomes were grade III-IV aGVHD and lower GI GVHD. The adjusted logistic regression model (full cohort) and time-to-event analysis (subcohort) included transplantation characteristics, GVHD risk factors, and adjunctive antibiotic exposures as covariates. The full cohort included 2550 patients at 36 centers; the subcohort included 1174 patients. In adjusted models, carbapenems were associated with an increased risk of grade II-IV aGVHD in the full cohort (adjusted odds ratio [aOR], 1.24; 95% confidence interval [CI], 1.02 to 1.51) and subcohort (sub hazard ratio [HR], 1.31; 95% CI, 0.99 to 1.72), as well as with an increased risk of grade III-IV aGVHD (subHR, 1.77; 95% CI, 1.25 to 2.52). Early carbapenem exposure (before day 0) especially impacted aGVHD risk. For antipseudomonal penicillins, the associations with aGVHD were in the direction of increased risk but were not statistically significant. There was no identified association between broad-spectrum cephalosporins and aGVHD. Carbapenems, more than other broad-spectrum antibiotics, should be used judiciously in pediatric HCT recipients to minimize aGVHD risk. Further research is needed to clarify the mechanism underlying this association.

Correlation of T Cell Subsets and Hypercholesterolemia of the Donor and Its Association with Acute Graft-versus-Host Disease.

Background: Acute graft-versus-host disease (aGVHD) is an important cause of death following allogeneic hematopoietic stem cell transplantation (allo-HSCT). The association between cholesterol and aGVHD was previously described potentially, resulting from pro-inflammatory responses associated with hypercholesterolemia. The aim of this study was to correlate T cell subsets in donor bone marrow (BM) samples with their levels of cholesterol and associate these results with recipients who developed aGVHD and those who did not. Materials and Methods: A prospective study was performed in 39 donor samples. T cell subsets were analyzed by flow cytometry. Results: Eleven (28%) donors had hypercholesterolemia. Donor samples with hypercholesterolemia had less Tregs compared to donors with normal levels of cholesterol (22.69 (IQR=30.6) cells/µL vs 52.62 (IQR=44.68) cells/µL, p=0.04). Among all individuals in the cohort, aGVHD was observed in 21%: 36% from donors with hypercholesterolemia versus 14% from donors with normal levels of cholesterol. Conclusion: As we described the association between hypercholesterolemia and diminished Tregs, our results might suggest that normalizing the levels of total cholesterol in the donor, prior performing allo-HSCT, might be an effective approach to diminish the risk of the receptor to develop aGVHD.

Association of recipient and donor hypercholesterolemia prior allogeneic stem cell transplantation and graft-versus-host disease.

Few authors have reported a decreased frequency of acute graft-versus-host disease (aGVHD) using statins, as these medications have anti-inflammatory effects, however, to date, the direct association between high cholesterol and GVHD has not been reported. The aim of his study was to investigate the association of recipient and donor hypercholesterolemia with the incidence of aGVHD. A retrospective analysis was performed identifying allogeneic hematopoietic stem cell transplantation (allo-HSCT) recipients and donors at the National Institute of Medical Sciences and Nutrition in Mexico City between May 1999 and August 2017. The final cohort included 113 consecutive patients undergoing allo-HSCT and 110 donors with complete data. Acute GVHD was present in 24% patients. A statistically significant increase in the frequency of aGVHD associated with hypercholesterolemia in the recipients or donors (p = 0.03 and p = 0.008, respectively). Hypercholesterolemia in both, donor and recipient, was also associated with increased aGVHD compared to either patient or donor having hypercholesterolemia or neither (p = 0.002). No statistical significance was observed for other variables. To date, this is the first study associating hypercholesterolemia with aGVHD. According to our results we conclude that hypercholesterolemia in the donor, or in both, the patient and donor, is an independent factor for the development of aGVHD, however, further prospective and larger studies are needed as our results are preliminary.