Post Hematopoietic Transplantation Research Articles

Abatacept-Prophylaxis Based Haploidentical Transplantation May Allow Sustained Engraftment and Offset Gvhd in Non-Malignant Disorders

Introduction: Non-malignant disorders (NMD) derive no benefit from graft-versus-host disease (GVHD) after allogeneic hematopoietic transplantation (HCT). Transplantation designed to optimize engraftment and immune reconstitution while minimizing GVHD and organ toxicities is desirable. Post-transplant cyclophosphamide (PTCy) decreases severe chronic GVHD risk in radiation-containing regimens for NMD when combined with serotherapy (anti-thymocyte globulin) (PMID: 22955919; PMID: 30500440). Abatacept effectively offsets GVHD following unrelated donor (URD) HCT (PMID: 33449816; PMID: 32813873). Based on these reports, NCT03128996 was developed for haploidentical transplantation in NMD after radiation-free reduced intensity conditioning (RIC) combining PTCy + Abatacept for GVHD prophylaxis. Serotherapy does not contribute to GVHD prophylaxis in this RIC approach. Outcomes focused on safety and GVHD- free donor engraftment in children with NMD and no better donor options. Method: Conditioning included alemtuzumab (day -22 to -19), fludarabine (day -8 to -4), melphalan (day -3) and thiotepa (day -4) as previously published (hydroxyurea prior to RIC was used in proliferative marrow disorders) (PMID: 32813873). Stem cell source was bone marrow (BM) or CD34-selected peripheral blood cells with a fixed CD3 cell dose of 1x10^6/kg recipient weight. GVHD prophylaxis included tacrolimus (day -1, with wean beginning 6-9 months post-HCT), PTCy on days +3 and +4, mycophenolate mofetil until day +28, and monthly abatacept doses until 13 months post-HCT. Survival, safety, donor chimerism, GVHD incidence, and incidence of post HCT infection were primary outcome measures. Results: Eleven patients 6 to 24 years old (median 9 years) underwent HCT for adrenoleukodystrophy ( 2), sickle cell disease (SCD) ( 6), aplastic anemia ( 1), second transplant for severe combined immunodeficiency ( 1) and STAT 5B gain-of-function mutation ( 1). Eight were trial participants and 3 treated according to trial guidelines. All patients tolerated abatacept infusions and engrafted with neutrophil recovery (>500/cu mm) at a median of 17 days (range 15-24 days). As of 7/31/2023, median follow up is 411 days (range 83-1979 days). The mean myeloid and T-cell chimerisms at D100 (n=10) and 1 year (n=6) were 97.8%+/- 5.3 and 96.3% +/- 6.4 and 98.6% +/- 3.3 and 95% +/-8.7, respectively. One patient with SCD has low myeloid engraftment (24%) at 7 months post-HCT (lymphoid 91%); a stem cell boost is planned after immune suppression with fludarabine/Cytoxan/ATG (PMID: 35798858). Viral reactivation was noted in 4 patients, none developed CMV disease or EBV post-transplant lymphoproliferation. One patient developed grade 1 acute GVHD. Of 6 evaluable patients, one developed localized chronic skin GVHD. This patient remains on systemic immune suppression beyond 13 months (ruxolitinib). The overall and GVHD-free survival at 1-year post HCT in evaluable patients (N=6) is 100 % and 83%, respectively. Conclusion: Promising preliminary results in this cohort support continuation of the trial with longer term follow-up. The effective suppression of GVHD without serious infectious complications in the first year post-HCT is encouraging. Virus reactivation was however noted early post-HCT and requires monitoring and prophylaxis. Incorporating abatacept into GVHD prophylaxis was successful in achieving desired outcomes; the phase 1 portion of the study is completed and the phase 2 trial and follow-up are ongoing.

Acute Kidney Injury in the Modern Era of Allogeneic Hematopoietic Stem Cell Transplantation.

AKI is a major complication of allogeneic hematopoietic stem cell transplantation, increasing risk of nonrelapse mortality. AKI etiology is often ambiguous due to heterogeneity of conditioning/graft versus host disease regimens. To date, graft versus host disease and calcineurin inhibitor effects on AKI are not well defined. We aimed to describe AKI and assess pre-/post-hematopoietic transplant risk factors in a large recent cohort. We performed a single-center, retrospective study of 616 allogeneic hematopoietic cell transplant recipients from 2014 to 2017. We defined AKI and CKD based on Kidney Disease Improving Global Outcomes (KDIGO) criteria and estimated GFR using the Chronic Kidney Disease Epidemiology Collaboration equation. We assessed AKI pre-/post-hematopoietic transplant risk factors using cause-specific Cox regression and association of AKI with CKD outcomes using chi-squared test. AKI was treated as a time-dependent variable in relation to nonrelapse mortality. Incidence of AKI by day 100 was 64%. Exposure to tacrolimus and other nephrotoxins conferred a higher risk of AKI, but tacrolimus levels were not associated with severity. Reduced-intensity conditioning carried higher AKI risk compared with myeloablative conditioning. Most stage 3 AKIs were due to ischemic acute tubular necrosis and calcineurin inhibitor nephrotoxicity. KRT was initiated in 21 out of 616 patients (3%); of these 21 patients, nine (43%) recovered and five (24%) survived to hospital discharge. T cell-depleted transplants, higher baseline serum albumin, and non-Hispanic ethnicity were associated with lower risk of AKI. CKD developed in 21% (73 of 345) of patients after 12 months. Nonrelapse mortality was higher in those with AKI (hazard ratio, 2.77; 95% confidence interval, 1.8 to 4.27). AKI post-hematopoietic cell transplant remains a major concern. Risk of AKI was higher with exposure to calcineurin inhibitors. T cell-depleted hematopoietic cell transplants and higher serum albumin had lower risk of AKI. Of the patients requiring KRT, 43% recovered kidney function. This article contains a podcast at https://www.asn-online.org/media/podcast/CJASN/2021_09_07_CJN19801220.mp3.

Inflammasomes and the Maintenance of Hematopoietic Homeostasis: New Perspectives and Opportunities.

Hematopoietic stem cells (HSCs) regularly produce various blood cells throughout life via their self-renewal, proliferation, and differentiation abilities. Most HSCs remain quiescent in the bone marrow (BM) and respond in a timely manner to either physiological or pathological cues, but the underlying mechanisms remain to be further elucidated. In the past few years, accumulating evidence has highlighted an intermediate role of inflammasome activation in hematopoietic maintenance, post-hematopoietic transplantation complications, and senescence. As a cytosolic protein complex, the inflammasome participates in immune responses by generating a caspase cascade and inducing cytokine secretion. This process is generally triggered by signals from purinergic receptors that integrate extracellular stimuli such as the metabolic factor ATP via P2 receptors. Furthermore, targeted modulation/inhibition of specific inflammasomes may help to maintain/restore adequate hematopoietic homeostasis. In this review, we will first summarize the possible relationships between inflammasome activation and homeostasis based on certain interesting phenomena. The cellular and molecular mechanism by which purinergic receptors integrate extracellular cues to activate inflammasomes inside HSCs will then be described. We will also discuss the therapeutic potential of targeting inflammasomes and their components in some diseases through pharmacological or genetic strategies.

Successful Treatment of Mature T-Cell Lymphoma with Allogeneic Stem Cell Transplantation: The Largest Multicenter Retrospective Analysis

Successful Treatment of Mature T-Cell Lymphoma with Allogeneic Stem Cell Transplantation: The Largest Multicenter Retrospective Analysis

Noninvasive Microscopic Imaging Reveals Increased Leukocyte Adhesion and Rolling in Skin of Acute Graft-Versus-Host Disease Patients Compared to Post-Transplant Controls

BACKGROUND. The nature and kinetics of leukocyte migration are intimately connected to the pathophysiology of acute graft-versus-host disease (aGVHD). Real-time visualization of leukocyte-endothelial interactions in patients post hematopoietic transplantation (HCT) may augment clinical diagnosis and patient management. In this cross-sectional pilot study, we aimed to test the feasibility of parameters characteristic of leukocyte motion in skin capillaries as potential imaging biomarkers to detect aGVHD. STUDY POPULATION. We enrolled 16 post-HCT patients: 8 patients with any organ aGVHD and 8 patients with no organ aGVHD on the day of the imaging. Diagnoses were retrospectively confirmed on day 100 by a transplant physician (MB), who was blinded to the results of the confocal imaging. 7 out of 8 aGVHD patients had only skin involvement: stage 1 (N=2 patients), stage 2 (N=2), and stage 3 (N=3) cutaneous aGVHD, whereas one patient had stage 1 gut aGVHD with no skin involvement. 6 out of 8 aGVHD group patients required systemic therapy with steroids. METHODS. To noninvasively visualize leukocyte motion in the topmost capillaries of post-HCT patients' skin, we used a clinical reflectance confocal microscope (Vivascope 1500, Caliber I.D.). It enables real-time en face view of individual cells at 9 frames per second. We took twenty 30-second videos of different capillaries in the left volar forearm and left upper chest. We counted the number of adherent and rolling leukocytes per 10 minutes of videos per patient. RESULTS. We found increased leukocyte rolling in the aGVHD group (average of 3 rolling leukocytes per patient), compared to post-HCT controls (average of 1 rolling leukocyte). Highest leukocyte rolling (>8 rolling leukocytes) was associated with increased all-cause mortality, but not predictive of transplant-related mortality (TRM). Similarly, we found increased leukocyte adhesion in the aGVHD group (average of 3 adherent leukocytes per patient), compared to post-HCT controls (average of 1 adherent leukocyte). Interestingly, we observed 7 adherent leukocytes in one patient who had isolated gut aGVHD and no skin involvement. DISCUSSION. Our preliminary results show altered leukocyte-endothelial interactions in the skin capillaries of aGVHD patients, suggesting that confocal microscopy may be an important tool in augmenting the clinical diagnosis and managing post-HCT patients. Future studies should test whether there is a difference in the pattern of change in parameter values between patients who did and who did not develop aGVHD. ACKNOWLEDGEMENT. This work was supported in part by Career Development Award Number IK2 CX001785 from the United Sates Department of Veterans Affairs Clinical Science R&D (CSRD) Service. Disclosures Byrne: Karyopharm: Research Funding. Jagasia:Janssen: Research Funding; Kadmon: Consultancy; Incyte: Consultancy. Tkaczyk:Incyte: Consultancy.

Fecal Calprotectin as a Diagnostic and Prognostic Biomarker for Gastrointestinal Graft Versus Host Disease: A Systematic Review of Literature.

The current practice for diagnosing graft versus host disease (GVHD) includes clinical or endoscopic evaluation of the patient. Clinical diagnosis is limited by an overlapping symptomatic spectrum with infectious causes, a common scenario in the post-transplant setting where an invasive procedure, such as endoscopy, is often impractical. We, therefore, evaluated the role of fecal calprotectin as a diagnostic as well as a prognostic biomarker for gastrointestinal GVHD (GI-GVHD) occurrence and severity in the post-hematopoietic transplant population. Following Prisma guidelines, we performed a systematic search of articles published after 2004 using the PubMed, Embase, Cochrane Library, and Web of Science databases. After a detailed screening, 10 studies involving a total of 494 patients were included. In the cohorts comparing median fecal calprotectin (mFC) level in GI-GVHD vs. non-GI-GVHD patients, the results indicated an increase in the mFC level in patients with GI-GVHD when compared to non-GI-GVHD patients. Similarly, an increase in the mFC level was seen in accordance with the severity of the disease. Moreover, corticosteroid-resistant patients had a higher mFC level as compared to corticosteroid-sensitive patients. Our study indicates that the mFC level can be used for diagnosing as well as predicting the treatment response to GI-GVHD. However, future randomized prospective trials involving larger populations are needed to further explore its significance.

Leucovorin Following Methotrexate Graft-Vs-Host Disease Prophylaxis Shortens the Duration of Mucositis, Time to Neutrophil Engraftment and Length of Hospitalization in Myeloablative Allogeneic Hematopoietic Transplantation

Introduction Grade (gr) 2-4 mucositis complicates myeloablative (MAC) allogeneic hematopoietic transplantation (HCT) in 90% of patients. Methotrexate (MTX) GVHD prophylaxis (ppx) contributes to mucositis and delays engraftment. Severe mucositis increases patient-controlled analgesia (PCA) and TPN (total parenteral nutrition) use, length of stay (LOS) and precludes completion of post HCT MTX. Leucovorin (LCV) following MTX is controversial. Objectives To address exacerbation of mucositis by MTX, we implemented routine LCV post MAC alloHCT and compared outcomes with historical controls. The primary endpoint was the duration of gr 2-4 mucositis. Secondary endpoints were the incidence of gr 3-4 mucositis, time to engraftment, TPN and PCA use, LOS, incidence of GVHD and relapse. Methods Thirty-two consecutive adults received MAC alloHCT and MTX ppx with LCV 15 mg PO q6h x 4 doses, 12 h post d+3, +6, and +11 MTX and were compared to 33 consecutive historical controls. Controls did not receive routine LCV, but may have received ≥ 1 dose post d+6 or +11 MTX at physician discretion. Results Baseline characteristics were similar between the groups. All patients received tacrolimus and MTX GVHD ppx, most with Cy/TBI conditioning. Forty percent received peripheral blood stem cells and 60% had a matched unrelated donor. The median age was 46 y and 50% had AML. Thirty percent of controls received ≥ 1 LCV dose (most post d+11 MTX). The mean duration of gr 2-4 mucositis was shorter with LCV vs. control (7.4 vs. 11 d, p = 0.02). A trend towards lower incidence of gr 3-4 mucositis was observed with LCV (34% vs. 58%, p = 0.08). TPN use was similar between groups (25% vs. 36%, p = 0.4), however duration was shorter with LCV (8.6 vs. 21 d, p = 0.02). A trend towards less frequent PCA use was observed with LCV (34% vs. 58%, p = 0.08). Neutrophil engraftment was faster with LCV (17.6 vs. 20 d, p = 0.04) and a trend towards faster platelet engraftment was observed (15 vs. 20 d, p = 0.06). LOS was shorter with LCV (27 vs. 33 d, p = 0.01). LCV did not impact the incidence of GVHD. Relapse was less common in the LCV group (3% vs. 27%, p = 0.01) albeit with a shorter follow up (mean 202 d vs. 560 d). Conclusion Routine LCV addition to MAC alloHCT with MTX ppx shortened the duration of gr 2-4 mucositis, expedited neutrophil engraftment, shortened TPN and LOS and showed trends towards reduction in clinically important endpoints related to mucositis incidence, severity and associated supportive care. Our small sample size and non-randomized design may limit the strength of these conclusions. Differences between the groups may have been biased towards the null as 30% of controls received ≥ 1 LCV dose. LCV ppx warrants prospective evaluation to assess effects on mucositis, engraftment, GVHD, resource utilization and LOS.

Magnesium alterations of graft immune cells (MAGIC).

e18541 Background: Magnesium plays important roles in cellular processes. Allogeneic hematopoietic stem cell transplantation provides potentially curative therapy to patients with a variety of hematologic malignancies. A common abnormality post transplant is hypomagnesaemia. Recent studies identified magnesium transporter 1 deficiency in association with XMEN syndrome. These patients exhibit low intracellular [Mg++] and low T cell and NK cell NKG2D expression. Therefore, we investigated this potential functional link between magnesium deficiency and immune function in patients post transplant. Methods: A pilot study investigated whether detectable changes in T and NK cells were associated with IV magnesium repletion post hematopoietic transplant. Blood was obtained before and after magnesium transfusions. Immunophenotypic analysis of T and NK cell NKG2D levels using commercially available monoclonal antibodies (CD3, CD16, CD56, CD4, CD8, CD314 [NKG2D]) was performed. Samples from patients demonstrating an increase in NKG2D expression were subsequently analyzed for ex vivo NK activity. Here, NK activity and cytotoxicity of Jurkat cells were assayed using standard laboratory techniques and changes > 2 fold were considered significant. A total of 7 patients were enrolled. Results: NKG2D expression on T and NK cell subsets were determined by flow cytometry using commercially available monoclonal antibodies. NK reactivity and killing of K562 were undertaken using standard cytotoxicity assays. Our findings indicate a significant response in five of the seven enrolled patients. NKG2D expression on CD8+ T cells increased by up to 31%. Conclusions: Our results suggest that magnesium infusions post allogeneic transplant are associated with augmented NKG2D expression on immune cells. Additional studies are needed to clarify the mechanisms by which therapeutic magnesium repletion restores the cytotoxicity of NK cells and T lymphocytes. Although the clinical utility of magnesium supplementation remains to be validated, our data indicate that NKG2D is regulated by magnesium post transplant and magnesium may play a key role in immune regulation.

Alloreactive Natural Killer Cells Initiate a Unique Cellular and Molecular Pathway That Greatly Accelerates Immune Reconstitution after Allogeneic Bone Marrow Transplantation

One outstanding issue in allogeneic hematopoietic transplantation is impaired immune reconstitution. As the primary site of T cell development, the thymus plays a key role in the generation of a strong yet self-tolerant adaptive immune response, essential in the face of the potential threat from pathogens or neoplasia. Allogeneic hematopoietic transplantation may acutely damage the thymus through the chemo or radiotherapy, antibody therapy of the conditioning regime, infections acquired by the immunosuppressed patient, and thymic graft versus host disease. To date, attempts to improve thymic reconstitution have been disappointing. Pre-clinical experiments and pilot clinical trials tried to assess the role of a variety of therapeutic approaches, such as transfer of lymphoid progenitor cells, thymic grafts, or enhancement of thymopoiesis by administration of hormonal or cytokine/growth factor-based therapies, such as sex-steroid blockade, and IL-7, IL-22, KGF, or Flt-3 ligand administration (reviewed in Chaudhry et al., Immunol Rev. 2016). In mouse MHC mismatched transplantation models (F1 H-2d/b→parent H-2b), we previously found that infusion of donor versus recipient alloreactive NK cells eradicated recipient-type lympho-hematopoietic lineage cells, thereby enhancing engraftment, protecting from GvHD and eradicating leukemia (Ruggeri et al., Science 2002). Here, in the same models we show that infusion of alloreactive NK cells greatly accelerates the post-hematopoietic transplant recovery of donor-type immune cells, i.e., dendritic cells (DCs) (p<0.001), B lineage cells (p<0.001) and thymocytes (p<0.001) and maturation to B (p<0.001) and T cells (p<0.001). By the use of recipient chimeric mice displaying different tissue (i.e., hematopoietic vs non-hematopoietic) susceptibility to donor alloreactive NK cell killing, we show that a specific interaction between donor alloreactive NK cells and recipient DCs is responsible for the accelerated immune rebuilding. We find that donor-versus-recipient alloreactive NK cells trigger recipient DCs to synthesize a protein factor in a DNA translation-depended fashion (i.e., blocking DNA transcription in DCs abrogated the DC ability to produce the factor), and release it. Infusion of NK/DC co-culture supernatants containing this factor induced bone marrow and thymic stromal cells to produce IL-7 (p<0.001) and c-Kit ligand (p<0.001) and, thereby, the extraordinarily accelerated maturation of donor DCs, B- and T-cell precursors. Interestingly, in vitro experiments with human thymic stromal cells that support human thymocyte proliferation and differentiation demonstrated the exact same mechanisms. Supernatants from human alloreactive NK cell clones and human (HLA-class I KIR ligand mismatched) allogeneic DCs induced IL-7 production by human thymic stromal cells which in turn supported accelerated proliferation and maturation of human thymocytes (p<0.001). The murine and the human "immune rebuilding" factors displayed biochemical similarities as they both are highly hydrophobic 12KDa molecular weight proteins. Mass spectrometry analysis by stable isotope labeling with amino acids in cell culture (SILAC) identified Beta-2 Microglobulin (B2M) as the newly synthesized protein sharing the above biochemical features and present both in murine and human samples. B2M-KO mice used as recipients of MHC mismatched bone marrow transplant and given donor versus recipient alloreactive NK cells were unable to undergo accelerated immune rebuilding. However, their defect was repaired and accelerated rebuilding of donor-type DCs (p<0.001), B lineage cells (p<0.001) and thymocytes (p<0.001) was restored by the administration of culture supernatants obtained from alloreactive NK cells and wild-type (non-KO) MHC mismatched DCs. Finally, RNA interference experiments that silenced the B2M gene in human DCs resulted in loss of biological activity of supernatants obtained from alloreactive NK cells and B2M-silenced DCs. B2M plays a key role in the immune system as it is known to be part of MHC class I molecules. However, its role in signaling for immune precursor cell development has never been recognized. Here we report the discovery of a novel cellular and molecular pathway initiated by alloreactive NK cells and mediated by B2M that leads to greatly accelerated rebuilding of B and T cells after hematopoietic transplantation. DisclosuresNo relevant conflicts of interest to declare.