Outcome Of Bone Marrow Transplantation Research Articles

The Potential Role of NOD2/CARD15 Genotype as a Prognostic Indicator for Bone Marrow Transplantation Outcomes in Patients With Acute Lymphoblastic Leukemia and Acute Myeloid Leukemia: A Systematic Review.

Hematopoietic stem-cell transplantation (HSCT) has emerged as a groundbreaking therapeutic option for acute myeloid leukemia (AML) and specific subtypes of acute lymphoblastic leukemia (ALL). The prognostic significance of the NOD2/CARD15 gene has been explored alongside various factors, encompassing diverse patient cohorts and gene variants. Siblings and unrelated donors used for stem cell transplantation exhibit significant associations between their genetic variations and graft-versus-host disease incidence. The transplantation of stem cells for leukemia patients involves numerous considerations, including patient survival, relapse rates, disease stage, donor and recipient ages, and compatibility. This study delved into research on the NOD2/CARD15 gene and its mutations to assess its suitability as a screening tool. A comprehensive literature search encompassing PubMed, ScienceDirect, and Google Scholararticles yielded 4,840 articles. After removing duplicates and applying inclusion and exclusion criteria, we narrowed the search results to 876 articles. Subsequent screening of abstracts and titles resulted in the selection of 230 relevant articles. Further exclusion of 198 articles unrelated to the research question led to the scrutinizing of 32 full-text articles, which were assessed against inclusion and exclusion criteria. Emphasis was placed on articles that specifically investigated the role of NOD2/CARD15 as a predictive factor for HSCT outcomes, ultimately resulting in the inclusion of 19 articles in this study.Single nucleotide polymorphisms (SNPs) such as NOD2 and CARD15 have demonstrated their potential as reliable genetic markers for predicting post-transplantation relapse and disease outcomes. Patients positive for these genetic markers have exhibited reduced overall survival and event-free survivaland increased transplant-related mortality. Interventions with interferon-gamma and muramyl tripeptide phosphatidylethanolamine have been considered to mitigate the inflammatory effects of these SNPs, thus enhancing the influence of natural killer cells on abnormal cells and potentially extending patient survival. NOD2/CARD15 typing may aid in identifying patients at higher risk for relapse and improving their clinical outcomes after allogeneic stem cell transplant, particularly in ALL patients. However, no remarkable change was observed in AML patients. Additionally, this study underscores the pivotal roles of adaptive and innate immune responses and their interplay in stem cell transplant immunology.

Second bone marrow transplantation into regenerating hematopoiesis enhances reconstitution of immune system.

In bone marrow transplantation (BMT), hematopoiesis-reconstituting cells are introduced following myeloablative treatment, which eradicates existing hematopoietic cells and disrupts stroma within the hematopoietic tissue. Both hematopoietic cells and stroma then undergo regeneration. Our study compares the outcomes of a second BMT administered to mice shortly after myeloablative treatment and the first BMT, with those of a second BMT administered to mice experiencing robust hematopoietic regeneration after the initial transplant. We evaluated the efficacy of the second BMT in terms of engraftment efficiency, types of generated blood cells, and longevity of function. Our findings show that regenerating hematopoiesis readily accommodates newly transplanted stem cells, including those endowed with a robust capacity for generating B and T cells. Importantly, our investigation uncovered a window for preferential engraftment of transplanted stem cells coinciding with the resumption of blood cell production. Repeated BMT could intensify hematopoiesis reconstitution and enable therapeutic administration of genetically modified autologous stem cells.

Myelodysplastic Syndromes Benefit from Iron-Restricted Bone Marrow Transplant in Preclinical Mouse Model

Background: Hematopoietic stem cell transplantation (HSCT) remains the only fully curative therapy for diseases such as b-thalassemia, sickle cell disease, MDS and AML and represents a relevant procedure for gene therapy .Tissue iron overload and non-transferrin bound iron (NTBI) have emerged as adverse prognostic factors for post-transplant survival. Recently, we showed that - in line with observations in transplanted individuals - preparatory chemotherapy conditioning with a combination of busulfan and fludarabine in wild-type mice resulted in elevated serum iron and transferrin (Tf) saturation and elicited NTBI formation. Tf saturation and NTBI levels increased proportionally with conditioning intensity. Interestingly, we found that pre-conditioning TMPRSS6 silencing through a GalNAc-conjugated siRNA - by increasing hepcidin levels - reduced the peri-transplant elevation of Tf saturation and limited the appearance of NTBI. These results proved that NTBI formation is induced by conditioning chemotherapy in the peri-trasplant period independently of the presence of pre-existing iron overload. Aims & Methods: Here we asked whether these observations hold true in conditions hallmarked by pre-existing iron overload and elevated NTBI and how iron restriction impacts on bone marrow transplant (BMT) outcome. To this aim, we applied the GalNAc TMPRSS6 siRNA as iron restriction strategy in a mouse model of myelodysplastic syndromes (MDS) and assessed its ability to reduce conditioning-elicited NTBI formation as well as its potential benefit on BMT. After TMPRSS6 siRNA administration, CD45.2 + MDS mice received conditioning chemotherapy and transplant of CD45.1 + wild-type bone marrow cells. Systemic iron parameters, engraftment and chimerism were then analyzed. Results: Importantly, conditioning exacerbated the elevated systemic iron levels in MDS mice, which was prevented by pre-conditioning TMPRSS6 silencing. Similar to our observation in wild-type mice, recipient CD45.2 + MDS mice showed a superior blood chimerism, resulting from increased bone marrow and spleen engraftment of CD45.1 + donor cells. Multilineage engraftment in bone marrow, spleen and peripheral blood was superior in iron-restricted recipient MDS mice compared to controls. Finally, donor chimerism of hematopoietic stem and progenitor cells (HSPCs) was higher in iron-restricted versus control MDS mice undergoing BMT. Conclusion: These findings demonstrate that pre-transplant conditioning profoundly alters iron homeostasis and restricting iron improves BMT outcome by increasing engraftment efficiency. Overall, these data suggest that conditioning-elicited NTBI is a critical mediator of peri-transplant toxicity and adversely impacts transplant outcome. Specifically, our observations prove that iron restriction improves donor HSPC engraftment in the recipient bone marrow, likely resulting in more efficient reconstitution of the HSPC pool and improved multilineage engraftment. Lastly, our results indicate that the peri-transplant application of novel iron restriction strategies aimed at reducing conditioning-induced NTBI - by positively affecting BMT outcome - is of potential benefit for patients undergoing BMT or gene therapy, presenting either with or without pre-transplant iron overload.

Superior Outcome of Bone Marrow Transplantation Using Fludarabine and Cyclophosphamide with ATG over Fludarabine and Cyclophosphamide without ATG Conditioning Regimen in Severe Aplastic Anemia

ABSTRACT Background: Matched sibling donor (MSD) allogenic hematopoietic stem cell transplantation (allo-HSCT) is the standard of care as the first line therapy for young fit patients with severe aplastic anemia (SAA). There are several conditioning regimens that have been evaluated and used for these patients. Fludarabine (Flu)/cyclophosphamide (Cy)/antithymocyte globulin (ATG) or Cy/ATG led to the best survival. The 5-year probabilities of survival after Flu/Cy/ATG, Cy/ATG, Cy ± Flu, and Busulfan /Cy were 91% (95% CI, 85-96), 91% (95% CI, 89-94), 80% (95% CI, 73-87), and 84% (95% CI, 75-91) (P = .001), (Bejanyan et al. 2019). Objective: To report the outcome of a homogeneous cohort of SAA patients who received a uniform transplantation conditioning regimen protocol (Flu/Cy/ATG) and compare it with the historical protocol (Flu/Cy) from the same transplant center for the same patients population. Methods: This study is a 10-year retrospective analysis of SAA patients who underwent MSD allo-HSCT at King Faisal Specialist Hospital & Research Centre, Riyadh City, Saudi Arabia during the period from 2013 to 2022. The conditioning regimen was consistent of Fludarabine 150 mg/m 2/Cyclophosphamide 200 mg/kg/ Anti-thymocyte globulin (rabbit) 2.5 mg/kg (Flu/Cy/ATG). Graft-versus-host disease (GVHD) prophylaxis was methotrexate (MTX)/tacrolimus (FK) or methotrexate/cyclosporin (CsA). The outcome was compared with the historical published data of 82 patients with SAA received Flu/Cy conditioning regimen for MSD allo-HSCT, (El Fakih et al. 2021). Results: The study included 28 patients with SAA, 46.2% (n=12) patients were females, paroxysmal nocturnal hemoglobinuria (PNH) clone detected in 57.1% (n=16). Total body irradiation (TBI) was added to the conditioning regimen in 25% (n=7) for patients with heavily blood products transfusion pre-HSCT. A total of twenty-six patients (93%) has fully MSD, while 2 (7%) has matched unrelated donors (MUD). Bone marrow (BM) was the stem cell source in 60.7% while 28.6% received peripheral blood (PB) stem cell and 10.7 % received combined (BM/PB). There were twelve patients with donor-recipient sex mismatch. Major ABO mismatch counted for 17.9% (n=5). The median age at transplantation was 25 ±10.2 years. During the observational period, there was no graft failure (GF) and none of the patients died. At 60 days of HSCT; 61 % (n=17) patients achieved full donor whole blood (WB) chimerism, while 11 (39 %) patients had mixed chimerism. At both 180 days and 365 days of HSCT; 69.2% (n=18) patients had full donor chimerism and 30.8% (n=8) of the patients had mixed donor chimerism. Acute GVHD occurs in five cases (17.9%) and none had developed grade III or IV, notably; 80% of GVHD cases (4/5) received PB or BM/PB stem cell source. Chronic GVHD was observed in six cases (21.7%), 66% (4/6) of them had received PB or PB/BM stem cell source. The use of PB was due to either donor related reasons like recipient-donor weight discrepancy or patient's medical condition to achieve early engraftment. Hemorrhagic cystitis occurred in one case (3.6%), none had sinusoidal obstruction syndrome (SOS). CMV viremia without disease occurs in eleven patients (39 %). The median follow-up time for survivors was 34 months, the 2 years overall survival (OS) was 100%, the 2 years event free survival (EFS) defined with GF, or death was 100% compared with the previous result published (El Fakih et al., 2021) which had showen lower OS and EFS of 87.8% and 82.9% respectively with median follow-up of 100 months, Figure1&2. Table-1 and table-2 summarize the comparison of patient's characteristics and outcome of both regimens (Flu/Cy vs Flu/Cy/ATG). Conclusions: The results of the present study showed better outcomes for SAA patients using Flu/Cy/ATG compared with the previous study (El Fakih et al., 2021). Early MSD allo-HSCT for SAA patients using Flu/Cy/ATG conditioning regimen is associated with a favorable outcome of excellent graft function, low rate GVHD and no GF. BM should be the stem cell source in severe aplastic anemia HSCT procedure. Key Words: Severe Aplastic Anemia, Matched Sibling Donor, Conditioning, Bone Marrow Graft, Flu/Cy/ATG.

HLA Class I Allele Loss and Bone Marrow Transplantation Outcomes in Immune Aplastic Anemia

In patients with immune-mediated acquired aplastic anemia (AA), HLA class I alleles often disappear from the surface of hematopoietic progenitor cells, potentially enabling evasion from cytotoxic T lymphocyte-mediated pathogenesis. Although HLA class I allele loss has been studied in AA patients treated with immunosuppressive therapy (IST), its impact on allogeneic bone marrow transplantation (BMT) has not been thoroughly investigated. The purpose of this study was to evaluate the clinical implications of HLA class I allele loss in patients with acquired AA undergoing allogeneic BMT. The study enrolled acquired AA patients who underwent initial BMT from unrelated donors through the Japan Marrow Donor Program between 1993 and 2011. The presence of HLA class I allele loss due to loss of heterozygosity (HLA-LOH) was assessed using pretransplantation blood DNA and correlated with clinical data obtained from the Japanese Transplant Registry Unified Management Program. A total of 432 patients with acquired AA were included in the study, and HLA-LOH was detected in 20 of the 178 patients (11%) available for analysis. Patients with HLA-LOH typically presented with more severe AA at diagnosis (P=.017) and underwent BMT earlier (P < .0001) compared to those without HLA-LOH. They also showed a slight but significant recovery in platelet count from the time of diagnosis to BMT (P=.00085). However, HLA-LOH status had no significant effect on survival, engraftment, graft failure, chimerism status, graft-versus-host disease, or other complications following BMT, even when the 20 HLA-LOH+ patients were compared with the 40 propensity score-matched HLA-LOH- patients. Nevertheless, patients lacking HLA-A*02:06 or HLA-B*40:02, the alleles most frequently lost and associated with a better IST response, showed higher survival rates compared to those lacking other alleles, with estimated 5-year overall survival (OS) rates of 100% and 44%, respectively (P=.0042). In addition, in a specific subset of HLA-LOH- patients showing clinical features similar to HLA-LOH+ patients, the HLA-A*02:06 and HLA-B*40:02 allele genotypes correlated with better survival rates compared with other allele genotypes, with estimated 5-year OS rates of 100% and 43%, respectively (P=.0096). However, this genotype correlation did not extend to all patients, suggesting that immunopathogenic mechanisms linked to the loss of certain HLA alleles, rather than the HLA genotypes themselves, influence survival outcomes. The survival benefit associated with the loss of these two alleles was confirmed in a multivariable Cox regression model. The observed correlations between HLA loss and the pretransplantation clinical manifestations and between loss of specific HLA class I alleles and survival outcomes in AA patients may improve patient selection for unrelated BMT and facilitate further investigations into the immune pathophysiology of the disease.

Efficacy of Cord Blood Transplantation for the Management of Chronic Myelomonocytic Leukemia: A Registry-Based Study in Japan

Introduction Chronic myelomonocytic leukemia (CMML) is a challenging malignancy that shares characteristics with myelodysplastic syndromes and myeloproliferative neoplasms. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only curative option for CMML. Numerous studies have examined transplant outcomes and prognostic factors; however, the focus has primarily been on bone marrow transplantation (BMT) and peripheral blood stem cell transplantation, while cord blood transplantation (CBT) is understudied. To address this knowledge gap, we conducted a nationwide retrospective study to comprehensively compare the outcomes of unrelated BMT and single-unit umbilical CBT in patients with CMML. Methods Data were collected from the Transplant Registry Unified Management Program 2 of the Japanese Data Center for Hematopoietic Cell Transplantation. The inclusion criteria were as follows: age of 16 years or higher, a diagnosis of CMML, and receipt of the first allo-HSCT between 2013 and 2019 through unrelated BMT from a human leukocyte antigen (HLA) 8/8-matched donor (UBMT group) or single-unit umbilical CBT (CBT group). The primary endpoint was overall survival (OS) after allo-HSCT, and the secondary endpoints included progression-free survival (PFS), cumulative incidences of relapse and non-relapse mortality (NRM), neutrophil engraftment, and graft-versus-host (GVHD)-free, relapse-free survival (GRFS). The impact of CBT versus unrelated BMT on outcomes was evaluated using inverse probability of treatment weighting (IPTW) analysis. In this analysis, the Cox proportional hazards regression model for OS, PFS, and GRFS and the Fine-Gray proportional hazards regression model for relapse, NRM, engraftment, and GVHD were used. Hazard ratios (HRs) and 95% confidence intervals (CIs) were calculated for all outcomes. The propensity score (PS) was calculated via logistic regression by using the following factors: age at allo-HSCT, sex, cytogenetic risk, disease status, days from diagnosis to allo-HSCT, and year of allo-HSCT. For the CBT group, the stabilized weights were calculated by dividing CBT prevalence by each individual's PS. Conversely, for the UBMT group, the weights were calculated by dividing the 1 minus prevalence of CBT by 1 minus each individual's PS. Results Between 2013 and 2021, 238 patients with CMML underwent their first allo-HSCT. Among them, 50 underwent HLA 8/8 allele-matched unrelated BMT (UBMT group) and 68 underwent single-unit umbilical CBT (CBT group). The median age at allo-HSCT was 58.5 (range: 26-75) years; the median follow-up period for survivors was 1,329.5 (range: 100-1,967) days. Intergroup differences in age at allo-HSCT, recipient sex, hematopoietic cell transplantation-specific comorbidity index, Eastern Cooperative Oncology Group performance status, cytogenetic risk, pre-transplant treatment, disease status at allo-HSCT, conditioning intensity, GVHD prophylaxis, and year of allo-HSCT were not significant. The transplantation outcomes are shown in Figure 1. There were no significant differences in the 3-year OS rate (UBMT: 51.0% [95% CI: 34.1-65.5%] vs CBT: 46.2% [95% CI: 33.2-58.1%]; P = 0.60; Figure 1A). IPTW analysis revealed no significant differences between the CBT and UBMT groups (HR: 1.21, 95% CI: 0.69-2.10, P = 0.51). PFS, relapse, NRM, and GRFS were not significantly different between the groups (Figure 1B-E). The neutrophil engraftment rate at 30 days was significantly higher in the UBMT group than in the CBT group (UBMT: 92.0% [95% CI: 78.9-97.1%] vs CBT: 79.4% [95% CI: 67.5-87.4%]; P &amp;lt; 0.01; Figure 1F). These observations associated with the secondary endpoints were concurrently corroborated by the IPTW analysis. Conclusion Our study suggests that CBT can serve as a viable treatment option, demonstrating efficacy comparable to that of unrelated BMT for patients with CMML. These findings underscore the improved outcomes over time after CBT in patients with CMML, which is a treatment previously perceived as inferior to other donor sources (Itonaga et al., Biol Blood Marrow Transplant. 2018;24:840-48). Further investigations are warranted to optimize transplantation strategies and improve outcomes in patients with CMML.

Favorable Outcomes for High-Risk MDS and Oligoblastic AML with MDS-Related Changes with Reduced Intensity Allogeneic Bone Marrow Transplantation with Post-Transplantation Cyclophosphamide

Introduction: Newer classification systems label myelodysplastic syndrome (MDS) with 10-20% blasts and acute myeloid leukemia (AML) with MDS-related changes (AML-MR) on a continuum as they share common biologic features. The prognostic implication of blast percentage is less clinically relevant particularly in diseases with high-risk molecular features such as MDS/AML with TP53 mutations. The recently introduced Molecular International Prognostic Scoring System (IPSS-M) allows for a more personalized prognosis for MDS patients (pts) than the prior revised IPSS (IPSS-R), but it is less clear if this translates to better therapeutic choices or outcomes for pts. Allogeneic bone marrow transplantation (BMT) remains the only potentially curative approach. Increasingly clinical trials of novel compounds combined with hypomethylating agents (HMAs) have not shown improved efficacy over single agent HMAs, necessitating further study of BMT approaches for durable disease control across the spectrum of disease biology. No uniform approach to the conditioning regimen and GVHD prophylaxis for this pt population has proven superior. In the current study, we describe the outcomes of BMT with post-transplant cyclophosphamide (PTCy) for patients with high-risk MDS and oligoblastic AML-MR, and analyze the prognostic implication of their chromosomal and molecular profiles within the international scoring systems. Patients and methods: This is a retrospective analysis of patients with 5-30% blasts and IPSS-R ≥3 who underwent BMT with PTCy at Johns Hopkins Hospital between 1/2013 and 12/2020. Next generation sequencing (NGS) was performed within 12 months of diagnosis. Kaplan-Meier analysis was performed to evaluate overall survival (OS) and relapse-free survival (RFS). Competing risks analysis was performed to evaluate the incidence of relapse and non-relapse mortality (NRM). Cox-regression models were performed in univariate analyses with false discovery rate adjustment and then subsequently conducted for multivariable analyses to identify statistically significant and independent prognostic factors. Results: Ninety-four consecutive patients (65% male) were identified with a median age of 61 (range 22 - 75) years at diagnosis. Median follow-up was 4.8 years (range 5 days-9.6 years) post BMT. Two-thirds (63%) underwent haploidentical transplantation, and 91% received reduced intensity conditioning with Fludarabine, Cyclophosphamide, and TBI. Median age at BMT was 65 years (range 22 - 76). The median time from diagnosis to BMT was 7.9 months. 52 (57%) received HMA based treatment and 36 (39%) intensive chemotherapy prior to BMT. Graft failure occurred in 6 patients (6.4%) The 5-year OS rate was 53% (95% CI: 43%-65%), the 5-year RFS rate was 47% (95% CI: 38%-59%) ( Figure 1A). The incidence of relapse was 21% (95% CI: 13% - 30%) at 1 year and 31% (95% CI: 21% - 40%) at 2 years after BMT ( Figure 1B). NRM rate in the first-year was 7% (95% CI: 2%-13%) and 11% (95% CI: 4%-17%) in the first two years ( Figure 1B). The incidence of acute GVHD (aGVHD) grades 3 to 4 was 3.2%, whereas chronic GVHD (cGVHD) presented in 12 patients (12.7%) with only 2 patients (2.1%) having moderate to severe disease based on the NIH grading criteria. Univariate analysis showed that higher IPSS-R (HR 1.68, 95% CI 1.39 - 2.04), poor or very poor karyotype risk (HR 7.48, 95% CI 3.49 - 16.02), complex karyotype (HR 3.72, 95% CI 2.01 - 6.88), higher IPSS-M (HR 1.72, 95% CI 1.38 - 2.14), TP53 mutations (HR 6.53, 95% CI 3.36 - 12.7) and RAS mutations (HR 4.96, 95% CI 1.7 - 14.5) were associated with worse OS. IPSS-R was significantly correlated with the patients' response to bridge therapy (OR 1.34, P=0.036). IPSS-R, IPSS-M, poor or very poor karyotype risk, complex karyotype, TP53 and any RAS mutations and response to bridge therapy were independently associated with worse OS and RFS ( Table 1, Figure 1C). Conclusion: This retrospective analysis supports that BMT with PTCy is a durable approach for patients with high-risk MDS and oligoblastic AML-MR leading to high OS and RFS associated with a promising safety profile with low NRM and severe GVHD incidence. IPSS-R, IPSS-M, karyotype and response to bridge therapy are independent predictors of OS and RFS post-BMT. The presence of TP53 and/or RAS mutations defines a sub-group of patients with dismal outcomes representing an unmet need within this patient population.

Developing an Allogeneic Hematopoietic Progenitor Cell Transplant Service in a Resource-Limited Country: Challenges and Outcomes.

Introduction Allogeneic stem cell transplant has curative potential for many hematological disorders. Building an allogeneic hematopoietic progenitor cell transplant (HPCT) unit requires huge investment, infrastructure, equipment, medical supplies, and training of health care professionals. The key objective of this study is to share our experience of developing an allogeneic HPCT service at our tertiary care cancer hospital in a low-middle-income country. In addition, this study presents the outcomes of the first 30 allogeneic HPCTs done at our center. Methods This retrospective observational study included adult patients 18 years old or older with hematological malignancies who underwent allogeneic HPCT between July 2019 and April 2023 at Shaukat Khanum Memorial Cancer Hospital and Research Centre. Result Of the 30 patients, 24 underwent matched sibling donor (MSD) transplants in which a myeloablative-conditioning regimen (MAC) was used in 19, and a reduced conditioning regimen (RIC) was used in one. Of the six haploidentical-related donor transplants, four received MAC, and two received RIC. The median recipient age at HPCT was 23 and 21 years for MSD and Haplo-related donor transplants, respectively. The median follow-up duration was 12 months (Range: 10 days - 33 months). The overall survival rate at one yearwas 71.3% among all allogeneic stem cell transplant patients, whereas the disease-free survival rate at one yearwas 63.7%. In the acute lymphoblastic leukemia group, the disease-free survival rate at one year post allograft was 51.5%, while in the acute myeloid leukemia group, it was 78.7%. Conclusion This study demonstrates the successful development of an allogeneic bone marrow transplant unit at our hospital despite significant financial constraints. This has allowed us to provide a potentially curative and life-saving treatment to a substantial number of cancer patients. The bone marrow transplant outcomes of this study are comparable to those reported by international bone marrow transplant registries.

Harvest Quality, Nucleated Cell Dose and Clinical Outcomes in Bone Marrow Transplantation: A Retrospective Study

Higher doses of infused nucleated cells (NCs) are associated with improved clinical outcomes in bone marrow transplantation (BMT) recipients. Most clinicians recommend infusing at least 2.0×108 NCs/kg. BMT clinicians request a target NC dose, but the harvested NC dose may be below the requested NC dose even before cell processing. We conducted this retrospective study to investigate the quality of bone marrow (BM) harvest and factors that influence infused NC doses at our institution. We also correlated infused NC doses with clinical outcomes. The study population included 347 BMT recipients (median age, 11 years; range, <1 to 75 years) at the University of Minnesota between 2009 and 2019. Underlying diagnoses mainly included 39% malignant and 61% nonmalignant diagnoses. Requested, harvested, and infused NC doses, as well as cell processing data, were obtained from the Cell Therapy Laboratory; clinical outcomes data were obtained from the University of Minnesota BMT Database. BM harvests were facilitated either by our institution (61%) or by the National Marrow Donor Program (39%). Associations of infused doses with baseline characteristics were assessed using the general Wilcoxon test/Pearson's correlation coefficient. The association of infused dose with neutrophil engraftment (absolute neutrophil count >500) by day 42, platelet engraftment (>20,000) by 6 months, acute graft-versus-host disease grade II-IV, and overall survival (OS) at 5 years were evaluated using regression and Kaplan-Meier curves. The median requested NC dose was 3.0×108/kg (range, 2 to 8×108/kg), and the median harvested and infused NC doses were 4.0×108/kg and 3.6×108/kg, respectively. Only 7% of donors had a harvested dose below the minimum requested dose. Moreover, the correlation between requested doses and harvested doses was adequate, with a harvested/requested dose ratio <.5 observed in only 5% of harvests. Additionally, the harvest volume and cell processing method were significantly correlated with the infused dose. Harvest volume exceeding the median of 948 mL was related to a significantly lower infused dose (P < .01). Moreover, hydroxyethyl starch (HES)/buffy coat processing (used to reduce RBCs with major ABO incompatibility) led to a significantly lower infused dose (P < .01). Donor age (median, 19 years; range, <1 to-70 years) and sex did not significantly influence the infused dose. Finally, the infused dose was significantly correlated with neutrophil and platelet engraftment (P < .05) but not with 5-year OS (P=.87) or aGVHD (P=.33). In our program's experience, BM harvesting is efficient and meets the requested minimum dose for 93% of recipients. Harvest volume and cell process play significant roles in determining the final infused dose. Minimizing harvest volume and cell processing could lead to increased infused dose and thus improved outcomes. Moreover, a higher infused dose leads to a better rate of neutrophil and platelet engraftment but not to improved OS, which may be linked to the sample size of our study.

Impact of percutaneous endoscopic gastrostomy on pediatric bone marrow transplantation outcomes: Retrospectice single-center cohort study.

Parenteral nutrition is commonly used to ensure nutrition support and prevent the harmful effects of malnutrition, which frequently occurs after allogeneic hematopoietic stem cell transplantation (aHSCT). Nevertheless, enteral nutrition supports the restoration of the gut barrier and microbiome as well as protects against infectious complications and acute graft-vs-host disease. Percutaneous endoscopic gastrostomy (PEG) may also be beneficial for gastric decompression and drug administration. We performed a retrospective cohort study to evaluate the impact of PEG on treatment outcomes in 75 children who underwent aHSCT with (n = 34) or without (n = 41) PEG from 2005 to 2016. In 34 patients, PEG was used to ensure enteral nutrition support (n = 30), oral drug intake (n = 28), and abdominal decompression (n = 2). During the study period, we observed a beneficial association between PEG placement and transplant-related mortality as well as 5-year overall survival compared with the non-PEG group (12.9% vs 59.0%, P = 0.000; 85.3% vs 35.1%, P = 0.000, respectively). The beneficial impact of PEG was most prominent on 5-year overall survival in older children (12-17 years) with grafts from matched unrelated donors. PEG placement had a positive association with transplant outcomes in pediatric patients undergoing aHSCT. To confirm these results, larger prospective studies are needed.

Iron Overload and Iron-Induced Microbiome Changes May Affect Lymphoid Levels Post Bone Marrow Transplant and Graft-Versus-Host Disease Survival in Mice

Introduction: Bone Marrow Transplant (BMT) represents the only cure for many malignant and non-malignant hematopoietic disorders. This treatment is often accompanied by multiple red blood cell transfusions, which can lead to iron overload. Iron overload is associated with poorer outcomes following BMT, such as increased risk of infections or Graft-versus-Host Disease (GVHD); however, it is not yet clear whether this correlation is caused by iron itself or is simply confounded by patients with more severe disease requiring more transfusions. Previous studies using mouse models, have shown that transfusional iron overload reduces the population of beneficial bacteria in the gut microbiome or decreases the production of microbiome-produced metabolites (i.e. indoles) that have been associated with BMT outcomes and GVHD risk. Here, we developed a reductionist mouse model of iron overload and BMT to investigate the effect of iron and iron-induced changes to gut microbiota and their metabolites on BMT outcomes and GVHD survival. Methods: Female Balb/C mice were infused twice a week, for two weeks, with a total of 50 mg of iron dextran or saline followed by lethal irradiation (9 Gy). Cohorts of mice (n=10 per group) received 1x10^10 T cell-depleted bone marrow cells with or without 5x10^6 T cells from UBC-GFP C57Bl6 mice to establish allogenic BMT and GHVD models, respectively. Mice were then monitored daily for survival with a formal severity score evaluated twice a week. Analysis of donor red blood cells, platelets, and white blood cells (neutrophils, B and T cells) from peripheral blood was also performed once a week to check for engraftment. To examine the impact of iron-induced microbiota changes, fecal microbiome transplant (FMT) was performed in mice receiving BMT. Briefly, mice were treated with an antibiotic cocktail (ampicillin 1 g/L, kanamycin 1 g/L, metronidazole 1g/L, vancomycin 0.5g/L) by gavage 2 days before the start of FMT. Feces from control and iron overloaded mice were collected freshly and weighed. Stool pellets were then resuspended in PBS (100 mg feces/ml PBS), vortexed and then spun down at 800g for 3 minutes. Mice received 100ul of supernatant by gavage. Additional cohorts of mice were administrated with indole-3 proprionic acid (IPA) in the drinking water at a concentration of 0.1 mg/ml prior to irradiation throughout the experiment and monitored for survival after receipt of allogeneic T cells to induce GVHD. All mice received enrofloxacin (10mg/kg) starting on the day of BMT and for the following 3 days by intraperitoneal injection. Results: There was a significant reduction in overall lymphoid (CD45+ CD172a- CD11b-/low GFP+), B (CD45+ CD172a- CD11b-/low CD19+ GFP+), and T (CD45+ CD172a- CD11b-/low CD3+ GFP+) cells in iron overloaded mice compared to control mice (p<0.0001 at day +28) by flow cytometry analysis in peripheral blood after BMT. Although FMT from control feces into iron overloaded mice did not affect lymphoid numbers, FMT from iron overloaded feces into control mice reduced the number of donor lymphoid and B cells (p<0.05 at day +28; Figure 1A). Iron overload reduced survival compared to control mice in the GHVD mouse model (p<0.05 Mantel-Cox test). Administration of IPA in iron overloaded mice did not significantly affect overall survival (Figure 1B). Conclusions: Our results show that iron overload negatively impacts lymphocyte engraftment in a mouse model of BMT. Furthermore, iron overload also reduces survival in mice with GVHD. Iron overload was previously shown to reduce indole levels in mice. Although we did not observe a significant impact of exogenous indole administration on GVHD survival, FMT from iron overloaded into control mice reduced levels of lymphoid and B cells after BMT implicating gut microbiota in this outcome. Taken together, these results suggest that iron overload is an important factor that negatively affects BMT outcomes. The extent to which iron-induced gut microbiota changes are involved in the mechanisms responsible for this effect requires further investigation. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

277 - Harvest Quality, Nucleated Cell Dose and Clinical Outcomes in Bone Marrow Transplantation: A Retrospective Study

277 - Harvest Quality, Nucleated Cell Dose and Clinical Outcomes in Bone Marrow Transplantation: A Retrospective Study

Mature Naive B Cells Regulate the Outcome of Murine Acute Graft-versus-Host Disease in an IL-10-Independent Manner

Graft-versus-host disease (GVHD) is the main complication of bone marrow transplantation (BMT). CD4+ T lymphocytes are the main effector cells for disease development, but other cell types can determine disease outcome through cytokine production and antigen presentation. B cells are abundant in BMT products and are involved in chronic GVHD immunopathogenesis. However, their role in acute GVHD is still unclear. Here we studied the role of donor resting B cells in a model of acute GVHD. Animals receiving transplants depleted of B cells developed more severe disease, indicating a protective role for B cells. Mice undergoing transplantation with IL-10 knockout B cells developed GVHD as severe as those receiving wild-type B cells. Moreover, mice that received MHC II-deficient B cells, and thus were unable to present antigen to CD4+ T cells, developed as severe GVHD as animals receiving transplants without B cells. This result suggests that the protection provided by mature naive B cells depends on antigen presentation and not on IL-10 production by B cells. Mice who underwent transplantation in the absence of donor B cells exhibited disorganized lymphoid splenic tissue. In addition, donor B cell depletion diminished the follicular T (Tfh)/effector T (Teff) cell ratio, suggesting that protection was correlated with a shift to Tfh differentiation, reducing the number of Teff cells. Importantly, the Tfh/Teff shift impacts disease outcome, with observed proinflammatory cytokine levels and tissue damage in target organs consistent with disease protection. The role of transplanted B cells in the outcome of BMT and the development of acute GVHD merits careful study, given that these cells are abundant in BMT products and are potent modulator and effector cells in the allogeneic response.

Promotion of Platelet Production by Co-Transplantation of Mesenchymal Stem Cells in Bone Marrow Transplantation.

Therapeutic strategies that can promote platelet production are in demand to enhance clinical outcomes of bone marrow transplantation (BMT). Our research group has studied human tonsil-derived mesenchymal stem cells (T-MSCs) and their effectiveness in promoting bone marrow (BM) engraftment. Here, we analyzed the effects of T-MSCs on platelet production and hemostasis. Donor BM cells (BMCs) were isolated from C57BL/6 mice and transplanted with or without T-MSCs to BALB/c recipient mice. Mice were sacrificed and blood cells were counted using an Auto Hematology Analyzer. Femur sections were stained with CD41 antibody to analyze megakaryocytes in the BM. Growth factor secretion from MSCs was analyzed using the Quantibody Array. Effects of T-MSC conditioned medium (CM) on megakaryopoiesis were investigated using the MegaCult assay. In a mouse model of BMT, T-MSC CM was injected with or without anti-placental growth factor (α-PlGF) blocking antibody, and blood cell numbers and coagulation were analyzed. T-MSC co-transplantation increased percent survival of BMT mice. Platelet numbers were significantly lower in the BMC-only group, whereas T-MSC co-transplantation restored circulating platelets to levels similar to those of the control group. Significantly reduced numbers of CD41 + megakaryocytes in Bu-Cy and BMC groups were increased by T-MSC co-transplantation. PlGF secretion from T-MSCs were detected and enhanced megakaryopoiesis, platelet production, and coagulation by T-MCS CM were disrupted in the presence of the α-PlGF blocking antibody. We demonstrated the effectiveness of T-MSC co-transplantation in promoting platelet production and coagulation after BMT. These findings highlight the potential therapeutic relevance of T-MSCs for preventing thrombocytopenia after BMT.

A Tale of Two Centers: Access to Private Care Improves Outcomes in Allogeneic Transplant Recipients

IntroductionAllogeneic bone marrow transplantation (BMT) is a potentially curative treatment for many hematological diseases. However, lack of availability of specialized centers and high costs limit access to the procedure in low and middle income countries. Previous research has shown that Latin-American patients with hematological malignancies have worse outcomes when treated in public health systems compared to patients treated in the private setting. Healthcare in Mexico is provided by three systems: the private sector (financed by a private insurance or out of pocket), social security (government-run health systems) and a public system (partially funded by the government), where the most vulnerable population is treated. To date, disparities in outcomes of BMT between patients in public and private health systems have not been widely studied.ObjectivesPrimary: to determine the impact of access to private healthcare in BMT recipients by comparing the outcomes of patients treated in two centers that are led by a single team. Secondary objectives were to determine differences in the overall treatment population and transplantation strategies in each context.Patients and MethodsWe performed a retrospective analysis of consecutive patients aged 15 and older who underwent BMT regardless of diagnosis form 2015-2021 in two Institutions: 1) A private hematology practice treating insured patients in conventional BMT units similar to those in high-income countries (Private) and 2) A public academic institution where an outpatient transplant strategy is common in the context of significant limitations in access to quality supportive care and high-cost medications (Public). Both programs are led by the same team of hematologists following similar transplant strategies with the salient features being the frequent use of peripheral blood stem cells, chemotherapy-based conditioning regimens, and the preferred use of haploidentical donors vs. matched unrelated donors. We excluded second transplants from this analysis and patients who received them were censored at the time of infusion. We compared baseline characteristics, overall survival (OS), event free survival (EFS), non-relapse mortality (NRM), and the incidence of GVHD in the two different treatment systems.ResultsA total of 219 patients underwent BMT from January 2015 to June 2021, n=166 (76%) were performed in the Public setting, and n=53 (24%) in the Private setting. Patients in the Private group were older, with a higher proportion of high/very high disease risk index (DRI), hematopoietic cell comorbidity index (HCT-CI) and more frequent use of myeloablative conditioning (Table 1). A similar proportion haploidentical donor grafts were performed (61 vs 57%) with a single matched unrelated donor transplant in the Private center. Median follow-up was 9.7 (0.2-71), and 10.3 (0.7-67.6) months, for Public and Private centers respectively (p=0.38). Median time to neutrophil and platelet engraftment were similar. Seventy patients (42%) in Public, and 15 patients (28%) in Private groups relapsed (p=0.049), with a median time to relapse of 17.5 vs. 47.6 months (p<0.017); there were no significant differences in non-relapse mortality at 2 years (27 vs. 18%) (Fig.1) and primary failure (9% vs 2%). Grade 2-4 aGVHD occurred in 31% patients in the Public setting vs. 19% in Private (p=0.08), without differences in grade 3-4 aGVHD (12% vs 8%). Moderate/severe cGVHD incidence was similar for both groups with (19 vs. 18%). Estimated 2-year EFS was 34% in Public vs. and 54% in Private (Fig. 2), with a median EFS of 8.8 vs 25.7 months (p= 0.024). There were no statistically significant differences in OS (p=0.65), with estimated 2-year OS of 51% for Public and 68% for Privately treated patients, and a median OS of 21.1 months vs. not reached (Fig. 3). When stratified by DRI, patients with Public BMT and a high/very high DRI had a median OS of only 9.7 months vs. not reached for the Private group (Fig. 4). Patients with high/very high DRI in the Private setting had similar outcomes to those with low/intermediate disease in the Public group with the best outcomes achieved by patients with low/intermediate disease treated privately.ConclusionPatients who undergo BMT in the public health system are at risk for significantly worse outcomes when compared to patients cared for in private systems even if a similar strategy is followed and are led by the same team. [Display omitted] DisclosuresGonzález López: AMGEN: Honoraria; JANSSEN: Honoraria. Gomez-Almaguer: Janssen: Honoraria, Speakers Bureau; Takeda: Honoraria, Speakers Bureau; Bristol-Myers-Squibb: Honoraria, Speakers Bureau; Roche: Honoraria, Speakers Bureau. Gomez-De Leon: ASH: Research Funding; Abbvie: Honoraria; Sanofi: Honoraria; Novartis: Honoraria.

Erythroid Chimerism Measurement As a Predictive Tool of Bone Marrow Transplantation Outcome in Sickle Cell Disease

Allogeneic bone marrow transplantation (BMT) is the only curative treatment available for sickle cell disease (SCD). The effectiveness of the treatment is primarily assessed by the chimerism rate, which is the fraction of cells derived from the donor's hematopoietic stem cells (HSCs) relative to the recipient's cells. New less toxic conditioning agents, described as reduced, nonmyeloablative (as opposed to myeloablative conditioning agents) are now used for BMT with sibling donors. Moreover, most sibling donors are sickle cell trait carriers (heterozygous A/S genotype). In the absence of a test performed on erythroid cells, the rate of chimerism is currently assessed either on different subpopulations of myeloid cells by molecular biology techniques or on hemoglobin measurement by chromatography (HPLC). However, the percentage of myeloid chimerism is rarely representative of the percentage of erythroid chimerism in these patients and HPLC cannot allow analysis at the single cell level.An easy-to-implement method for accurate measurement of fetal hemoglobin (HbF) content in individualized red blood cells (RBCs) has recently been developed (Hebert, et. al, AJH 2020). This method relies on flow cytometry for cell individualization and intracellular labeling of HbF with a fluorescent monoclonal antibody for quantification. Based on the same approach, we propose the use of monoclonal antibodies directed against adult hemoglobins (HbA0 and HbA2) or hemoglobin S (HbS). This strategy allows to discriminate, from a blood sample, the different RBC subpopulations coming from either the recipient or the donor, or from the transfusion associated with this procedure, thus allowing us to precisely determine the rates of erythroid chimerism in patients.As a proof of concept, we show here the evaluation of the percentage of erythroid chimerism during a longitudinal follow-up performed on 3 patients with SCD (P1 - 18 months, P2 - 4 months, and P3 - 4 months). The 3 patients received allogeneic BMT from a sibling A/S donor following nonmyeloablative conditioning consisting of the administration of an anti-CD52 antibody associated with radiotherapy (300 cGy TBI - total-body irradiation). We observed for P1 and P2 donor erythroid chimerism >90% and >95%, 1 and 2 months after BMT, respectively (measured after exclusion of transfused RBCs). These results are in accordance with the presence of most RBCs HbA- and HbS-positive, as determined by flow cytometry. In addition, the analysis of the reticulocyte (retic) subpopulations (CD71-positive cells) of these patients provides precise information on the RBC production of the bone marrow following transplantation. Retic donor chimerism levels were >99% for both P1 and P2, 1 month after BMT procedure. P1 and P2 showed stable donor reticulocyte and donor erythroid chimerism levels during the follow-up. On the contrary, P3 presented a different response with 43%, 41% and 5% donor erythroid chimerism 1, 2 and 3 months after BMT, respectively. Retrospectively, this bad response in P3 would have been predicted by low donor reticulocyte chimerism levels (5%, 6% and 0.5% at 1, 2 and 3 months after BMT, respectively). Our erythroid specific chimerism results were also compared to the assessment of chimerism performed on either total white blood cells (WBCs) or lineage specific subpopulations after sorting. Over 18 months after BMT in P1, the correlation between retic chimerism and total WBCs or T lymphocytes (CD3-positive cells) chimerism were not statistically significant (P = 1 and P = 0.833, respectively - Spearman test) (Figure 1).This new tool to monitor BMT outcome to treat SCD shows great precision and potentially a stronger predictive capability than total WBCs or lineage specific chimerism assessed by molecular biology. Our results highlight the importance of analyzing reticulocyte population to better assess BMT efficacy and predict the outcome a few days after the transplant. Its use could also be considered in the context of the various gene therapy strategies currently under development using modified β-globin gene addition or direct mutation correction through CRISPR/Cas9 and homologous recombination system. [Display omitted] DisclosuresBencheikh: Innovhem: Current Employment. Bartolucci: Innovhem: Current holder of individual stocks in a privately-held company.

Polyvinyl alcohol hydrolysis rate and molecular weight influence human and murine HSC activity ex vivo

Ex vivo expansion of hematopoietic stem cells (HSCs) is one of the most promising strategies to increase the availability of transplantable HSCs and improve bone marrow transplantation outcomes. We recently demonstrated that mouse HSCs could be efficiently expanded in polyvinyl alcohol (PVA)-containing culture medium using only recombinant stem cell factor and thrombopoietin cytokines. However, the behavior of human HSCs in these simple PVA-based media was not fully elucidated. In this study, we analyzed the compatibility of PVA of different hydrolysis rates (HR) and molecular weights (MW) to support functional human and mouse HSCs ex vivo. Human and mouse HSCs proliferated more frequently in media containing PVA with lower HR than with higher HR, but both PVA types supported HSC multilineage reconstitution potential. Importantly, human HSCs cultured in PVA-containing media engrafted not only in irradiated recipients but also in non-irradiated recipients. Our results demonstrate that human HSCs can be maintained ex vivo using PVA-based culture systems and suggest approaches for future optimization of human HSC expansion.

Intestinal outcome of bone marrow transplantation for monogenic inflammatory bowel disease.

Some monogenic inflammatory bowel diseases (IBDs) are known to be refractory to conventional treatments. Although allogeneic hematopoietic stem-cell transplantation (allo-HSCT) has become a curative therapeutic option for certain monogenic IBDs, its effectiveness regarding endoscopic improvements has not been clarified. The clinical course and endoscopic findings of patients with monogenic IBDs who were treated with allo-HSCT between December 2017 and November 2018 at the National Center for Child Health and Development, were retrospectively reviewed. The clinical disease activity was assessed using the weighted Pediatric Crohn's Disease Activity Index (wPCDAI) and the endoscopic finding was evaluated using the Simple Endoscopic Score for Crohn's Disease (SES-CD). Clinical remission was defined as a wPCDAI <10 and endoscopic remission was defined as an SES-CD of 2 or less. Four patients with severe monogenic IBDs, including three with X-linked inhibitors of apoptosis protein (XIAP) deficiency and one with interleukin-10 signaling defect, were treated with allo-HSCT with reduced-intensity conditioning. In four patients, the maximum scores of wPCDAI and SES-CD before allo-HSCT ranged from 67.5 to 120 and 20 to 34, respectively. After allo-HSCT, all four patients showed a significant improvement in intestinal inflammation and achieved both clinical and endoscopic remission. Although patients with XIAP deficiency presented with post-transplant hemophagocytic lymphohistiocytosis and a relatively late engraftment, all patients achieved prolonged clinical remission, and IBD medications were successfully discontinued in all patients. Allo-HSCT for monogenic IBD resulted in complete clinical resolution with endoscopically confirmed mucosal healing.

Tonsil-derived mesenchymal stem cells enhance allogeneic bone marrow engraftment via collagen IV degradation

BackgroundCo-transplantation of bone marrow cells (BMCs) and mesenchymal stem cells (MSCs) is used as a strategy to improve the outcomes of bone marrow transplantation. Tonsil-derived MSCs (TMSCs) are a promising source of MSCs for co-transplantation. Previous studies have shown that TMSCs or conditioned media from TMSCs (TMSC-CM) enhance BMC engraftment. However, the factors in TMSCs that promote better engraftment have not yet been identified.MethodsMice were subjected to a myeloablative regimen of busulfan and cyclophosphamide, and the mRNA expression in the bone marrow was analyzed using an extracellular matrix (ECM) and adhesion molecule-targeted polymerase chain reaction (PCR) array. Nano-liquid chromatography with tandem mass spectrometry, real-time quantitative PCR, western blots, and enzyme-linked immunosorbent assays were used to compare the expression levels of metalloproteinase 3 (MMP3) in MSCs derived from various tissues, including the tonsils, bone marrow, adipose tissue, and umbilical cord. Recipient mice were conditioned with busulfan and cyclophosphamide, and BMCs, either as a sole population or with control or MMP3-knockdown TMSCs, were co-transplanted into these mice. The effects of TMSC-expressed MMP3 were investigated. Additionally, Enzchek collagenase and Transwell migration assays were used to confirm that the collagenase activity of TMSC-expressed MMP3 enhanced BMC migration.ResultsMice subjected to the myeloablative regimen exhibited increased mRNA expression of collagen type IV alpha 1/2 (Col4a1 and Col4a2). Among the various extracellular matrix-modulating proteins secreted by TMSCs, MMP3 was expressed at higher levels in TMSCs than in other MSCs. Mice co-transplanted with BMCs and control TMSCs exhibited a higher survival rate, weight recovery, and bone marrow cellularity compared with mice co-transplanted with BMCs and MMP3-knockdown TMSCs. Control TMSC-CM possessed higher collagenase activity against collagen IV than MMP3-knockdown TMSC-CM. TMSC-CM also accelerated BMC migration by degrading collagen IV in vitro.ConclusionsCollectively, these results indicate that TMSCs enhance BMC engraftment by the secretion of MMP3 for the modulation of the bone marrow extracellular matrix.

The Human Bone Marrow Is Host to the DNAs of Several Viruses.

The long-term impact of viruses residing in the human bone marrow (BM) remains unexplored. However, chronic inflammatory processes driven by single or multiple viruses could significantly alter hematopoiesis and immune function. We performed a systematic analysis of the DNAs of 38 viruses in the BM. We detected, by quantitative PCRs and next-generation sequencing, viral DNA in 88.9% of the samples, up to five viruses in one individual. Included were, among others, several herpesviruses, hepatitis B virus, Merkel cell polyomavirus and, unprecedentedly, human papillomavirus 31. Given the reactivation and/or oncogenic potential of these viruses, their repercussion on hematopoietic and malignant disorders calls for careful examination. Furthermore, the implications of persistent infections on the engraftment, regenerative capacity, and outcomes of bone marrow transplantation deserve in-depth evaluation.