Unrelated Donor Bone Marrow Research Articles

Uncommon HLA Alleles Observed in a Population of Istanbul Province

Objective: New polymorphisms are formed in human leucocyte antigen (HLA) genes with point mutations, gene conversions, and duplication, and the diversity continues to increase. Various new HLA alleles have significant roles in transplantation, and epidemiologic and population studies. The aim of our study was to determine the status of HLA alleles in the Turkish population, which is uncommon, well-defined, and non-defined in the world population according to the international ImMunoGeneTics information system® (IMGT) database. Methods: We performed HLA-A, -B, -C, -DQB1, and DRB1 loci at the four-field resolution level, using Sanger- sequence-based typing (SBT) for 5592 healthy, unrelated bone marrow donor volunteers from Istanbul Province. The uncommon alleles were also confirmed using high-throughput next-generation sequencing (NGS). Results: Uncommon alleles were determined at five loci as follows: HLA-A*01:155, 02:66, 02:90, 02:110, 02:343, 03:82, 24:28, 24:146, 24:276, 24:356, 31:23,33:33, 68:38; HLA-B *07:240, 18:19, 35:193, 40:303, 51:69, 51:169; HLA-C*04:39, 06:40, 07:93, 12:149, 15:73; HLA-DRB1*11:149, 13:14:02 and HLA-DQB1*03:27. All alleles were arranged according to the common and well‐documented (CWD) 3.0.0 catalog. Conclusion: This is the first study to show uncommon alleles in our population. These reported data increase the knowledge of HLA polymorphisms in the Turkish population and provide a basis for further studies in population genetics. This information may also be useful in determining whether a matched, unrelated donor is unlikely to be found so that a mismatch strategy, an extended family search, or alternate therapy, can be pursued, thus saving time and cost for patients.

Allogeneic Hematopoietic cell Transplantation Using Alemtuzumab in Asian Patients with Inborn Errors of Immunity.

Alemtuzumab is used with reduced-toxicity conditioning (RTC) in allogeneic hematopoietic cell transplantation (HCT), demonstrating efficacy and feasibility for patients with inborn errors of immunity (IEI) in Western countries; however, the clinical experience in Asian patients with IEI is limited. We retrospectively analyzed patients with IEI who underwent the first allogeneic HCT with alemtuzumab combined with RTC regimens in Japan. A total of 19 patients were included and followed up for a median of 18 months. The donors were haploidentical parents (n = 10), matched siblings (n = 2), and unrelated bone marrow donors (n = 7). Most patients received RTC regimens containing fludarabine and busulfan and were treated with 0.8mg/kg alemtuzumab with intermediate timing. Eighteen patients survived and achieved stable engraftment, and no grade 3-4 acute graft-versus-host disease was observed. Viral infections were observed in 11 patients (58%) and 6 of them presented symptomatic. The median CD4+ T cell count was low at 6 months (241/µL) but improved at 1 year (577/µL) after HCT. Whole blood cells continued to exhibit > 80% donor type in most cases; however, 3/10 patients exhibited poor donor chimerism only among T cells and also showed undetectable levels of T-cell receptor recombination excision circles (TRECs) at 1 year post-HCT. This study demonstrated the efficacy and safety of alemtuzumab; however, patients frequently developed viral infections and slow reconstitution or low donor chimerism in T cells, emphasizing the importance of monitoring viral status and T-cell-specific chimerism. (238 < 250 words).

Association of HLA-C*07:359 with HLA-A, -B, and -DRB1 alleles in Taiwanese.

It is thought that Taiwanese indigenous people were the "first people" to populate Taiwan (Formosa) having been there for over 5000 years, preceding the Dutch colonization (from 1624 to 1662) and Spanish colonization (from 1626 to 1642). Taiwan's indigenes, represented by Austronesian language speakers, currently constitute approximately 2% of the total population in Taiwan. It is unknown whether they evolved from Taiwan's Paleolithic or Neolithic cultures, arrived during or after the Neolithic period from China or Southeast Asia or both. HLA studies on the Taiwanese indigenous population have found several intriguing genetic information showing one or two relatively frequently observed alleles and a small number of relatively less frequently observed ones. We report here a relatively frequently observed HLA-C*07:359 allele in the Taiwanese indigenous population, its linkage with HLA-B*39:01, and its probable associated HLA haplotype in two Taiwanese indigenous families. HLA-C*07:359 is a rarely observed allele in the HLA-C locus in the world populations. The objective of this study is to report the allele HLA-C*07:359 that is more frequently found in the Taiwanese population, especially in the Taiwanese indigenous people, to demonstrate that it has a close linkage with HLA-B*39:01 allele in the HLA-B locus and to show the plausible deduced HLA-A-C-B-DRB1-DQB1 haplotypes in association with HLA-C*07:359 in two families of Taiwanese indigenous unrelated individuals. The samples were peripheral whole blood, with dipotassium ethylenediaminetetraacetic acid and/or acid citrate dextrose anticoagulation additives. The sequence-based typing method was employed to confirm the low incidence of the allele of HLA-C*07:359 observed in Taiwanese. Polymerase chain reaction was carried out to amplify exons 2, 3, and 4 of the HLA-A,-B,-C,-DRB1 and-DQB1 loci with group-specific primer sets. Amplicons were sequenced using the BigDye Terminator Cycle Sequencing Ready Reaction Kit in both directions according to the manufacturer's protocol. C*07:359 is an uncommon allele in the HLA-C locus in the world general population, according to our literature review. However, in this study, it is observed in the general Taiwanese population (frequency 0.41%), especially in the Taiwanese indigenous people at a frequency of 0.23%. In addition, we deduced two probable HLA haplotypes in association with C*07:359 in two indigenous families: A*24:02-C*07:359-B*39:01-DRB1*04:36 and A*24:02-C*07:359-B*39:01-DRB1*04:04. The two deduced HLA haplotypes associated with the uncommon C*07:359 allele that we report here are valuable for HLA tissue typing laboratories for reference purposes and for stem cell transplantation donor search coordinators to determine the likelihood of finding compatible donors in unrelated bone marrow donor registries for patients bearing the uncommon HLA allele. Since C*07:359 was found mostly in the Taiwanese indigenous population, we think the allele and its haplotypes we report here are important in population and anthropological studies.

The novel HLA-C*12:392 allele was identified in four unrelated bone marrow donors.

One nucleotide substitution in codon 83 of HLA-С*12:02:02:01 results in the novel allele, HLA-C*12:392.

Alemtuzumab, Dual Graft-versus-Host Disease Prophylaxis, and Lower CD3+ T Cell Doses Equalize Rates of Acute and Chronic Graft-versus-Host Disease in Pediatric Patients Receiving Allogeneic Hematopoietic Stem Cell Transplantation with Matched Unrelated Donor Peripheral Blood Stem Cells or Bone Marrow Grafts

Data comparing hematopoietic stem cell transplantation (HSCT) using bone marrow (BM) or peripheral blood stem cell (PBSC) grafts in children after alemtuzumab-based conditioning are lacking. We investigated whether in vivo T cell depletion using alemtuzumab could reduce the risk of severe acute graft-versus-host disease (aGVHD) and chronic GVHD (cGVHD) after HSCT with matched unrelated donor (MUD) BM or PBSCs. This retrospective multicenter study included 397 children (BM group, n = 202; PBSC group, n = 195) who underwent first MUD HSCT at 9 pediatric centers in the United Kingdom between 2015 and 2019. The median age at transplantation was 7.0 years (range, .1 to 19.3 years), and the median duration of follow-up was 3.1 years (range, .3 to 7.5 years). The 3-year overall survival was 81% for the entire cohort (BM group, 80%; PBSC group, 81%). The incidence of grade II-IV aGVHD was significantly higher in the PBSC group (31%) compared to the BM group (31% versus 19%; P = .003), with no difference in the incidence of grade III-IV aGVHD (BM, 7%; PBSC, 12%; P = .17). CD3+ T cell dose >5 × 108/kg and the use of PBSCs were independent predictors of grade II-IV aGVHD. When considering CD3+ T cell dose and GVHD prophylaxis, PBSC transplantation with a calcineurin inhibitor (CNI) and mycophenolate mofetil (MMF) and a CD3+ T cell dose ≤5 × 108/kg had a comparable grade II-IV aGVHD to BM transplantation plus a CNI (20% versus 18%; P = .52). PBSC transplantation was associated with a lower incidence of cGVHD compared to BM transplantation (6% versus 11%; P = .03). Within the limits of this study, we identified a potential strategy to reduce the risk of severe GVHD in pediatric PBSC recipients that includes a combination of in vivo T cell depletion using alemtuzumab and dual GVHD prophylaxis (with a CNI and MMF) and limiting the CD3+ T cell dose to ≤5 × 108/kg.

Hematopoietic Stem Cell Transplantation for Fanconi Anemia: Outcome and Prognostic Factors for Survival and Subsequent Neoplasms

Introduction Fanconi anemia (FA) is an inherited disorder of impaired DNA repair, associated with progressive bone marrow failure (BMF), leukemia, and cancer predisposition. Hematopoietic cell transplantation (HCT) may be curative for FA associated BMF, and changes in HCT practice over the past decades may have impacted outcomes in FA patients. Methods We used CIBMTR database to perform a retrospective cohort study of children and adults undergoing first allogeneic HCT for FA BMF (patients with MDS/AML excluded) from 2000-2018. We hypothesized that contemporary HCT approaches, including decreases in radiation and increase in use of serotherapy and ex-vivo graft manipulation, would associate with improved outcomes in FA patients receiving HCT. The primary endpoints were overall survival (OS) and early subsequent malignant neoplasm (SMN; by 5 yr post-HCT). Secondary endpoints included event free survival (EFS; death, SMN or graft failure), treatment related mortality (TRM), graft failure, and acute (grade II-IV) and cGVHD. Univariate and multivariate Cox regression models were created to evaluate the impact of conditioning, donor and graft source, treatment era, and patient characteristics on outcomes, adjusting for other significant covariates. Due to confounding factors, we were not able to create a single multivariate model that included radiation, ex-vivo graft manipulation, and donor and graft source; therefore, we performed subgroup univariate analysis for patients receiving unrelated donor bone marrow (BM) or peripheral blood stem cell (PBSC) HCT. Results We analyzed 652 HCTs from 2000-07 (n=353) and 2008-18 (n=293). 223 (35%) patients had a matched related donor (MRD) with any graft type, 244 (38%) had an unrelated donor BM or PBSC grafts, 123 (19%) had an unrelated cord blood graft, and 44 (7%) had a related haploidentical donor with any graft type. Between the two treatment eras, 5-yr OS has dramatically improved (78.8% 2008-18 v 62.5% 2000-07, p&amp;lt;0.01; figure 1) driven by lower 5-yr TRM (15.2% v 25.5%, p=0.004) and lower rates of primary (7.9% v 12.9% at 42 days, p=0.038) and secondary (4.1% v 8.9% at 1 year, p=0.014) graft failure. No statistically significant change was observed for aGVHD, cGVHD, or early SMN. Assessing the entire cohort, use of total body irradiation (TBI) &amp;gt;300cGy was associated with lower OS (HR=1.75, 95% CI 1.24-2.47) and increased TRM (HR=3.36, 95% CI 1.54-7.34; figure 2) on multivariable analysis. For patients that received unrelated BM or PBSC HCT, use of TBI was associated with inferior OS survival (HR=1.93, 95% CI 1.18-3.17), EFS (HR=1.78, 95% CI 1.13-2.80), and TRM (HR=2.31, 95% CI 1.20-4.46) on univariate analysis. Among all patients, CD34 selection or T-cell depletion ( ex vivo depletion) were not associated with better OS (95% HR=1.014, 95% CI 0.84-1.56), EFS (HR=1.163, 95% CI 0.86-1.58), or TRM (HR=0.79, 95% CI 0.52-1.19) but was associated with less aGVHD (HR=0.35 95% CI 0.22-0.57) and lower but not-statistically significant cGVHD (HR=0.54, 95% CI 0.29-1.01) on multivariable analysis. Within the unrelated donor setting, univariate analysis showed reduced aGVHD (HR=0.39 95% CI 0.23-0.68) and cGVHD (HR=0.47 95% CI 0.28-0.79) but did not have a significant effect on OS, EFS, or TRM. Early SMN occurred in 3.2% of participants by 5-years post-HCT and was associated with cGVHD occurrence (HR=2.14, 95% CI 1.06-4.33) and older age (&amp;gt;18 v &amp;lt;9 years, HR=6.35, 95% CI 2.66-15.14), but not ex-vivo depletion or use of ATG or alemtuzumab. Early SMN risk was not impacted by the use of TBI (HR=1.60, 95% CI 0.77-3.34), but secondary analysis demonstrated a trend towards a relationship with doses &amp;gt;300 cGy (HR=2.24, 95% CI 0.99-5.08). Conclusions Outcomes for patients with FA BMF have improved significantly in more recent HCT eras, driven by reductions in TRM and graft failure risk. Although radiation doses &amp;gt;300cGy were associated with greater toxicity, doses of 100-300cGy were not associated with inferior outcomes in our cohort. Use of CD34 selection or T-cell depletion may reduce the risk of GVHD in transplant FA patients, but does not appear to impact 5-yr OS. Risk for early SMN development in FA patients receiving HCT associates with older age and cGVHD.

Prevalence and Outcome of Vexas Syndrome in Unrelated Hematopoietic Stem Cell Transplantation

Introduction Vacuoles, E1 enzyme, X-linked, Autoinflammatory, Somatic (VEXAS) syndrome is a newly identified monogenic disorder defined by somatic UBA1 mutations in hematopoietic progenitor cells, resulting in bone marrow failure (BMF) and systemic inflammation. This syndrome is frequently associated with myelodysplastic syndrome (MDS) or plasma cell dyscrasia. Although high-dose corticosteroids alleviate symptoms and cytopenia, allogeneic hematopoietic stem cell transplantation (HSCT) has been suggested as a potentially curative treatment. This study evaluated the safety and efficacy of unrelated HSCT for treating VEXAS syndrome and its prevalence in patients with hematologic diseases. Methods We conducted retrospective screening for UBA1 mutations in patients with BMF or plasma cell neoplasms who underwent their first HSCT from unrelated donors in Japan between 1995 and 2020. Pre-HSCT blood DNA samples and clinical data were collected from the Japanese Data Center for Hematopoietic Cell Transplantation. Multiplex real-time PCR was used to detect the most common pathogenic UBA1 mutations, including p.M41L (c.121A&amp;gt;C), p.M41V (c.121A&amp;gt;G), and p.M41T (c.122T&amp;gt;C), with detection limits of 0.13%, 0.5%, and 1%, respectively. The presence of these mutations was confirmed using Sanger sequencing and droplet digital PCR. In addition, we examined UBA1 mutations in 2 patients previously diagnosed with MDS but suspected of having VEXAS syndrome based on specific clinical features. Results Among 4340 patients with either BMF or plasma cell neoplasms who underwent unrelated HSCT, 1771 had available pre-HSCT blood DNA samples. The diagnoses of the 1771 patients included MDS (n = 1264), acquired aplastic anemia (n = 375), inherited BMF syndromes (n = 89), plasma cell neoplasms (n = 23), and other acquired BMF syndromes (n = 20). The median patient age at HSCT was 46 (range, 0-75; interquartile range, 20-59) years old, and the male-to-female ratio was 1.6:1. Pathogenic UBA1 mutations were detected in 2 of the 1771 patients (0.11%), both of whom were male MDS patients older than the median age (cases 1 and 2); thus, the prevalence of VEXAS syndrome within this patient subgroup was 0.36%. Both patients received chemotherapy before unrelated donor bone marrow transplantation (BMT), including induction chemotherapy and azacitidine, and achieved complete and partial responses, respectively. In addition, UBA1 mutations were detected in the 2 MDS patients at Kanazawa University Hospital (cases 3 and 4). Case 3 also underwent unrelated BMT after experiencing cytopenia relapse and inflammatory symptoms during corticosteroid reduction. The clinical characteristics and outcomes of the 4 patients with VEXAS syndrome are summarized in Table 1. Variant allele frequencies (VAFs) of UBA1 mutations ranged from 2% to 71%. The 3 patients who underwent unrelated BMT received fludarabine and busulfan-based conditioning regimens and graft-versus-graft disease (GVHD) prophylaxis with tacrolimus + methotrexate or cyclosporine + methotrexate. All 3 patients achieved neutrophil and platelet engraftment. One patient required systemic corticosteroids for acute skin GVHD. One patient died of idiopathic interstitial pneumonia 5 months after BMT. The other 2 patients were alive without relapse, chronic GVHD, or other complications at 18 and 28 months after BMT. Discussion Our results support the curative potential of allogeneic HSCT for VEXAS syndrome without excessive toxicity. The occurrence of transplant-related mortality due to idiopathic interstitial pneumonia may be related to the reported poor outcome of the p.M41V mutation following non-transplant therapy. The low prevalence of VEXAS syndrome in the current cohort may be due to the late age of onset and the rare progression to advanced MDS. The low VAFs of UBA1 mutations in the 2 patients who responded to chemotherapy before HSCT may indicate the efficacy of chemotherapy in selectively eradicating UBA1 mutant clones. Our rapid, sensitive, and cost-effective screening method for VEXAS syndrome can help optimize treatment strategies for this rare disease in future studies.

Allogeneic Hematopoietic Cell Transplantation for Juvenile Myelomonocytic Leukemia with a Busulfan, Fludarabine, and Melphalan Regimen: JPLSG JMML-11

Juvenile myelomonocytic leukemia (JMML), which is classified as a myelodysplastic/myeloproliferative neoplasm, is a rare hematologic malignancy of childhood. Most patients with JMML require allogeneic hematopoietic cell transplantation (HCT) as a curative therapy. A Japanese retrospective analysis demonstrated favorable outcomes for a busulfan (BU)+fludarabine (FLU)+melphalan (MEL) regimen, with an overall survival (OS) of 72% and an event-free survival (EFS) of 53%. To further validate the efficacy and safety of this regimen, the Japan Pediatric Leukemia/Lymphoma Study Group (JPLSG) conducted a nationwide prospective study, JMML-11. Between July 2011 and June 2017, 28 patients with newly diagnosed JMML were enrolled in JMML11. Low-dose chemotherapy for tumor control before HCT was recommended, and patients treated with AML-type chemotherapy and azacitidine were excluded. The conditioning regimen comprised i.v. BU, 16 doses administered every 6 h, with dose adjustment based on pharmacokinetic (PK) studies on days -11 to -8; FLU, 30 mg/m2/day or 1 mg/kg/day for patients <10 kg or age <1 year on days -7 to -4; and MEL, 90 mg/m2/day or 3 mg/kg/day for patients <10 kg or <1 year on days -3 to -2. The donor was selected by the physician in charge. A family donor was available for 7 patients (3 HLA-matched siblings, 3 HLA-1-antigen mismatched parents, and 1 haploidentical father). Overall, 21 patients received grafts from unrelated donors, including 8 HLA-matched donors and 13 HLA-mismatched donors. The graft source was related bone marrow (BM) for 7 patients, unrelated BM for 14 patients, and unrelated cord blood for 7 patients. Neutrophil engraftment was achieved in 21 of 28 patients (75%), with a median of 20.5 days (range, 11 to 39 days) after transplantation. The 3-year OS, 3-year EFS, 3-year relapse rate, and 3-year transplantation-related mortality were 63% (95% confidence interval [CI], 42% to 78%), 52% (95% CI, 32% to 69%), 18% (95% CI, 6% to 34%), and 21% (95% CI, 9% to 38%), respectively. WBC count before the conditioning regimen (≥7.0×109/L) was significantly associated with inferior EFS and OS. Body surface area ≥.5 m2, spleen size <4 cm before conditioning, and HLA-matched unrelated BM donors were significantly associated with better OS. Adverse effects related to the conditioning regimen included febrile neutropenia (86%), diarrhea (39%), hypoxemia (21%), and mucositis (18%). BU-associated toxicity, including sinusoidal obstruction syndrome (SOS) and thrombotic microangiopathy (TMA), occurred in 7 patients (25%; SOS, n=6; TMA, n=2). Retrospective analysis of PK data after the first BU dose in 23 patients, including 6 with SOS and 17 without SOS, did not show significant differences between groups. The JMML-11 study confirms the positive results of previous retrospective analyses. BU+FLU+MEL might become a standard conditioning regimen for patients with JMML.

Genotypic Frequencies of Mutations Associated with Alpha-1 Antitrypsin Deficiency in Unrelated Bone Marrow Donors from the Murcia Region Donor Registry in the Southeast of Spain.

Alpha-1 antitrypsin (AAT1) deficiency (AAT1D) is an inherited disease with an increased risk of chronic obstructive pulmonary disease (COPD), liver disease, and skin and blood vessel problems. AAT1D is caused by mutations in the SERPINE1 gene (Serine Protease Inhibitor, group A, member 1). Numerous variants of this gene, the Pi system, have been identified. The most frequent allelic variants are Pi*M, Pi*S, and Pi*Z. The development of COPD requires both a genetic predisposition and the contribution of an environmental factor, smoking being the most important. Studies on this deficiency worldwide are very scarce, and it is currently considered a rare disease because it is underdiagnosed. The aim of this study was to analyze the genotypic frequencies of mutations associated with AAT1 deficiency in unrelated bone marrow donors from the donor registry of the Region of Murcia in southeastern Spain due to the high risk of presenting with different pathologies and underdiagnosis in the population. A total of 112 DNA-healthy voluntary unrelated bone marrow donors from different parts of the Region of Murcia were analyzed retrospectively. AAT1 deficiency patient testing involved an automated biochemical screening routine. The three main variants, Pi*M, Pi*Z, and Pi*S, were analyzed in the SERPINE1 gene. Our results showed a frequency of 3.12% of the Pi*Z (K342) mutation in over 224 alleles tested in the healthy population. The frequency of Pi*S (V264) was 11.1%. The frequency of the haplotype with the most dangerous mutation, EK342 EE264, was 4.46%, and the frequency of EK342 EV264 was 1.78% in the healthy population. Frequencies of other EE342 EV264-mutated haplotypes accounted for 18.7%. As for the EE342 VV264 haplotype, 0.89% of the total healthy population presented heterozygous for the EV264 mutation and one individual presented homozygous for the VV264 mutation. In conclusion, the frequencies of Pi mutations in the healthy population of the Region of Murcia were not remarkably different from the few studies reported in Spain. The genotype and haplotype frequencies followed the usual pattern. Health authorities should be aware of this high prevalence of the Pi*S allelic variant and pathological genotypes such as Pi*MZ and Pi*SZ in the healthy population if they consider screening the smoking population.

Nomenclature for factors of the HLA system, update January, February, and March 2023.

Nomenclature for factors of the HLA system, update January, February, and March 2023.

Efficacy and safety of allogeneic hematopoietic cell transplantation in acute myeloid leukemia patients aged &gt; 65 years with unfavorable cytogenetics

Unrelated donor bone marrow transplantation (UR-BMT), unrelated donor cord blood stem cell transplantation (UR-CBT), and haploidentical peripheral blood stem cell transplantation (Haplo-PBSCT) are the main alternative stem cell sources for allogeneic hematopoietic cell transplantation (HCT) in Japan. The present study aimed to identify factors associated with the outcomes of UR-BMT, UR-CBT, and Haplo-PBSCT in older patients with acute myeloid leukemia (AML) and intermediate- or poor-risk cytogenetics to improve the clinical efficacy and safety of allogeneic HCT. We retrospectively analyzed data for 448 AML patients aged > 65 years who received UR-BMT (n = 102), UR-CBT (n = 250), or Haplo-PBSCT (n = 96) between 2014 and 2020. Overall survival (OS) in the UR-BMT group was superior (P = 0.033) to that in the other groups. However, all patients without complete remission (non-CR) who had Karnofsky performance status (KPS) < 80 at HCT and poor-risk cytogenetics died within 1 year after HCT. Multivariate Cox regression analysis identified KPS <80 at HCT and poor-risk cytogenetics as independent predictors of worse OS in non-CR patients. KPS < 80 may be an alternative indicator for non-CR AML patients with poor-risk cytogenetics during the selection of HCT, alternative treatments, or best supportive therapy, and the optimal KPS is important for the success of HCT.

Detection and characterization of the novel HLA-DPA1*02:66:02N allele, with a premature stop codon in exon 2.

The failure to identify HLA null alleles in bone marrow transplantation could be life-threatening because this could result in an HLA mismatch with the ability to trigger the graft-vs-host disease (GVHD) and to reduce patient's survival. In this report we describe the identification and characterization of the novel HLA-DPA1*02:66:02N allele with a non-sense codon in exon 2. This new allele was discovered in two unrelated bone marrow donors during routine HLA-typing using next-generation sequencing (NGS). DPA1*02:66:02N is homologous to DPA1*02:01:01:03 with a single nucleotide difference in exon 2, codon 50, where the replacement of C located at genomic position 3825 by T, causes the formation of a premature stop codon (TGA), resulting in a null allele. This description illustrates the benefits of HLA typing by NGS since it permits to reduce ambiguities, identify new alleles, analyze multiple HLA loci and improve transplantation outcome.

A review of HLA matching to improve clinical outcome in bone marrow transplants and the role of haplotyping in unrelated donor transplants

When the first allogeneic bone marrow transplants in humans was attempted, little was known of the major histocompatibility complex (HLA) and its role in ensuring success of the graft. Therefore, many of the early attempts resulted in failure. During the 1970s just a small number of alleles were detected serologically and the mixed lymphocyte reaction (MLR) served as a surrogate for class II matching, the DR series of alleles not being formally serologically defined until 1978. Nevertheless, selection of nuclear family members who were two HLA haplotype matched became possible using serology and the MLR and transplant successes were reported. The breakthrough in technology was undoubtedly the introduction of DNA techniques, particularly DNA sequencing. This enabled fully matched unrelated individuals to be found as potential bone marrow donors. Genetic phasing or haplotyping of unrelated donors who are found on bone marrow registries has remained elusive until recently. The development of individual chromosome sequencing will permit haplotype matching in the absence of family studies and will expedite studies aimed at identifying other genes which are important matching factors. AIM: The aim of this review is to outline the technological advances that have been made in HLA genotyping over the last 50 years. The progress from serology to DNA sequence as part of the matching algorithm is one of the reasons clinical results have improved so much during this period. This review outlines the DNA techniques that have been utilised for this purpose, concluding with a discussion of the value of haplotype matching in unrelated donor transplants, where the donor has been sourced from a bone marrow registry. Haplotype matching represents one of the last remaining hurdles to be overcome in unrelated donor bone marrow transplants.

Three-Year Outcomes in Recipients of Mismatched Unrelated Bone Marrow Donor Transplants Using Post-Transplantation Cyclophosphamide: Follow-Up from a National Marrow Donor Program-Sponsored Prospective Clinical Trial

The use of post-transplantation cyclophosphamide (PTCy) as graft-versus-host disease (GVHD) prophylaxis has resulted in reductions in GVHD and improved outcomes in allogeneic hematopoietic cell transplantation (HCT) using HLA-mismatched related donors. We report the 3-year outcomes of the first multicenter prospective clinical trial using PTCy in the setting of mismatched unrelated donor (MMUD) bone marrow HCT. The study enrolled 80 patients, treated with either myeloablative conditioning (MAC; n=40) or reduced-intensity conditioning (RIC; n=40), with the primary endpoint of 1-year overall survival (OS). The median follow-up for this study was 34 months (range, 12 to 46 months) in the RIC group and 36 months (range, 18 to 49 months) in the MAC group. Three-year OS and nonrelapse mortality were 70% and 15%, respectively, in the RIC group and 62% and 10% in the MAC group. No GVHD was reported after 1 year. The incidence of relapse was 29% in the RIC group and 51% in the MAC group. OS did not differ based on HLA match grade (63% in the 7/8 strata and 71% in the 4 to 6/8 strata). These encouraging outcomes, which were sustained for 3 years post-HCT, support the continued exploration of MMUD HCT using a PTCy platform. Important future areas to address include relapse reduction and furthering our understanding of optimal donor selection based on HLA and non-HLA factors.

The novel HLA-C*04:441:01:02 allele was identified in three unrelated bone marrow donors.

The novel HLA-C*04:441:01:02 allele was characterized using next generation sequencing technology.

A survey on preoperative autologous blood donation policy in bone marrow stem cell donors in Italy.

The high safety of homologous blood components, together with the introduction of the Patient Blood Management strategy, has led to the progressive abandonment of preoperative autologous blood donation (PAD) in surgery. Furthermore, recent scientific publications provide evidence about the non-usefulness of PAD in the collection of hematopoietic stem cells (HSC) from bone marrow (BM), also in consideration of harvest procedure safety. Nevertheless, no conclusive studies have been published yet. Blood Establishments (BE) and Bone Marrow Collection Centers (BMCC) participated in a specific qualitative survey proposed by Italian National Blood and Transplant centers with the support of the relevant Italian Scientific Societies. The survey aimed at evaluating the policy adopted for PAD in related and unrelated adult HSC donors in Italy during the period 2018-2020. Forty-one BE corresponding to 37 BMCC filled in the questionnaire. Of 830 BM donors, 661 (80%) underwent 1063 PAD (mean 1.6 PAD/donor). The remaining 169 donors (20%) underwent BM harvest without PAD. No serious adverse events were reported for either donor group. In the case of ineligibility of donors for the PAD program, due to low hemoglobin values, 7/10 centers shifted donors to peripheral blood stem cell collection and three centers chose a different donor. Remarkably, only 51% of the PAD units requested were eventually transfused during the BM harvest process. Finally, the iron support policy among centers was heterogeneous. The results of this survey show that PAD is heterogeneously applied in Italian BMCC, as in other countries. However, all BMCC except two are willing to adopt a Patient Blood Management strategy as an alternative approach to adult related and unrelated BM donor harvests.

Identification of the novel HLA-B*07:458 allele, detected in two unrelated bone marrow donors.

HLA-B*07:458 has one non-synonymous nucleotide compared with HLA-B*07:02:01 in codon-21.

Recruitment of Marrow Donors in the Non-governmental Sector of the Republic of North Macedonia, 2022

BACKGROUND: Bone marrow transplantation is a well-established lifesaving treatment procedure for many patients with various diseases, such as blood cell cancers (leukemia), some severe hematological diseases (aplastic anemia) and some rare diseases, as well as immune deficiency disorders. Allogeneic hematopoietic stem cell transplantation (HSCT) represents a potentially curative therapy for various hematologic disorders and even the only curative treatment for some diseases (e.g., myelodysplastic syndromes). As an alternative donor strategy, in patients lacking a human leukocyte antigen-matched donor or haploidentical donor, mismatched unrelated donors offer the option of using HSCT in this group of patients. Since the establishment of the first registry in the world for recruitment of unrelated bone marrow donors, the number of newly formed registries has increased over the years. In our country, in addition to the government-funded registry, housed at the Institute of Immunobiology and Human Genetics, Faculty of Medicine in Skopje, an additional non-governmental registry was established, Scientific Foundation SPIROSKI - Marrow Donor Registry (MK-SFSMDR). This will contribute to the increase of the number of newly registered donors in the Republic of North Macedonia and will be a great benefit for the patients searching for a bone marrow transplantation. AIM: We aimed to present the establishment of the first MDR in the non-governmental sector of the Republic of North Macedonia and the recruitment strategies of new donors. METHODS: The core activity was the creation, realization, and implementation of the Marrow Donors Recruitment Program. The constructed program for increasing the awareness of the population about HSCT and the importance of becoming a marrow donor encompasses the following activities: (1) creation of a web portal for bone MDR in English, Macedonian, and Albanian language; (2) application of standards from the World Marrow Donor Association for educational material preparation and informed consent creation; (3) composing printed and electronic materials for recruitment activities; (4) organization of public debates for marrow donation in governmental and non-governmental sectors; (5) inclusion of students and other non-governmental organizations for recruitment and registration of marrow donors; and (6) submission the obtained results from Macedonian donors and the donors from other communities (Albanians, Roma, Serbs, Turks, Vlachs, and others) in the database for marrow donors. RESULTS: In 2022, 988 submissions from marrow donors were registered, from which 678 (68.62%) were females and 310 (31.37%) were males. The biggest number of submissions for marrow donors are in the age group of 26–40 years (567 or 57.38%), followed by the age group of 18–25 (270 or 27.32%) and the age group of 41–55 (151 or 15.28%). The most efficient method for new submissions was through social networks, 538 or 54.45% out of 988 registered donors. The second was through word of mouth (200 or 20.24%), the third was onsite marrow donor recruitment actions (145 or 14.68%), followed by information from the websites (19 or 1.92%), and other (86 or 8.70%). From the total Macedonian population (2,062,044), we were able to recruit 988 marrow donors, which is 0.048% of the population. The biggest recruitment percentage was in the Skopje region (0.067%), followed by the Pelagonia region (0.059%) and the Southwest region (0.043%). The lowest participation was in the Polog region (0.011%). The biggest percentage of recruited marrow donors (988) were Macedonians (94.13%), and the rest of them were from the minorities: Albanians (1.92), Turks (0.71%), Roma (0.1%), Croats (0.2%), Bosniaks (0.51%), Vlachs (0.81%), Serbs (1.21%), Macedonian Muslims (0%), and others (0.4%). CONCLUSION: With the establishment of the SFS-MDR (MK-SFSMDR), one of the goals of SFS was achieved, which, together with the publication of scientific journals, enables and contributes to the scientific research in the field of the main tissue-compatible complex and transplantation of cells, tissues, and organs. The results will be used by patients who need bone marrow treatment in our country, in neighboring countries, and around the world.

Radiation Sparing Reduced Intensity Conditioning for Allogeneic Cell Transplantation in Patients with Dyskeratosis Congenita Comparable to Standard of Care: A Phase III Single Institution Trial

Introduction Dyskeratosis congenita (DKC) is a rare form of inherited multiorgan disease caused by mutations affecting telomere maintenance. Complications include bone marrow failure (BMF), pulmonary and liver fibrosis, and predisposition to malignancy. Allogeneic hematopoietic cell therapy (alloHCT) can rescue affected individuals from hematologic complications. However, toxicity from alkylator and radiation exposure can be lethal or potentially expedite DKC-specific organ complications. In this single institution phase III clinical trial, we aimed to demonstrate non-inferior short-term alloHCT outcomes with a radiation sparing reduced intensity conditioning (RIC) regimen while monitoring for long-term reductions in systemic toxicity. Methods We compared two consecutive cohort of patients with DKC requiring alloHCT for BMF or myelodysplastic syndrome (MDS). Both received RIC with fludarabine 200 mg/m2, cyclophosphamide 50 mg/kg, and alemtuzumab 1 mg/kg. The earlier cohort (4/2006-8/2012) additionally received 200 cGy total body irradiation ("TBI"), while the latter (11/2015-2/2021) "Non-TBI" cohort did not. We restricted our analysis to bone marrow (BM) and peripheral blood stem cell (PBSC) recipients. AlloHCT milestones including neutrophil engraftment, graft failure, disease progression, acute and chronic graft-versus host disease (GvHD), serious infections, and overall survival were prospectively recorded. The cumulative incidence of time-dependent events was compared between cohorts using the Fine-Gray test with death as a competing risk. Infection density over the first 100 days after alloHCT was calculated using the Mantel-Haenszel chi-square test for incidence density. Estimates of overall survival were calculated by Kaplan-Meier curves. Results The TBI cohort included 12 patients aged 2.2-52.2 years (median 20.5), 11 with SAA and 1 with MDS. Stem cell sources were unrelated donor BM in 8, sibling donor PBSCs in 2, and sibling BM in 1, with a median follow-up of 1531 days (range 83-3960). The comparable Non-TBI group included 10 patients aged 1.7-65.9 years (median 9.8), 8 with SAA and 2 with MDS/RAEB. Stem cell donor sources were unrelated donor BM in 8 and sibling BM in 2, with a median follow-up of 870 days (range 303-2420). Neutrophil engraftment occurred at a median of 13 days in the TBI cohort and 8.5 days in the Non-TBI cohort (p<0.001; Fig 1a). Cumulative incidence of acute GvHD by day 100 for the TBI cohort was 0.08 (95% CI <0.01, 0.32) compared to 0.1 (95% CI <0.01, 0.37) for the Non-TBI cohort (p=0.74). Cumulative incidence of chronic GvHD by 3 years for the TBI cohort was 0.17 (95% CI 0.02, 0.43) compared to 0.1 (95% CI <0.01, 0.37) for the Non-TBI cohort (p=0.7). With regard to serious infections the first 100 days after alloHCT, there were no fungal infections in either group and infection density of viral infections, bacteremia, and pneumonia episodes was equivalent between cohorts. In the TBI cohort, one case was recorded as a late graft failure at day +1371, however upon retrospective review, this patient maintained 10% BM cellularity with 100% donor chimerism and experienced cytopenias and death likely resulting from Epstein-Barr viremia and possible post-transplant lymphoproliferative disease. In the Non-TBI cohort, one patient experienced non-neutropenic graft failure/rejection at day +28. A second Non-TBI patient demonstrated graft failure in the form of MDS recurrence at day +359. There was no statistically significant difference in overall survival between the two groups (p=0.4; Fig 1b, Table 1), with the 5.5 years median survival of the TBI cohort and the median survival not achieved for the Non-TBI cohort. Conclusions A successful alloHCT conditioning regimen for DKC must provide myeloablation to allow for sustained donor engraftment and immunosuppression to decrease GvHD risk, while limiting an unfavorable toxicity profile to which DKC patients are more susceptible. Elimination of TBI from the RIC regimen demonstrated equivalent overall survival, acute and chronic GvHD incidence, and day +100 infection density with an expedited neutrophil engraftment and no increase in graft failure risk. While there appears to be a trend toward less lethal DKC late effects like pulmonary failure and secondary malignancy, definitive assessment of such outcomes requires further longitudinal follow-up of both cohorts. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Identification of the novel HLA-A*11:01:115 allele, detected in two unrelated bone marrow donors.

HLA-A*11:01:115 has one synonymous nucleotide compared to HLA-A*11:01:01:01 in codon -5.