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Impact of recipient abh secretor status on outcome in minor aboincompatible hematopoietic stem cell transplantationImpact of recipient ABH secretor status on outcome
in minor ABO-incompatible hematopoietic
stem cell transplantation
Andreas Holbro,1,2 Martin Stern,1 Laura Infanti,1,2 Alix O'Meara,1 Beatrice Drexler,1 Beat M. Frey,3 Jean-Marie Tiercy,4 Jakob R. Passweg,1 Christoph Gassner,3 Andreas Buser,1,2 and Joerg-Peter Sigle2,5 BACKGROUND: The impact of ABO incompatibility on
hematopoietic stem cell transplantation (HSCT) tion (HSCT) is a potentially curative treatmentapproach for different malignant and nonma- outcome is controversial. As ABH substances are lignant diseases.1 Several given factors such expressed on tissues and secreted in body fluids, they as patient age, comorbidities, donor type, and donor- could drive an immune response in minor ABO- recipient sex combinations have been shown to affect sur- incompatible HSCT. The aim of the study was to inves- vival and major outcomes after HSCT.2 Scoring systems tigate the prognostic role of the recipients' ABH secretor integrate these pretransplant determinants into a global transplant risk assessment.3 STUDY DESIGN AND METHODS: Patients who under-
ABO incompatibility is not considered an obstacle for went minor ABO-incompatible HSCT were included.
HSCT and occurs in approximately 30% to 50% of trans- Secretor status was determined either serologically or plants.4 Different types of donor-recipient ABO incompat- by molecular genetics.
ibilities exist and are classified as either major, minor, or RESULTS: Between March 1996 and June 2012, a
bidirectional.5 In major ABO-incompatible HSCT the total of 176 patients received minor ABO-incompatible patient has preformed antibodies (i.e. isohemagglutinins) HSCT and 150 (85%) were secretors. Incidence and against A and/or B antigens expressed on donor red blood severity of acute graft-versus-host disease (GVHD) and cells (RBCs). Minor ABO-incompatible HSCT is character- chronic GVHD did not differ between secretors and ized by the transfer of donor isohemagglutinins against nonsecretors (cumulative incidences ± standard errors: recipient RBC antigens and of the corresponding immune acute GVHD on Day 100, 41 ± 11 and 46 ± 5%, cells (i.e., lymphocytes). A bidirectional blood group p = 0.59; chronic GVHD at 2 years, 52 ± 13 and56 ± 5%, p = 0.62, for secretors and nonsecretors, ABBREVIATIONS: HR = hazard ratio; HSCT = hematopoietic
respectively). Additionally, nonrelapse mortality (NRM) stem cell transplantation; NRM = nonrelapse mortality; and overall survival (OS) were similar in the two groups OS = overall survival.
(2-year NRM, 27 ± 9 and 23 ± 3%, p = 0.45; 4-year OS,64 ± 10 and 55 ± 4%, p = 0.28, for secretors and nonse- From the 1Division of Hematology, University Hospital, and the 2Blood Transfusion Centre, Swiss Red Cross, Basel, Switzerland; CONCLUSION: The recipients' ABH secretor status in
the 3Blood Transfusion Centre, Swiss Red Cross, Zurich, minor ABO-incompatible HSCT has no prognostic Switzerland; the 4National Reference Laboratory for impact on major transplant outcomes.
Histocompatibility, Department of Internal Medicine, University Hospitals, Geneva, Switzerland; and the 5Blood Transfusion Centre, Swiss Red Cross, Aarau, Switzerland.
Address reprint requests to: Joerg-Peter Sigle, MD, Blood Transfusion Centre, Swiss Red Cross, Kantonsspital Aarau, 5001 Aarau, Switzerland; e-mail: firstname.lastname@example.org.
Received for publication March 14, 2014; revision received May 19, 2014, and accepted May 22, 2014.
doi: 10.1111/trf.12768 HOLBRO ET AL.
barrier is a combination of major and minor ABO incom- MATERIALS AND METHODS
patibility. Various specific complications—for example, All adult patients that underwent minor or bidirectional pure RBC aplasia in major ABO-incompatible HSCT or ABO-incompatible allogeneic HSCT at our institution delayed hemolysis through passenger lymphocyte syn- between March 1996 and June 2012 were included in this drome in minor ABO-incompatible HSCT—can occur.
retrospective study. Patients with blood group A1, who Several approaches to prevent complications after ABO- received a HSCT from an A2 donor, were also included in incompatible HSCT have been proposed, including pre- the analysis. We excluded patients who received more ventive measures in the recipient and different graft than one HSCT, cord blood as stem cell source, or highly processing steps.5 T-cell-depleted haploidentical HSCT. Patient, disease, and Several studies have addressed the impact of ABO transplant characteristics were collected by chart review incompatibility on HSCT outcome. A large study found no and through the electronic database of our institution. All difference in overall survival (OS), transplant-related mor- patients provided written informed consent to have their tality, and Grade II to IV acute graft-versus-host disease data on disease, treatment, and outcome reported.
(GVHD) after ABO-identical, major, minor, or bidirec-tional ABO-incompatible HSCT from HLA-identical sib-lings.6 On the other hand, Kanda and coworkers7 found a ABH secretor status
lower OS in a subgroup of patients after minor ABO-incompatible, unrelated HSCT with bone marrow as stem According to the Lewis (LE) phenotype and the secretor cell source. As A and B antigens and their precursor, the gene (α1,2-l-fucosyltransferase; FUT2), individuals can be H glycoprotein, are expressed not only by RBC, but also classified as secretors and nonsecretors.11 Thus Se and se many other tissues including vascular endothelium (the two alleles of FUT2, Se being dominant over se) deter- ("histo-blood group"), one may speculate that in the pres- mine the presence or absence of the ABH substance in ence of an ABO barrier the tissue expression of ABH anti- body fluids.
gens can trigger or sustain an inflammatory reaction The ABH secretor status was assessed through deter- similar to that occurring in GVHD.8 In particular, in minor mination of LE phenotype or by Se genotyping. At our ABO-incompatible HSCT a humoral immune response institution serologic typing for all clinically relevant blood mediated by antibodies produced by donor lymphocytes groups, including LE, is routinely performed before HSCT against recipient ABH antigens may trigger GVHD by in all patients by gel test (Gel Test ID-system, Bio-Rad binding to and thus damaging the recipient's endo- Laboratories DiaMed GmbH, Cressier, Switzerland) or by thelium. Data regarding the effect of minor ABO- conventional agglutination test in tubes (antisera from incompatible HSCT on rate and severity of GVHD are Immucor, Inc., Norcross, GA). Se genotyping was per- conflicting. Some studies have shown an increased risk of formed in patients where serologic LE phenotyping was GVHD in minor ABO-incompatible HSCT.5,9 The study by missing, equivocal (mixed field after recent transfusions), Stussi and coworkers9 showed similar OS after minor ABO- or negative for both Lea and Leb.
incompatible HSCT compared to ABO-compatible HSCT,but a higher incidence of acute GVHD (Grade I-IV). Otherstudies failed to demonstrate a significant effect of minor Molecular determination of secretor status
ABO-incompatible HSCT on either rate or severity of Genomic DNA was isolated from peripheral blood mononuclear cells with the use of a DNA isolation However, the above-mentioned studies did not kit (MagnaPure LC, Roche Diagnostics, Mannheim, include the recipients' secretor status, which could Germany). The classic human secretor locus (Se) FUT2 explain the conflicting results. Eighty percent of all indi- encodes α1,2-l-fucosyltransferase and is located on Chro- viduals, who are defined as secretors, do not only express mosome 19. A nonsense mutation involving Codon 143 their ABH antigens on tissues, but are also capable of (numbered from the putative initiator methionine of the secreting soluble ABH substance in their body fluids, short FUT2 protein) is responsible for the nonsecretor including plasma.10 phenotype.12 The nonsense mutation is due to a G-to-A Soluble A/B antigens in the recipient's plasma poten- transition at Nucleotide 428. Wild-type (Se, 428G) and tially neutralize in vivo circulating anti-A and/or anti-B mutant (se, 428A) alleles of FUT2 gene were detected by derived from donor lymphocytes in minor ABO- polymerase chain reaction using sequence specific incompatible HSCT and thus mitigate possible immuno- priming technology in two independent reactions. Het- logic and/or inflammatory responses. This could affect erozygous individuals would give positive amplification incidence and severity of GVHD, disease relapse, and OS.
in both reactions, and homozygous individuals in one The aim of this retrospective study was to investigate the reaction only. Primers for the wild-type allele (428G) prognostic role of the recipients' ABH secretor status after were FUT2-all+523R (CCGGCTCCCGTTCACCTG-3′) and minor ABO-incompatible HSCT.
FUT2-Se+428G-F (CCGGCTACCCCTGCTCGTG-3′), and ABH-SECRETOR STATUS AND TRANSPLANTATION
FUT2-all+523R and FUT-se+428A-F (ACCGGCTACCCCTGCTCGTA-3′) for the mutant allele (428A), respectively.
TABLE 1. Patient, disease, and transplant
Concentrations of the primers in the final reaction volume characteristics according to the secretor status
were 200 nmol/L, and those of the control primers 90 nmol/L. Sequences of the control primers, reaction, and cycling conditions have been described previously.13 Mean age at HSCT (years) Patient, disease, and transplant characteristics were com- pared between secretors and nonsecretors using Pear- son's chi-square tests for categorical variables. For acute and chronic GVHD and nonrelapse mortality (NRM), competing risks analysis was used. For univariate analysis of OS, the Kaplan-Meier method was used. Multivariable Cox analysis was used to adjust for donor type and disease Conditioning regimen stage. All comparisons were two-sided, and p values of less than 0.05 were considered significant. All analyses were carried out with computer software (Stata, Version 12, StataCorp, College Station, TX).
Mismatched related Between March 1996 and June 2012, a total of 788 adult patients underwent allogeneic HSCT at our institution. A total of 201 patients received a minor ABO-incompatible HSCT. Patients who received more than one HSCT (n = 19) or a highly T-cell-depleted haploidentical HSCT (n = 5) ALL = acute lymphoblastic leukemia; AML = acute myeloid were excluded from the analysis. One additional patient leukemia; CMV = cytomegalovirus; CYA = cyclosporine; had to be excluded because of missing LE phenotyping D = donor; LPD/PCD = lymphoproliferative disorders/plasma and no available DNA for molecular testing. Overall, 176 cell diseases; MDS = myelodysplastic syndrome;MMF = mycophenolate; MPN = myeloproliferative disorder; patients were included in the analysis. ABO donor- MTX = methotrexate; R = recipient; RIC = reduced-intensity recipient combinations were as follows: O/A1/2 91; O/AB 3; O/B 22; A/B 11; B/A 17; A2/A1 19; A2/A1B 3; B/AB 3; andA/AB 7. A total of 112 patients (64%) were male, and64 (36%) were female. Mean age at transplantation was quent (n = 64; 36%). Diagnosis distribution and disease 40 years and was not different between secretors and stage at HSCT were not different between secretors and nonsecretors. Most patients (n = 140; 80%) received amyeloablative conditioning regimen. Ninety-five patients(54%) received a HSCT from an unrelated donor. GVHD Secretors and nonsecretors
prophylaxis consisted primarily of cyclosporine and A total of 150 patients (85%) were secretors and 26 (15%) methotrexate. Table 1 summarizes patient, disease, and were nonsecretors. Of the nonsecretors, 20 patients (11%) transplant characteristics, according to the secretor had their secretor status determined by molecular analy- sis because of missing LE phenotype (n = 6) or becausethe phenotype was Le(a–b–) (n = 14). Seven of thesepatients were homozygous (Se/Se) and 10 heterozygous (Se/se) in the secretor gene locus and three were nonse- Cumulative incidence of acute (≥Grade II) GVHD on cretors. Overall, 22 nonsecretors (85%) were male.
Day 100 was 41 ± 11% for secretors and 46 ± 5% for non-secretors (p = 0.59). Incidence of chronic GVHD at 2 yearswas 52 ± 13% for secretors and 56% ± 5% for nonsecretors Disease and transplant characteristics
(p = 0.62; Fig. 1).
The main indication for HSCT was acute leukemia (n = 93; Adjusted multivariable Cox analysis confirmed that 53%), with acute myeloid leukemia being the most fre- secretor status was not predictive of GVHD development:
HOLBRO ET AL.
Fig. 1. Acute (≥Grade II) and chronic GVHD according to secretor status. (—) Nonsecretor; (- - -) secretor.
Fig. 2. NRM and OS according to secretor status. (—) Nonsecretor; (- - -) secretor.
hazard ratio (HR) secretor versus nonsecretor for acute the other hand, strongly affects NRM, incidence and GVHD 0.85, 95% confidence interval (CI) 0.42-1.72, severity of GVHD, and OS.14 For several decades, HSCT has p = 0.64; and HR for chronic GVHD 1.08, 95% CI 0.58-2.01, been performed—if unavoidable—across the ABO blood group barrier. While in major ABO-incompatible HSCT,acute hemolysis and pure RBC aplasia are the major short- NRM and OS
term complications, passenger lymphocyte syndromeis a possible complication in minor ABO-incompatible Two-year NRM was similar between secretors and nonse- HSCT, with varying clinical course from asymptomatic cretors (27 ± 9% and 23 ± 3% for secretors and nonsecre- laboratory finding to severe and even life-threatening tors, respectively; p = 0.45).
condition.15,16 Besides the above-mentioned immune- The same was seen in the 4-year OS, which was hematologic complications, the data on the impact of 64 ± 10% for secretors and 55 ± 4% for nonsecretors minor ABO blood group incompatibility on OS as well as (p = 0.28; Fig. 2). Again, adjusted multivariable Cox analy- incidence and severity of GVHD is controversial.5 As sis confirmed that secretor status was not predictive of ABO blood group antigens are expressed on different NRM and OS: HR secretor versus nonsecretor for NRM tissues, the presence of antibodies against these antigens 1.17, 95% CI 0.54-2.55, p = 0.70; and HR for OS 0.79, 95% could have an impact on the underlying disease, GVHD, CI 0.41-1.55, p = 0.50.
and OS. In particular, antibodies produced by donorlymphocytes in minor ABO-incompatible HSCT are directed against recipient ABO antigens and could In contrast to solid organ transplantation ABO incompat- induce a humoral immune response. Endothelial cells ibility is of minor importance for HSCT. HLA matching, on which express A and/or B substances could be a ABH-SECRETOR STATUS AND TRANSPLANTATION
possible target of this immune response, as there is a per- Even though we found no impact of secretor status in sistence of recipient type endothelium after HSCT.17 minor ABO-incompatible HSCT, the role of secretor status Recipients, who are ABH secretors could—on the other in ABO-incompatible solid organ transplants should be hand—neutralize these donor antibodies ("in vivo" investigated. It is widely recognized that anti-A and anti-B adsorption), thus mitigating a possible immunologic isohemagglutinins can cause hyperacute rejection of reaction in both directions (graft-versus-leukemia and incompatible transplants. However, the current organ graft-versus-host reaction). To the best of our knowledge, shortage has driven new incentives and strategies, includ- the impact of ABH secretor status in recipients of minor ABO-incompatible HSCT has not been investigated Together with immunosuppression and different prepara- tive protocols including rituximab and immunoad- In this study, 85% of the 176 patients with minor ABO- sorption and/or plasma exchange in ABO-incompatible incompatible HSCT were ABH secretors, which is consis- solid organ transplantation, ABH secretion could have an tent with the known prevalence in a Caucasian impact on short- and long-term transplant outcomes, population. We did not find any differences in major trans- acting as an "in vivo" adsorption mechanism.
plant outcomes, including acute (≥Grade II) and chronic In conclusion, the role of the recipients' ABH secretor GVHD, NRM, and OS between secretors and nonsecretors.
status in minor ABO-incompatible HSCT appears to be Our data therefore do not suggest a clinically significant not relevant for clinical outcome and is not an explanation effect of secretor status on a donor-derived humoral for the discrepant results of the published literature. This immune response against recipients' ABO antigens. One may not be the case for ABO-incompatible solid organ possible explanation for this finding could be the lack transplantation, an expanding field that deserves further of a general increase of donor derived anti-A/B after minor ABO-incompatible HSCT.18 Additionally one mayfurther speculate that transplant-associated micro- angiopathy as a manifestation of GVHD also representsan antibody-mediated endothelial cell activation and We thank Sonja Sigurdardottir for development and implemen- damage. However, in a previous study we did not find ABO tation of the molecular determination of secretor status. AH, JPS, incompatibility to be a risk factor for the development of and AB designed the study and drafted the manuscript; AH and JPS collected the data; MS performed statistical analysis; CG per- From a pathophysiologic viewpoint, the transfusion formed molecular genetic analysis; and all authors contributed to of ABO-compatible nonidentical plasma can mimic a sce- data analysis, data interpretation, and writing of the manuscript.
nario similar to the one described above. Soluble ABHsubstance in plasma obtained from secretors can interact CONFLICT OF INTEREST
with recipients' isohemagglutinins, if transfused in anABO-compatible but nonidentical manner. A large retro- The authors have disclosed no conflicts of interest.
spective study has described a potentially negative impactof ABO-compatible nonidentical plasma transfusions on survival.20 The authors hypothesized that immunecomplex formation could be a possible cause. However, 1. Passweg JR, Baldomero H, Bregni M, et al. Hematopoietic our finding on NRM and OS suggests that immune SCT in Europe: data and trends in 2011. Bone Marrow complex formation, which could occur in secretors in minor ABO-incompatible HSCT has a negligible impact 2. Gooley TA, Chien JW, Pergam SA, et al. Reduced mortality after allogeneic hematopoietic-cell transplantation. N Engl Our study has various limitations. These include the J Med 2010;363:2091-101.
relatively small overall number of nonsecretors, the lack of 3. Gratwohl A, Stern M, Brand R, et al. Risk score for outcome data regarding recipient isohemagglutinin titers before after allogeneic hematopoietic stem cell transplantation: and during the course of HSCT and the lack of additional a retrospective analysis. Cancer 2009;115:4715-26.
data on immunohematologic complications after minor 4. Rowley SD, Donato ML, Bhattacharyya P. Red blood cell- ABO-incompatible HSCT, including passenger lympho- incompatible allogeneic hematopoietic progenitor cell cyte syndrome in secretors and nonsecretors. An addi- transplantation. Bone Marrow Transplant 2011;46:1167-85.
tional limitation is the power of our retrospective analysis, 5. Booth GS, Gehrie EA, Bolan CD, et al. Clinical guide to which would detect differencies in the defined outcome ABO-incompatible allogeneic stem cell transplantation.
variables with a HR of 2 to 2.5. ABH secretor status was Biol Blood Marrow Transplant 2013;19:1152-8.
determined either through determination of LE pheno- 6. Seebach JD, Stussi G, Passweg JR, et al.; GVHD Working type or by genotyping. Therefore, we could not further Committee of Center for International Blood and analyze the influence of zygosity of the secretor gene.
Marrow Transplant Research. ABO blood group barrier in HOLBRO ET AL.
allogeneic bone marrow transplantation revisited. Biol 14. Petersdorf EW. The major histocompatibility complex: a Blood Marrow Transplant 2005;11:1006-13.
model for understanding graft-versus-host disease. Blood 7. Kanda J, Ichinohe T, Matsuo K, et al. Impact of ABO mis- matching on the outcomes of allogeneic related and unre- 15. Kimura F, Sato K, Kobayashi S, et al.; Japan Marrow Donor lated blood and marrow stem cell transplantations for Program. Impact of AB0-blood group incompatibility on hematologic malignancies: IPD-based meta-analysis of the outcome of recipients of bone marrow transplants cohort studies. Transfusion 2009;49:624-35.
from unrelated donors in the Japan Marrow Donor 8. Socie G, Blazar BR. Acute graft-versus-host disease: Program. Haematologica 2008;93:1686-93.
from the bench to the bedside. Blood 2009;114:4327-36.
16. Daniel-Johnson J, Schwartz J. How do I approach ABO- 9. Stussi G, Muntwyler J, Passweg JR, et al. Consequences of incompatible hematopoietic progenitor cell transplanta- ABO incompatibility in allogeneic hematopoietic stem cell tion? Transfusion 2011;51:1143-9.
transplantation. Bone Marrow Transplant 2002;30:87-93.
17. Mueller RJ, Stussi G, Puga Yung G, et al. Persistence of 10. Henry S, Oriol R, Samuelsson B. Lewis histo-blood group recipient-type endothelium after allogeneic hematopoietic system and associated secretory phenotypes. Vox Sang stem cell transplantation. Haematologica 2011;96: 11. Grubb R. Correlation between Lewis blood group and 18. Stussi G, Huggel K, Schanz U, et al. Levels of anti-A/B anti- secretor character in man. Nature 1948;162:933.
bodies after ABO-incompatible hematopoietic stem cell 12. Kelly RJ, Rouquier S, Giorgi D, et al. Sequence and expres- transplantation. Transplant Proc 2005;37:1385-7.
sion of a candidate for the human secretor blood group 19. Martinez MT, Bucher C, Stussi G, et al. Transplant- alpha(1,2)fucosyltransferase gene (FUT2). Homozygosity associated microangiopathy (TAM) in recipients of alloge- for an enzyme-inactivating nonsense mutation commonly neic hematopoietic stem cell transplants. Bone Marrow correlates with the non-secretor phenotype. J Biol Chem 20. Shanwell A, Andersson TM, Rostgaard K, et al.
13. Kormoczi GF, Wagner T, Jungbauer C, et al. Genetic diver- Post-transfusion mortality among recipients of sity of KELnull and KELel: a nationwide Austrian survey.
ABO-compatible but non-identical plasma. Vox Sang
International Journal of Risk & Safety in Medicine 27 (2015) 85–91 DOI 10.3233/JRS-150645IOS Press Suicidal risk from TADS study was higherthan it first appeared G¨oran H¨ogberga,b,∗, David O. Antonuccioc and David Healyda Department of Women's and Children's Health, Child and Adolescent Psychiatric Unit,Karolinska Institutet, Astrid Lindgren Children's Hospital, Stockholm, SwedenbStockholm Child and Adolescent Psychiatry, BUP Huddinge, Stockholm, SwedencDepartment of Psychiatry and Behavioral Sciences, University of Nevada School of Medicine,Reno, NV, USAdBangor University, Wales, UK