Generation of Unexpected Allele-Specific Anti-HLA Antibodies after the Transplantation of a Fully-Matched Kidney Allograft and the Diagnostic Approaches Required for Excluding Harmful Effects after Subsequent Renal Regrafts

The specification of anti-human leukocyte antigen (HLA) antibodies is an important task for patients awaiting kidney allografts. Especially the patients immunized in previous transplantations, transfusions, or pregnancies must be carefully observed, since grafting patients with HLA antigens/phenotypes recognized by their pre-formed antibodies are the main cause of harmful hyperacute and acute rejection. The complement-dependent lymphocytotoxicity-based de facto (physical) crossmatching (CDC-CM) has thus been implemented as the last diagnostic obstacle before kidney allografting. Here, an assay is performed by incubating the donors’ lymphocytes with the sera of the prospective recipients, and a negative outcome was desired for eligibility of the underlying organ allocation. Furthermore, valid antibody specification has to be performed at least quarterly for each patient on the kidney waiting list, as defined by certain guidelines, for example, the Eurotransplant guidelines. Based on the exclusion of these specificities, also referred to as virtual crossmatching, certain donors are a priori listed as unacceptable for these recipients. In this case report, we showed that defining unacceptable antigens may be difficult if the recipients’ antibodies are allele-specific after being generated in the patient who is expressing the HLA-class II antigen DQ6 and also developing antibodies against this antigen. Low resolution (two-digit) typing is used before kidney allografting. Thus, these antibodies are generally not definable, as donors and recipients share the same antigen (allelic group). Here, we demonstrate the diagnostic approaches required to exclude inadequate kidney donors for a patient exhibiting antibodies only against the HLA-DQB1*06:04 allelic variant and not against the common phenotype HLA-DQ6. In practice, the patient’s HLA-class II high resolution (four-digit) typing, as well as his antibody specification at the highest (single antigen) resolution, are included. Furthermore, we critically discuss, according to the Eurotransplant guidelines, the missing possibility to declare own HLA-antigens unacceptable, which may be very helpful for recipients who exhibit allele-specific antibodies.

Neuromyelitis optica is an HLA associated disease different from Multiple Sclerosis: a systematic review with meta-analysis

Neuromyelitis Optica and Multiple Sclerosis are idiopathic inflammatory demyelinating diseases of the central nervous system that currently are considered distinct autoimmune diseases, so differences in genetic susceptibility would be expected. This study aimed to investigate the HLA association with Neuromyelitis Optica by a systematic review with meta-analysis. The STROBE instrument guided research paper assessments. Thirteen papers published between 2009 and 2020 were eligible. 568 Neuromyelitis Optica patients, 41.4% Asians, 32.4% Latin Americans and 26.2% Europeans were analyzed. Only alleles of the DRB1 locus were genotyped in all studies. Neuromyelitis Optica patients have 2.46 more chances of having the DRB1*03 allelic group than controls. Ethnicity can influence genetic susceptibility. The main HLA association with Neuromyelitis Optica was the DRB1*03:01 allele in Western populations and with the DPB1*05:01 allele in Asia. Differences in the Multiple Sclerosis and Neuromyelitis Optica genetic susceptibility was confirmed in Afro descendants. The DRB1*03 allelic group associated with Neuromyelitis Optica has also been described in other systemic autoimmune diseases.

Population Analysis of the HLA-a* and -B* Allele Frequency in Bone Marrow Donors, Republic of Tatarstan, Russia

Introduction HLA genotyping is a critical stage in finding the best matched bone marrow donor for transplantation. Kazan Federal University (KFU) is involved in the «Rusfond» Charity «Rusfond. Register» program to generate the regional register of the bone marrow donors in the Republic of Tatarstan (RT). Russians and Tatars represent the majority of the RT population. Calculation of the HLA allelic group and haplotype profile of donors in RT could be useful to identify the closely related populations, which could contain the best-matched donors. This will facilitate selection of donors which could be located in far distances. The goal of this study is to characterize the potential bone marrow donors in the RT based on HLA-A* and -B* allelic group frequencies and to identify related populations in Russia and neighboring countries, as well as in European and non-European countries. Materials and methods HLA-A* and -B* allelic group frequencies in a group of 483 potential bone marrow donors from RT reported in our previous study (Davidyuk et al, 2017) were used to calculate the genetic distances between RT and 50 other populations. Data on HLA-allele frequencies in Caucasians, Mongoloid and Negroid populations were obtained from The Allele Frequency Net Database - website http://www.allelefrequencies.net (Gonzalez-Galarza et al, 2015). Analysis was conducted using R language for statistical computing and Rstudio (R Core Team, 2018). Packages ade4 and adegenet were used to calculate Reynolds' distances for HLA-A and HLA-B allelic group frequencies, perform clusterization with Ward's method and construct a dendrogram. Results and discussion The HLA-allele and haplotype frequencies in RT donors were reported in our previous study (Davidyuk et al, 2017). The comparative analysis of HLA-A* and -B* allelic group frequencies showed that the RT population and other populations from Russia as well as Slovakian and Polish populations are belong to one cluster on the dendrogram (Fig. 1). This cluster also contained the Caucasian populations from Western and Northern Europe. We also found that the RT population is more related to the populations from Samara region and Moscow. Two populations from north-western region of Russia and two Tatar populations were also closely related to the RT population. All populations listed above had low genetic distances from the RT population, while the genetic distance between the RT population and Russians from Chelyabinsk region was high (Fig. 2). Interestingly, Russians from Chelyabinsk region were more distantly related to the populations of European part of Russia as compared to RT population. Collected data suggest that the RT population could be used as potential bone marrow donors for recipients from closely related populations in the neighboring regions. However, in future, as the number of donors in the RT regional register will increase, the genetic relationship between RT population and other populations in Russia could be changed. Conclusion Our data revealed that potential bone marrow donors studied in the RT are closely related to the Russians and Tatars populations from European part of Russia. This observation could be important for selection of the hematopoietic stem cell transplant recipients from these populations to find a phenotypically matched donor within the register of RT. The Russian Government Program of Competitive Growth of KFU and «Rusfond» supported this study. Albert Rizvanov was supported by state assignment 20.5175.2017/6.7 of the Ministry of Education and Science of Russian Federation. ReferencesDavidyuk et al (2017). HLA Alleles and Haplotypes Frequency in the Population of the Republic of Tatarstan, Russia. Blood. 130:4942.Gonzalez-Galarza et al (2015). Allele frequency net 2015 update: new features for HLA epitopes, KIR and disease and HLA adverse drug reaction associations. Nucleic Acid Research 39, 28, D784-8.R Core Team (2018). R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL https://www.R-project.org/. Disclosures No relevant conflicts of interest to declare.

HLA Class I-Specific Effects in AML with Mutated Nucleophosmin

Background We have recently identified a skewed distribution of class I human leukocyte antigens (HLA) among patients with AML and mutated nucleophosmin gene (NPMc+ AML). A lower frequency of several HLA allelic groups in the NPMc+ AML patient cohort was in good correlation with the results of theoretical predictions for high-affinity immunopeptides derived from NPM1, suggesting that an anti-NPM1 immune response could prevent leukemia development in patients with a suitable HLA class I type. Aim We here present more detailed analysis of HLA-specific features, in a much larger cohort (N = 357) of AML patients with NPM1 mutation (type A/D) from several centers in the Czech Republic, Germany and Poland. Results Most importantly, we confirmed the lower incidence of B´40 and C´07 allelic groups in NPMc+ AML compared to the normal values. The moderate decrease of A´02 allelic group frequency became statistically significant in this larger patient cohort (see Table). On the other hand, no difference in HLA class II frequencies was found compared to the normal distribution. Furthemore, patients with B´07 allelic group had a significantly better prognosis, but only in the absence of Flt-3-ITD mutations (p = 0.049, see Figure). HLA typing was performed by molecular methods in a subgroup of patients (N = 73), allowing for discrimination between C´07 alleles. In this subgroup, C*07:01, but not C*07:02/04 expression was associated with better overall survival (p = 0.036), in agreement with theoretical predictions. Conclusion Our results support the hypothesis that anti-NPM1 immune response reduces AML development and contributes to a better outcome of NPMc+ AML patients with suitable HLA class I type (including at least A´02, B´07, B´40 and C*07:01). Furthermore, NPMc+ immunogenicity is caused rather by its aberrant localization than by the generation of a unique aminoacid sequence. Figure Figure. Disclosures Schetelig: Sanofi: Honoraria. Thiede:AgenDix: Employment, Other: Ownership.

Rio Grande do Norte, Brazil, voluntary bone marrow donors registry analysis

Objective: this study aimed to report the allele and haplotype frequencies of volunteer bone marrow donors (VBMD) from the state of Rio Grande do Norte (RN) who were enrolled in the Brazilian Volunteer Bone Marrow Donor Registry (REDOME). Methods: the sample comprised 12,973 VBMD who had their allele and haplotype frequencies calculated by Arlequin 3.5.1.2. A multivariate analysis of the data was obtained through a principal component analysis (PCA) and hierarchical cluster analysis (HCA) performed with SPSS 8.0. Results: the most frequent allelic group was HLA-A*02, followed by -DRB1*13, -DRB1*04, -DRB1*07, -B*44, -B*35, -A*24 and -DRB1*01. Of the 2,701 haplotypes observed, the three most frequent were HLA-A*01 B*08 DRB1*03 (1.62%), -A*29 B*44 DRB1*07 (1.56%) and -A*02 B*44 DRB1*04 (1.29%). These haplotypes were in linkage disequilibrium. RN allele and haplotype frequencies were very similar to those in other Brazilian states in which similar studies have been performed. The PCA revealed that RN is highly genetically similar to Caucasian populations, especially those from Iberian countries, which strongly influenced the state’s ethnic composition. Africans and Amerindians also influenced the RN population structure, to a lesser extent. Conclusion: the HCA reinforced the conclusion that, despite its highly admixed profile, the RN population is genetically similar to European and European-descended populations. The PCA also showed that RN cities do not contribute to the same extent to REDOME, with less populous cities being underrepresented, indicating the need to enroll more VBMD from these smaller cities to faithfully depict the state’s population structure in the database.

Genetic diversity among volunteer donors of bone marrow in southeastern Brazil, according to the HLA system

CONTEXT AND OBJECTIVE: Checking the histocompatibility of the molecules of the human leukocyte antigen (HLA) system is vital for performing bone marrow transplantation with allogeneic material. The objective of this study was to characterize bone marrow donors according to gender, age, ethnicity and HLA groups at a regional hemotherapy center in Brazil.DESIGN AND SETTING:Descriptive study on registered donors at a regional hemotherapy center in a public university hospital in the southeastern region of Brazil.METHODS: The records of 66,780 donors who were registered between 2005 and June 2011 were consulted, and the variables studied were tabulated.RESULTS:There were equal numbers of male and female donors and 82.8% of them were under 45 years of age. In terms of ethnicity, 77.3% declared themselves to be white, 15.0% mixed race, 5.7% black and 2% others. In terms of immunogenetic characterization, the most frequent HLA-A allelic group was HLA-A*02, with 39.20% of the donors; in the HLA-B allelic group, the most common was HLA-B*35, with 14.18%; while in the HLA-DRB1 allelic group, the most frequent was HLA-DRB1*03, with 17.03%. Comparison between these results and data from the Brazilian Bone Marrow Donor Registry (REDOME) showed that there were demographic and immunogenetic differences due to the history of immigration in the region of Ribeirão Preto, in southeastern Brazil.CONCLUSIONS: The results reinforce the importance of understanding the demographic and immunogenic profile of regions of Brazil, in order to reduce the waiting time for a histocompatible donor.

Earliness per se and its dependence upon temperature in diploid wheat lines differing in the major gene Eps-Am1 alleles

Differences in development among wheat cultivars are not only restricted to photoperiod and vernalization responses. When both requirements are fully satisfied differences may still arise due to earliness per se. It is not clear at present to what extent this trait is ‘intrinsically’ expressed (a constitutive trait) independently of the environmental conditions so that it might be selected under any thermal condition or if it may be altered to the extent of showing a crossover interaction with temperature in which the ranking of wheat genotypes may be altered. The present study assessed the influence of temperature on the intrinsic earliness for lines of diploid wheat characterized for their differences in a major gene for intrinsic earliness, but also possibly differing in their genetic background for other factors controlling this polygenic trait. To do so the lines were grown individually in two temperature regimes (16 and 23°C) under long days having previously been fully vernalized. Multiple comparisons analyses were carried out among lines of the same allelic group for the Eps-Am1 gene. Results indicated that within each group there were lines that did not differ in their earliness per se, others differed but without exhibiting any line×temperature interaction and finally different types of interaction were shown, including cases where the ranking of lines was altered depending on the growing temperature. It is thus possible that the selection of a genotype based on its earliness per se in an environment might not represent the same performance in another location where temperature varied significantly.

Comparative gene genealogical analyses of strains of serotype AD identify recombination in populations of serotypes A and D in the human pathogenic yeast Cryptococcus neoformans

Cryptococcus neoformans is a major pathogen of humans throughout the world. Using commercial monoclonal antibodies to capsular epitopes, strains of C. neoformans manifest five serotypes: A, B, C, D and AD. Previous studies demonstrated significant divergence among serotypes A, B, C and D, which are typically haploid. In contrast, most strains of serotype AD are diploid or aneuploid and result from recent hybridization between strains of serotypes A and D. Whether serotypes A, B, C and D represent strictly asexual lineages is not known. Using comparative genealogical analyses of two genes, the authors investigated whether recombination occurred among strains within serotypes A and D. For each of 14 serotype AD strains, a portion (642 bp) of the orotidine monophosphate pyrophosphorylase (URA5) gene was cloned and sequenced. Each of these 14 strains contained two different alleles and sequences for both alleles were obtained. The URA5 gene genealogy was compared to that derived from the laccase (LAC) gene, which was reported recently for the same 14 strains. For both genes, each of the 14 serotype AD strains contained two phylogenetically distinct alleles: one allele was highly similar to those from serotype A strains and the other to alleles from serotype D strains. However, within both the serotype A allelic group and the serotype D allelic group, there was significant incongruence between genealogies derived from URA5 and LAC. The results suggest recombination in natural populations of both serotypes A and D.