Impact of peptides on the recognition of HLA class I molecules by human HLA antibodies

MHC class I molecules expressed on cell surfaces are composed of H chain, beta2-microglobulin and any of a vast array of peptides. The role of peptide in the recognition of HLA class I by serum HLA Abs is unknown. In this study, the solid-phase assay of a series (n = 11) of HLA-A2-reactive, pregnancy-induced, human mAbs on a panel (n = 12) of recombinant monomeric HLA-A2 molecules, each containing a single peptide, revealed peptide selectivity of the mAbs. The flow cytometry membrane staining intensities on the HLA-A2-transduced cell line K562, caused by these mAbs, correlated with the number of monomer species detected by the mAbs. Flow cytometry staining on HLA-A2-bearing cell lines of a variety of lineages was indicative of tissue selectivity of these HLA-A2 mAbs. This tissue selectivity suggests that the deleterious effect on allografts is confined to alloantibodies recognizing only HLA class I loaded with peptides that are derived from tissue-specific and household proteins. Since Abs that are only reactive with HLA loaded with irrelevant peptides are expected to be harmless toward allografts, the practice of HLA Ab determination on lymphocyte-derived HLA deserves reconsideration.     

HLA 1990

The HLA system encompasses approximately one thousandth of the human genome and contains a series of closely linked loci coding for molecules which provide the context for the recognition of antigens by T lymphocytes. Within the HLA system, several phenotypically expressed and genomic polymorphisms can be distinguished. These polymorphisms are described and the main objectives for the future research are discussed.     

Selection of HLA compatible donors for patients with HLA antibodies

A strain of 1,507 typised donors enables 15.97 average donors to be selected for a thrombocyte transfusion in one of 75 patients selected at random with HLA antibodies being previously determined (1 to 66 donors per patient). HLA compatible donors were found for 72 patients (97.33 per cent). More than 5 HLA compatible donors could be found for 58 patients. The high number of compatible donors is based on the fact that among 1,507 donors there were 156 HLA homozygotes and 556 donors with 3 HLA-A or B-antigens respectively. Compatibility in the ABO-system was not taken into account.     

Allogeneic transplantation across the HLA barriers

In high-risk acute leukemia patients, a 10-fold increase in the dose of extensively T-cell-depleted hematopoietic stem cells ensures sustained full-donor engraftment of one-haplotype-mismatched transplants without graft-vs.-host disease. Since our first successful pilot study, which exploited the principle of a megadose stem cell transplant, our efforts have concentrated on developing new conditioning regimens, optimizing graft processing and improving the post-transplant immunologic recovery. The results so far achieved in more than 100 high-risk acute leukemia patients show that haploidentical transplantation is now a clinical reality. Because virtually all patients in need of a hematopoietic stem cell transplant have a full-haplotype-mismatched family donor, a T-cell-depleted mismatched transplant can be offered with curative intent, thus extending allogeneic transplantation procedures to virtually all candidates.     

Interesting relations in the HLA system

There exists no absolute binding between the antigens HLA-Cw 2, Cw 3 and Cw 4, on the one hand, and HLA-B 27, HLA-B 15 and HLA-Bw 35, on the other hand. Even if 91% of human beings with HLA Cw 4 will simultaneously have the antigen HLA-Bw 35, another antigen as HLA-B 27 or HLA-B 15 can be identified in approximately 55% of individuals with HLA-Cw 2 and Cw 3. In this connection, the joint presence of some pairs of cross-reacting HLA antigens (A 2 and A 28, B 5 and Bw 35, B 7 and B 27, B 8 and B 14, B 12 and Bw 2) could be proved and their frequency be determined. 2 cases of a simultaneous presence of two subtypes of HLA-A 10 antigen, A 25 and A 26, could be found in family examinations. Moreover, two atypical bindings of anti-HLA-Bw 4 and anti-HLA-Bw 6 cytotoxins with HLA antigens could be identified: 7,49% of HLA-Bw 35 positive lymphocytes no positive response with anti-HLA-B 4 and 1,69% of HLA-B 12 with anti-HLA Bw 6. The importance of the findings for determining HLA in practice is discussed.     

HLA antibody responses in HLA class I transgenic mice

In a previous report we described how cross-immunizations of pairs of transgenic mice expressing different HLA class I antigens led to the production of antibodies directed exclusively at polymorphic epitopes. This was ascribed to self-tolerance of HLA that prevents immune responses to monomorphic epitopes and focuses responses on polymorphic ones. In the present report we extend our findings and demonstrate that immunizations of class I transgenic mice with HLA transfected mouse fibrosarcoma as well as with human lymphoblastoid cells also preferentially yield antibodies to polymorphic epitopes. This was the case whether or not immunizations were carried out across locus barriers [e.g., Tg (HLA-A *0201) or Tg (HLA-Cw*0301) transgenic mice immunized with HLA-B27 transfectants] or within the same locus [e.g., Tg (HLA-B*1302) transgenic mice immunized with HLA-B27 transfectants or B27-expressing lympho-blastoid cell]. Use of an extended immunization protocol with four or more booster injections favored antibodies of IgG isotype with affinities high enough to lyse normal peripheral blood lymphocytes (PBLs) in complement-dependent cytotoxicity assays and to immunoprecipitate HLA antigens. The specificities covered by the monoclonal antibodies (mAbs) could be either broad or narrow, depending on the genetic distance of the HLA antigens or alleles involved. For instance, a Tg(HLA-B*1302) transgenic mouse immunized with B27 produced both broad B7/B27-specific antibodies, Bw4-specific antibodies, and one antibody reacting with all B alleles except B13 and with some C alleles. On the other hand, a Tg(HLA-B*1302) transgenic mouse immunized with Bw47 transfectants responded narrowly with an antibody to Bw60 and Bw47. Thus it appears that by choosing appropriate recipient mice and closely related or more distant HLA antigens, antibodies of a programmed specificity can be generated. Address correspondence and offprint requests to: U. Hämmerling.     

HLA and the probability of paternity.

In cases of disputed paternity, blood tests are often used to obtain an estimate of the probability that the accused male is the true father. The interpretation of the genetic data is usually based upon a statistic called the paternity index. This paper shows that the paternity index method cannot be applied to data from compound loci in the absence of information on linkage phase. Since phenotypic data from compound loci, such as HLA, MNSs, and Rh, are often useful in disputed paternity proceedings, they should be analyzed with available alternative statistics.     

HLA Typing for the Next Generation

Allele-level resolution data at primary HLA typing is the ideal for most histocompatibility testing laboratories. Many high-throughput molecular HLA typing approaches are unable to determine the phase of observed DNA sequence polymorphisms, leading to ambiguous results. The use of higher resolution methods is often restricted due to cost and time limitations. Here we report on the feasibility of using Pacific Biosciences’ Single Molecule Real-Time (SMRT) DNA sequencing technology for high-resolution and high-throughput HLA typing. Seven DNA samples were typed for HLA-A, -B and -C. The results showed that SMRT DNA sequencing technology was able to generate sequences that spanned entire HLA Class I genes that allowed for accurate allele calling. Eight novel genomic HLA class I sequences were identified, four were novel alleles, three were confirmed as genomic sequence extensions and one corrected an existing genomic reference sequence. This method has the potential to revolutionize the field of HLA typing. The clinical impact of achieving this level of resolution HLA typing data is likely to considerable, particularly in applications such as organ and blood stem cell transplantation where matching donors and recipients for their HLA is of utmost importance.     

Soluble HLA class I and HLA class II antigens in the blood of HIV infected patients

The levels of HLA, class I, antigen and HLA-DR antigen in the blood sera of HIV-infected persons were determined by the enzyme immunoassay. The levels of soluble antigens HLR-DR and of HLA, class I, in serum samples containing only antibodies to HIV were elevated, respectively, 1.8- and 1.3-fold in comparison with the norm. The level of soluble HLA-DR antigen in samples containing the markers of other infections (HBsAg, antibodies to hepatitis C virus, anti-IgM antibodies to cytomegalovirus) was significantly higher in comparison with samples containing only antibodies to HIV and serum samples from healthy donors (p < 0.05). The concentration of soluble HLA antigen, class I, in the samples containing antibodies to HIV and markers of other infections was also elevated, but the statistically authentic increase in comparison with the normal level was observed only in the presence of the markers of HIV infection, hepatitis C and cytomegalovirus infection.     

HLA haplotype discordance

Previous work on the inheritance of disease has often used certain measures of HLA haplotype concordance (such as the number of haplotypes "identical by descent," IBD) among affected siblings from each of a number of sibships, each of which contains at least two affected siblings. Here we introduce a new measure of HLA haplotype discordance between the affected and unaffected siblings of each sibship (provided there is at least one of each). We show how the measure can be used to give a simple test for inheritance, which we exemplify with data.     

HLA and leukemia

HLA-A, -B, -C characteristics were determined in 85 AML patients and in 47 ALL patients as well as in 95 pairs of parents of leukemic patients. The detected antigen frequencies of patients were compared with those of 2,947 donors from Leipzig. A significant increase of frequency was found for Cw4 and in the group of ALL patients (p = 0.05), an increase of frequency could be observed for B22 and in the group of PR/NR patients for pcor = 0.05. A correlation between A9 and longer survival time in leukemic patients could not be found. Parents of leukemic patients (but not the patients themselves) showed a limited genetic heterogeneity with respect to the HLA system.     

Localization of the Bf locus within the HLA region

Summary Since the estimation of recombination fractions is only an arbitrary means to map genetic loci on chromosomal regions, family studies of cases with informative cross-overs within the region in question are of decisive importance. This paper reports the study of a family with a HLA-B to HLA-D cross-over which is informative in the Bf system. The possible reasons for conflicting published data on Bf gene mapping are discussed.Supported in part by grants from Deutsche Forschungsgemeinschaft (Ri 164/11, Sonderforschungsbereich 37, TPB3, München)     

The HLA loci of the Hopi and Navajo

The purpose of this paper is to present the genetic distribution at the HLA-A,B,C, and DR loci in the Hopi and the Navajo. A sample of 100 outpatients from each tribe was selected at the Public Health Service Indian Hospital in Keam's Canyon, Arizona, and was typed for the antigens at the four loci. The distributions of the alleles and the haplotypes are similar in each tribe. A distance measure, f, confirms the genetic similarity of the two populations. It is concluded that the great cultural diversity of the Hopi and the Navajo is the result of a cultural evolution and diversification that has greatly outstripped the genetic evolution at the major histocompatibility loci over the past 20,000 years.     

HLA genes in populations of the Aleutian islands

We typed a subset of the Aleut population for HLA loci (HLA-A, HLA-B, HLA-DRB1, HLA-DQB1) to obtain an HLA profile, which was compared to other Eurasian and Amerindian populations for studying Aleut origin and its significance on the peopling of the Americas. Allele frequencies at the four loci were identified in an Aleut sample using standard indirect DNA sequencing methods. Genetic distances with Amerindians and Eurasians were obtained by comparing Aleut allele frequencies with a worldwide population database (13,164 chromosomes). The most frequently extended HLA haplotypes were also calculated. We also generated Aleut relatedness dendrograms and calculated correspondence relatedness in a multidimensional scale. Both neighbor-joining dendrograms and correspondence analysis separated Aleuts from Eskimos and Amerindians. Aleuts are closer genetically to Europeans, including Scandinavians and English. Our results are concordant with those obtained by Y-chromosome analysis, suggesting that most male Aleut ancestors of our sample came mainly from Europe.     

Evidence for heterosis in the HLA system

The number of persons with homozygous HLA haplotypes in several groups of South American Indians was 39% less than that expected assuming unmodified equilibrium. In a subpopulation of 122 persons whose parents' HLA constitutions were known, there were 56% fewer homozygous persons than expected. This deficit was widely distributed in different haplotypes and different tribal groups.