Soo Young Yang

Bone marrow-allograft rejection by T lymphocytes recognizing a single amino acid difference in HLA-B44.

BONE marrow from unrelated donors is increasingly being transplanted for the treatment of patients with leukemia, aplastic anemia, and lethal congenital disorders of hematopoiesis and immunologic function. National and international registries of HLA-typed volunteer donors have been developed to optimize HLA matching between unrelated donors and recipients.1 , 2 The fate of marrow grafts depends on many factors, including conditioning regimens, the composition of the cells in the graft, immunosuppression after transplantation, and the degree of histocompatibility between donor and recipient. Clinically serious graft-versus-host disease occurs in 20 to 50 percent of patients receiving marrow grafts from HLA-identical siblings, and some measure .xa0.xa0.

The Natural History of Kaposi's Sarcoma in the Acquired Immunodeficiency Syndrome

Kaposis sarcoma is a multifocal systemic neoplasm histologically characterized by proliferating fibroblastic and microvascular elements. Initial signs include macules, papules, or nodules on the skin or mucosal surface. Lesions are frequently found on the trunk, arms, and head and neck. In general, sites of involvement and tumor load do not correlate with prognosis. A general decrease in the functional capacities of T and B cells is found in most patients. Kaposis sarcoma is reported as the initial manifestation of the acquired immunodeficiency syndrome (AIDS) in approximately 30% of cases. Most cases are in men, although it has been reported in all risk groups. Kaposis sarcoma in AIDS is more frequent among whites and homosexuals than blacks and intravenous drug abusers. Overall mortality is approximately 41%, with over 60% of patients alive at 1 year and 50% at 22 months. Overall survival is 18 months; however, some patients who have had the disease for 3 to 4 years are still doing well.

Genetic Mapping of the 21-Hydroxylase-Deficiency Gene within the HLA Linkage Group

To document further the proposed genetic linkage between congenital adrenal hyperplasia due to 21-hydroxylase deficiency and HLA, 34 unrelated families from New York and Zurich, with a total of 48 patients, 48 siblings and their parents, were studied. All patients were HLA genotypically different from the healthy sibs; when two or more children were affected in the same sibship they were always HLA-B identical. The gene for 21-hydroxylase deficiency was separated by genetic recombination from the HLA-A locus and from the locus for glyoxalase I-polymorphism. No HLA-A, HLA-B or HLA-C antigen was selectively increased among the 34 unrelated patients. Lod-score analysis for HLA-B:21-hydroxylase deficiency gave a peak for theta approximately 0.00 at 5.20 for females and 4.30 for males, giving a total peak lod score of 9.5 at theta approximately 0.00 when male and female lod scores were combined. Close genetic linkage between HLA-B and 21-hydroxylase deficiency was thus established.

Description of the Reference Panel of B-Lymphoblastoid Cell Lines for Factors of the HLA System: The B-Cell Line Panel Designed for the Tenth International Histocompatibility Workshop

The major objective for the collaborative studies within the Tenth International Histocompatibility Workshop was to compare biochemical and molecular genetic techniques for characterization of polymorphic HLA determinants with HLA specificities defined by conventional serological and cellular techniques. It was, for this purpose, attractive to develop an experimental design that would focus on the in-depth characterization of a limited number of target cells common for all Workshop components and shared by the participating laboratories.

Comparison of one-dimensional IEF patterns for serologically detectable HLA-A and B allotypes

A new mouse monoclonal antibody (MoAb) 4E, which detects an epitope shared by HLA-B locus antigens, together with the MoAb W6/32, detecting a common HLA, B, C, determinant, and the MoAb 4B, detecting HLA-A2 and A28, were used to isolate HLA-A and -B antigens in sequential immunoprecipitation. The HLA antigens obtained from metabolically labeled cell extracts of B-lymphoblastoid cell lines or from phytohemagglutinin (PHA) activated peripheral blood lymphocytes were compared by one-dimensional isoelectric focusing (1D-IEF). The IEF banding patterns obtained with native HLA antigens segregated in a family with HLA. Neuraminidase treatment of isolated antigens reduced the number of bands to one or two, simplifying the analysis of characteristic patterns. Thus, we have cataloged IEF banding patterns for the majority of the serologically recognized HLA-A and -B allotypes obtained from 57 unrelated American Caucasians. While no HLA-A locus or HLA-B locus specific banding patterns were observed, the HLA-A antigens had, in general, slightly higher pl values than the HLA-B antigens. HLA-C antigens could not be detected in this assay system. The polymorphism detected by IEF banding patterns was as extensive as the serologically detected polymorphism identified by classical HLA serology. Moreover, variants for some HLA allotypes could be detected by this method. In addition to previously recognized A2 variants, new variants were identified for HLA-A1, A26, and Bw44. Each A1 and Bw44 variant was associated with particular haplotypes. The HLA-A2 antigens occurred on 43 HLA haplotypes in the unrelated Caucasian population. Only one of each A2 variants was identified in this population.

Alloantibodies to pha-activated lymphocytes detect human Qa-like antigens

Platelet-absorbed sera were obtained from placental clots after delivery by multiparous women. These sera contained antibodies that react with PHA-activated lymphocytes after the latter are separated from peripheral blood and expanded with interleukin 2. These alloantibodies did not react with resting T lymphocytes, but reacted with B lymphocytes, PHA-activated lymphocytes, or both types of cells obtained from some but not all of the T lymphocyte donors. Reactions against B lymphocytes were associated with anti-Ia-like antibodies on the basis of blockage by turkey antibodies against human Ia. Reactions against PHA-activated lymphocytes that were blocked by turkey anti-beta 2m were classified as HT. Several antibodies were found to give reactions to HT determinants in separate panels of lymphocytes from Tel Aviv and Boston. The reproducibility of the cytotoxicity reactions was 89%. Altogether, 23 of 1100 sera were found to contain these reactions when screened by a panel of cells obtained from 30 individuals of known HLA phenotypes. Correlation coefficients were determined for all reactions, determining three clusters of significant reactivities: sera 965 and 1032 defined HT-2; sera SF48 and 1642 defined HT-3; and sera 1136, 1605, 1014, and 1227 defined HT-4. HT-2 was found to be inherited with HLA in 11 siblings from four families. Some of these antibodies react with antigens (non-HLA) containing beta 2m that were expressed on activated lymphocytes, but not on resting T lymphocytes, and did not react with thymocytes from the same donors of the peripheral lymphocytes. Our findings suggest that the HT alloantigens expressed on lectin-activated lymphocytes are class I differentiation antigens of a system analogous to the murine Qa system.

Advances in DNA sequencing technologies for high resolution HLA typing.

This communication describes our experience in large-scale G group-level high resolution HLA typing using three different DNA sequencing platforms - ABI 3730 xl, Illumina MiSeq and PacBio RS II. Recent advances in DNA sequencing technologies, so-called next generation sequencing (NGS), have brought breakthroughs in deciphering the genetic information in all living species at a large scale and at an affordable level. The NGS DNA indexing system allows sequencing multiple genes for large number of individuals in a single run. Our laboratory has adopted and used these technologies for HLA molecular testing services. We found that each sequencing technology has its own strengths and weaknesses, and their sequencing performances complement each other. HLA genes are highly complex and genotyping them is quite challenging. Using these three sequencing platforms, we were able to meet all requirements for G group-level high resolution and high volume HLA typing.

Identification of 2127 new HLA class I alleles in potential stem cell donors from Germany, the United States and Poland.

We describe 2127 new human leukocyte antigen (HLA) class I alleles found in registered stem cell donors. These alleles represent 28.9% of the currently known class I alleles. Comparing new allele sequences to homologous sequences, we found 68.1% nonsynonymous nucleotide substitutions, 28.9% silent mutations and 3.0% nonsense mutations. Many substitutions occurred at positions that have not been known to be polymorphic before. A large number of HLA alleles and nucleotide variations underline the extreme diversity of the HLA system. Strikingly, 156 new alleles were found not only multiple times, but also in carriers of various parentage, suggesting that some new alleles are not necessarily rare. Moreover, new alleles were found especially often in minority donors. This emphasizes the benefits of specifically recruiting such groups of individuals.

8/8 and 10/10 High-Resolution Match Rate for the Be The Match Unrelated Donor Registry

The National Marrow Donor Programs Be The Match Registry(®) facilitates the worldwide utilization of unrelated donor (URD) grafts for patients in need of a hematopoietic cell transplantation. In this study, we estimate the URD match rate for patients of White (WH), Hispanic (HIS), Asian/Pacific Islander (API), and African American/Black (AFA) race and ethnic groups. We chose 1344 URD at random as pseudo-patients (PP) to estimate the likelihood of finding an 8/8 or 10/10 high-resolution HLA-A,-B,-C,-DRB1 (and -DQB1) matched URD. Searches were conducted in the Be The Match Registry database for each PP at 2 time points: 2009 and 2012. URD who were a potential match for a PP by low/intermediate resolution were HLA typed by sequence-based typing to resolve the matching status. The 8/8 match rate for WH PP improved from 68% in 2009 to 72% in 2012. Corresponding match rates were 41% to 44% for HIS, 44% to 46% for API, and 27% to 30% for AFA, for 2009 and 2012, respectively. The 2012 10/10 match rates were 67% for WH, 38% for HIS, 41% for API, and 23% for AFA. These results provide baseline 8/8 and 10/10 match rate estimates by race for patients seeking an URD.

IEF patterns of HLA-1313 antigens from Orientals and Caucasians

HLA-B 13 antigens were isolated from metabolically labeled cell extracts from Caucasian and Oriental donors by means of an HLA-B13-specific monoclonal antibody, SY1. Ethnic differences in B13 molecules were identified by one-dimensional isoelectric focusing in which the pI of desialated Oriental B13 molecules was found to be higher than that of Caucasians. An additional Caucasian variant pattern was detected by peptide mapping using limited proteolysis in sodium dodecyl sulfate analyzed by polyacrylamide gel electrophoresis. Dual allotypic determinants for B13 molecules were recognized by two HLA-B13-specific monoclonal antibodies, SY1 and TU110, as determined by their sensitivity to complement-dependent cell lysis. Whereas the SY1 target epitope was shared by both ethnic B13 molecules, the two ethnic B13 molecules carried different Tu110 target structures. The Caucasian variant molecules appear to carry altered allotypic determinants which are recognized by both SY1 and Tu110 antibodies. This study suggests that the HLA-1313 private structure may comprise two epitopes recognized by SY1 and Tü 110 antibodies, respectively, whose binding sites overlap. Present data also suggest that the private determinant was already present when the two racial groups diverged, and thus the mutations which gave rise to the variants may be of relatively recent origin.