HLA-B44 Supertype Research Articles

HLA-B*44 and the Bw4-80T motif are associated with poor outcome of relapse-preventive immunotherapy in acute myeloid leukemia.

HLA-B alleles are associated with outcomes in various pathologies, including autoimmune diseases and malignancies. The encodedHLA-B proteins are pivotal in antigen presentation to cytotoxic T cells, and some variants containing a Bw4 motif also serve as ligands to the killer immunoglobulin-like receptors (KIR) 3DL1/S1 of NK cells. We investigated the potential impact of HLA-B genotypes on the efficacy of immunotherapy for relapse prevention in acute myeloid leukemia (AML). Seventy-eight non-transplanted AML patients receiving HDC/IL-2 in the post-consolidation phase were genotyped for HLA-B and KIR genes. HLA-B*44 heralded impaired LFS (leukemia-free survival) and overall survival (OS), but the negative association with outcome was not shared across alleles of the HLA-B44 supertype. Notably, HLA-B*44 is one of few HLA-B44 supertype alleles containing a Bw4 motif with a threonine at position 80, which typically results in weak binding to the inhibitory NK receptor, KIR3DL1. Accordingly, a strong interaction between KIR3DL1 and Bw4 was associated with superior LFS and OS (p = 0.014 and p = 0.027, respectively). KIR3DL1+ NK cells from 80T-Bw4 donors showed significantly lower degranulation responses and cytokine responses than NK cells from 80I-Bw4 donors, suggesting impaired KIR3DL1-mediated education in 80T-Bw4 subjects. We propose that presence of a strong KIR3DL1+-Bw4 interaction improves NK cell education and thus is advantageous in AML patients receiving HDC/IL-2 immunotherapy for relapse prevention.

Analysis of the different subpeptidomes presented by the HLA class I molecules of the B7 supertype

MHC-I molecules of the HLA-B7 supertype preferentially bind peptides with proline at position 2. HLA-B*51:01 and B*51:08 present two predominant subpeptidomes, one with Pro2 and hydrophobic residues at P1, and another with Ala2 and Asp enriched at position 1. Here, we present a meta-analysis of the peptidomes presented by molecules of the B7 supertype to investigate the presence of subpeptidomes across different allotypes. Several allotypes presented subpeptidomes differing in the presence of Pro or another residue at P2. The Ala2 subpeptidomes preferred Asp1 except in HLA-B*54:01, where ligands with Ala2 contained Glu1. Sequence alignment and the analysis of crystal structures allowed us to propose positions 45 and 67 of the MHC heavy chain as relevant for the presence of subpeptidomes. Deciphering the principles behind the presence of subpeptidomes could improve our understanding of antigen presentation in other MHC-I molecules.Running title: HLA-B7 supertype subpeptidomes.

Association of HLA class I genotype with outcomes of gastrointestinal cancer patients with immunotherapy.

e16551 Background: Chowell et al. showed that HLA (human leukocyte antigen) class I (HLA-I) heterozygosity may avail to survival benefit from immune checkpoint blockade (ICB) in advanced cancer patients. Among their cohorts of melanoma patients, patients with the HLA-B44 supertype had improved survival, while the individuals with HLA-B62 supertype had worse outcome. However, these observations were not supported by another study of NSCLC patients treated with immune checkpoint inhibitors (ICIs). Now whether HLA-I genotype can serve as predictive marker for ICB treatment of other malignancies such as gastrointestinal cancer remains unclear. Methods: Tumor tissue samples before immunotherapy as well as related clinical data were collected from 91 Chinese gastrointestinal cancer patients receiving anti-PD-1/PD-L1 inhibitors and/or anti-CTLA-4 antibodies. Genomic features of tumor specimens were characterized by whole exome sequencing (WES) and HLA-I genotyping of normal DNA from peripheral blood cells were performed using WES data. Tumor immune microenvironment signatures were examined by RNA-sequencing targeting 395 immune related genes. Results: We observed that the individuals with HLA-B62 supertype (n = 15) had a significantly shorter PFS (median PFS, 1.83 months versus 3.53 months; HR, 2.3; 95% CI, 1.03-5.0; p= 0.009) than that of those without HLA-B62 supertype (n = 35) in a subgroup of patients with microsatellite stability (MSS, n = 50) and it was not so in other patients with microsatellite instability-high (MSI-H, n = 17) or unknown MSI status (n = 24). Moreover, the patients with B62 supertype and TP53 mutations (n = 15) exhibited more markedly shortened PFS (mPFS, 1.8 months versus 5.57 months; HR, 3.9; 95% CI, 1.5-9.6; p< 0.0001) and OS (mOS, 4.2 months versus 17.3 months; HR, 4.5; 95% CI, 1.5-13.2; p< 0.0001) compared to those patients with non-B62 supertype and TP53 wildtype (n = 26). Furthermore, gene expression levels of certain immune activation related pathways or markers (IFN-γ signature, cytolytic marker, etc.) in the patients with B62 supertype and without B44 supertype (n = 23) were statistically lower ( p< 0.05) than in those patients without B62 supertype (n = 62). Chi-square test revealed no correlation between HLA-B62 supertype and TP53 mutation, MSI status, tumor mutation burden (TMB) or PD-L1 expression. Conclusions: HLA-B62 supertype in combination with TP53 mutations may define a subset of gastrointestinal cancer patients who probably fail to benefit from ICIs, especially in MSS patients.

A prospective cohort study of HLA-B44 supertype as predictor of response to novel immune checkpoint combinations.

e15071 Background: To date there are no clear predictors of response to novel immune checkpoint inhibitor (ICI) combinations. Recent reports suggest that HLA-I (Human Leukocyte Antigen Class I) supertype B44 may predict survival to PD1(L)1 inhibition in patients with melanoma and non-small cell lung cancer (NSCLC) ( Chowell et al, Science 2018). We sought to study if this effect is exclusive for PD(L)1 inhibition alone and the impact of HLA-B44 on ICI combinations with new immunotherapeutic agents. Methods: Between 2017 and 2019, 77 patients treated across 11 different early phase clinical trials (NSCLC: 64: (83,1%); melanoma: 4: (5,2%), bladder 8 (10,4%); and one nasopharynx carcinoma), ICI-naive were prospectively evaluated for HLA-I subclass. All patients were ECOG 0 or 1 (50 and 27 respectively). Most patients had ≤ 2 metastatic sites (55: 71,4%); 24 (31,2%) patients were treated with PD(L)1 inhibitors as monotherapy, and 53 (68.8%) with PD(L)1-containing combinations including ICI and agonists such as ICOS, OX40 or GITR monoclonal antibodies. HLA-I was classified for supertypes and the variables of progression-free survival (PFS) and overall survival (OS) were compared for patients with HLA-B44+ and B44-, in both groups (monotherapy and combinations). Results: 44 patients were HLA-B44+ and 33 were negative. For patients treated with anti-PD(L)1 agents as monotherapy, those that were HLA-B44+ had a longer PFS: [medianPFS: 22,38 months (m) (95% CI: 13,39 – 31,37); vs 3,8m (95% IC: 2,05 – 5,65); p: 0,002] and OS: [median: 33,2 m (95% CI: 23,7 – 42,6) vs 14,46 m (95% IC: 7,86 – 21,1); p: 0,13]. In contrast, these marked differences were not evident in patients receiving PD1 combination therapies [HLA-B44+ mPFS: 7,17m (95% CI: 4,41 - 9,93) vs 9,46 m (IC95% 6,8 m – 12,07); p:0,290], and OS: 11,53m (95% IC: 8,33 – 14,7) vs 18,9 m (IC 95% 13,3 – 24,5) p: 0,24. Lastly, response rate was also better for patients with HLA-B44 positive treated with PD1 monotherapy (66,7% vs 16,7% in HLA-B44 negative patients) p: 0,048, whilst when administering ICI combinations, HLA-B44 negative patients seemed to have a better response rate (44% vs 27% in HLA-B44 positive patients; p = 0,148). Conclusions: These results confirm in a prospective cohort the predictive impact of HLA-B44 supertype for patients treated with anti-PD(L)1 agent and suggest a potential benefit of combining checkpoint inhibitors in patients with other HLA supertypes (non-HLA-B44+).

P1.04-14 HLA B44 Supertype Associated with Less Favorable Neoantigen Binding in Non-Small Cell Lung Cancer Treated with Immunotherapy

P1.04-14 HLA B44 Supertype Associated with Less Favorable Neoantigen Binding in Non-Small Cell Lung Cancer Treated with Immunotherapy

Association of serum 25-hydroxyvitamin D concentration with HLA-B, -DRB1 and -DQB1 genetic polymorphisms.

The human leukocyte antigen (HLA) gene region associates with the risk for several autoimmune diseases, including type 1 diabetes. An association between vitamin D deficiency and several autoimmune diseases has been suggested. We tested the association between serum 25-hydroxyvitamin D (25OHD) concentrations and HLA alleles in pregnant Finnish women. HLA-B (n=395), HLA-DRB1 (n=501) and HLA-DQB1 (n=475) alleles were genotyped in pregnant women (mothers of children who later developed type 1 diabetes and mothers of non-diabetic children). HLA-B alleles were divided into supertypes that share similar peptide-binding specificity. Serum 25OHD concentration had been previously measured in these women from sera collected during the first trimester of pregnancy. Multiple testing was controlled for using the false discovery rate method. An association was found between 25OHD concentration and HLA-B44 supertype (P=0.009); women with HLA-B44 supertype (B*18, B*37, B*40 and B*44 alleles) had lower 25OHD concentrations. No association was found between HLA-DRB1 or -DQB1 alleles and 25OHD concentration. In this study we found for the first time an association between HLA genetic polymorphisms and 25OHD concentration. In future studies, the mechanistic background of this association and the role of vitamin D in the regulation of HLA gene expression should be investigated.

HLA Class I Supertype Associations With Clinical Outcome of Secondary Dengue Virus Infections in Ethnic Thais.

Human leukocyte antigen (HLA) supertypes are groups of functionally related alleles that present structurally similar antigens to the immune system. To analyze HLA class I supertype associations with clinical outcome in hospitalized Thai children with acute dengue illness. Seven hundred sixty-two patients and population-matched controls recruited predominantly in Bangkok were HLA-A and -B typed. HLA supertype frequencies were compared and tested for significant dengue disease associations using logistic regression analyses. Multivariable models were built by conducting forward stepwise selection procedures. In the final logistic regression model, the HLA-B44 supertype was protective against dengue hemorrhagic fever (DHF) in secondary infections (odds ratio [OR] = 0.46, 95% confidence interval [CI], .30-.72), while the HLA-A02 supertype (OR = 1.92, 95% CI, 1.30-2.83) and the HLA-A01/03 supertype (OR = 3.01, 95% CI, 1.01-8.92) were associated with susceptibility to secondary dengue fever. The B07 supertype was associated with susceptibility to secondary DHF in the univariate analysis (OR = 1.60, 95% CI, 1.05-2.46), whereas that was not retained in the final model. As the HLA-B44 supertype is predicted to target conserved epitopes in dengue, our results suggest that B44 supertype-restricted immune responses to highly conserved regions of the dengue proteome may protect against secondary DHF.

HLA specificities are related to development and prognosis of diffuse large B-cell lymphoma

AbstractDiffuse large B-cell lymphoma (DLBCL) is an aggressive disease influenced by genetic and environmental factors. The role of the HLA system in tumor antigen presentation could be involved in susceptibility and disease control. We analyzed the phenotypic frequencies of HLA-A, HLA-B, HLA-C, HLA-DRB1, and HLA-DQB1 in 250 DLBCLs, comparing them with 1940 healthy individuals. We also evaluated the influence of HLA polymorphisms on survival in those patients treated with curative intention using cyclophosphamide, doxorubicin, vincristine, and prednisolone (CHOP)-like regimen without (n = 64, 26%) or with (n = 153, 61%) rituximab. DLBCL patients have a higher phenotypic frequency of HLA-DRB1*01 (29% vs 19.5%, P = .0008, Pc = .0104) and a lower frequency of HLA-C*03 (6.4% vs 17.9%, P < .0005, Pc = .007) compared with healthy individuals. Irrespective of the age-adjusted International Prognostic Index, those patients receiving a CHOP-like plus rituximab regimen and carrying the HLA-B44 supertype had worse 5-year progression-free (54% vs 71%, P = .019) and 5-year overall (71% vs 92%, P = .001) survival compared with patients without this supertype. Our data suggest that some HLA polymorphisms influence the development and outcome of DLBCL, allowing the identification of an extremely good-risk prognostic subgroup. However, these results are preliminary and need to be validated in order to exclude a possible population effect.

Recognition of Class I MHC by a Rat Ly49 NK Cell Receptor Is Dependent on the Identity of the P2 Anchor Amino Acid of Bound Peptide

Members of the rodent Ly49 receptor family control NK cell responsiveness and demonstrate allele specificity for MHC class I (MHC-I) ligands. For example, the rat Ly49i2 inhibitory NK cell receptor binds RT1-A1(c) but not other rat MHC class Ia or Ib molecules. RT1-A1(c) preferentially binds peptides with proline at the second, or P2, position, which defines it as an HLA-B7 supertype MHC-I molecule. Previously, our laboratory showed that mutations within the MHC-I supertype-defining B-pocket of RT1-A1(c) could lead to alterations in P2 anchor residues of the peptide repertoire bound by RT1-A1(c) and loss of recognition by Ly49i2. Although suggestive of peptide involvement, it was unclear whether the peptide P2 anchor residue or alteration of the RT1-A1(c) primary sequence influenced Ly49i2 recognition. Therefore, we directly investigated the role of the P2 anchor residue of RT1-A1(c)-bound peptides in Ly49i2 recognition. First, fluorescent multimers generated by refolding soluble recombinant RT1-A1(c) with individual synthetic peptides differing only at the P2 anchor residue were examined for binding to Ly49i2 NK cell transfectants. Second, cytotoxicity by Ly49i2-expressing NK cells toward RMA-S target cells expressing RT1-A1(c) bound with peptides that only differ at the P2 anchor residue was evaluated. Our results demonstrate that Ly49i2 recognizes RT1-A1(c) bound with peptides that have Pro or Val at P2, whereas little or no recognition is observed when RT1-A1(c) is complexed with peptide bearing Gln at P2. Thus, the identity of the P2 peptide anchor residue is an integral component of MHC-I recognition by Ly49i2.

Functional analysis of frequently expressed Chinese rhesus macaque MHC class I molecules Mamu-A1*02601 and Mamu-B*08301 reveals HLA-A2 and HLA-A3 supertypic specificities

The Simian immunodeficiency virus (SIV)-infected Indian rhesus macaque (Macaca mulatta) is the most established model of HIV infection and AIDS-related research, despite the potential that macaques of Chinese origin is a more relevant model. Ongoing efforts to further characterize the Chinese rhesus macaques’ major histocompatibility complex (MHC) for composition and function should facilitate greater utilization of the species. Previous studies have demonstrated that Chinese-origin M. mulatta (Mamu) class I alleles are more polymorphic than their Indian counterparts, perhaps inferring a model more representative of human MHC, human leukocyte antigen (HLA). Furthermore, the Chinese rhesus macaque class I allele Mamu-A1*02201, the most frequent allele thus far identified, has recently been characterized and shown to be an HLA-B7 supertype analog, the most frequent supertype in human populations. In this study, we have characterized two additional alleles expressed with high frequency in Chinese rhesus macaques, Mamu-A1*02601 and Mamu-B*08301. Upon the development of MHC–peptide-binding assays and definition of their associated motifs, we reveal that these Mamu alleles share peptide-binding characteristics with the HLA-A2 and HLA-A3 supertypes, respectively, the next most frequent human supertypes after HLA-B7. These data suggest that Chinese rhesus macaques may indeed be a more representative model of HLA gene diversity and function as compared to the species of Indian origin and therefore a better model for investigating human immune responses.Electronic supplementary materialThe online version of this article (doi:10.1007/s00251-010-0502-8) contains supplementary material, which is available to authorized users.

A CD8+ T-Cell Clone Directed Against AML Blasts that Recognizes a Tumor Specific Antigen Expressed Also by Solid Tumors

AbstractAbstract 4289The aim of this study was to generate cytotoxic T-lymphocytes (CTL) clones directed against AML cells and to identify new immunogenic antigens with the following properties: i) to be overexpressed by leukemic cells and not by normal tissues; ii) to be shared among different AML subtypes; iii) to play a role in leukemic growth/survival; iv) to be expressed by leukemic stem cells.To this end, we loaded normal dendritic cells (DC) from a healthy donor with apoptotic bodies from primary AML cells and used loaded DC to stimulate autologous lymphocytes (i.e. lymphocytes from the donor). Donor was selected to be partially matched with the leukemic patient for MHC-class I, in that he shared two MHC-class I alleles at the HLA-supertype level (HLA-B7 and HLA-B44). With this strategy, a CD8+ T-cell line was generated that recognized both loaded DC and AML cells used for loading, but not DC loaded with normal cells derived from the patient (PHA-blasts). This CTL line was cloned by limiting dilutions and 180 clones were screened by IFN-g elispot against 2 different AML samples that were expressing both HLA-B7 and HLA-B44; one additional mismatched AML sample was used as negative control. Two clones were selected that recognized HLA-B7+/HLA-B44+ AML cells but not the negative control (namely, clone 31D3 and 8E12). To determine the HLA-restriction element and to verify shared antigen expression among AML subtypes, we tested both clones against a panel of 18 HLA-typed primary AML samples. We found that clone 31D3 was restricted by HLA-B7 and clone 8E12 by HLA-B44. Each clone recognized 5/5 HLA-matched AML samples of different subtype. In addition, the clones did not recognize both resting and activated normal myeloid, nor lymphoid and CD34+ cells expressing the proper HLA-restriction allele. In addition to elispot activity, both clones showed killing of AML cells in a CFSE-based cytotoxic assay. To confirm the lack of reactivity against normal tissues, we analyzed the activity of both clones against HLA matched or mismatched fibroblasts and we found that clone 8E12 displayed a low reactivity activity against normal matched fibroblasts, while clone 31D3 was not reactive.We then focused the analysis on clone 31D3 and tested whether its activity was restricted to leukemia or also directed against a panel of HLA-B7 positive (N=3) or negative (N=5) solid tumors. Interestingly, we found that the clone 31D3 could recognize one colon carcinoma and one melanoma cell line expressing the HLA-B7 supertype. In particular, the melanoma cell line (G4-mel) was HLA-B35+ (HLA-B35 belongs to the HLA-B7 supertype) and was recognized at very high levels. The availability of a tumor cell line expressing adequate amounts of antigen will make the generation of a cDNA library for antigen identification more feasible than with primary leukemic cells. Finally, to further confirm that HLA-B35 is the proper MHC restriction element and to determine to what extent the antigen is shared among different tumors, we transduced 8 tumor cell lines, 3 normal B-lymphoblastoid and 1 fibroblast cell line with the HLA-B35 allele. Clone 31D3 efficiently recognized 6/8 HLA-B35 transduced tumor cell lines (1 AML, 2 melanomas, 2 colon and 1 breast carcinoma) but none of the control cell lines.In conclusion, clone 31D3 is restricted by the HLA-B7 supertype, it recognizes an antigen that is expressed by leukemic cells and not by normal tissues and, most importantly, is shared among tumors of different histology. Since the vast majority of malignant cells tested (both primary cells and in vitro established cell lines) were targeted by this clone, the putative antigen might be a protein playing an important role in tumor growth or survival. We are now testing the activity of the clone against purified leukemic stem cells and attempting to identify this shared tumor antigen. Disclosures:No relevant conflicts of interest to declare.

Essential differences in ligand presentation and T cell epitope recognition among HLA molecules of the HLA‐B44 supertype

Human leukocyte antigens (HLA) have long been grouped into supertypes to facilitate peptide-based immunotherapy. Analysis of several hundreds of peptides presented by all nine antigens of the HLA-B44 supertype (HLA-B*18, B*37, B*40, B*41, B*44, B*45, B*47, B*49 and B*50) revealed unique peptide motifs for each of them. Taking all supertype members into consideration only 25 out of 670 natural ligands were found on more than one HLA molecule. Further direct comparisons by two mass spectrometric methods--isotope labeling as well as a label-free approach--consistently demonstrated only minute overlaps of below 3% between the ligandomes of different HLA antigens. In addition, T cell reactions of healthy donors against immunodominant HLA-B*44 and HLA-B*40 epitopes from EBV lacked promiscuous T-cell recognition within the HLA-B44 supertype. Taken together, these results challenge the common paradigm of broadly presented epitopes within this supertype.

35-OR: Large-scale sequencing of endogenous peptides from three members of the HLA-B7 supertype reveals novel shared peptide anchor positions

35-OR: Large-scale sequencing of endogenous peptides from three members of the HLA-B7 supertype reveals novel shared peptide anchor positions

Identification of immunogenic HLA-B7 “Achilles’ heel” epitopes within highly conserved regions of HIV

Genetic polymorphisms in class I human leukocyte antigen molecules (HLA) have been shown to determine susceptibility to HIV infection as well as the rate of progression to AIDS. In particular, the HLA-B7 supertype has been shown to be associated with high viral loads and rapid progression to disease. Using a multiplatform in silico/in vitro approach, we have prospectively identified 45 highly conserved, putative HLA-B7 restricted HIV CTL epitopes and evaluated them in HLA binding and ELISpot assays. All 45 epitopes (100%) bound to HLA-B7 in cell-based HLA binding assays: 28 (62%) bound with high affinity, 6 (13%) peptides bound with medium affinity and 11 (24%) bound with low affinity. Forty of the 45 peptides (88%) stimulated a IFN-gamma response in PBMC from at least one subject. Eighteen of these 40 epitopes have not been previously described; an additional eight epitopes have not been previously described as restricted by B7. The HLA-B7 restricted epitopes discovered using this in silico screening approach are highly conserved across strains and clades of HIV as well as conserved in the HIV genome over the 20 years since HIV-1 isolates were first sequenced. This study demonstrates that it is possible to select a broad range of HLA-B7 restricted epitopes that comprise stable elements in the rapidly mutating HIV genome. The most immunogenic of these epitopes will be included in the GAIA multi-epitope vaccine.

Cross-Species Dependence of Ly49 Recognition on the Supertype Defining B-Pocket of a Class I MHC Molecule

Ly49 recognition of MHC class I (MHC I) can be allele specific. However, the site of interaction on MHC I consists of highly conserved solvent-exposed amino acids, leaving it unclear how allele specificity occurs. In examining the specificity of mouse and rat Ly49, we noticed that MHC I ligands for mouse Ly49G and W, and the rat Ly49i2, typically share the HLA-B7 supertype, defined by a B-pocket that prefers a proline at position 2 in bound peptides. Through mutagenesis, we show that the supertype-defining B-pocket of RT1-A1(c) controls its allele-specific recognition by the syngeneic rat Ly49i2 inhibitory receptor and xenogeneic mouse inhibitory Ly49G and activating Ly49W receptors. Single amino acid substitutions in the B-pocket that did not prevent peptide binding disrupted Ly49 recognition. In contrast, single mutations in other regions of the peptide-binding groove had no effect. We provide a model whereby the B-pocket dictates the conformation of conserved residues at the Ly49 interaction site below, defining Ly49 allele specificity for MHC I. Therefore, at least some Ly49 may recognize supertypes, detectable even across species, and are sensitive to polymorphisms in the supertype-defining B-pocket. This would ensure that expression of specific MHC I supertypes capable of Ag presentation to T cells is sensed by NK cells, and if lacking, targets a cell for elimination, suggesting a supertype-mediated link between innate and adaptive immunity.

Immunogenic HLA-B7-restricted peptides of hTRT

Telomerase reverse transcriptase (TRT) is the first bona fide common tumor antigen. While several 9mer peptides of the human TRT have been identified for HLA-A2, little information exists on peptides for the remaining HLA types. Here, we used a multi-step approach to select and characterize a panel of HLA-B7 9mer peptides as candidate immunogens. In sequence, we used algorithm-based predictions, in vivo immunization of HLA-B7 transgenic (Tg) mice, in vitro immunization of human blood lymphocytes from two normal donors and two cancer patients, in vivo processing in HLA-B7 Tg mice and HLA-B7 supertype binding. We found a correlation between the in vivo immunogenicity and the actual HLA-B7 binding avidity of the seven predicted peptides. Furthermore, endogenous processing correlated with in vitro immunogenicity in human PBMC and HLA-B7 supertype binding. Peptide (1123)LPSDFKTIL(1131) (p1123) with the wider spectrum of supertype binding displayed the highest immunogenicity overall and was endogenously processed in several human lymphoblastoid cells. Since no single step of the screening/selection process could substitute for the whole approach, we conclude that the identification of MHC class I-restricted peptides for potential vaccination of cancer patients remains, by and large, an empirical process.

Identification of Vaccinia CD8 + T-Cell Epitopes Conserved among Vaccinia and Variola Viruses Restricted by Common MHC Class I Molecules, HLA-A2 or HLA-B7

Immunization with vaccinia virus results in long-lasting protection against smallpox and is an approach that has been successfully used to eliminate natural smallpox infections worldwide. Today, vaccinia virus is very important not only as a vaccine virus to protect human against smallpox, but also as an expression vector for immunization against other infectious diseases, such as HIV and cancer. In this article, we identify three new vaccinia human CD8 + T-cell epitopes conserved among vaccinia and variola viruses restricted by HLA-A2, HLA-B7, or HLA-B*3502, which belongs to the HLA-B7 supertype. Identification of these CD8 + T-cell epitopes restricted by common HLA alleles will help to quantitate human CD8 + T-cell responses to licensed and experimental smallpox vaccines and to vaccinia virus vectors. CD8 + T-cell responses specific to these epitopes can also be used to quantitate cellular immune responses, especially with new smallpox vaccines that do not induce a “take,” such as the modified vaccinia virus Ankara strain. Combined with previous reports by us and others, these results show that there are some vaccinia viral proteins containing multiple epitopes restricted by different MHC molecules of humans and mice.

Mycobacterium tuberculosisgenome-wide screen exposes multiple CD8+ T cell epitopes

Mounting evidence suggests human leucocyte antigen (HLA) class I-restricted CD8(+) T cells play a role in protective immunity against tuberculosis yet relatively few epitopes specific for the causative organism, Mycobacterium tuberculosis, are reported. Here a total genome-wide screen of M. tuberculosis was used to identify putative HLA-B*3501 T cell epitopes. Of 479 predicted epitopes, 13 with the highest score were synthesized and used to restimulate lymphocytes from naturally exposed HLA-B*3501 healthy individuals in cultured and ex vivo enzyme-linked immunospot (ELISPOT) assays for interferon (IFN)-gamma. All 13 peptides elicited a response that varied considerably between individuals. For three peptides CD8(+) T cell lines were expanded and four of the 13 were recognized permissively through the HLA-B7 supertype family. Although further testing is required we show the genome-wide screen to be feasible for the identification of unknown mycobacterial antigens involved in immunity against natural infection. While the mechanisms of protective immunity against M. tuberculosis infection remain unclear, conventional class I-restricted CD8(+) T cell responses appear to be widespread throughout the genome.

Impact of disparity of minor histocompatibility antigens HA-1, CD31, and CD49b in hematopoietic stem cell transplantation of patients with chronic myeloid leukemia with sibling and unrelated donors.

Despite human leukocyte antigen (HLA) identity between donor and recipient, several patients develop acute graft-versus-host disease (aGVHD) after hematopoetic stem cell transplantation (HSCT) because of minor histocompatibility antigen (mHag) incompatibilities. The impact of multiple mHag disparities on the clinical outcome after HSCT still remains to be determined. We studied the genomic polymorphisms of HA-1, CD31, and CD49b and correlated mHag distribution with the occurrence of aGVHD after HSCT from HLA-matched sibling and unrelated donors. All 163 patients examined in our single-center study underwent HSCT for chronic myeloid leukemia in the first chronic phase. HA-1 and CD31 disparities are associated with increased aGVHD incidence in a subgroup of patients who test HLA-B44 supertype positive in univariate analysis. However, in a multivariate analysis, only increased patient age was confirmed as an independent aGVHD risk factor. Our findings indicate that the impact of mHag disparity on aGVHD development in HSCT from HLA-matched sibling and unrelated donors seems to be subordinated to classic aGVHD risk factors.

Simultaneous prediction of binding capacity for multiple molecules of the HLA B44 supertype.

We selected for study a set of B44-supertype molecules collectively represented in >40% of the individuals in all major ethnicities (B*1801, B*4001, B*4002, B*4402, B*4403, and B*4501). The peptide-binding specificity of each molecule was characterized using single amino acid substitution analogues and nonredundant peptide libraries. In all cases, only peptide ligands with glutamic acid in position 2 were preferred. At the C terminus, each allele was associated with a unique but broad pattern of preferences, but all molecules tolerated hydrophobic/aliphatic (leucine, isoleucine, valine, methionine), aromatic (tyrosine, phenylalanine, tryptophan), and small (alanine, glycine, threonine) residues. Secondary anchor motifs were also defined for all molecules. Together, these features were used to define a B44 supermotif and a novel algorithm for calculating degeneracy scores that can be used to predict B44-supertype degenerate binders. Approximately 90% of the peptides with a B44 supermotif degeneracy score of >10 bound at least three of the six B44-supertype molecules studied with high affinity. Finally, a number of peptides derived from hepatitis B and C viruses, HIV, and Plasmodium falciparum have been identified that have degenerate B44 supertype-binding capacity. Taken together, these findings have important implications for epitope-based approaches to vaccination, immunotherapy, and the monitoring of immune responses.