MHC Alleles Research Articles

State of the art and challenges in sequence based T-cell epitope prediction.

Sequence based T-cell epitope predictions have improved immensely in the last decade. From predictions of peptide binding to major histocompatibility complex molecules with moderate accuracy, limited allele coverage, and no good estimates of the other events in the antigen-processing pathway, the field has evolved significantly. Methods have now been developed that produce highly accurate binding predictions for many alleles and integrate both proteasomal cleavage and transport events. Moreover have so-called pan-specific methods been developed, which allow for prediction of peptide binding to MHC alleles characterized by limited or no peptide binding data. Most of the developed methods are publicly available, and have proven to be very useful as a shortcut in epitope discovery. Here, we will go through some of the history of sequence-based predictions of helper as well as cytotoxic T cell epitopes. We will focus on some of the most accurate methods and their basic background.

Inflammatory arthritis ‐ an exciting confluence of human and animal research

Inflammatory arthritis ‐ an exciting confluence of human and animal research

Loss of Non-Inherited Maternal MHC and Materno-Fetal Transmission of p190 Type BCR-ABL Leukemia.

Abstract 980Poster Board I-2 Introduction:Materno-fetal transmission of malignant clone is one of the rare causes of leukemia in infants. It has been recorded some 20 times over the past 100 years but definitive, genetic evidence for a maternal origin has been lacking. Here we present a detailed genetic analysis of materno-fetal transmission in a case with p190 type BCR-ABL leukemia. Patients course:A mother was found to have p190 type BCR-ABL positive acute lymphoblastic leukemia (ALL) one month after delivery of a baby, and died during induction therapy by uncontrolled bacillus cereus encephalitis. Her baby developed p190 type BCR-ABL positive B-precursor lymphoblastic lymphoma (B-LBL stage3 jaw tumor and pleural effusion) at 11-month old. Non-Hodgkin's lymphoma type chemotherapy according to Japanese Pediatric Leukemia/Lymphoma Study Group (JPLSG) ALB-NHL03, followed by Hyper-CVAD imatinib therapy without stem cell transplantation were given to her, leading to complete remission for more than 31 months. Material and methods:We first cloned the BCR-ABL genomic breakpoint region from the infant's pleural effusion (PE). The breakpoint was designated as a fusion between BCR intron 1 and ABL intron1. Then, We conducted Q-PCR by using BCR-ABL genomic breakpoint specific primer and sequencing. Fifteen polymorphic STR markers were amplified in the paternal blood DNA and Patient's PE and PB DNA samples using the Powerplex-16 system. HLA serotyping was by microdroplet lymphocyte cytotoxity. Genotyping was carried out using a reversed SSO HLA DNA typing method. The infant jaw tumor was further analyzed by Affymetrix high resolusion (250K) SNP arrays. Results:Short tandem repeat profiles indicated that the infant jaw tumor was, unambiguously, of maternal origin. Amplification of BCR-ABL genomic breakpoints on maternal leukemic cells and infant lymphoma cells revealed that these leukemia clones shared the same unique BCR-ABL genomic sequence, indicating a shared, single cell origin. Positivity of BCR-ABL specific PCR on Guthrie card blood spot also indicated that the leukemic clone was present in the blood at birth. Additionally the infant, maternal-derived leukemic cells had uniparental disomy of chromosome 6p with a deletion of non-inherited maternal MHC allele (NIMA), suggesting a mechanism for immune evasion in the baby. Conclusions:This is the first case with definite evidence of in utero materno-fetal transmission of mother's leukemic cells. This case also suggests that materno-feto transfer of leukemic cells is more common than as reflected in the frequency of clinically diagnosed cases, where the immuno-surveillance, predominantly directed against major HLA locus antigens, may be the principal constraint. Disclosures:No relevant conflicts of interest to declare.

MHC diversity in bottlenecked populations: a simulation model

The depletion of variation at MHC loci, which play a crucial role in pathogen recognition, has been postulated to be one of important extinction risk factors for endangered populations. Thus, it is important to understand how selection affects the level of polymorphism in these genes when populations undergo a reduction in size. We followed MHC diversity in computer simulations of population bottlenecks. The fates of MHC alleles in the simulations were determined either by drift, or by balancing selection resulting from host–parasite coevolution. We found that the impact of selection on MHC polymorphism in bottlenecked populations was dependent upon the timescales involved. Initially, selection maintained lower number of alleles than drift, but after ~40 generations of hosts selection maintained higher MHC diversity, as compared to drift. The adverse effects of decreased MHC polymorphism on population viability may be, to some extent, compensated for if selection helps to retain MHC alleles which show high functional diversity, which should allow protection against a broader range of pathogens. Our simulation shows, however, that the mean divergence of alleles retained under selection in bottlenecked populations is not, on average, significantly higher than the divergence due to drift.

Modeling the repertoire of true tumor-specific MHC I epitopes in a human tumor.

DNA replication has a finite measurable error rate, net of repair, in all cells. Clonal proliferation of cancer cells leads therefore to accumulation of random mutations. A proportion of these mutational events can create new immunogenic epitopes that, if processed and presented by an MHC allele, may be recognized by the adaptive immune system. Here, we use probability theory to analyze the mutational and epitope composition of a tumor mass in successive division cycles and create a double Pölya model for calculating the number of truly tumor-specific MHC I epitopes in a human tumor. We deduce that depending upon tumor size, the degree of genomic instability and the degree of death within a tumor, human tumors have several tens to low hundreds of new, truly tumor-specific epitopes. Parenthetically, cancer stem cells, due to the asymmetry in their proliferative properties, shall harbor significantly fewer mutations, and therefore significantly fewer immunogenic epitopes. As the overwhelming majority of the mutations in cancer cells are unrelated to malignancy, the mutation-generated epitopes shall be specific for each individual tumor, and constitute the antigenic fingerprint of each tumor. These calculations highlight the benefits for personalization of immunotherapy of human cancer, and in view of the substantial pre-existing antigenic repertoire of tumors, emphasize the enormous potential of therapies that modulate the anti-cancer immune response by liberating it from inhibitory influences.

190 The Immune Response to HIV: Friend or Foe

Our work aims to investigate a newly discovered link between inflammatory events that characterize early stages of HIV infection and a link to an infected person's MHC alleles. Hypothesis: We propose that it is the inflammation characteristic of an innate immune response that is responsible for the loss of CD4+ T cells in HIV. Experiments and Results: We base this proposition on five observations: (1) There is evidence that, in the absence of functional T regulatory cells (Tregs), the impact of polyclonal B cell activation is chronic inflammation and the activation of αβT cells*; αβ T cells have been shown to kill CD4 T cells during HIV infection**; (2) It has been shown that as T reg numbers decline, there is a concomitant rise in viral load in the HIV-infected patient*** (3) We have discovered that the B cells from an HIV infected lymph node have high levels of an important self peptide on their surfaces that indicates the lymph node B cells have been non-specifically activated. (4) Our data suggests peptide loading of non-specifically-activated B cells with targeted peptides can stimulate the activity of Tregs in an MHC allele-dependent manner; and (5) Treatment with a mixture of targeted peptides, reduces viral load in approximately 25 to 30% of patients treated, a result potentially explained by differences in MHC between patients. Conclusions: The consequences of unraveling the link between inflammation, cellular activation, and MHC may result in understanding a mechanism that may lead to development of targeted peptides that can redirect the immune response to, and interfere with, the inflammation-dependent pathology of HIV. Future Directions: Our exciting findings suggest that, with the proper predictions, we can synthesize targeted peptides to treat people with all different types of MHC alleles providing an inexpensive and potentially efficacious biological therapy for HIV.

Many different Vbeta CDR3s can reveal the inherent MHC reactivity of germline-encoded TCR V regions.

We have hypothesized that in the prenegative selection TCR repertoire, many somatically generated complementary-determining region (CDR) 3 loops combine with evolutionarily selected germline Valpha/Vbeta CDR1/CDR2 loops to create highly MHC/peptide cross-reactive T cells that are subsequently deleted by negative selection. Here, we present a mutational analysis of the Vbeta CDR3 of such a cross-reactive T-cell receptor (TCR), YAe62. Most YAe62 TCRs with the mutant CDR3s became less MHC promiscuous. However, others with CDR3s unrelated in sequence to the original recognized even more MHC alleles than the original TCR. Most importantly, this recognition was still dependent on the conserved CDR1/CDR2 residues. These results bolster the idea that germline TCR V elements are inherently reactive to MHC but that this reactivity is fine-tuned by the somatically generated CDR3 loops.

Development of inflammatory arthritis in DR4 transgenic and wild type DBA/1J mice immunized with human fibrinogen (49.9)

Abstract The human MHC allele DR4 is associated with susceptibility for rheumatoid arthritis (RA). Citrullinated proteins, derived from the conversion of peptidyl-arginine to peptidyl-citrulline, are present in the joints of RA patients, who also uniquely produce high levels of anti-citrullinated protein antibodies (ACPAs). ACPAs enhance damage in animal models of autoimmune arthritis. DR4 molecules have been shown to preferentially bind citrullinated peptides over arginine controls. Recently, the DR4 allele was shown to be essential to develop arthritis in C57Bl/6 mice immunized with human citrullinated fibrinogen, but these mice demonstrated delayed arthritis at relatively low incidence and severity as compared to other arthritis models. To potentially obtain a more susceptible strain, we backcrossed the DR4 gene onto the DBA1/J strain. Following immunization with either human citrullinated or noncitrullinated fibrinogen, these mice developed inflammatory arthritis around day 24 regardless of whether they were DR4+ or DR4- littermates. Serum from both strains demonstrated ACPA generation as defined by reactivity to a citrullinated but not control peptide, as well as a splenic proliferative response to both citrullinated and noncitrullinated human, but not mouse, fibrinogen. The DR4 DBA/1J transgenic strain may provide a useful tool for the analysis of ACPA-related arthritis. Grant U19 AI146374

Epidermal Langerhans cells in actinic prurigo: a comparison between lesional and non-lesional skin

Actinic Prurigo (AP) is a chronic pruritic dermatosis of unknown cause affecting sun exposed skin in defined ethnic groups with characteristic MHC alleles. However, the cutaneous dendritic cells have not been assessed. To assess in situ the epidermal Langerhans Cell (LC) status in Actinic Prurigo. Fresh skin samples from three AP patients were used to evaluate in situ the epidermal LC, comparing lesional and non-lesional sites in each subject. AP patients attending the Dermatology Department at the Hospital M. Gea-Gonzalez in Mexico city. Lesional and non-lesional skin samples were taken from each subject to prepare both epidermal sheets and conventional tissue sections. Three markers restricted to LC in epidermis (CD1a, ATPase, MHC-II) were used to quantify the LC per area in epidermal sheets. Compared to non-lesional skin from the same subject, a significant reduction in the number of LC per area of epidermis was found in lesional skin; with any of the three markers evaluated. The frequency of epidermal LC decreases importantly in lesional skin from AP patients.

Mamu-A⁎01/K b transgenic and MHC Class I knockout mice as a tool for HIV vaccine development

We have developed a murine model expressing the rhesus macaque (RM) Mamu-A⁎01 MHC allele to characterize immune responses and vaccines based on antigens of importance to human disease processes. Towards that goal, transgenic (Tg) mice expressing chimeric RM (α1 and α2 Mamu-A⁎01 domains) and murine (α3, transmembrane, and cytoplasmic H-2K b domains) MHC Class I molecules were derived by transgenesis of the H-2K bD b double MHC Class I knockout strain. After immunization of Mamu-A⁎01/K b Tg mice with rVV-SIVGag–Pol, the mice generated CD8 + T-cell IFN-γ responses to several known Mamu-A⁎01 restricted epitopes from the SIV Gag and Pol antigen sequence. Fusion peptides of highly recognized CTL epitopes from SIV Pol and Gag and a strong T-help epitope were shown to be immunogenic and capable of limiting an rVV-SIVGag–Pol challenge. Mamu-A⁎01/K b Tg mice provide a model system to study the Mamu-A⁎01 restricted T-cell response for various infectious diseases which are applicable to a study in RM.

Overlapping synthetic peptides encoding TPD52 as breast cancer vaccine in mice: Prolonged survival

Peptide-based vaccines, one of several anti-tumor immunization strategies currently under investigation, can elicit both MHC Class I-restricted (CD8 +) and Class II-restricted (CD4 +) responses. However, the need to identify specific T-cell epitopes in the context of MHC alleles has hampered the application of this approach. We have tested overlapping synthetic peptides (OSP) representing a tumor antigen as a novel approach that bypasses the need for epitope mapping, since OSP contain all possible epitopes for both CD8 + and CD4 + T cells. Here we report that vaccination of inbred and outbred mice with OSP representing tumor protein D52 (TPD52-OSP), a potential tumor antigen target for immunotherapy against breast, prostate, and ovarian cancer, was safe and induced specific CD8 + and CD4 + T-cell responses, as demonstrated by development of specific cytotoxic T cell (CTL) activity, proliferative responses, interferon (IFN)-γ production and CD107a/b expression in all mice tested. In addition, TPD52-OSP-vaccinated BALB/c mice were challenged with TS/A breast carcinoma cells expressing endogenous TPD52; significant survival benefits were noted in vaccine recipients compared to unvaccinated controls ( p < 0.001). Our proof-of-concept data demonstrate the safety and efficacy of peptide library-based cancer vaccines that obviates the need to identify epitopes or MHC backgrounds of the vaccinees. We show that an OSP vaccination approach can assist in the disruption of self-tolerance and conclude that our approach may hold promise for immunoprevention of early-stage cancers in a general population.

High resolution human leukocyte antigen (HLA) class I and class II allele typing in Mexican mestizo women with sporadic breast cancer: case-control study

BackgroundThe development of breast cancer is multifactorial. Hormonal, environmental factors and genetic predisposition, among others, could interact in the presentation of breast carcinoma. Human leukocyte antigen (HLA) alleles play an important role in immunity (cellular immunity) and may be important genetic traits. HLAAllele-specific interaction has not been well established. Recently, several studies had been conducted in order to do so, but the results are controversial and in some instances contradictory.MethodsWe designed a case-control study to quantify the association of HLA class I and II genes and breast cancer. HLA typing was performed by high resolution sequence-specific oligotyping after DNA amplification (PCR-SSOP) of 100 breast cancer Mexican mestizo patients and 99 matched healthy controls.ResultsHLA-A frequencies that we were able to observe that there was no difference between both groups from the statistical viewpoint. HLA-B*1501 was found three times more common in the case group (OR, 3.714; p = 0.031). HLA-Cw is not a marker neither for risk, nor protection for the disease, because we did not find significant statistical differences between the two groups. DRB1*1301, which is expressed in seven cases and in only one control, observing an risk increase of up to seven times and DRB1*1602, which behaves similarly in being present solely in the cases (OR, 16.701; 95% CI, 0.947 – 294.670). DQ*0301-allele expression, which is much more common in the control group and could be protective for the presentation of the disease (OR, 0.078; 95% CI, 0.027–0.223, p = 0.00001).ConclusionOur results reveal the role of the MHC genes in the pathophysiology of breast cancer, suggesting that in the development of breast cancer exists a disorder of immune regulation. The triggering factor seems to be restricted to certain ethnic groups and certain geographical regions since the relevant MHC alleles are highly diverse. This is the first study in Mexican population where high resolutions HLA typing has been performed in order to try to establish an association with malignancy.

MHCBN 4.0: A database of MHC/TAP binding peptides and T-cell epitopes

BackgroundMany databases housing the information about MHC binders and non-binders have been developed in the past to help the scientific community working in the field of immunology, immune-informatics or vaccine design. As the information about these MHC binding and non-binding peptides continues to grow with the time and there is a need to keep the databases updated. So, in order to provide the immunological fraternity with the most recent information we need to maintain and update our database regularly. In this paper, we describe the updated version of 4.0 of the database MHCBN.FindingsMHCBN is a comprehensive database comprising over 25,857 peptide sequences (1053 TAP binding peptides), whose binding affinity with either MHC or TAP molecules has been assayed experimentally. It is a manually curated database where entries are collected & compiled from published literature and existing immunological public databases. MHCBN has a number of web-based tools for the analysis and retrieval of information like mapping of antigenic regions, creation of allele specific dataset, BLAST search, various diseases associated with MHC alleles etc. Further, all entries are hyper linked to major databases like SWISS-PROT, PDB etc. to provide the information beyond the scope of MHCBN. The latest version 4.0 of MHCBN has 6080 more entries than previously published version 1.1.ConclusionMHCBN database updating is meant to facilitate immunologist in understanding the immune system and provide them the latest information. We feel that our database will complement the existing databases in serving scientific community.

Safety, immunogenicity and efficacy of peptide‐pulsed cellular immunotherapy in macaques

Simple and effective delivery methods for cellular immunotherapies are needed. We recently published on the effectiveness of using ex vivo pulsing of overlapping SIV Gag 15mer peptides onto fresh peripheral blood cells in 32 SIV(mac251)-infected pigtail macaques. We now report on the safety of this approach, analysis of a novel assay for immunogenicity, the effect of an MHC allele, Mane-A*10, on CD8 T cell escape occurring and disease outcome. The vaccine strategy was safe, with no perturbations in weight or hematological profiles in comparison to controls. The high levels of SIV-specific T cell immunogenicity of this approach was confirmed using a novel assay measuring upregulation of surface CD134 of CD4 T cells. A substantial effect of the Mane-A*10 allele in reducing SIV viral load of pigtail macaques was observed in both vaccinees and controls; the virologic efficacy of the immunotherapy in comparison to controls was greatest in Mane-A*10- animals. Escape mutations at several new CD8 T cell epitopes throughout the SIV proteome were observed, primarily in animals with poorer virologic control. In summary, we provide further information that peptide-pulsed PBMC are a safe, immunogenic and effective immunotherapy. The observed influence of MHC alleles and immune escape allows us to design more insightful future immunotherapy studies.

Multispecific and heterogeneous recognition of the gag protein by cytotoxic T lymphocytes (CTL) from HIV-infected patients: factors other than the MHC control the epitopic specificities.

The HIV gag polyprotein is a major target for recognition by CTL in infected humans. Using recombinant vaccinia viruses (rVV) expressing truncations of the p24gag, and the p18gag, p15gag and HIV-2 p56gag proteins, the characterization of epitope regions recognized by in vitro-stimulated peripheral blood mononuclear cells (PBMC) from 18 infected patients has been studied. The gag-specific response of most individuals is polyclonal and multispecific, and interindividual variations between target epitope regions were frequently observed, despite shared MHC alleles. As CTL may play an important role in the control of HIV replication in infected hosts, these results have important implications for designing vaccine strategies.

Host Genetic Factors and Treatment of Hepatitis C

Infection with the hepatitis C virus (HCV) is a major health problem worldwide due to the associated risk of developing liver cirrhosis and its sequelae. Approximately 200 million persons are chronically infected worldwide. Furthermore, about one third of HIV-infected individuals in Europe and the US are co-infected with HCV. Currently pegylated interferon-alpha in combination with ribavirin represents the backbone of HCV-specific therapy. However, with interferon-based combination therapy sustained virologic response (SVR) is achieved in only about 50% of HCV-infected patients in clinical studies and may be even lower in clinical practice. HCV genotype and viral load are major determinants of treatment response in HCV infection. However, emerging data suggest host genetic factors also influence response to treatment. These data might hold the keys to better understand and predict outcome of HCV-specific therapy and might help to develop novel anti-HCV strategies. Here, we review the role of genetic aspects including the role of cytokines, chemokines/ chemokine receptors, and MHC alleles with respect to HCV therapy that have been elucidated so far and offer suggestions for how to use these observations as platforms for future research to further understand differential response to antiviral therapy in HCV-infected patients.

Host Genetic Factors and Treatment of Hepatitis C

Infection with the hepatitis C virus (HCV) is a major health problem worldwide due to the associated risk of developing liver cirrhosis and its sequelae. Approximately 200 million persons are chronically infected worldwide. Furthermore, about one third of HIV-infected individuals in Europe and the US are co-infected with HCV. Currently pegylated interferon-?? in combination with ribavirin represents the backbone of HCV-specific therapy. However, with interferon-based combination therapy sustained virologic response (SVR) is achieved in only about 50% of HCV-infected patients in clinical studies and may be even lower in clinical practice. HCV genotype and viral load are major determinants of treatment response in HCV infection. However, emerging data suggest host genetic factors also influence response to treatment. These data might hold the keys to better understand and predict outcome of HCV-specific therapy and might help to develop novel anti-HCV strategies. Here, we review the role of genetic aspects including the role of cytokines, chemokines/ chemokine receptors, and MHC alleles with respect to HCV therapy that have been elucidated so far and offer suggestions for how to use these observations as platforms for future research to further understand differential response to antiviral therapy in HCV-infected patients. Keywords: Hepatitis C, interferon α, therapy, gene polymorphism, HLA, interleukin, chemokine, cytokine

Widespread Unilateral Pain Associated With Herpes Simplex Virus Infections

We describe a patient group with unexplained widespread pain on one side of the body and pain exacerbations during active labial or genital herpes and during herpetic central nervous system infections. The patients had no visible lesion of the central nervous system on magnetic resonance imaging or abnormality in electrophysiological studies. To understand the nature of the pain and its possible relation to herpes simplex virus (HSV) infections, a clinical neurological examination was performed and quantitative sensory testing and skin biopsies were assessed in 17 patients. The levels of serum total immunoglobulins and IgG subclasses and the frequencies of the immune response genes at the IGH@, HLA-A, -B, -DRB1, C4A, and C4B loci were analyzed in the patients and in control subjects. The patients manifested a uniform clinical syndrome with unilateral pain that was best described as neuropathic and that was exacerbated by HSV reactivations. Low plasma IgG3 concentrations, the presence of either low plasma IgG1 or IgG3 or both, and high anti-HSV-2-IgG titers were more common in the patients than in the control subjects, which rendered the patients more vulnerable to HSV recurrences. We suggest that low immunoglobulin subclass levels and certain MHC alleles render the patients susceptible to recurring HSV infections. HSV reactivations and the accompanying inflammatory process cause dysfunction of the central nervous system that manifests as neuropathic pain. Studies using functional brain imaging are needed to clarify this syndrome.

Alloreactivity and Association of Human Natural Killer Cells with the Major Histocompatibility Complex

All NK cells potentially lytic for autologous cells but not expressing self-major histocompatibility complex (MHC)-reactive receptors could be eliminated by a negative selection mechanism during ontogeny. This idea is based on the existence of a NK cell subset expressing a specific inhibitory receptor for allogeneic MHC alleles. As ancestral haplotypes of the MHC appear to define identical MHC haplotypes in unrelated individuals, unrelated individuals having the same ancestral haplotype should also have the same NK-defined allospecificities that have been shown to map to the human MHC. To test this prediction, multiple cell lines from unrelated individuals having the same ancestral haplotypes were tested for the NK-defined allospecificities. It was found that cells having the same ancestral haplotypes do have the same NK-defined specificities. Furthermore, the NK-defined phenotype of cells that possess two different ancestral haplotypes can be predicted from the NK-defined phenotypes of unrelated cells that are homozygous for the ancestral haplotypes concerned. Although the group 1 and 2 NK-defined allospecificities can be explained to some extent by HLA-C alleles, evidence is presented that additional genes may modify the phenotype conferred by HLA-C.

Regulation of the NK Cell Alloreactivity to Bone Marrow Cells by the Combination of the Host NK Gene Complex and MHC Haplotypes

Host NK cells can reject MHC-incompatible (allogeneic) bone marrow cells (BMCs), suggesting their effective role for graft-vs leukemia effects in the clinical setting of bone marrow transplantation. NK cell-mediated rejection of allogeneic BMCs is dependent on donor and recipient MHC alleles and other factors that are not yet fully characterized. Whereas the molecular mechanisms of allogeneic MHC recognition by NK receptors have been well studied in vitro, guidelines to understand NK cell allogeneic reactivity under the control of multiple genetic components in vivo remain less well understood. In this study, we use congenic mice to show that BMC rejection is regulated by haplotypes of the NK gene complex (NKC) that encodes multiple NK cell receptors. Most importantly, host MHC differences modulated the NKC effect. Moreover, the NKC allelic differences also affected the outcome of hybrid resistance whereby F1 hybrid mice reject parental BMCs. Therefore, these data indicate that NK cell alloreactivity in vivo is dependent on the combination of the host NKC and MHC haplotypes. These data suggest that the NK cell self-tolerance process dynamically modulates the NK cell alloreactivity in vivo.