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1.
Immunohorizons ; 8(6): 415-430, 2024 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-38885041

RESUMO

The individual HLA-related susceptibility to emerging viral diseases such as COVID-19 underscores the importance of understanding how HLA polymorphism influences peptide presentation and T cell recognition. Similar to HLA-A*0101, which is one of the earliest identified HLA alleles among the human population, HLA-A*2601 possesses a similar characteristic for the binding peptide and acts as a prevalent allomorph in HLA-I. In this study, we found that, compared with HLA-A*0101, HLA-A*2601 individuals exhibit distinctive features for the T cell responses to SARS-CoV-2 and influenza virus after infection and/or vaccination. The heterogeneous T cell responses can be attributed to the distinct preference of HLA-A*2601 and HLA-A*0101 to T cell epitope motifs with negative-charged residues at the P1 and P3 positions, respectively. Furthermore, we determined the crystal structures of the HLA-A*2601 complexed to four peptides derived from SARS-CoV-2 and human papillomavirus, with one structure of HLA-A*0101 for comparison. The shallow pocket C of HLA-A*2601 results in the promiscuous presentation of peptides with "switchable" bulged conformations because of the secondary anchor in the median portion. Notably, the hydrogen bond network formed between the negative-charged P1 anchors and the HLA-A*2601-specific residues lead to a "closed" conformation and solid placement for the P1 secondary anchor accommodation in pocket A. This insight sheds light on the intricate relationship between HLA I allelic allomorphs, peptide binding, and the immune response and provides valuable implications for understanding disease susceptibility and potential vaccine design.


Assuntos
COVID-19 , Epitopos de Linfócito T , SARS-CoV-2 , Humanos , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/genética , SARS-CoV-2/imunologia , SARS-CoV-2/genética , COVID-19/imunologia , COVID-19/virologia , Antígenos HLA-A/imunologia , Antígenos HLA-A/genética , Antígenos HLA-A/metabolismo , Antígenos HLA-A/química , Peptídeos/imunologia , Peptídeos/química , Alelos , Antígeno HLA-A1
2.
J Biomed Sci ; 29(1): 80, 2022 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-36224625

RESUMO

BACKGROUND: Human Papillomavirus type 18 (HPV18) is a high-risk HPV that is commonly associated with cervical cancer. HPV18 oncogenes E6 and E7 are associated with the malignant transformation of cells, thus the identification of human leukocyte antigen (HLA)-restricted E6/E7 peptide-specific CD8 + T cell epitopes and the creation of a HPV18 E6/E7 expressing cervicovaginal tumor in HLA-A2 transgenic mice will be significant for vaccine development. METHODS: In the below study, we characterized various human HLA class I-restricted HPV18 E6 and E7-specific CD8 + T cells mediated immune responses in HLA class I transgenic mice using DNA vaccines encoding HPV18E6 and HPV18E7. We then confirmed HLA-restricted E6/E7 specific CD8 + T cell epitopes using splenocytes from vaccinated mice stimulated with HPV18E6/E7 peptides. Furthermore, we used oncogenic DNA plasmids encoding HPV18E7E6(delD70), luciferase, cMyc, and AKT to create a spontaneous cervicovaginal carcinoma model in HLA-A2 transgenic mice. RESULTS: Therapeutic HPV18 E7 DNA vaccination did not elicit any significant CD8 + T cell response in HLA-A1, HLA-24, HLA-B7, HLA-B44 transgenic or wild type C57BL/6 mice, but it did generate a strong HLA-A2 and HLA-A11 restricted HPV18E7-specific CD8 + T cell immune response. We found that a single deletion of aspartic acid (D) at location 70 in HPV18E6 DNA abolishes the presentation of HPV18 E6 peptide (aa67-75) by murine MHC class I. We found that the DNA vaccine with this mutant HPV18 E6 generated E6-specific CD8 + T cells in HLA-A2. HLA-A11, HLA-A24 and HLA-b40 transgenic mice. Of note, HLA-A2 restricted, HPV18 E7 peptide (aa7-15)- and HPV18 E6 peptide (aa97-105)-specific epitopes are endogenously processed by HPV18 positive Hela-AAD (HLA-A*0201/Dd) cells. Finally, we found that injection of DNA plasmids encoding HPV18E7E6(delD70), AKT, cMyc, and SB100 can result in the development of adenosquamous carcinoma in the cervicovaginal tract of HLA-A2 transgenic mice. CONCLUSIONS: We characterized various human HLA class I-restricted HPV18 E6/E7 peptide specific CD8 + T cell epitopes in human HLA class I transgenic mice. We demonstrated that HPV18 positive Hela cells expressing chimeric HLA-A2 (AAD) do present both HLA-A2-restricted HPV18 E7 (aa7-15)- and HPV18 E6 (aa97-105)-specific CD8 + T cell epitopes. A mutant HPV18E6 that had a single deletion at location 70 obliterates the E6 presentation by murine MHC class I and remains oncogenic. The identification of these human MHC restricted HPV antigen specific epitopes as well as the HPV18E6/E7 expressing adenosquamous cell carcinoma model may have significant future translational potential.


Assuntos
Carcinoma Adenoescamoso , Proteínas Oncogênicas Virais , Infecções por Papillomavirus , Vacinas de DNA , Animais , Ácido Aspártico , Linfócitos T CD8-Positivos , Carcinoma Adenoescamoso/complicações , Epitopos de Linfócito T/genética , Feminino , Antígenos HLA-A , Antígeno HLA-A1 , Antígeno HLA-A11 , Antígeno HLA-A2/genética , Antígeno HLA-A24 , Antígeno HLA-B40 , Antígeno HLA-B44 , Antígeno HLA-B7 , Células HeLa , Papillomavirus Humano 18 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas Oncogênicas Virais/genética , Infecções por Papillomavirus/complicações , Peptídeos , Proteínas Proto-Oncogênicas c-akt , Linfócitos T Citotóxicos , Vacinas de DNA/genética
3.
Hepatology ; 74(4): 2032-2046, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-33971035

RESUMO

BACKGROUND AND AIMS: Genetic predisposition to autoimmune hepatitis (AIH) in adults is associated with possession of human leukocyte antigen (HLA) class I (A*01, B*08) and class II (DRB1*03, -04, -07, or -13) alleles, depending on geographic region. Juvenile autoimmune liver disease (AILD) comprises AIH-1, AIH-2, and autoimmune sclerosing cholangitis (ASC), which are phenotypically different from their adult counterparts. We aimed to define the relationship between HLA profile and disease course, severity, and outcome in juvenile AILD. APPROACH AND RESULTS: We studied 236 children of European ancestry (152 female [64%], median age 11.15 years, range 0.8-17), including 100 with AIH-1, 59 with AIH-2, and 77 with ASC. The follow-up period was from 1977 to June 2019 (median 14.5 years). Class I and II HLA genotyping was performed using PCR/sequence-specific primers. HLA B*08, -DRB1*03, and the A1-B8-DR3 haplotype impart predisposition to all three forms of AILD. Homozygosity for DRB1*03 represented the strongest risk factor (8.8). HLA DRB1*04, which independently confers susceptibility to AIH in adults, was infrequent in AIH-1 and ASC, suggesting protection; and DRB1*15 (DR15) was protective against all forms of AILD. Distinct HLA class II alleles predispose to the different subgroups of juvenile AILD: DRB1*03 to AIH-1, DRB1*13 to ASC, and DRB1*07 to AIH-2. Possession of homozygous DRB1*03 or of DRB1*13 is associated with fibrosis at disease onset, and possession of these two genes in addition to DRB1*07 is associated with a more severe disease in all three subgroups. CONCLUSIONS: Unique HLA profiles are seen in each subgroup of juvenile AILD. HLA genotype might be useful in predicting responsiveness to immunosuppressive treatment and course.


Assuntos
Colangite Esclerosante/genética , Hepatite Autoimune/genética , População Branca/genética , Adolescente , Criança , Pré-Escolar , Feminino , Predisposição Genética para Doença , Antígenos HLA/genética , Antígeno HLA-A1/genética , Antígeno HLA-B8/genética , Antígeno HLA-DR3/genética , Cadeias HLA-DRB1/genética , Humanos , Lactente , Masculino , Índice de Gravidade de Doença
4.
Nat Commun ; 11(1): 5332, 2020 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-33087697

RESUMO

Cytotoxic T lymphocyte (CTL)-based cancer immunotherapies have shown great promise for inducing clinical regressions by targeting tumor-associated antigens (TAA). To expand the TAA landscape of pancreatic ductal adenocarcinoma (PDAC), we performed tandem mass spectrometry analysis of HLA class I-bound peptides from 35 PDAC patient tumors. This identified a shared HLA-A*0101 restricted peptide derived from co-transcriptional activator Vestigial-like 1 (VGLL1) as a putative TAA demonstrating overexpression in multiple tumor types and low or absent expression in essential normal tissues. Here we show that VGLL1-specific CTLs expanded from the blood of a PDAC patient could recognize and kill in an antigen-specific manner a majority of HLA-A*0101 allogeneic tumor cell lines derived not only from PDAC, but also bladder, ovarian, gastric, lung, and basal-like breast cancers. Gene expression profiling reveals VGLL1 as a member of a unique group of cancer-placenta antigens (CPA) that may constitute immunotherapeutic targets for patients with multiple cancer types.


Assuntos
Antígenos de Neoplasias/imunologia , Neoplasias da Mama/imunologia , Proteínas de Ligação a DNA/imunologia , Neoplasias Pancreáticas/imunologia , Fatores de Transcrição/imunologia , Antígenos de Neoplasias/genética , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Neoplasias da Mama/genética , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/imunologia , Carcinoma Ductal Pancreático/terapia , Linhagem Celular Tumoral , Citotoxicidade Imunológica , Proteínas de Ligação a DNA/genética , Feminino , Perfilação da Expressão Gênica , Antígeno HLA-A1/imunologia , Humanos , Imunoterapia Adotiva , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/terapia , Placenta/imunologia , Gravidez , Prognóstico , Linfócitos T Citotóxicos/imunologia , Fatores de Transcrição/genética , Neoplasias Pancreáticas
5.
Clin Transplant ; 34(12): e14110, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33053214

RESUMO

Human leukocyte antigen (HLA) class I presentation pathway plays a central role in natural killer (NK) cell and cytotoxic T-cell activities against BK polyomavirus (BKPyV) DNAemia. We determined the risk of sustained BKPyV DNAemia in 175 consecutive renal transplant recipients considering the simultaneous effect of donor/recipient HLA class I antigens and pre- or post-transplant variables. Median (IQR) age was 53 (44-64) years, and 37% of patients were female. 40 patients (22.9%) developed sustained BKPyV DNAemia [median (IQR) viral load: 9740 (4350-17 125) copies/ml]. In the Cox proportional hazard analysis, HLA-A1 (HR: 3.06, 95% CI: 1.51-6.17) and HLA-B35-Cw4 (HR: 4.63, 95% CI: 2.12-10.14) significantly increased the risk of sustained BKPyV DNAemia, while 2 HLA-C mismatches provided a marginally protective effect (HR: 0.32, 95% CI: 0.10-0.98). HLA-Cw4 is a ligand for NK cell inhibitory receptor, and HLA-B35 is in strong linkage disequilibrium with the HLA-Cw4 allele. The association between HLA-B35-Cw4 expression and sustained BKPyV DNAemia supports the important role of cytotoxic T cells and NK cells that would normally control BKPyV activation through engagement with immunoglobulin-like killer receptors (KIRs). Further studies are required to investigate the effect of HLA-C alleles along with NK cell activity against BKPyV DNAemia.


Assuntos
Vírus BK , Transplante de Rim , Infecções por Polyomavirus , Adulto , Vírus BK/genética , Feminino , Antígeno HLA-A1 , Humanos , Transplante de Rim/efeitos adversos , Masculino , Pessoa de Meia-Idade , Infecções por Polyomavirus/etiologia , Transplantados
6.
Virol J ; 17(1): 128, 2020 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-32831108

RESUMO

BACKGROUND: Heterozygosity at HLA class I loci is generally considered beneficial for host defense. We report here an element of HLA class I homozygosity that may or may not help preserve its existence in populations but which could indicate a new avenue for antiviral research. METHODS: Lymphocytes from serologically HLA-homozygous or -heterozygous donors were examined for synthesis of influenza virus proteins and RNA after exposure to virus as peripheral blood mononuclear cells. The virus-exposed lymphocytes were also examined for internalization of the virus after exposure, and for susceptibility to virus-specific cytotoxic T lymphocytes in comparison with virus-exposed monocytes/macrophages and unseparated peripheral blood mononuclear cells. Results were compared using two-tailed Fisher's exact test. RESULTS: Serologically-defined HLA-A2-homozygous lymphocytes, in contrast to heterozygous lymphocytes, did not synthesize detectable influenza virus RNA or protein after exposure to the virus. HLA-A2-homozygous lymphocytes, including both homozygous and heterozygous donors by genetic sequence subtyping, did internalize infectious virus but were not susceptible to lysis by autologous virus-specific cytotoxic T lymphocytes ("fratricide"). Similar intrinsic resistance to influenza virus infection was observed with HLA-A1- and HLA-A11-homozygous lymphocytes and with HLA-B-homozygous lymphocytes. CONCLUSIONS: A significant proportion of individuals within a population that is characterized by common expression of HLA class I alleles may possess lymphocytes that are not susceptible to influenza virus infection and thus to mutual virus-specific lysis. Further study may identify new approaches to limit influenza virus infection.


Assuntos
Genes MHC Classe I/imunologia , Influenza Humana/genética , Influenza Humana/imunologia , Macrófagos/virologia , Linfócitos T Citotóxicos/imunologia , Alelos , Feminino , Antígeno HLA-A1/imunologia , Antígeno HLA-A11/imunologia , Antígeno HLA-A2/imunologia , Homozigoto , Humanos , Leucócitos Mononucleares/virologia , Macrófagos/imunologia , Masculino
7.
Immunogenetics ; 72(3): 143-153, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31970435

RESUMO

Specificity analyses of peptide binding to human leukocyte antigen (HLA)-A molecules have been hampered due to a lack of proper monoclonal antibodies (mAbs) for certain allomorphs, such as the prevalent HLA-A1 for Caucasians and HLA-A11 for Asians. We developed a mAb that recognizes a conformational epitope common to most HLA-A allomorphs. The mAb, named A-1, does not discriminate peptides by amino acid sequences, making it suitable for measuring peptide binding. A stabilization assay using TAP-deficient cell lines and A-1 was developed to investigate the specificity of peptide binding to HLA-A molecules. Regarding the evolution of HLA-A genes, the A-1 epitope has been conserved among most HLA-A allomorphs but was lost when the HLA-A gene diversified into the HLA-A*32, HLA-A*31, and HLA-A*33 lineages together with HLA-A*29 after bifurcating from the HLA-A*25 and HLA-A*26 branchs. The establishment of A-1 is expected to help researchers investigate the peptide repertoire and develop computational tools to identify cognate peptides. Since no HLA-A locus-specific mAb has been available, A-1 will also be useful for analyzing the locus-specific regulation of the HLA gene expression.


Assuntos
Anticorpos Monoclonais/metabolismo , Antígenos HLA-A/imunologia , Antígeno HLA-A1/imunologia , Sequência de Aminoácidos , Anticorpos Monoclonais/imunologia , Linhagem Celular Tumoral , Epitopos/imunologia , Antígenos HLA-A/química , Antígeno HLA-A1/química , Humanos , Modelos Moleculares , Peptídeos/imunologia , Ligação Proteica/imunologia , Conformação Proteica
8.
Int J Nanomedicine ; 14: 2069-2089, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30988609

RESUMO

PURPOSE: Melanoma is the most aggressive form of skin cancer. Chemotherapy at a late stage fails due to low accumulation in tumors, indicating the need for targeted therapy. MATERIALS AND METHODS: To increase drug uptake by tumor cells, we have targeted doxorubicin-containing liposomes using a T-cell receptor (TCR)-like antibody (scFv G8 and Hyb3) directed against melanoma antigen A1 (MAGE-A1) presented by human leukocyte antigen A1 (M1/A1). With the use of flow cytometry and confocal microscopy, we have tested our formulation in vitro. In vivo pharmacokinetics was done in tumor-free nu/nu mice, while biodistribution and efficacy study was done in nu/nu mice xenograft. RESULTS: We demonstrated two to five times higher binding and internalization of these immunoliposomes by M1+/A1+ melanoma cells in vitro in comparison with nontargeted liposomes. Cytotoxicity assay showed significant tumor cell kill at 10 µM doxorubicin (DXR) for targeted vs nontargeted liposomes. In vivo pharmacokinetics of nontargeted and targeted liposomes were similar, while accumulation of targeted liposomes was 2- to 2.5-fold and 6.6-fold enhanced when compared with nontargeted liposomes and free drug, respectively. Notably, we showed a superior antitumor activity of MAGE-A1-targeted DXR liposomes toward M1+/A1+ expressing tumors in mice compared with the treatment of M1-/A1+ tumors. Our results indicate that targeted liposomes showed better cytotoxicity in vitro and pharmacokinetics in vivo. CONCLUSION: Liposomes decorated with TCR-mimicking scFv antibodies effectively and selectively target antigen-positive melanoma. We showed that DXR-loaded liposomes coupled to anti-M1/-A1 scFv inflict a significant antitumor response. Targeting tumor cells specifically promotes internalization of drug-containing nanoparticles and may improve drug delivery and ultimately antitumor efficacy. Our data argue that targeting MAGE in A1 context, by nanosized carriers decorated with TCR-like antibodies mimicking scFv, can be used as a theragnostic platform for drug delivery, immunotherapy, and potentially imaging, and diagnosis of melanoma.


Assuntos
Apresentação de Antígeno/imunologia , Doxorrubicina/administração & dosagem , Sistemas de Liberação de Medicamentos , Antígeno HLA-A1/imunologia , Lipossomos/administração & dosagem , Melanoma/tratamento farmacológico , Nanopartículas/administração & dosagem , Animais , Antibióticos Antineoplásicos/administração & dosagem , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/farmacocinética , Doxorrubicina/química , Doxorrubicina/farmacocinética , Humanos , Lipossomos/química , Lipossomos/imunologia , Melanoma/imunologia , Camundongos Nus , Nanopartículas/química , Receptores de Antígenos de Linfócitos T/imunologia , Anticorpos de Cadeia Única/imunologia , Distribuição Tecidual , Células Tumorais Cultivadas
9.
Arch Dermatol Res ; 311(4): 277-285, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30826962

RESUMO

To verify whether PsA-associated HLA alleles proposed in other populations are also related to PsA in Chinese Han population, a study of PsA susceptible alleles in the HLA-A, HLA-B, HLA-C and HLA-DRB1 alleles was presented for Chinese Han population. Genotyping was performed by Illumina Miseq platform (Illumina, USA). 50 subtypes and 77 subtypes of HLA-A, HLA-B, HLA-C and HLA-DRB1 with minor allele frequency (MAF) > 1% were genotyped from two-digit and four-digit resolution analysis in 111 PsA and 207 HCs (healthy controls) collected from Chinese Han population, respectively. Data handling, quality control and association analysis were performed using SPSS 25.0 software. In risk estimate, by mean of Bonferroni correction, a newfound four-digit allele HLA-A*01:01 [P = 5.5 × 10-4, OR 3.35 (1.69-6.66)], four-digit allele HLA-C*06:02 [P = 8.5 × 10-7, OR 3.80 (2.23-6.47)] and six two-digit alleles HLA-A*01 [P = 5.2 × 10-5, OR 3.43 (1.89-6.23)], HLA-B*13 [P = 4.0 × 10-6, OR 2.65 (1.76-4.01)], HLA-B*27 [P = 7.5 × 10-4, OR 5.84 (2.09-16.29)], HLA-B*57 [P = 5.8 × 10-5, OR 20.10 (4.65-86.83)], HLA-C*03 [P = 2.1 × 10-4, OR 0.40 (0.25-0.65)], HLA-C*06 [P = 1.9 × 10-12, OR 4.48 (2.95-6.81)] showed statistical significance by the univariate binary logistic regression analysis. Besides, in the binary logistic regression analysis with multiple variables, when the two alleles HLA-A*01:01 and HLA-C*06:02 were considered as covariates, HLA-A*01:01 [P = 2.7 × 10-3,OR 2.95 (1.46-5.98)] also showed significant association for PsA as risk factor, but may be not the main risk factor [HLA-C*06:02, P = 3.0 × 10-6, OR 3.68 (2.13-6.37)]. When all the above two-digit alleles were included as covariates, HLA-A*01 [P = 4.8 × 10-2, OR 2.00 (1.01-3.94)], HLA-B*13 [P = 4.2 × 10-5, OR 2.62 (1.65-4.16)], HLA-B*27 [P = 1.7 × 10-4, OR 7.62 (2.64-21.96)], HLA-B*57 [P = 2.97 × 10-4, OR 15.90 (3.55-71.18)], HLA-C*06 [P = 6.1 × 10-5, OR 2.70 (1.66-4.40)] showed significant for PsA as risk factors, HLA-C*03 [OR 0.65 (0.39-1.09), P = 0.10] showed no association with PsA. In conclusion, we assessed HLA-A, HLA-B, HLA-C and HLA-DRB1 alleles in PsA cohort of Chinese Han population, found HLA-A*01:01 and HLA-A*01 may be the susceptible genes associated with PsA, and also confirmed the association of four loci with PsA in Chinese Han population. These findings may extend the susceptibility HLA alleles of PsA and help in developing possible genetic markers to predict PsA.


Assuntos
Artrite Psoriásica/genética , Genótipo , Antígeno HLA-A1/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , China , Estudos de Coortes , Feminino , Frequência do Gene , Estudos de Associação Genética , Marcadores Genéticos , Predisposição Genética para Doença , Humanos , Complexo Principal de Histocompatibilidade/genética , Masculino , Pessoa de Meia-Idade , Polimorfismo Genético , Risco , Adulto Jovem
11.
Sci Rep ; 9(1): 842, 2019 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-30696911

RESUMO

Cell surface antigen discovery is of great interest for biomedical research both for isolation of rare cell populations and therapeutic targeting. We developed a rapid, cost-effective, fully in vitro technology which facilities the simultaneous target discovery and human antibody generation on the surface of virtually any cell population of interest. We apply our technique to human colorectal cancer-initiating cells (CICs) and identify hundreds of unique human antibodies. We characterized the top three antibody candidates targeting these CICs and identify their protein targets as integrin α7 (ITGA7), HLA-A1 and integrin ß6 (ITGB6). We demonstrate that these antibodies can be used to isolate self-renewing colorectal CICs, and that the integrin α7 antibody can prospectively identify glioblastoma brain tumor initiating cells as well as human muscle stem cells. We also demonstrate that genetic ablation of integrin ß6 impedes colorectal CIC function. The methodology can be readily applied to other cell populations including stem cells, cancer, or immune cells to facilitate the rapid identification of novel targets and simultaneous generation of potent and specific antibodies with therapeutic potential.


Assuntos
Anticorpos/imunologia , Antígenos de Superfície/imunologia , Biomarcadores Tumorais/imunologia , Neoplasias Colorretais/imunologia , Glioblastoma/imunologia , Antígenos CD/imunologia , Células CACO-2 , Células HCT116 , Células HEK293 , Antígeno HLA-A1/imunologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Cadeias alfa de Integrinas/imunologia , Cadeias beta de Integrinas/genética , Cadeias beta de Integrinas/imunologia , Células MCF-7 , Células PC-3 , Interferência de RNA , RNA Interferente Pequeno/genética , Células Tumorais Cultivadas
12.
Int J Immunogenet ; 46(1): 31-37, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30474304

RESUMO

The opportunity for the highly efficient recovery of immune receptor recombination data from cancer specimens, including the ready assessment of immune receptor V and J usage, raises the issue of establishing precise values of assessing the immune receptor status as opposed to obtaining basic information regarding lymphocyte infiltration, in the cancer setting. In this report, we obtained the lymphocyte infiltration percentages from the cancer digital slide archive representing uterine corpus endometrial carcinoma (UCEC) and correlated these data with recovery of the immune receptor recombination reads from corresponding UCEC exome files. Results indicated a basic correlation of the recovery of productive T-cell receptor beta (TRB) recombination reads with lymphocyte infiltration percentages. However, the recovery of specific immune receptor recombination reads did not indicate the same survival outcomes as microscope detection of lymphocyte infiltrate percentages. To further exploit the value of recovery of the TRB recombination reads from the UCEC exome files, we determined the survival outcomes for combinations of TRB gene segment usage and HLA class I alleles, with the most important result being that the combination of HLA-A*01:01 and TRB-J1 segment usage reflected a strikingly high survival rate. Overall, this report emphasized the increased value of the knowledge of the immune receptor recombinations, in comparison with basic lymphocyte infiltration percentages, in assessing cancer survival rates.


Assuntos
Neoplasias do Endométrio/genética , Antígeno HLA-A1/genética , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Adulto , Idoso , Alelos , Intervalo Livre de Doença , Neoplasias do Endométrio/epidemiologia , Neoplasias do Endométrio/patologia , Exoma/genética , Feminino , Humanos , Estimativa de Kaplan-Meier , Linfócitos/patologia , Pessoa de Meia-Idade
13.
J Immunol ; 202(2): 451-459, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30559321

RESUMO

The intracellular pathway of cross-presentation, which allows MHC class I-restricted presentation of peptides derived from exogenous Ags, remains poorly defined and may vary with the nature of the exogenous Ag and the type of APC. It can be cytosolic, characterized by proteasome and TAP dependency, or vacuolar, usually believed to be proteasome and TAP independent. Cross-presentation is particularly effective with long synthetic peptides, and we previously reported that the HLA-A2-restricted cross-presentation of a long peptide derived from melanoma Ag gp100 by human monocyte-derived immature dendritic cells occurred in a vacuolar pathway, making use of newly synthesized HLA-A2 molecules that follow a nonclassical secretion route. In this article, we show that the HLA-A1-restricted cross-presentation of a long peptide derived from tumor Ag MAGE-A3 by human monocyte-derived immature dendritic cells also follows a vacuolar pathway. However, as opposed to the HLA-A2-restricted peptide, cross-presentation of the HLA-A1-restricted peptide is TAP dependent. We show that this paradoxical TAP-dependency is indirect and reflects the need for TAP to load HLA-A1 molecules with peptides in the endoplasmic reticulum, to allow them to escape the endoplasmic reticulum and reach the vacuole, where peptide exchange with the cross-presented peptide likely occurs. Our results confirm and extend the involvement of the vacuolar pathway in the cross-presentation of long peptides, and indicate that TAP-dependency can no longer be used as a key criterion to distinguish the cytosolic from the vacuolar pathway of cross-presentation. They also stress the existence of an alternative secretory route for MHC class I, which will be worthy of further studies.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Antígenos de Neoplasias/metabolismo , Células Dendríticas/imunologia , Retículo Endoplasmático/metabolismo , Antígeno HLA-A1/metabolismo , Proteínas de Neoplasias/metabolismo , Linfócitos T Citotóxicos/imunologia , Vacúolos/metabolismo , Apresentação de Antígeno , Linhagem Celular , Apresentação Cruzada , Citosol/metabolismo , Antígeno HLA-A2/metabolismo , Humanos , Peptídeos/metabolismo , Antígeno gp100 de Melanoma/metabolismo
14.
J Immunol ; 202(2): 618-624, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30530481

RESUMO

Adenoviruses are a major cause of infectious mortality in children following allogeneic hematopoietic stem cell transplantation, with adoptive transfer of adenovirus-specific T cells being an effective therapeutic approach. We have previously shown that T cells specific for the peptide epitope LTDLGQNLLY were protective. In this study, we aimed to establish a viral dissemination assay to measure the antiviral capacity of T cells specific for this and other peptide epitopes in an infectious setting. We used replication-competent adenovirus 11 (Ad11pGFP) and adenovirus 5 containing adenovirus 35 fiber (Ad5F35GFP) viruses and T cells specific for HLA-A*01-restricted LTDLGQNLLY, HLA-B*07-restricted KPYSGTAYNAL, and HLA-A*02-restricted LLDQLIEEV peptide epitopes. T cells in PBMC from healthy donors were expanded with peptide and IL-2 or treated with IL-2 alone to serve as nonstimulated control cells, and then these expanded or nonstimulated CD8+ cells were purified and cocultured with autologous monocytes infected with adenovirus at low multiplicity of infection. After 3 d, the number of infected GFP+ monocytes and, hence, viral dissemination was quantified by flow cytometry. T cells expanded with LTDLGQNLLY peptide from multiple HLA-A*01+ donors prevented adenovirus dissemination, and nonstimulated T cells did not prevent dissemination, thus, indicating that LTDLGQNLLY-specific T cells have high antiviral capacity. Similarly, expanded KPYSGTAYNAL- and LLDQLIEEV-specific T cells could prevent viral dissemination. However, the frequency of expanded T cells specific for these last two epitopes was variable between donors with consequent variable prevention of adenoviral dissemination. Taken together, we demonstrate that T cells specific for three peptide epitopes, from both structural and nonstructural proteins, can prevent adenoviral dissemination and provide a novel method to measure the antiviral capacity of adenovirus-specific T cell responses.


Assuntos
Infecções por Adenoviridae/imunologia , Adenoviridae/imunologia , Linfócitos T CD8-Positivos/imunologia , Epitopos de Linfócito T/imunologia , Antígenos Virais/imunologia , Criança , Citotoxicidade Imunológica , Citometria de Fluxo , Antígeno HLA-A1/imunologia , Antígeno HLA-A2/imunologia , Antígeno HLA-B7/imunologia , Humanos , Interleucina-2/farmacologia , Leucócitos Mononucleares/imunologia , Peptídeos/imunologia
15.
J Clin Invest ; 127(7): 2705-2718, 2017 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-28628042

RESUMO

Preferentially expressed antigen in melanoma (PRAME) is a cancer-testis antigen that is expressed in many cancers and leukemias. In healthy tissue, PRAME expression is limited to the testes and ovaries, making it a highly attractive cancer target. PRAME is an intracellular protein that cannot currently be drugged. After proteasomal processing, the PRAME300-309 peptide ALYVDSLFFL (ALY) is presented in the context of human leukocyte antigen HLA-A*02:01 molecules for recognition by the T cell receptor (TCR) of cytotoxic T cells. Here, we have described Pr20, a TCR mimic (TCRm) human IgG1 antibody that recognizes the cell-surface ALY peptide/HLA-A2 complex. Pr20 is an immunological tool and potential therapeutic agent. Pr20 bound to PRAME+HLA-A2+ cancers. An afucosylated Fc form (Pr20M) directed antibody-dependent cellular cytotoxicity against PRAME+HLA-A2+ leukemia cells and was therapeutically effective against mouse xenograft models of human leukemia. In some tumors, Pr20 binding markedly increased upon IFN-γ treatment, mediated by induction of the immunoproteasome catalytic subunit ß5i. The immunoproteasome reduced internal destructive cleavages within the ALY epitope compared with the constitutive proteasome. The data provide rationale for developing TCRm antibodies as therapeutic agents for cancer, offer mechanistic insight on proteasomal regulation of tumor-associated peptide/HLA antigen complexes, and yield possible therapeutic solutions to target antigens with ultra-low surface presentation.


Assuntos
Antígenos de Neoplasias/imunologia , Antígeno HLA-A1/imunologia , Imunoglobulina G/farmacologia , Neoplasias Experimentais , Receptores de Antígenos de Linfócitos T/imunologia , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Imunoglobulina G/imunologia , Masculino , Camundongos , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Nucleic Acids Res ; 45(W1): W344-W349, 2017 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-28407117

RESUMO

Peptides are extensively used to characterize functional or (linear) structural aspects of receptor-ligand interactions in biological systems, e.g. SH2, SH3, PDZ peptide-recognition domains, the MHC membrane receptors and enzymes such as kinases and phosphatases. NNAlign is a method for the identification of such linear motifs in biological sequences. The algorithm aligns the amino acid or nucleotide sequences provided as training set, and generates a model of the sequence motif detected in the data. The webserver allows setting up cross-validation experiments to estimate the performance of the model, as well as evaluations on independent data. Many features of the training sequences can be encoded as input, and the network architecture is highly customizable. The results returned by the server include a graphical representation of the motif identified by the method, performance values and a downloadable model that can be applied to scan protein sequences for occurrence of the motif. While its performance for the characterization of peptide-MHC interactions is widely documented, we extended NNAlign to be applicable to other receptor-ligand systems as well. Version 2.0 supports alignments with insertions and deletions, encoding of receptor pseudo-sequences, and custom alphabets for the training sequences. The server is available at http://www.cbs.dtu.dk/services/NNAlign-2.0.


Assuntos
Algoritmos , Redes Neurais de Computação , Peptídeos/química , Software , Sequência de Aminoácidos , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/química , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Sítios de Ligação , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Bases de Dados de Proteínas , Fatores de Transcrição Forkhead/química , Fatores de Transcrição Forkhead/metabolismo , Antígeno HLA-A1/química , Antígeno HLA-A1/metabolismo , Antígeno HLA-B7/química , Antígeno HLA-B7/metabolismo , Antígeno HLA-B8/química , Antígeno HLA-B8/metabolismo , Cadeias HLA-DRB1/química , Cadeias HLA-DRB1/metabolismo , Humanos , Internet , Ligantes , Peptídeos/metabolismo , Ligação Proteica , Alinhamento de Sequência , Transativadores/química , Transativadores/metabolismo
17.
J Immunol ; 198(5): 1838-1845, 2017 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-28148736

RESUMO

Initial studies associated the HLA class I A*01 and B*08 alleles with celiac disease (CD) susceptibility. Subsequent analyses showed a primary association with HLA class II alleles encoding for the HLA DQ2.5 molecule. Because of the strong linkage disequilibrium of A*01 and B*08 alleles with the DR3-DQ2.5 haplotype and a recent genome-wide association study indicating that B*08 and B*39 are predisposing genes, the etiologic role of HLA class I in CD pathogenesis needs to be addressed. We screened gliadin proteins (2α-, 2ω-, and 2γ-gliadin) using bioinformatic algorithms for the presence of peptides predicted to bind A*0101 and B*0801 molecules. The top 1% scoring 9- and 10-mer peptides (N = 97, total) were synthesized and tested in binding assays using purified A*0101 and B*0801 molecules. Twenty of ninety-seven peptides bound B*0801 and only 3 of 97 bound A*0101 with high affinity (IC50 < 500 nM). These 23 gliadin peptides were next assayed by IFN-γ ELISPOT for recognition in peripheral blood cells of CD patients and healthy controls carrying the A*0101 and/or B*0801 genes and in A*0101/B*0801- CD patients. Ten of the twenty-three peptides assayed recalled IFN-γ responses mediated by CD8+ T cells in A*0101/B*0801+ patients with CD. Two peptides were restricted by A*0101, and eight were restricted by B*0801. Of note, 50% (5/10) of CD8+ T cell epitopes mapped within the γ-gliadins. Our results highlight the value of predicted binding to HLA molecules for identifying gliadin epitopes and demonstrate that HLA class I molecules restrict the anti-gluten T cell response in CD patients.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Doença Celíaca/imunologia , Gliadina/imunologia , Antígeno HLA-A1/imunologia , Antígeno HLA-B8/imunologia , Peptídeos/imunologia , Adolescente , Adulto , Algoritmos , Proteínas de Transporte/imunologia , Proteínas de Transporte/fisiologia , Doença Celíaca/genética , Doença Celíaca/fisiopatologia , Criança , Pré-Escolar , Biologia Computacional , ELISPOT , Epitopos de Linfócito T/imunologia , Feminino , Genes MHC Classe I , Glutens/imunologia , Antígeno HLA-A1/genética , Antígeno HLA-A1/metabolismo , Antígeno HLA-B8/genética , Antígeno HLA-B8/metabolismo , Humanos , Interferon gama/genética , Interferon gama/imunologia , Masculino , Pessoa de Meia-Idade , Peptídeos/metabolismo , Adulto Jovem
18.
Cytotherapy ; 19(1): 107-118, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27793552

RESUMO

BACKGROUND AIMS: Herpes simplex virus (HSV) reactivation and infection is common in patients undergoing hematopoietic stem cell transplant (HSCT) and requires routine antiviral prophylaxis. Drug-resistant strains are increasingly common, and effective alternative therapy is currently unavailable. We generated and characterized HSV-1-specific T cells for use in adoptive cellular immunotherapy following allogeneic stem cell transplantation. METHODS: Peripheral blood mononuclear cells from HLA-A1 and HLA-A2 HSV-seropositive hereditary hemochromatosis donors were used as the antigen source. Three HLA-A1 and four HLA-A2 specific epitopes were used for stimulation of T cells. Cells were stimulated with antigen-pulsed dendritic cells and cultured for 21 days in medium with interleukin (IL)-2. Cultured cells were phenotyped and tested for cytokine production, proliferation and cytotoxicity. RESULTS: There was a 5.3-fold expansion in total cell numbers over 21 days of culture, with 35% of T cells being CD8 positive. Thirty-five percent, 21% and 5% of CD8 cells secreted interferon-γ, tumor necrosis factor-α and IL-2 upon HSV antigen re-stimulation. More than 50% of antigen-specific T cells secreted multiple cytokines. Cultured T cells proliferated upon antigen re-stimulation and lysed HSV-1 peptide and virus-infected targets. CONCLUSIONS: It is feasible to generate functional HSV-1 specific T cells from the blood of HLA-A1 and HLA-A2 HSV-seropositive donors using specific peptides. The utility of these cells in preventing and treating HSV-1 reactivation in allogeneic HSCT will need to be tested clinically.


Assuntos
Antígeno HLA-A1/sangue , Antígeno HLA-A2/sangue , Herpesvirus Humano 1/imunologia , Imunoterapia Adotiva/métodos , Linfócitos T/imunologia , Linfócitos T CD8-Positivos/imunologia , Técnicas de Cultura de Células , Células Dendríticas/imunologia , Epitopos , Antígeno HLA-A1/imunologia , Antígeno HLA-A2/imunologia , Humanos , Interferon gama/metabolismo , Interleucina-2/farmacologia , Leucócitos Mononucleares/imunologia , Linfócitos T/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo
19.
J Immunol Res ; 2016: 6829283, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27999823

RESUMO

Malignant melanoma, a very common type of cancer, is a rapidly growing cancer of the skin with an increase in incidence among the Caucasian population. The disease is seen through all age groups and is very common in the younger age groups. Several studies have examined the risk factors and pathophysiological mechanisms of malignant melanoma, which have enlightened our understanding of the development of the disease, but we have still to fully understand the complex immunological interactions. The examination of the interaction between the human leucocyte antigen (HLA) system and prognostic outcome has shown interesting results, and a correlation between the down- or upregulation of these antigens and prognosis has been seen through many different types of cancer. In malignant melanoma, HLA class Ia has been seen to influence the effects of pharmaceutical drug treatment as well as the overall prognosis, and the HLA class Ib and regulatory T cells have been correlated with tumor progression. Although there is still no standardized immunological treatment worldwide, the interaction between the human leucocyte antigen (HLA) system and tumor progression seems to be a promising focus in the way of optimizing the treatment of malignant melanoma.


Assuntos
Antígeno HLA-A1/genética , Antígeno HLA-A1/imunologia , Antígenos HLA-G/genética , Antígenos HLA-G/imunologia , Melanoma/diagnóstico , Melanoma/etiologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Animais , Suscetibilidade a Doenças , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Imunoterapia , Melanoma/terapia , Gravidez , Prognóstico , Fatores de Risco , Raios Ultravioleta/efeitos adversos
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