Project description:Analysis of the peptide repertoires eluted from different HLA-DP molecules expressed in HeLa cells co-expressing Invariant chain either with or without HLA-DM as components of the HLA class II processing machinery. Divergence of the immunopeptidomes and the impact of HLA-DM were investigated in relation to the capacity of HLA-DP molecules to elicit alloreactive T-cell responses.
Project description:In the context of HLA-DP-mismatched allogeneic stem cell transplantation, mismatched HLA-DP alleles can provoke profound allo-HLA-DP-specific immune responses from the donor T-cell repertoire leading to graft-versus-leukemia effect and/or graft-versus-host disease in the patient. The magnitude of allo-HLA-DP-specific immune responses has been shown to depend on the specific HLA-DP disparity between donor and patient and the immunogenicity of the mismatched HLA-DP allele(s). HLA-DP peptidome clustering (DPC) was developed to classify the HLA-DP molecules based on similarities and differences in their peptide-binding motifs. To investigate a possible categorization of HLA-DP molecules based on overlap of presented peptides, we identified and compared the peptidomes of the thirteen most frequently expressed HLA-DP molecules. Our categorization based on shared peptides was in line with the DPC classification. We found that the HLA-DP molecules within the previously defined groups DPC-1 or DPC-3 shared the largest numbers of presented peptides. However, the HLA-DP molecules in DPC-2 segregated into two subgroups based on the overlap in presented peptides. Besides overlap in presented peptides within the DPC groups, a substantial number of peptides was also found to be shared between HLA-DP molecules from different DPC groups, especially for groups DPC-1 and -2. The functional relevance of these findings was illustrated by demonstration of cross-reactivity of allo-HLA-DP-reactive T-cell clones not only against HLA-DP molecules within one DPC group, but also across different DPC groups. The promiscuity of peptides presented in various HLA-DP molecules and the cross-reactivity against different HLA-DP molecules demonstrate that these molecules cannot be strictly categorized in immunogenicity groups.
Project description:A high-throughput mass spectrometry analysis was used to identify more than 16,000 cell peptides bound to several HLA-DR and -DP class II molecules isolated from large amounts of two human cell lines (HOM-2 and JY).
Project description:Cord blood (CB)-derived chimeric antigen receptor (CAR)-natural killer (NK) cells targeting CD19 has been shown to be effective against B cell malignancies. While human CD56+ NK cells can be expanded in vitro, it is also known that NK cells can be differentiated from hematopoietic progenitor cells. It is still unclear whether CAR-NK cells are originated from mature NK cells or NK progenitor cells in CB. Here we found that CAR-NK cells are predominantly derived from the CD56- NK progenitor cells. We first found that substantial numbers of CD19 CAR-NK cells were produced from the CD56- CB mononuclear cells after in vitro culture for two weeks. Single cell RNAseq analysis of CD56-CD3-CD14-CD19- CB mononuclear cells revealed that those cells were subdivided into three subpopulations based on the expression of CD34 and HLA-DR. NK cells were predominantly produced from the CD34-HLA-DR- cells. In addition, in the CD34-HLA-DR- cells, only CD7+ cells could differentiate into NK cells. These results indicate that CD56-CD7+CD34- HLA-DR-lineage marker (Lin)- cells are the major origin of human CB-derived CAR-NK cells, indicating that we need to develop methods to enhance the quality and quantity of NK cells produced from these NK cell progenitor cells.
Project description:HLA-E molecules can present self and pathogen-derived peptides to both NK-cells and T-cells. T-cells that recognize HLA-E peptides via their T-cell receptor (TCR) are termed donor-unrestricted T-cells due to restricted allelic variation of HLA-E. The composition and repertoire of HLA-E TCRs is not known so far. We performed TCR sequencing on CD8+ T-cells from 21 individuals recognizing HLA-E tetramers (TM) folded with 2 Mtb HLA-E restricted peptides. We sorted HLA-E Mtb TM+ and TMCD8+ T-cells directly ex vivo and performed bulk RNA-sequencing and single cell TCR sequencing. The identified TCR repertoire was diverse and showed no conservation between and within individuals. TCRs selected from our single cell TCR sequencing data could be activated upon HLA-E/peptide stimulation, although not robust, reflecting potentially weak interactions between HLA-E peptide complexes and TCRs. Thus, HLA-E Mtb specific T-cells have a highly diverse TCR repertoire.
Project description:CD8+ T cells from HLA-B*2705 HIV+ chronic progressors Hi and Lo samples are CD8+ T cells bound by an HLA-B*2705 tetramer specific for the Gag p24 peptide KK10 PBMCs were incubated in the presence and absence of KK10 peptide for 6 days, and CD8+ and KK10-specific CD8+ T cell were sorted by flow cytometry
Project description:CD8+ T cells from HLA-B*2705 HIV+ elite controllers Hi and Lo samples are CD8+ T cells bound by an HLA-B*2705 tetramer specific for the Gag p24 peptide KK10 PBMCs were incubated in the presence and absence of KK10 peptide for 6 days, and CD8+ and KK10-specific CD8+ T cell were sorted by flow cytometry