The nature of peptides presented by an HLA class I low expression allele

Hinrichs, Jan B.; Föll, Daniel; Bade‐Doeding, Christina; Huyton, Trevor; Blasczyk, Rainer

doi:10.3324/haematol.2009.016089

Cited by 10 publications

(12 citation statements)

References 40 publications

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“…The importance of classifying alleles evading such mismatches is also supported by our findings (6, 7) that the peptides presented by HLA‐A*30:14L are very similar to those presented by its relative HLA‐A*30:01. So far two cases of mismatch by non‐detection of an L allele have been reported.…”

Section: Discussionsupporting

confidence: 70%

Closing the gap: discrimination of the expression profile of HLA questionable alleles by a cytokine‐induced secretion approach using HLA‐A*32:11Q

et al. 2012

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Matching of human leukocyte antigen (HLA) alleles between donors and recipients plays a major role in hematopoietic stem cell transplantation (HSCT). Null or questionably expressed HLA allelic variants are a major issue in HLA matching, because the aberrant expression of such alleles can have a major impact on the outcome of HSCT and/or its complications such as graft-versus-host disease. The goal of this study was to investigate the potential of a recently developed cytokine-induced secretion assay to differentiate the expression levels of HLA-A*32:11Q (questionable) into a null (N) or low (L) expression variant. An amino acid mutation at position 164 of HLA-A*32:11Q disrupts the disulfide bridge in the α2 domain. HLA-A*32:11Q is not detectable by standard microlymphocytotoxicity assay. To this end, we cloned soluble HLA-A*32:11Q and a reference allele (HLA-A*32:01) into expression vectors and transfected/transduced HEK293 and K562 cells. Allele-expressing K562 cells were simultaneously transfected/transduced with a β2-microglobulin (B2M)-encoding vector to ensure the intact HLA structure with B2M. After treatment with proinflammatory cytokines, secreted soluble HLA molecules were determined by enzyme-linked immunosorbent assay in the supernatant and intracellular accumulation of the recombinant proteins by flow cytometry. HLA-A*32:11Q was nearly undetectable in untreated transfectants. Cytokine treatment increased the secretion of HLA-A*32:11Q to detectable levels and resulted in intracellular accumulation of the allele. There was no difference in mRNA transcription between the A*32 alleles. On the basis of these results, we recommend reclassification of HLA-A*32:11Q as a low expression (L) variant.

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Section: Discussionsupporting

confidence: 70%

Closing the gap: discrimination of the expression profile of HLA questionable alleles by a cytokine‐induced secretion approach using HLA‐A*32:11Q

et al. 2012

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“…Since HLA mismatches may produce severe graft –vs host disease (GvHD) and graft rejection, a misinterpretation of these variants could strongly affect transplant‐related mortality. Hence, several studies proposed assays and methods to discriminate between low expressed or non‐expressed HLA alleles, mainly based on EBV‐transformed cell lines or on recombinant HLA‐molecules' cloning and transfection in established cell lines, not easily reproducible in most laboratories. Gerritsen et al demonstrated that HLA‐A*23:19Q , which has a single polymorphism at position 619 (G > A) compared with HLA‐A*23:01:01 , has no expression by serology and flow cytometry, therefore, it should be reclassified as a null allele .…”

Section: Discussionmentioning

confidence: 99%

Expression profile of HLA‐B38:55Q* allele

Fusco¹,

Cervelli

Mas

et al. 2020

HLA

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The identification of null or questionably expressed HLA allelic variants is a major issue in HLA diagnostics, because the mistyping of the aberrant expression of such alleles can have a major impact on the outcome of both hematopoietic stem cell transplantation (HSCT) and solid organ transplants. It is debated how questionable (Q) alleles, because of their unknown expression profile, should be considered in an allogenic HSCT setting. The HLA-B*38:55Q allele was detected as an HLA-B blank specificity; DNA sequencing identified a single polymorphism at position 373 in exon 3 (TGC > CGC), which results in the replacement of cysteine 101 with an arginine in the HLA-B heavy chain, thus, impairing disulfide bridge formation in the alpha-2 domain, essential for the normal expression of the HLA molecules. In order to determine the RNA and protein expression profile of this allelic variant, we analyzed antigenic expression at different levels, transcriptional and transductional, using a combination of cellular methods, such as serological testing and flow cytometric analysis, polymerase chain reaction (PCR) sequence-specific primer (SSP) cDNA group-specific amplification and immunocytochemical assay, demonstrating the prevalent cytoplasmatic distribution of the HLA-B*38:55Q protein.Our findings suggest that in matching process the HLA-B*38:55Q allele needs to be considered as a low expressed allele, able to elicit an allogenic T-cell response in vivo and impair the transplant outcome. K E Y W O R D S hematopoietic stem cell transplantation, HLA expression variants, HLA-B*38:55Q, null alleles, questionable expression allele

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“…As a result of this indirect allorecognition pathway, GvHD or graft rejection might be promoted in the event of a severe mismatch. It was shown for the first time that an HLA low expression allele (HLA-A*30:14L) presents peptides with identical features to those of its most closely related relative, HLA-A*30:01 (Hinrichs et al 2010). The results indicate that a mismatch at amino acid position 164 might be permissive.…”

Section: Discussionmentioning

confidence: 99%

“…Identification and comparison of allele-specific peptide-binding motifs provide important information for donor-recipient matching and prediction of HLA subtype allogenicity in allogeneic HSCT. In order to determine the functionality of HLA low-expression alleles, peptides from recombinant truncated HLA-A*30:14L molecules secreted in the supernatant of a human cell line were eluted and sequenced (Hinrichs et al 2010). The suitability of the monoclonal anti-HLA class I antibody W6/32 for purifying recombinant HLA-A*30:14L molecules suggested its proper folding and assembly.…”

Section: The Nature Of Peptides Presented By Hla Class I Expression Vmentioning

confidence: 99%

“…The preference for lysine as the top amino acid at the P position of the bound peptides, like described by positional scanning combinatorial peptide libraries (PSCPL) analysis, could be consolidated by the obtained peptide sequence data (Lamberth et al 2008;Sidney et al 2008). Position P3 of the peptides was identified as a primary-secondary anchor showing a high preference for the basic amino acids K and R. Six amino acids are reportedly favored at position P2: phenylalanine ( To verify the presentation of naturally presented peptides from recombinant HLA-A*30:01/30:14L molecules, peptide binding was analyzed by flow cytometry ( Storkus et al 1993;Zeh et al 1994;Maeurer et al 1996) in three EBV-transformed B-LCLs expressing either HLA-A*30:14L,*02:01 (Ulm-241539), HLA-A*30:01,*02:01 (EBRCC-256) or HLA-A*02:01 (EBRCC-2296) ( Warburton et al 1994;Hirv et al 2006;Hinrichs et al 2009;2010). Homology-based modeling for each HLA-A*30 alleles with the shared 9, 11 and 12-mer peptide epitopes revealed only marginal differences between the two HLA-A*30 alleles.…”

Section: The Nature Of Peptides Presented By Hla Class I Expression Vmentioning

confidence: 99%

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Relevance of HLA Expression Variants in Stem Cell Transplantation

Blasczyk¹

2012

New Advances in Stem Cell Transplantation

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The nature of peptides presented by an HLA class I low expression allele

Cited by 10 publications

References 40 publications

Closing the gap: discrimination of the expression profile of HLA questionable alleles by a cytokine‐induced secretion approach using HLA‐A*32:11Q

Closing the gap: discrimination of the expression profile of HLA questionable alleles by a cytokine‐induced secretion approach using HLA‐A*32:11Q

Expression profile of HLA‐B38:55Q* allele

Relevance of HLA Expression Variants in Stem Cell Transplantation

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The nature of peptides presented by an HLA class I low expression allele

Cited by 10 publications

References 40 publications

Closing the gap: discrimination of the expression profile of HLA questionable alleles by a cytokine‐induced secretion approach using HLA‐A*32:11Q

Closing the gap: discrimination of the expression profile of HLA questionable alleles by a cytokine‐induced secretion approach using HLA‐A*32:11Q

Expression profile of HLA‐B*38:55Q allele

Relevance of HLA Expression Variants in Stem Cell Transplantation

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Product

Resources

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Expression profile of HLA‐B38:55Q* allele