title: Correlation of PD-L1 Surface Expression on Leukemia Cells with the Ratio of PD-L1 mRNA Variants and with Electrophoretic Mobility.
abstract: The expression on the surface of tumor cells of ligands for the PD-1 inhibitory receptor prevents the antitumor immune response and is considered to be a negative prognostic factor in a variety of solid tumors as well as in hematologic malignancies. To determine if it were possible to analyze PD-L1 with PCR-based methods, we assessed the expression of PD-L1 in primary samples from patients with acute myeloid leukemia, in healthy donors, and in a panel of cell lines, by means of flow cytometry, RT-PCR, and Western blotting. Although the surface density of the protein was not correlated with the amount of expressed full-length mRNA, we found a statistically significant positive correlation between PD-L1 surface density and the ratio of two transcript variants (variant 1/variant 2). Our PCR-based method allows for retrospective examination of PD-L1 surface expression from frozen cDNA samples, without the need for a reference gene. Our results also suggest that variant 2, which is produced by alternative splicing, negatively regulates PD-L1 protein expression on the cell surface. In addition, PD-L1 exposition on the cell surface is clearly associated with a shift of electrophoretic mobility, observed on Western blots. This finding can explain the relatively large variability in PD-L1 apparent molecular weight reported in the literature and offers an alternate means for the assessment of PD-L1 surface expression. Cancer Immunol Res; 4(10); 815-9. ©2016 AACR.
authors.affiliation: Department of Proteomics, Institute of Hematology and Blood Transfusion, Prague, Czech Republic.
authors.forename: Barbora
authors.initials: B
authors.lastname: Brodská
authors.affiliation: Department of Proteomics, Institute of Hematology and Blood Transfusion, Prague, Czech Republic.
authors.forename: Petra
authors.initials: P
authors.lastname: Otevřelová
authors.affiliation: Department of Proteomics, Institute of Hematology and Blood Transfusion, Prague, Czech Republic. kuzel@uhkt.cz.
authors.forename: Kateřina
authors.initials: K
authors.lastname: Kuželová
mesh_terms: D017398:Alternative Splicing; D060890:B7-H1 Antigen / Q000096:biosynthesis* / Q000097:blood / Q000235:genetics; D014408:Biomarkers, Tumor / Q000096:biosynthesis* / Q000097:blood / Q000235:genetics; D015153:Blotting, Western / Q000379:methods; D015972:Gene Expression Regulation, Neoplastic / Q000276:immunology; D014644:Genetic Variation; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000235:genetics / Q000276:immunology*; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000097:blood / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics; D014407:Tumor Cells, Cultured
chemical_list: D060890:B7-H1 Antigen; D014408:Biomarkers, Tumor; C423236:CD274 protein, human; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm