[Eukaryotic expression and functional characterization of PD-1 extracellular domain].
The negative signal provided by interactions of costimulatory molecules, programmed death-1 (PD-1) and its ligands, PD-L1 (also B7-H1) and PD-L2 (also B7-DC), is involved in the mechanisms of tumor immune evasion. To block PD-Ls-PD-1 interactions by a soluble receptor of PD-1, we constructed a eukaryotic expression plasmid that expresses extracellular region (aa1-aa167) of murine PD-1 (pPD-1A) and, another version of pPD-1A, pPD-1B, carrying cDNAs encoding for both extracellular region of PD-1 and green fluorescent protein (GFP) reporter gene, which was inserted downstream of PD-1. Experiment of BHK cells transfected with pPD-1B determined that most expression product (sPD-1) in the cells was secreted out. FACS analysis revealed that sPD-1 was specific and bound efficiently to PD-1 ligands. Cytotoxicity assay showed that blocking PD-Ls on either tumor cells or spleen cells by sPD-1 mediated enhanced lysis of H22 cells by Hsp70-H22 peptides complexstimulated spleen cells. The constructed plasmid vector would provide a novel method of tumor gene therapy of blocking PD-Ls-PD-1 interactions by expression of soluble receptor of PD-1 in tumor sites, which could increase the antitumor activity.
Department of Biochemistry & Molecular Biology, Tongji Medical College, Huazhong University of Science & Technology, Wuhan 430030, China.
Yu-Fei
YF
He
Gui-Mei
GM
Zhang
Xiao-Hong
XH
Wang
Hui
H
Zhang
Ye
Y
Yuan
Dong
D
Li
Zuo-Hua
ZH
Feng
D000818:Animals; D015703:Antigens, CD / Q000502:physiology; D000954:Antigens, Surface / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D051017:Apoptosis Regulatory Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D060908:CTLA-4 Antigen; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D005434:Flow Cytometry; D015316:Genetic Therapy; D051379:Mice; D009369:Neoplasms / Q000628:therapy; D010957:Plasmids; D061026:Programmed Cell Death 1 Receptor; D017434:Protein Structure, Tertiary; D014162:Transfection
D015703:Antigens, CD; D000954:Antigens, Surface; D051017:Apoptosis Regulatory Proteins; D060908:CTLA-4 Antigen; C556707:Ctla4 protein, mouse; C491383:Pdcd1 protein, mouse; D061026:Programmed Cell Death 1 Receptor