Expression and bioactivity of recombinant human serum albumin and dTMP fusion proteins in CHO cells.
The 14-amino acid (IEGPTLRQWLAARA) thrombopoietin mimetic peptide (TMP) shares no sequence homology with native thrombopoietin (TPO). When dimerized, it displays a high-binding affinity for the TPO receptor and has equipotent bioactivity with recombinant human TPO (rhTPO) in stimulating proliferation and maturation of megakaryocytes in vitro. However, TMP is limited for clinical usage because of its short half-life in vivo. In this study, fusion proteins that composed of tandem dimer of TMP (dTMP) genetically fused at the C- or N-terminus of human serum albumin (HSA) were separately expressed in Chinese hamster ovary (CHO) cells. In vitro bioactivity assays showed that purified fusion proteins promoted the proliferation of megakaryocytes in a dose-dependent manner and activated signal transducer and activator of transcription (STAT) pathway in TPO receptor-dependent manner. Following subcutaneous administration, both HSA-dTMP and dTMP-HSA significantly elevated peripheral platelet counts in normal mice in a dose-dependent manner. In addition, fusion with HSA successfully prolonged dTMP half-life in mice. However, when HSA was fused at the C-terminus of dTMP, the bioactivity of dTMP-HSA was about half of that of HSA-dTMP. In conclusion, these results suggested that HSA/dTMP fusion proteins might be potential drugs for thrombocytopenia and, when HSA was fused at the N-terminus of dTMP, the fusion protein had a higher activity.
Gansu Key Laboratory of Biomonitoring and Bioremediation for Environmental Pollution, Institute of Microbiology, School of Life Sciences, Lanzhou University, Tianshui South Road No. 222, Lanzhou, People's Republic of China.
Yi
Y
Ru
Gansu High Throughput Screening and Creation Center for Health Products, School of Pharmacy, Lanzhou University, Lanzhou, People's Republic of China.
Dejuan
D
Zhi
Gansu High Throughput Screening and Creation Center for Health Products, School of Pharmacy, Lanzhou University, Lanzhou, People's Republic of China.
Dingding
D
Guo
Gansu Key Laboratory of Biomonitoring and Bioremediation for Environmental Pollution, Institute of Microbiology, School of Life Sciences, Lanzhou University, Tianshui South Road No. 222, Lanzhou, People's Republic of China.
Yong
Y
Wang
Gansu High Throughput Screening and Creation Center for Health Products, School of Pharmacy, Lanzhou University, Lanzhou, People's Republic of China.
Yang
Y
Li
Gansu Key Laboratory of Biomonitoring and Bioremediation for Environmental Pollution, Institute of Microbiology, School of Life Sciences, Lanzhou University, Tianshui South Road No. 222, Lanzhou, People's Republic of China.
Meizhu
M
Wang
Gansu High Throughput Screening and Creation Center for Health Products, School of Pharmacy, Lanzhou University, Lanzhou, People's Republic of China.
Suzhen
S
Wei
Gansu High Throughput Screening and Creation Center for Health Products, School of Pharmacy, Lanzhou University, Lanzhou, People's Republic of China.
Haiqing
H
Wang
Gansu High Throughput Screening and Creation Center for Health Products, School of Pharmacy, Lanzhou University, Lanzhou, People's Republic of China.
Na
N
Wang
Gansu High Throughput Screening and Creation Center for Health Products, School of Pharmacy, Lanzhou University, Lanzhou, People's Republic of China.
Jingmin
J
Che
Gansu Key Laboratory of Biomonitoring and Bioremediation for Environmental Pollution, Institute of Microbiology, School of Life Sciences, Lanzhou University, Tianshui South Road No. 222, Lanzhou, People's Republic of China. lihy@lzu.edu.cn.
Hongyu
H
Li
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects*; D006224:Cricetinae; D003412:Cricetulus; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D015870:Gene Expression; D006801:Humans; D008297:Male; D008533:Megakaryocytes / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D010455:Peptides / Q000235:genetics* / Q000378:metabolism; D010976:Platelet Count; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000494:pharmacology*; D050791:STAT Transcription Factors / Q000378:metabolism; D012709:Serum Albumin / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D013921:Thrombocytopenia / Q000188:drug therapy
D010455:Peptides; D011993:Recombinant Fusion Proteins; D050791:STAT Transcription Factors; D012709:Serum Albumin; C515728:thrombopoietin mimetic peptide
Chinese hamster ovary cells; Fusion protein; Human serum albumin; Thrombopoietin mimetic peptide