title: A novel and simple hollow-fiber assay for in vivo evaluation of nonpeptidyl thrombopoietin receptor agonists.
abstract: Preclinical in vivo assessment of the pharmacologic activity of nonpeptidyl thrombopoietin receptor (TPOR) agonists is very difficult because of the high species specificity of such agonists. In this study, we have developed a novel and simple in vivo hollow-fiber assay to preclinically evaluate TPOR agonists. The 32D-mpl cell line was generated by stable transfection of human TPOR into 32D lymphoblast cells and shown to be a specific model for nonpeptide TPOR agonists in vitro. Stably transfected 32D-mpl cells were then sealed in hollow fibers and implanted into nude mice. Cells in hollow fibers specifically responded to TPOR agonists, including thrombopoietin and eltrombopag, a nonpeptide small-molecule TPOR agonist, but not to granulocyte colony-stimulating factor or erythropoietin. Oral administration of eltrombopag stimulated 32D-mpl cell proliferation, prevented 32D-mpl cell apoptosis, and stimulated the phosphorylation of cellular signaling transducers and activators of transcription in a TPOR- and dose-dependent manner. These results indicate that the hollow-fiber assay is a specific and efficient model for rapidly evaluating the in vivo activity of small-molecule TPOR agonists.
authors.affiliation: Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Cheng-Ying
authors.initials: CY
authors.lastname: Xie
authors.forename: Yong-Ping
authors.initials: YP
authors.lastname: Xu
authors.forename: Hong-Bing
authors.initials: HB
authors.lastname: Zhao
authors.forename: Li-Guang
authors.initials: LG
authors.lastname: Lou
mesh_terms: D000818:Animals; D001565:Benzoates / Q000494:pharmacology; D002460:Cell Line / Q000637:transplantation; D003114:Colony-Forming Units Assay; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation* / Q000379:methods; D004921:Erythropoietin / Q000494:pharmacology; D005260:Female; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D006834:Hydrazines / Q000494:pharmacology; D020007:Implants, Experimental; D007377:Interleukin-3 / Q000494:pharmacology; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D010766:Phosphorylation / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011720:Pyrazoles / Q000494:pharmacology; D053628:Receptors, Thrombopoietin / Q000819:agonists*; D011993:Recombinant Fusion Proteins / Q000819:agonists; D011994:Recombinant Proteins / Q000494:pharmacology; D050796:STAT3 Transcription Factor / Q000378:metabolism; D013926:Thrombopoietin / Q000494:pharmacology; D014162:Transfection
chemical_list: D001565:Benzoates; C554270:EPO protein, human; D006834:Hydrazines; D007377:Interleukin-3; D011720:Pyrazoles; D053628:Receptors, Thrombopoietin; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; D004921:Erythropoietin; D016179:Granulocyte Colony-Stimulating Factor; C082218:MPL protein, human; D013926:Thrombopoietin; C520809:eltrombopag