a novel bioassay for the activity determination of therapeutic human brain natriuretic peptide (bnp)一种新型生物测定的活动确定治疗人脑利钠肽(bnp).pdfVIP

A Novel Bioassay for the Activity Determination of Therapeutic Human Brain Natriuretic Peptide (BNP) 1 1 1 1 1 2 1 Lei Yu , Chunming Rao , Xinchang Shi , Yonghong Li , Kai Gao , Xuguang Li , Junzhi Wang * 1 National Institute for the Control of Pharmaceutical and Biological Products, Beijing, People’s Republic of China, 2 Biologics and Genetic Therapies Directorate, Health Canada, Tunney’s Pasture, Ottawa, Canada Abstract Background: Recombinant human brain natriuretic peptide (rhBNP) is an important peptide-based therapeutic drug indicated for the treatment of acute heart failure. Accurate determination of the potency of therapeutic rhBNP is crucial for the safety and efficacy of the drug. The current bioassay involves use of rabbit aortic strips, with experiments being complicated and time-consuming and markedly variable in results. Animal-less methods with better precision and accuracy should be explored. We have therefore developed an alternative cell-based assay, which relies on the ability of BNP to induce cGMP production in HEK293 cells expressing BNP receptor guanylyl cyclase-A. Methodology/Principal Findings: An alternative assay based on the measurement of BNP-induced cGMP production was developed. Specifically, the bioassay employs cells engineered to express BNP receptor guanylyl cyclase-A (GCA). Upon rhBNP stimulation, the levels of the second messager cGMP in these cells drastically increased and subsequently secreted into culture supernatants. The quantity of cGMP, which corresponds to the rhBNP activity, was determined using a competitive ELISA developed by us. Compared with the traditional assay, the n