Biological characterization of recombinant human follicle stimulating hormone isoforms.
It has been established that follicle stimulating hormone (FSH) circulates in the bloodstream as a heterogeneous population of molecules. Individual FSH isoforms, while displaying identical amino acid sequences, differ in their extent of post-translational modification. As a result of these variations, the FSH isoforms exhibit differences in overall charge, degree of sialic acid or sulphate incorporation, receptor binding affinity and plasma half-life. Taking advantage of the fact that these forms can be separated from each other on the basis of their charge, we have evaluated in rats the metabolic clearance rates of the acidic [with an isoelectric point (pI) </=ISOdia</= 4.8] and the less acidic (pI > 4.8) isoforms of recombinant human FSH (rhFSH) obtained after chromatofocusing. The less acidic isoform group was found to have a faster clearance from the circulation in rats as compared with the acidic isoform group. This finding is in agreement with the lower bioactivity in vivo (as determined by the Steelman-Pohley assay) of the less acidic isoform group, compared with the acidic one. The mass spectra of the two groups of isoforms showed a difference in the sialic acid content thus highlighting the importance of these residues on the in-vivo activity of FSH. Conversely, when the two groups of isoforms were tested in vitro by using the Y1 human FSH receptor (Y1 hFSHR) assay and a reporter gene assay, no significant differences in the biological activities between these preparations were detected when test concentrations were based on mass.
D000818:Animals; D001681:Biological Assay; D016466:CHO Cells; D006224:Cricetinae; D005640:Follicle Stimulating Hormone / Q000737:chemistry* / Q000378:metabolism / Q000493:pharmacokinetics; D017930:Genes, Reporter; D006207:Half-Life; D006801:Humans; D007525:Isoelectric Focusing; D013058:Mass Spectrometry; D008657:Metabolic Clearance Rate; D020033:Protein Isoforms / Q000737:chemistry* / Q000378:metabolism / Q000493:pharmacokinetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000493:pharmacokinetics