Regulatory considerations for replacing in vivo bioactivity assays with in vitro methods for recombinant glycoprotein hormones
LI Yi-jun
WEI Wei
CHENG Su-yuan
Abstract：In vivo animal testing has long been the standard method for assessing the biological activity of recombinant glycoprotein hormone products.In alignment with the 3R(replacement,reduction,refinement)principles for animal welfare,both the pharmaceutical industry and regulatory agencies are actively seeking in vitro bioactivity assays to replace animal testing.Recombinant glycoprotein hormones exhibit high structural heterogeneity,with complex glycosylation patterns where glycosylation and sialic acid levels are critically linked to their biological activity and pharmacokinetics.When variants with different sialic acid contents are analyzed using both in vivo animal models and in vitro cell-based assays,the results can be inversely correlated.This negative correlation complicates the development of in vitro methods intended to replace in vivo bioactivity testing.The Chinese Pharma-copoeia(2025)has incorporated a reporter gene assay as an alternative method for determining the biological activity of follicle-stimulating hormone(FSH)drug substance,further promoting the transition from in vivo animal tests to in vitro potency assays.However,such alternative approaches require extensive product-specific data for support.From a regulatory perspective,this article discusses the key considerations for replacing in vivo animal testing with in vitro cell-based bioassays,using FSH bioactivity assessment as a case study.
Keywords：recombinant glycoprotein hormonesin vitro biological activityanalytical method changeinternational active unit
Publication Date：2025-12-30
Online Publishing Date：2026-08-14(First online date of this platform, not the publication date of the document)
Pages：5( 2652-2656 )