Recombinant Human FSH/LH (r-hFSH/r-hLH) vs. Highly Purified Human Menopausal Gonadotropin (HP-hMG): A Comprehensive Analysis of Protein Composition and Quantification

초록

Study question: What are the differences in protein composition between r-hFSH/r-hLH and HP-hMG? Summary answer: r-hFSH/r-hLH is composed only of FSH and LH, whereas HP-hMG contains FSH, LH, hCG, and other non-gonadotropin proteins, underscoring significant compositional differences. What is known already: r-hFSH/r-hLH and HP-hMG are commonly used in ovarian stimulation for assisted reproductive technologies. These gonadotropins have similar clinical purposes but differ in their composition and biological activity. r-hFSH/r-hLH contains pure r-hFSH and r-hLH, while HPhMG includes hCG, which mimics LH-like activity. LH promotes granulosa cell survival through anti-apoptotic activity, whereas hCG, with a longer halflife, induces pro-apoptotic signaling. Although previous research has noted these differences, the application of both modern proteomics methods and conventional analytical techniques for a detailed qualitative and quantitative analysis of r-hFSH/r-hLH and HP-hMG remains underexplored. Study design, size, duration: Commercially available batches of r-hFSH/rhLH, HP-hMG A, and HP-hMG B from different manufacturers were analyzed. The study included three batches each of r-hFSH/r-hLH and HP-hMG A, and two batches of HP-hMG B for validation. Participants/materials, setting, methods: Samples underwent in-solution digestion followed by peptide analysis using nanoscale liquid chromatography-tandem mass spectrometry (nano LC-MS/MS). Protein identification was based on Homo sapiens (UniProt 9606) and Cricetulus griseus (CHO; UniProt 10029) databases. Enzyme-linked immunosorbent assay (ELISA) was performed to measure protein levels in r-hFSH/r-hLH and HPhMG products. Main results and the role of chance: Qualitative analysis using nano LCMS/MS revealed that r-hFSH/r-hLH contained only FSH and LH, with no additional proteins detected, highlighting its stable and pure composition. In contrast, HP-hMG A included FSH, LH, hCG, and various other non-gonadotropin proteins such as complement component C7 and peptidoglycan recognition protein 1. Similarly, HP-hMG B, examined for the first time in this study, also contained FSH, LH, hCG, and additional proteins like clusterin. ELISA was used to quantify hCG levels, and results showed that r-hFSH/rhLH had no detectable hCG. However, both HP-hMG A and HP-hMG B contained significant amounts of hCG. These findings demonstrate that rhFSH/r-hLH offers a high-purity composition with minimal batch-to batch variation. Additionally, considering the detected levels of hCG, it can be inferred that the hCG present in HP-hMG products is derived from an external source. Limitations, reasons for caution: While compositional differences exist between r-hFSH/r-hLH and HP-hMG products, the effect of the non-gonadotropin proteins in HP-hMG products on clinical outcomes and the reproductive system remains unexplored, necessitating further investigation. Wider implications of the findings: This study demonstrated that rhFSH/r-hLH contains high-purity r-hFSH and r-hLH with a stable composition, confirming its advantage in enabling personalized treatment with predictable and accurate dosing. Trial registration number: No