The Critical Need for Implementing RRF in the Accurate Assessment of Impurities in Peptide Therapeutics

Abstract

The Relative Response Factor (RRF) is a critical parameter in the quantification of impurities in peptides and other pharmaceutical substances, particularly during High-Performance Liquid Chromatography (HPLC) analysis with UV detection. Impurity profiling is crucial for ensuring the safety, efficacy, and quality of peptide drugs. Impurities (process/degradation) in peptide drug products originate due to insertion, truncation, deamidation, isomerization, oxidation, and manufacturing processes that may not have equal responses (RRF ≥ 1 or RRF ≤ 1) compared to their main analyte; hence, the RRF determination is necessary to ensure accurate impurity quantification by accounting for differences in detector responses between the main analyte (e.g., peptide) and its impurities. The use of RRF allows for quantifying trace-level impurities relative to the peptide API and ensuring compliance with stringent quality standards for therapeutic peptides. RRF ensures that the quantification of impurities is not skewed by differences in molecular weight, detector sensitivity, or chromatographic conditions. By applying the RRF value by default as 1, impurity estimation results may be overestimated (actual RRF > 1) or underestimated (actual RRF < 1) in peptide therapeutics. Failure to accurately quantify impurities could result in noncompliance with ICH guidelines and other regulatory requirements related to synthetic peptides, potentially affecting drug approval. RRF-based impurity analysis is particularly important for assessing the immunogenicity risk of peptide therapeutics since impurities in peptide drugs can potentially trigger unwanted immune responses. Accurate quantification of these impurities using RRF helps in evaluating their potential impact on the drug’s immunogenicity profile.