Root cause analysis investigation of visible particulates in therapeutic protein drug products using morphologically directed Raman spectroscopy

Abstract

Abstract Visible particulate (VP) matter in therapeutic protein drug products (DPs) is a quality attribute that can impact product efficacy and patient safety across the product lifecycle. This study evaluates Morphologically Directed Raman Spectroscopy (MDRS) for detecting and characterizing VPs in therapeutic protein DPs. MDRS combines morphological and chemical analyses, which are essential for root cause analysis (RCA) during incident investigations of VP formation. We evaluated MDRS using polystyrene (PS) microsphere standards, candidate reference materials from the US National Institute of Standards and Technology (NIST), and stressed therapeutic protein DPs, insulin lispro and rituximab. Using MDRS, VPs were imaged and chemically characterized in insulin or rituximab drug products with varying morphologies and ranged from 100 to 400 μm in diameter. The intensity of Raman spectral peaks varied with VP size, transparency, morphological complexity, and between therapeutic protein drug classes. Principal component analysis (PCA) of morphological features (e.g., size, shape, and transparency) revealed that NIST ethylene tetrafluoroethylene (ETFE) particles displayed a range of morphologies, akin to stressed therapeutic protein VPs. These results highlight the potential of combining morphological analysis with Raman spectroscopy to support RCA in therapeutic protein DPs for both pre- and post-market quality assessments.