PEGylation is a favorable strategy for half-life extension, potency attenuation and function adjustment for biologics for ~30 years. Quality of PEGs is critical since polymer impurities significantly complicate downstream purification and analytical control. Beyond the conventional SEC/GPC and NMR combination, we have developed a platform PEG characterization strategy mainly utilizing RPLC-UV-CAD and MALDI that guides PEG purification process, thus ensuring the purest possible PEG to be used for bioconjugation.

It is critical to characterize charge heterogeneity and profiles of biotherapeutic proteins closely throughout the development and manufacturing process. In this presentation, we will discuss how to achieve speed and efficiency in high-throughput charge variant analysis and screening of biotherapeutic proteins, by applying LabChip® microfluidic technologies.

We will discuss how LC-MS MAM methods were developed for protein therapeutics, to improve overall in-process analysis efficiency and potential to displace conventional testing assays. The method development involved three stages of quantitation: (i) >600 peptide attributes for broad coverage, (ii) 10-20 targeted intact/subunit proteoforms for throughput, and (iii) a few attributes for sensitivity and specificity. Automations for sample preparation and data analysis are the focus for successful MAM implementation.

For products registered on the market, analytical technologies used for quality control testing often do not represent the best options currently available. To replace an analytical method used in registration-relevant studies with a current state-of-the art method, a comprehensive method bridging study is required to ensure continuity of quality attribute monitoring and demonstration that the new method is equally, or more, sensitive to detect the relevant critical quality attribute.

Over the lifespan of a biotherapeutic, analytical characterization will support early development, clinical evaluations and commercial manufacturing in different jurisdictions. This presentation will discuss the workflow progression as each of these stages evolve and the challenges at each step.

Recent development in coupling native size exclusion chromatography (SEC) and ion exchange chromatography (IEX) to native MS detection has gained a lot of interest from the biopharmaceutical industry in studying the size and charge heterogeneity of protein drug products. However, routine implementation of native LC-MS analysis in industrial labs has been limited due to the difficulties and complexities in establishing a sensitive and robust native LC-MS platform. Here, we describe the development of a novel native LC-MS platform that can be easily integrated with various LC separation techniques and features great sensitivity, robustness, and versatility. In particular, a dopant desolvation gas with different modifiers could be easily applied using this platform to achieve charge-reduced native MS in order to improve the characterization of both heterogeneous and labile biomolecules. The new platform was built with all commercially available parts and can be readily implemented by other labs.

Historically, there were precedents with product safety, which led to the development of current testing methods. Thus, before implementing new alternative analytical technology, we need to be sure it still detects known safety issues. A common strategy must be applied in Analytical Development, not to create tools to provide patches for some novel, sometimes hypothetical, angle of the problem, but rather to revisit why at first place something was created.

We develop homogeneous ELISAs using upconverting nanoparticles (UCNPs) for a potency measurement of a therapeutic monoclonal antibody (mAb). The UCNP is the best donor material for LRET which offers long-lasting excited states and increased S/N ratio due to low autofluorescence and light scattering from NIR excitation. Our immunoassay format has potential to achieve high-throughput screenings (HTS) for evaluating MOA of the model mAb in QC laboratory.

Developability assessments are being increasingly adapted in early biologic drug discovery because of our improved understanding of the connection between molecular liabilities and drug development hurdles. Incorporating <i>in silico</i> and biophysical assessments at the lead optimization stages can help smooth the transition between research and development, potentially saving both time and money. I will present a case study of how predictive modelling has helped improve developability of a lead candidate.

We will discuss a MAM with subunit mass analysis that has the potential to replace three conventional methods widely used for mAb release testing including identification assay, glycosylation profiling, and ratio determination for co-formulated mAbs. The suitability of this MAM for use in a QC setting is demonstrated through assessment of specificity for identity, as well as accuracy, precision, linearity and robustness for glycan profiling and ratio determination.

Drug development timelines are increasingly being shortened, leaving little time for iterative sequence modification and improvement based on experimental data. The ability to rapidly identify sequence liabilities experimentally and earlier would allow for targeted hotspot correction while maintaining timelines. We evaluated several clinical monoclonal antibodies with available stress data and compared this to analyses with a novel technology that has the potential to decrease timelines and reduce cost.