This talk will give an overview of how scientists at Merck Biodevelopment, are overcoming the scientific, technological, and methodological challenges encountered to routinely apply Multi-attribute Method (MAM) approaches during the development and manufacturing of novel biotherapeutics.

Biologics have achieved outstanding successes in oncology and will continue to become more significant in the pharmaceutical landscape. In recent years, the interest in NMR for their analysis has sharply increased as it presents a huge potential for the monitoring of their High Order Structure (HOS). This nondestructive method allows the fingerprinting of the proteins and is a powerful addition to existing biophysical tools to assess the biochemical parameters of the proteins from their PTMs to their structural configuration.

Cellular therapies have shown considerable clinical benefit recently. A goal of CAR T process development is to optimize the process conditions through levers that achieve the quality target product profile. To reach this goal, we employ process analytical technologies that can provide data on the upstream process, downstream process, and product quality attributes of the final product. This talk focuses on utilizing mass spectrometry for characterization and monitoring of those attributes.

• ​NEW DATA - This Presentation Contains New/Unpublished Data

Global study with 21 sites (pharmaceutical and IVD companies), along with a regulatory agency that compared the precision and robustness of an imaged cIEF method. The NIST mAb and a fusion protein were analyzed on the iCE3 and Maurice instruments. Information on the comparability of the results obtained by two instruments widely requested. It will help companies better understand how to evaluate the instruments with key parameters/ transition to newer technology.

We present iMAM, an intact mass LC-MS based multi-attribute monitoring workflow for the characterisation of monoclonal antibodies and therapeutic proteins. The platform is based on native and denaturing separations coupled to high resolution mass spectrometry and uses a discovery phase, followed by a retention time and charge envelope based targeted workflow for routine monitoring of quality attributes on the intact and sub unit levels.

• NEW DATA - This Presentation Contains New Data
• NEW TALK - This Presentation Will be Given for the First Time

Recent years have seen a multitude of separation techniques successfully hyphenated to MS instruments for in-depth, high confidence characterization in biopharmaceutical development. Likewise, software tools for organizing, processing, and reporting information-rich MS data have progressed substantially in recent years, to support the explosion in MS data generation. Here, recent progress in MS-based characterization workflows at Symphogen will be presented and illustrated through case studies of biopharmaceuticals in development.

In biotherapeutic discovery, traditional hybridoma and phage approaches now both contend with and are complemented by NGS and synthetic strategies to meet candidate timelines measured in weeks rather than years. Despite substantial market pressures driving more sophisticated and differentiated approaches, there are stages in these workflows where turnkey solutions can be implemented to eliminate bottlenecks. In this talk HT-SPR technology will be highlighted as a tool in biotherapeutics discovery that is deployable in any workflow.

Mass photometry is a novel bioanalytical technology that measures mass of native biomolecules on a single-molecule level. It is suitable for rapid assessments of sample purity, binding affinities or structural integrity of proteins, DNA and small viruses (AAVs). In the talk we will show how mass photometry can answer different biological questions and introduce a new mass photometry product that will massively improve performance and ease of use of the technology.

Aggregates and fragments are critical product-related impurities in monoclonal-antibody (mAb)-based therapeutics. Identification and quantification of these low-level impurities, and investigation on their interplays have been an important topic to better understand and predict drug stability. As a significant part of mAb aggregates is stabilized by non-covalent molecular interactions, we utilized size exclusion chromatography coupled with native mass spectrometry (SEC-nMS) to characterize structural heterogeneity of aggregates in their native form to gain an advanced understanding of the mechanism of aggregation formation.

• NEW DATA - This Presentation Contains New Data 
• NEW TALK - This Presentation Will be Given for the First Time

Fixed-dose combinations of antibodies can offer therapeutic benefits. Compatibility between the proteins is an important prerequisite for developing combination products. In this presentation, I will show you how to identify compatible antibodies during early-stage development by assessing several biophysical parameters. The feasibility of the approach will be demonstrated with a study on binary mixtures of model antibodies with different physicochemical properties.

• ​NEW TALK - This Presentation Will be Given for the First Time

Pre-existing responses to a human IgG4 monoclonal antibody therapeutic (REGN-X) were detected in approximately 20% of patients using a bridging anti-drug antibody (ADA) assay. High pre-existing assay signal could potentially hinder detection of treatment-emergent ADA, confounding immunogenicity assessment for REGN-X. This presentation will describe the extensive characterization of the pre-existing response at baseline to identify the specific binding region in an IgG4 molecule. A novel mitigation strategy was implemented using a modified version of REGN-X as a reagent in the bridging ADA assay. This approach enabled straightforward assessment of drug-specific treatment-emergent immunogenicity for REGN-X.

Efgartigimod is an investigational antibody fragment designed to reduce disease-causing IgG antibodies and block IgG recycling. Efgartigimod binds to FcRn which plays a central role in rescuing IgG antibodies from degradation. Blocking FcRn reduces IgG levels representing a logical potential therapeutic approach for severe autoimmune diseases. This presentation will cover the key highlights in the development journey of an Fc fragment and successful use of established and innovative analytical methodologies.

A phase appropriate and risk-based control strategy is implemented at Biogen to support antisense oligonucleotide programs from the start of clinical development to commercialization. Due to structural and process similarities among the molecules in the pipeline, platform analytical methods are applied as much as possible, while continuous innovation in technologies drives improvements in analytical capabilities. Examples will be discussed to demonstrate the challenges and progress.

The analytical control strategy for manufacturing of large molecules ensures consistent and high quality during manufacturing leading to a safe and efficacious product. A thorough analytical characterization provides the basis for identifying critical quality attributes (CQAs) which need to be tightly controlled, as well as justification of reduced control for (CQAs) with lower criticality.  Using this science-driven approach providing a thorough understanding of the molecule enables the setup of a comprehensive and smart control strategy, which allows manufacturing flexibility without compromising quality and safety of the product.