Monitoring of PTMs mAbs is essential during their production in both upstream and downstream processes. However, characterization of PTMs using conventional peptide mapping procedure requires time-consuming and labor-intensive offline sample preparation steps. This work describes for the first time, the implementation of a Protein-A affinity chromatography column as the first dimension (1D) in a multi-dimensional LC setup for the automated characterization of mAb variants from harvest cell culture fluid.

Continuous manufacturing is gaining acceptance in the biopharmaceutical industry. With constant production, however, comes the need for rapid analytical techniques to ensure product quality at the production line. To address this need, we are focusing on rapid techniques for monitoring critical in-process attributes during production runs. This presentation will review approaches for at-line measurement of aggregates and bioburden to improve control of continuous processes

The biopharmaceutical industry is transitioning towards continuous biomanufacturing processes that require advanced analytical tools to monitor and control bioprocesses to produce high-quality biologics. Post-translational modification of therapeutic proteins, such as N-linked glycosylation, are critical quality attributes that affect biologics safety and efficacy, requiring close monitoring during biomanufacturing. We have developed an analytical toolkit to monitor and control protein glycosylation during biologics manufacturing. Here we highlight this toolkit and provide examples of its use for monitoring an upstream process for producing a commercially relevant Trastuzumab biosimilar.

Digital Twins allow us to understand the behavior of the in silico replicated process, to optimize or manipulate it. DTs require that the underlying models are predictive. Hybrid modeling is a cost-effective method for capturing process behavior. Due to its nature, it balances the need for data with very attractive extrapolation capabilities. We showcase how hybrid modeling rendered possible the development of DTs for challenging downstream scenarios.

Measuring kinetics of hundreds to thousands of protein interactions is currently an expensive and time-consuming task, but critical to resolving protein function, and unraveling disease pathways. To bridge this gap, INanoBio has developed the first-of-kind protein biosensor array platform termed SPOC (sensor-integrated proteome on chip) – customizable array of >500 full-length folded proteins on a gold biosensor chip (spoc.bio). Surface plasmon resonance (SPR) outputs kinetic data with multiple quantitative parameters Rmax, Ka, and Kd for each protein. Using machine learning-based analytics, SPOC can be used to resolve biological networks, characterize phenotypes, and discover novel biomarkers for early diagnosis and precision medicine. 

The Institute for Protein Design has developed powerful processes for computational protein design, most recently the Diffusion model, which combines structural prediction networks with generative diffusion with the ability to generate highly accurate designs optimized for soluble expression. This results in a high numbers of  proteins requiring experimental validation.  The IPD has developed methods to express, purify, and characterize these designed proteins that can keep pace with design velocity. These methods include utilization of open source robotics, intentional scaling of culture volumes and fractionation for streamlined operations, and high throughput analytical methods for the evaluation of protein aggregation and oligomerization.

The Just-Evotec Biologics CLD platform is optimized to decrease development timelines and increase throughput by using automation from transfection through RCB creation. Our high-throughput transfection method allows us to simultaneously screen 96 transfectants in stable pools to identify more manufacturable molecules with a reduced timeline. Using automation, we are capable of screening over 350 clones allowing us to identify cell lines with high productivity and favorable product quality attributes.

The expression screening of large numbers of protein constructs can be automated utilizing the baculovirus expression system (BEVS) and TECAN automation. Biophysical characterization of small-scale screening samples, such as aSEC and nanoDSF,  provides a more robust screening funnel with better prediction of successful clones than simple SDS-page analysis. Additionally, expanding to an automated “midi-scale” screen allows for production of sufficient material to perform more in-depth POC studies such as Biacore, MST, and LC-MS.

Clearance of residual host DNA is an important part of the biopharmaceutical process as host DNA can pose a potential risk to the patient. Using the KingFisher Presto integrated into a Hamilton liquid handling system, we have automated the residual DNA assay from sample preparation through qPCR plate preparation, significantly reducing FTE labor and allowing for a walk-up system for quicker turnaround and high-throughput for residual DNA results.

Generating data on a large number of variants is crucial for advanced analytics and modeling. For this purpose, we are building LabDroid, a highly automated platform for variant characterization. On a floor area of ~300 m2 (~3200 sqft), we have installed eight robotic cells for production of proteins and peptides, quality assessment as well as functional analysis, and biophysical characterization. The system is fully integrated with our scientific data management platform and all results are automatically registered in a cloud-based repository. During this talk, we will share initial results, highlight some of our challenges, and show how we addressed them.

Given the need to dose bi- and multi-specific antibodies in the range of classic mAbs, expression levels similar to mAbs are required to maintain a reasonable cost of goods. The presentation describes a cell line development and upstream process created by Ichnos Sciences that allows > 5 g/L expression of bi- and multi-specific BEAT and TREAT molecules with desired product quality attributes, focusing on specific improvements such as process intensification.

Looking at the continuum from process development to commercial production, it is obvious that different stages require different approaches. Initially, speed from gene to first clinical batch counts; later, the space and time yield is relevant. This presentation gives a comprehensive overview on strategies where, how, and when to implement process intensification and quantifies the benefits like plant occupancy time and capacity optimization based on successful examples and case studies.