Base editing offers a fundamentally new strategy for precise gene editing. We combine base editing technology with available delivery methods to provide innovative gene editing and cell therapies. Multiple critical materials (such as gRNA, mRNA, plasmid, etc,), and delivery vehicles (such as lentivirus, adeno-associated virus AAV, and lipid nanoparticles) mean the processes and products are very complex. As a result, unprecedented analytical needs arise from such complex processes and drug products. Here, I will discuss the overall analytical development strategy in developing base editing drug candidates. 

Pertaining to cell-based therapies, monitoring transfection vector payloads has become important. While several techniques are described, studies show specific advantages conferred by Analytical Ultra-Centrifugation.

In contrast, often absent is direct cell transduction monitoring; analyses do occur, but further downstream. Data are presented on the Coulter Principle as uniquely suited for transduction monitoring.  Together, the two platforms lend efficiencies in cell therapy development.

The characterization and testing of cellular therapeutic products (CTPs) is a critical aspect of product development, translation and release. Here I will describe recent efforts in the standardization of the characterization and testing of CTPs. I will also describe recently published standards on cell counting as well as NIST technical programs for increasing confidence in cell count and viability assays.

The quality and performance of complex analytical reagents are arguably as critical to product quality as raw materials.  Even different batches of some materials may require re-evaluation of the analytical method at some level, especially where custom made.  This talk will discuss what is expected along with tips and advice on how to avoid issues.

QbD concept is extended to our cell therapy process and analytical method design as a systematic approach to add value, improve efficiencies in production and process, and prevent problems later on (e.g. frequently non-confirmed. out-of-specification results due to the non-robustness of the process or method).Here, we will discuss the determination of process and method goals for the improvement of robustness of analytical method for cell-based therapies, risk assessment, specification of a design space, implementing a control strategy and continuous improvement to increase method robustness and knowledge.

Our team is developing an allogeneic cell therapy to treat painful disc degeneration called IDCT (rebonuputemcel). Despite regenerative capacity demonstration across donors, each production lot comes from a new donor cell line with unique quality attributes, which poses a significant challenge in the development of robust biomanufacturing practices. Here we discuss practical quality by design, process modeling, and cross-validation approaches for multi-donor process characterization. By designing a multivariate design space with special considerations for donor variability we link raw material, donor, and process inputs to quality outputs, increase yield 1000-fold, and maintain batch-to-batch consistency for our allogeneic cell therapy.

Rubius Therapeutics is developing a novel cell therapy platform, called the RED PLATFORM®, by genetically engineering donor-derived precursor cells to express therapeutic proteins inside or on the cell surface and culturing them into red blood cells, called Red Cell Therapeutics. RCT products lack a nucleus and are unable to divide in the body. Analytical characterization of product properties and impurities to understand the critical quality attributes of RCTs requires a multi-pronged approach with a comprehensive toolbox. Technical challenges in assessing cellular phenotype, biophysical properties, and biological activity of RCTs will be discussed.

Cell-based gene therapy drug products require comprehensive analytical assays to characterize their safety and efficacy. This seminar will focus on the GMP release tests implemented across multiple phase I/II and III clinical trials and the additional non-GMP characterization assays performed to address heterogeneity of drug products. Correlative analyses between the results of different analytical assays will be discussed.

The ability to deliver transgenes into the human genome using viral vectors is a major enabler of the gene-modified cell therapy field. However, controlling viral transduction is difficult and can lead to product heterogeneity, impacting efficacy and safety. We generated and qualified a novel assay to measure vector copy distribution at a single cell level, label-free transduction efficiency, and bulk vector copy number, demonstrating its applicability on clinically relevant lentiviral-based hematopoietic stem cell therapies. The ability to characterise cell-to-cell variability provides a powerful high-resolution approach for product characterisation which could ultimately allow improved control over product quality, safety and consistency. 

Improving the manufacturing processes used in CGT requires an understanding of certain critical parameters. One such parameter, osmolality, has long been considered a critical tool in bioprocessing and recently has been implicated across the entire gene therapy process workflow. This seminar will focus on the impact of osmolality on improving the quality and yield of an AAV manufacturing process, showing a significant shift on the relative extracellular vs intracellular vector yield.

The Lonza Cocoon® is a closed, automated platform for end-to-end cell therapy manufacturing. As manufacturing processes evolve, Lonza continues to upgrade the Cocoon® Platform with new functionalities. The soon-to-be-released "Integrated, in-line magnetic cell separation" will allow the automated isolation of T cells as a unit operation within the system, resulting in standardized cellular input material and greater control over closed cell therapy manufacturing processes.

The Connell & O’Reilly Families CMCF at the DFCI has 15 separate ISO-7 clean rooms, 2 method development and 1 QC Lab. The facility focusses on translating prototype products into cGMP-compliant manufacturing processes. The presentation will cover a variety of IND-application and CMC requirements that are encountered during development of a manufacturing process for autologous iPSC-based therapies.

Cellino’s vision is to make personalized regenerative medicines viable at large scale. Cellino’s platform combines label-free imaging and high-speed laser editing with machine learning to automate cell reprogramming, expansion, and differentiation in a closed cassette format, enabling thousands of patient samples to be processed in parallel in a single facility.

The era of gene editing has unlocked the potential to cure diseases with few treatment options. In particular, gene-edited stem cell therapies have shown great promise in the treatment of beta-thalassemia and sickle cell disease, and could change the treatment landscape in hematological malignancies. This talk will discuss some points to consider when developing a gene-edited stem cell manufacturing process.

GEN-011 is an autologous, neoantigen-specific T cell therapy. GEN-011 is produced by the PLANET (Proliferation of Lymphocytes Activated by Neoantigens Endogenous in Tumors) manufacturing process, and T cells are specifically expanded on neoantigens identified by Genocea’s proprietary antigen discovery platform ATLAS™. This presentation will provide an insight into the PLANET process and go over our strategies and considerations to overcome manufacturing challenges associated with personalized therapy.