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8:15 am 8:30 am (15 mins)

Opening Remarks

8:30 am 9:15 am (45 mins)

Platform Development Incorporating Sony SH800 Cell Sorter, ClonePix 2, and ambr15 Technologies for Monoclonal Antibody Expression in CHO-DG44

We have utilized FACS-based enrichment, ClonePix 2 clone selection and ambr15 microbioreactor technologies to standardize and improve our clone generation and top clone selection protocols for monoclonal antibody development in the CHO-DG44 platform.

  • Scientist Elizabeth H. Scheideman, Scientist, Cell Line Development, Vaccine Production Program, NIH

9:15 am 9:45 am (30 mins)

Meta-analysis Supports Aggressive Timeline Reduction for Cell Line Construction (CLC)

Meta-analysis of CLC data supports improvements to our miniature bioreactor model, host, transfection and selection strategies. These improvements reduced the timeline for CLC by greater than a month, while benefitting overall quality. Screening cell lines in miniature bioreactors provides quality process information which supports selection of cell lines better suited for cGMP manufacture and achieving critical quality attributes

  • Associate Principal Scientist Elaine Lidington, Associate Principal Scientist, Lonza Biologics
  • Group Leader Megan Mason, Group Leader, Lonza Biologics

9:45 am 10:15 am (30 mins)

Networking Refreshment Break in Poster & Exhibit Hall

10:15 am 10:45 am (30 mins)

To Fucosylate or Not: Utilizing FX Knockout CHO Lines to Express Either WT

During antibody dependent cell cytotoxicity (ADCC) the target cells are killed by monocytes and natural killer cells. ADCC is enhanced when the antibody heavy chain's core glycan lacks the fucose molecule(s). Several strategies have been utilized to generate afucosylated antibodies. A commonly used and efficient approach has been knocking out the Fut8 gene of the Chinese hamster ovary (CHO) host cells, which results in expression of antibody molecules with completely afucosylated glycan profiles. However, a major drawback of the Fut8-KO host is the requirement for undertaking two separate cell line development (CLD) efforts in order to obtain both WT and afucosylated antibody species for comparative studies in vitro and in vivo. Even more challenging is obtaining WT and Fut8-KO clones with similar enough product quality attributes to ensure that any observed advantage(s) can be strictly attributed to afucosylation. Here, we report the generation and use of a FX knockout (FXKO) CHO host cell line that is capable of expressing antibody molecules with either afucosylated or WT glycan profiles with similar product quality attributes, dependent upon the addition of fucose to the media. Hence, the FXKO host not only obviates the requirement for undertaking two separate CLD efforts, but it also averts the need for screening many colonies to identify clones with comparable product qualities. Finally, FXKO clones can express antibodies with the desired ratio of WT to afucosylated glycans by titrating fucose to achieve therapeutically optimal levels of ADCC for an antibody.

  • Scientist Shahram Misaghi, Scientist, Genentech

10:45 am 11:15 am (30 mins)

Engineering CHO Cell Factories by Directed Evolution

  • Engineered CHO cells with a significantly increased biomass synthesis capability
  • Simple and effective methodology
  • Detailed analysis of mechanism
  • Professor David C. James, Ph.D., Professor, Bioprocess Engineering, University of Sheffield

11:15 am 11:45 am (30 mins)

GlycoExpress™: A Toolbox for High Yield Production of Glycooptimized Fully Human Biopharmaceuticals in Perfusion Bioreactors at Different Scales

Glycosylation is one of the major post-translational modifications of biotherapeutics important for bioactivity, bioavailability, immunogenicity and patient coverage. By establishment of the GlycoExpress™ toolbox (GEX™) we have generated a set of glycoengineered human cell lines for high yield production of fully human glycoproteins to optimize the glycosylation of antibodies and non-antibody biotherapeutics for improvement of the clinical efficacy and side effects. The system shows high yield for different classes of glycoproteins including different antibody formats and hormones as well as highly similar glycan structure when comparing lab scales perfusion bioreactors of 10 mL working volume to large scale production reactors including a continuous 1000L single-use bioreactor.

11:45 am 12:15 pm (30 mins)

Technology Workshop - CHO Media Optimization Using ambr15 and DoE

Combining bioreactors with Design of Experiment (DoE) approach can provide a systematic solution for biopharmaceutical process development. Sartorius Lonza will present results from experiments with the ambr®15 automated micro bioreactor system and an integrated DoE software (MODDE). These findings demonstrate an enabling technology to provide complete basal and feed media and fed-batch protocol for a given CHO clone.

  • Senior Scientist Michael Gillmeister, Ph.D., Senior Scientist, Lonza

12:15 pm 1:25 pm (70 mins)

Networking Luncheon in the Poster & Exhibit Hall

1:25 pm 1:55 pm (30 mins)

Optimizing Techniques for Mass Spectrometry Detection of Sequence Variants in Support of Cell Line Development

Sequence variants refer to product-related impurities whose amino acid sequences differ from that derived from the expected eDNA sequence. These are of interest during cell line development as they may represent an error in the DNA sequence of the expression vector. This presentation will review applications of liquid chromatography-mass spectrometry to identify sequence variants in support of cell line development.

  • Scientist I Thomas Slaney, Ph.D., Scientist I, Molecular and Analytical Development, Biologics Development & Operations, Bristol-Myers Squibb

1:55 pm 2:25 pm (30 mins)

Evaluation of siRNA Technology as a Bioprocessing Tool to Improve Productivity and Product Quality Challenges in CHO

  • siRNA technology is an effective tool that can be applied to CHO bioprocessing
  • siRNA targets were successfully temporally regulated in 3 CHO case studies: LDHA, β1,4 galactosyl transferase and cathepsin D protease
  • siRNA targets can be temporally regulated to improve productivity and control product quality attributes in CHO
  • Associate Research Fellow Rodney Combs, M.S., , Associate Research Fellow, Bioprocess R&D, Culture Process Development, World Wide Pharmaceutical Sciences, Pfizer Inc

2:25 pm 3 pm (35 mins)

Engineering of CHO Cell Lines for Enhanced Process Robustness

High quality production cell lines secreting maximal levels of recombinant proteins require stable integration of the recombinant DNA, elevated gene transcription, optimized secretion machinery to handle increased protein secretion and folding loads and, ideally, be easily tracked during manufacturing. Using the data from our CHO-K1 genome and transcriptome, we have engineered our CHO-K1 to address these issues, particularly for difficult-to-express proteins, as well as provide detailed genomic analysis packages for manufacturing cell lines.

  • CEO Igor Fisch, PhD, CEO, Selexis

3:30 pm 4 pm (30 mins)

The Control of Antibody Glycosylation During a Production Bioprocess

The glycoform profile of a monoclonal antibody (Mab) determines many functional properties that affect therapeutic efficacy. Common variations of the conserved Fc glycan include galactosylation, fucosylation and sialylation. The observed glycan profile of the final product can depend upon the producer cell line, the growth media, the culture conditions as well as the Mab protein structure. The presentation will review the parameters that can be controlled in order to minimize batch to batch variation of Mab glycosylation. Strategies will also be discussed to produce Mabs with pre-defined glycan structures.

  • Distinguished Professor Michael Butler, Ph.D., Distinguished Professor, Microbiology, University of Manitoba, Canada

4 pm 4:30 pm (30 mins)

Utilization of a Small Molecule Cell Cycle Inhibitor to Achieve Cell Cycle Arrest, Improves Specific Productivity and Desired Product Quality of Recombinant Proteins in CHO Cell Cultures

Increasing specific productivity for biologics cell culture processes is critical to enabling cost-effective commercial development. In this study, we demonstrate the use of cell cycle inhibitors in multiple recombinant mAb producing CHO cell lines to achieve growth arrest and increase specific productivity by 2-3 fold. When comparing this approach to other commonly used methods of growth arrest, such as temperature shift, we demonstrate specific benefits in key product quality attributes. Under thermal induced growth arrest we observe an increase in non-desirable high mannose glycans, whereas mAbs produced in the presence of the small molecule cell cycle inhibitor resulted in further processing of glycans with a decrease in high mannose species offset with an increase in galactosylated species. The difference in glycosylation during temperature shifts was attributed to decreased mRNA levels of multiple regulatory genes of the N-linked glycosylation pathway which was not evident in the presence of these small molecule inhibitors. In summary, the use of these small molecule cell cycle inhibitors can optimize cell culture processes resulting in optimal cell growth, specific productivity, and glycosylation profiles of the target recombinant antibody.

  • Scientist Tharmala Tharmalingam, Ph.D., Scientist, Cell Line Development, Amgen

4:30 pm 5 pm (30 mins)

Human-like Sialylation of IgGs in CHO Cells

To evaluate the impact of Fc sialylation on IgG’s effector functions, we produced glycovariants of Herceptin in CHO cells through transient co-expression of Herceptin with various combinations of glycosyltransferases. The glycovariants interactions with FcγRIIIa were analyzed by surface plasmon resonance (SPR). For both highly and poorly fucosylated IgGs, galactosylation and sialylation resulted in the stabilization of IgG/FcγRIIIa complexes.

  • Céline Raymond, PhD., USP Development Engineer, Development Mammalian Unit, Millipore Sigma

5 pm 7 pm (120 mins)

Networking Cocktail Reception in the Poster & Exhibit Hall