Alison Ping Lee

Background As the field of cultivated meat and seafood progresses, traditional approaches to risk assessment, particularly whole-food animal feeding studies, require critical evaluation. Animal testing, especially long-term feeding studies, are resource and time-intensive, poses unreliable interpretation, and are ethically contentious. Given the controlled production environment of cultivated foods and the availability of compositional, molecular, and exposure data, this paper examines the applicability of a weight-of-evidence framework as a scientifically robust approach to safety assessment. Scope and approach The objective is to critically evaluate the scientific justification for in vivo toxicological studies in the safety assessment of cultivated food products and examine alternative, non-animal based methods. Key findings and conclusions Drawing on regulatory precedents, analytical frameworks, and case studies, this review demonstrates that a tiered, weight-of-evidence framework integrating compositional analysis, exposure assessment, and New Approach Methodologies offers a scientifically rigorous and proportionate strategy in comparison to applying uniform requirements for animal studies.

How pathogenesis of inflammatory bowel disease (IBD) depends on the complex interplay of host genetics, microbiome and the immune system is not fully understood. Here, we showed that Downstream of Kinase 3 (DOK3), an adapter protein involved in immune signaling, confers protection of mice from dextran sodium sulfate (DSS)-induced colitis. DOK3-deficiency promotes gut microbial dysbiosis and enhanced colitis susceptibility, which can be reversed by the transfer of normal microbiota from wild-type mice. Mechanistically, DOK3 exerts its protective effect by suppressing JAK2/STAT3 signaling in colonic neutrophils to limit their S100a8/9 production, thereby maintaining gut microbial ecology and colon homeostasis. Hence, our findings reveal that the immune system and microbiome function in a feed-forward manner, whereby DOK3 maintains colonic neutrophils in a quiescent state to establish a gut microbiome essential for intestinal homeostasis and protection from IBD.

A robust monoclonal antibody (mAb) bioprocess requires physiological parameters such as temperature, pH, or dissolved oxygen to be well‐controlled as even small variations in them could potentially impact the final product quality. For instance, pH substantially affects N‐glycosylation, protein aggregation, and charge variant profiles, as well as mAb productivity. However, relatively less is known about how pH jointly influences product quality and titer. In this study, we investigated the effect of pH on culture performance, product titer, and quality profiles by applying longitudinal multi‐omics profiling, including transcriptomics, proteomics, metabolomics, and glycomics, at three different culture pH set points. The subsequent systematic analysis of multi‐omics data showed that pH set points differentially regulated various intracellular pathways including intracellular vesicular trafficking, cell cycle, and apoptosis, thereby resulting in differences in specific productivity, product titer, and quality profiles. In addition, a time‐dependent variation in mAb N‐glycosylation profiles, independent of pH, was identified to be mainly due to the accumulation of mAb proteins in the endoplasmic reticulum disrupting cellular homeostasis over culture time. Overall, this multi‐omics‐based study provides an in‐depth understanding of the intracellular processes in mAb‐producing CHO cell line under varied pH conditions, and could serve as a baseline for enabling the quality optimization and control of mAb production.

A robust monoclonal antibody (mAb) bioprocess requires physiological parameters such as temperature, pH, or dissolved oxygen (DO) to be well-controlled as even small variations in them could potentially impact the final product quality. For instance, pH substantially affects N-glycosylation, protein aggregation and charge variant profiles, as well as mAb productivity. However, relatively less is known about how pH jointly influences product quality and titer. In this study, we investigated the effect of pH on culture performance, product titer and quality profiles by applying longitudinal multi-omics profiling, including transcriptomics, proteomics, metabolomics and glycomics, at three different culture pH set points. The subsequent systematic analysis of multi-omics data showed that pH set points differentially regulated various intracellular pathways including intracellular vesicular trafficking, cell cycle, and apoptosis, thereby resulting in differences in specific productivity, product titer and quality profiles. In addition, a time-dependent variation in mAb N-glycosylation profiles, independent of pH was identified to be mainly due to the accumulation of mAb proteins in the endoplasmic reticulum (ER) over culture time, disrupting cellular homeostasis. Overall, this multi-omics-based study provides an in-depth understanding of the intracellular processes in mAb-producing CHO cell line under varied pH conditions and could serve as a baseline for enabling the quality optimization and control of mAb production.

Chinese hamster ovary (CHO) cells are the most prevalent mammalian cell factories for producing recombinant therapeutic proteins due to their ability to synthesize human‐like post‐translational modifications and ease of maintenance in suspension cultures. Currently, a wide variety of CHO host cell lines has been developed; substantial differences exist in their phenotypes even when transfected with the same target vector. However, relatively less is known about the influence of their inherited genetic heterogeneity on phenotypic traits and production potential from the bioprocessing point of view. Herein, we present a global transcriptome and proteome profiling of three commonly used parental cell lines (CHO‐K1, CHO‐DXB11, and CHO‐DG44) in suspension cultures and further report their growth‐related characteristics, and N‐ and O‐glycosylation patterns of host cell proteins (HCPs). The comparative multi‐omics and subsequent genome‐scale metabolic network model‐based enrichment analyses indicated that some physiological variations of CHO cells grown in the same media are possibly originated from the genetic deficits, particularly in the cell‐cycle progression. Moreover, the dihydrofolate reductase deficient DG44 and DXB11 possess relatively less active metabolism when compared to K1 cells. The protein processing abilities and the N‐ and O‐glycosylation profiles also differ significantly across the host cell lines, suggesting the need to select host cells in a rational manner for the cell line development on the basis of recombinant protein being produced.

Chinese hamster ovary (CHO) cells are the most prevalent mammalian cell factories for producing recombinant therapeutic proteins due to their ability to synthesize human-like post-translational modifications and ease of maintenance in suspension cultures. Currently, a wide variety of CHO host cell lines have been developed; substantial differences exist in their phenotypes even when transfected with the same target vector. However, relatively less is known about the influence of their inherited genetic heterogeneity on phenotypic traits and production potential from the bioprocessing point of view. Herein, we present a global transcriptome and proteome profiling of three commonly used parental cell lines (CHO-K1, CHO-DXB11 and CHO-DG44) in suspension cultures and further report their growth-related characteristics, and N- and O-glycosylation patterns of host cell proteins (HCPs). The comparative multi-omics analysis indicated that some physiological variations of CHO cells grown in the same media are possibly originated from the genetic deficits, particularly in the cell cycle progression. Moreover, the dihydrofolate reductase deficient DG44 and DXB11 possess relatively less active metabolism when compared to K1 cells. The protein processing abilities and the N- and O-glycosylation profiles also differ significantly across the host cell lines, suggesting the need to select host cells in a rational manner for the cell line development on the basis of recombinant protein being produced.