FIGURE LEGENDS

Figure 1. Cell culture profiles of a mAb-expressing CHO cell line cultured at various pH set points. (A) Viable cell density, (B) viability, (C) cumulative viable cells, (D) titer, (E) specific growth rate, and (F) specific productivity (qP) at pH 6.7 (low), 6.9 (medium), 7.1 (high). Error bars denote mean ± SD (n=3).
Figure 2. Groups of correlated product quality attributes identified through hierarchical clustering. Heatmap of hierarchical clustering of product quality attributes (rows) across cell culture samples (columns). The product quality attributes include N-glycoforms, charge variants (Total Acidic, Main, Total Basic) and aggregation (Total LMW, Monomer, Total HMW). The samples are grouped by a horizontal bar with increasingly darker shades of red (low pH), green (medium pH) and blue (high pH) at increasing time-points.
Figure 3. Global analysis of multi-omics data. (A) PCA of individual omics datasets. Each data point is a single replicate sampled on a particular day of a pH set point. A polynomial spline trend line is added for each pH set point. The numbers in each plot denote culture day for the sample. (B) Overview of data analysis steps. Overlapping pathways across significantly enriched Reactome pathways are identified by individual omics analyses. The numbers of enriched pathways are shown in the Venn diagrams.
Figure 4. Localization of glycan variant profiles and associated differential omics features in the N-glycosylation pathway among the three different pH set-points. The process of N-glycosylation takes place in the ER and Golgi complex along a luminal pH gradient of increasing acidity. Accumulation of group 1 “early glycans” at the more basic cis- to medial-Golgi were observed with increased expression of features associated with precursor glycan assembly (at the ER), demannosylation and GlcNAc extension in the low-pH cultures. Culture pH had minimal effect on distribution of group 2 medial-Golgi “intermediate glycans” which include the dominant G0F variant. Group 3 “complex glycans” at the more acidic trans-Golgi mainly decreased with time but with a slightly higher proportion observed in high-pH cultures; associated omics features reflected the same trend over time but were not differentially expressed by culture pH. Omics features are differentially expressed at FDR < 0.01 (except for those marked with ‘*’) between cultures of different pH.1,2,3Glycan: Group 1, 2 or 3 of mAb quality attributes in the samples that were clustered according to their distributions over pH and time. ER: endoplasmic reticulum; TGN: trans-Golgi network; T: transcriptomics, P: proteomics, M: metabolomics.