Maleknia, S. | bioRxiv Science

bioRxiv2020

BNrich: A Bayesian network approach to the pathway enrichment analysis

MotivationOne of the most popular techniques in biological studies for analyzing high throughput data is pathway enrichment analysis (PEA). Many researchers apply the existing methods without considering the topology of pathways or at least they have overlooked a significant part of the structure, which may reduce the accuracy and generalizability of the results. Developing a new approach while considering gene expression data and topological features like causal relations regarding edge directions will help the investigators to achieve more accurate results. ResultsWe proposed a new pathway enrichment analysis based on Bayesian network (BNrich) as an approach in PEA. To this end, the cycles were eliminated in 187 KEGG human signaling pathways concerning intuitive biological rules and the Bayesian network structures were constructed. The constructed networks were simplified by the Least Absolute Shrinkage Selector Operator (LASSO), and their parameters were estimated using the gene expression data. We finally prioritize the impacted pathways by Fishers Exact Test on significant parameters. Our method integrates both edge and node related parameters to enrich modules in the affected signaling pathway network. In order to evaluate the proposed method, consistency, discrimination, false positive rate and empirical P-value criteria were calculated, and the results are compared to well-known enrichment methods such as signaling pathway impact analysis (SPIA), bi-level meta-analysis (BLMA) and topology-based pathway enrichment analysis (TPEA). AvailabilityThe R package is available on carn.

bioinformatics↗

bioRxiv2019

Implementation of Design of Experiments (DOE) for Optimization of Feeding Strategy and Glyco-Engineering of Trastuzumab Biosimilar

Fed-batch cell culture is the most commonly used process for antibody production in biopharmaceutical industries. Basal media, feed, feeding strategy and glycan structures are always among the most important concerns during process development and optimization. In this study, first, a traditional screening study was performed to identify the top media/feed combinations by evaluating the cell culture performance including cell growth and protein titre. Optimization of the process was also performed using response surface methodology in order to find the most optimum feeding strategy and glucose set point regarding final titre of the recombinant monoclonal antibody being produced in Chinese hamster ovary cell line. The focus of this study is not only on titre, but also on product quality and comparability especially protein glycosylation. The prediction model of product titre as a function of feeding percentage and glucose set point was successfully applied for the second set of experiments that was performed for glycan improvement. Statistical design of experiments was applied to determine the most important factors and their effects on galactosylated and afucosylated glycans. Uridine, manganese, galactose and fucosyltransferase inhibitor were chosen to evaluate if their presence can affect glycans and to obtain their best combination for fed-batch culture supplementation. We determined that 2.5 % daily feeding combined with maintaining the glucose set point on 2.5{+/-}0.2 g/L could achieve final titre of 2.5{+/-} 0.1 g/L. Galactosylation of antibody was increased about 25% using MnCl2 and galactose while afucosylation was increased about 8% in presence of fucosyltransferase inhibitor. Galactose and Mn2+ led to a shift from G0F to G1F and presence of Fucosyltransferase inhibitor caused to an increase in G0 compared to its absence. These results demonstrated that supplementation of culture with all these components can provide exact control of antibody galactosylation and fucosylation with minimal impact on culture characteristics and product quality attributes. Subsequently, validation experiments were also carried out in 5L STR bioreactors which showed that similar results could be achieved in bioreactors compared to shake flasks regarding both titre and quality.

bioengineering↗