Laboratory of Mammalian Cell Bioengineering

Research directions

1. To study the fundamental mechanisms controlling the biosynthesis and secretion of heterologous proteins and the vital activity of cultured mammalian cells.Practical application:Obtaining highly effective producers of biologically active substances of a protein nature.
2. Studies of the organization of the cellular genome and regulation of gene expression on a model of cultured ovarian cells of a Chinese hamster.Practical application:Identification and elimination of factors that limit the overproduction of secreted heterologous proteins by CHO cells, as well as factors that impede the industrial cultivation of dense CHO cell culture, including pathways of programmed cell death and autophagy.
3. Fundamental research in the field of protein chemistry and methods of mathematical planning of experiments.Practical application:Development of cost-effective and fully scalable technologies for isolation and purification of pharmaceutical-grade therapeutic proteins. Preparation of conjugates of therapeutic and vaccine proteins with desired properties.

The laboratory implements a full cycle of biotechnological drug development — from obtaining highly effective producers to developing methods for isolating target proteins and controlling their quality. This cycle includes both standard and specific (including laboratory-developed) methods and approaches. The laboratory conducts research on factors that limit cell productivity at different levels – transcription, translation, secretion, factors affecting cell survival in culture, factors affecting the stability of protein products in the medium and during isolation. The properties of plasmid vectors and their regulatory elements are studied using model fluorescent proteins by flow cytometry and lysate fluorometry. Bioinformatics methods are used to analyze integrated genes of target proteins and optimize cDNA - selection of codons effective for producer cells, determination of the optimal number of introns, as well as to analyze the state of the genomes of the obtained producer cells. Producer lines are being investigated for the selection of optimal cultivation conditions and stability. In the resulting producer cells, the copyicity of the target gene, the effectiveness of its transcription, translation, and the course of post-translational processing of the target protein are determined. In addition, the laboratory is developing methods for purifying target proteins from the culture medium to pharmacopoeial parameters.

Experimental methods:

• molecular cloning and analytical methods of work with NC — PCR-RV, southern blot;
• editing of the genome of CHO cells by the CRISPR-Cas9 system;
• obtaining, cultivating and researching the effectiveness and stability of producer lines, selecting optimal conditions for the expression of target proteins;
• microscopy, flow cytometry of CHO cells;
• immunochemical methods (ELISA, Western blot, isoelectrofocusing);
• preparative and analytical chromatography of proteins and pDNA.

Brief history of the laboratory: The laboratory was founded in 2013 with the aim of combining basic and applied research in the field of pharmaceutical biotechnology. The head of the laboratory is D.B.N. Vorobyov I.I.