Torsten Schulz - Academia.edu (original) (raw)
Papers by Torsten Schulz
Cells and Culture, 2010
ABSTRACT The majority of biopharmaceuticals is currently being expressed in mammalian cells, most... more ABSTRACT The majority of biopharmaceuticals is currently being expressed in mammalian cells, mostly Chinese hamster ovary (CHO) and mouse myeloma (NS0) cells. The two most prominent challenges for mammalian cell-based systems relate to (i) product titer in the cell culture fluid at the end of cultivation and (ii) development times from final drug candidate selection to an established process to produce clinical grade material. The newest generations of mammalian cell culture production processes evolve rapidly to overcome these limitations. Boehringer Ingelheim’s current high expression (BI HEX™) cell line generation concept combines many improvements including novel or modified genetic elements to improve transcription rate, high throughput screening concepts to obtain highly productive clones reliably, and host cell lines that grow to high densities in serum-free chemically defined media. Specific productivities above 50 pg per cell and day for monoclonal antibodies have been achieved in CHO cells and were successfully translated into product titers of up to 4 g/L in an 11-day process. Furthermore, a strategy is presented for supply of material for toxicological studies produced from CHO cells in as little as 15 months starting with cloning of product-encoding genetic sequences into BI HEX vectors. At the same time it ensures that the production cell generated during this program will have the high expression potential needed to avoid a change in production cell line at later stages in development.
Genomics and Systems Biology of Mammalian Cell Culture, 2011
Development of efficient bioprocesses is essential for cost-effective manufacturing of recombinan... more Development of efficient bioprocesses is essential for cost-effective manufacturing of recombinant therapeutic proteins. To achieve further process improvement and process rationalization comprehensive data analysis of both process data and phenotypic cell-level data is essential. Here, we present a framework for advanced bioprocess data analysis consisting of multivariate data analysis (MVDA), metabolic flux analysis (MFA), and pathway analysis for mapping of large-scale gene expression data sets. This data analysis platform was applied in a process development project with an IgG-producing Chinese hamster ovary (CHO) cell line in which the maximal product titer could be increased from about 5 to 8 g/L.Principal component analysis (PCA), k-means clustering, and partial least-squares (PLS) models were applied to analyze the macroscopic bioprocess data. MFA and gene expression analysis revealed intracellular information on the characteristics of high-performance cell cultivations. By MVDA, for example, correlations between several essential amino acids and the product concentration were observed. Also, a grouping into rather cell specific productivity-driven and process control-driven processes could be unraveled. By MFA, phenotypic characteristics in glycolysis, glutaminolysis, pentose phosphate pathway, citrate cycle, coupling of amino acid metabolism to citrate cycle, and in the energy yield could be identified. By gene expression analysis 247 deregulated metabolic genes were identified which are involved, inter alia, in amino acid metabolism, transport, and protein synthesis.
Nucleic Acids Research, 2010
The arrival of next-generation sequencing (NGS) technologies has led to novel opportunities for e... more The arrival of next-generation sequencing (NGS) technologies has led to novel opportunities for expression profiling and genome analysis by utilizing vast amounts of short read sequence data. Here, we demonstrate that expression profiling in organisms lacking any genome or transcriptome sequence information is feasible by combining Illumina's mRNA-seq technology with a novel bioinformatics pipeline that integrates assembled and annotated Chinese hamster ovary (CHO) sequences with information derived from related organisms. We applied this pipeline to the analysis of CHO cells which were chosen as a model system owing to its relevance in the production of therapeutic proteins. Specifically, we analysed CHO cells undergoing butyrate treatment which is known to affect cell cycle regulation and to increase the specific productivity of recombinant proteins. By this means, we identified sequences for >13 000 CHO genes which added sequence information of $5000 novel genes to the CHO model. More than 6000 transcript sequences are predicted to be complete, as they covered >95% of the corresponding mouse orthologs. Detailed analysis of selected biological functions such as DNA replication and cell cycle control, demonstrated the potential of NGS expression profiling in organisms without extended genome sequence to improve both data quantity and quality.
Biotechnology and Bioengineering, 2010
Increase in both productivity and product yields in biopharmaceutical process development with re... more Increase in both productivity and product yields in biopharmaceutical process development with recombinant protein producing mammalian cells can be mainly attributed to the advancements in cell line development, media, and process optimization. Only recently, genome-scale technologies enable a system-level analysis to elucidate the complex biomolecular basis of protein production in mammalian cells promising an increased process understanding and the deduction of knowledge-based approaches for further process optimization. Here, the use of gene expression profiling for the analysis of a low titer (LT) and high titer (HT) fed batch process using the same IgG producing CHO cell line was investigated. We found that gene expression (i) significantly differed in HT versus LT process conditions due to differences in applied chemically defined, serum-free media, (ii) changed over the time course of the fed batch processes, and that (iii) both metabolic pathways and 14 biological functions such as cellular growth or cell death were affected. Furthermore, detailed analysis of metabolism in a standard process format revealed the potential use of transcriptomics for rational media design as is shown for the case of lipid metabolism where the product titer could be increased by about 20% based on a lipid modified basal medium. The results demonstrate that gene expression profiling can be an important tool for mammalian biopharmaceutical process analysis and optimization.
Analytical and Bioanalytical Chemistry, 2010
The "Pharmaceutical Current Good Manufacturing Practices (CGMPs) for the 21st Century-A Risk Base... more The "Pharmaceutical Current Good Manufacturing Practices (CGMPs) for the 21st Century-A Risk Based Approach" initiative announced by the FDA in August 2002 to improve and modernize pharmaceutical manufacturing facilitated adoption of process analytical technology (PAT) by the pharmaceutical industry. The potential for improved operational control and compliance resulting from continuous real-time quality assurance was highlighted as a likely benefit that would result from PAT implementation. A considerable amount of work has been done on this topic by academic and industrial contributors in the last decade. In this paper, we will start with a brief overview of evolution of PAT concepts and a review of their application in the wider pharmaceutical industry. The rest of the paper focuses on PAT applications for biotech processes with emphasis on developments in the last five years. It is our observation that while significant advances have been accomplished with regard to our ability to analyze/monitor key process and quality attributes in the biotech industry, much more needs to be done with regard to utilizing the collected data for subsequent control of the process, to achieve optimum yield and product quality. The latter is necessary to achieve the benefits that will result from PAT implementation.
Cells and Culture, 2010
ABSTRACT The majority of biopharmaceuticals is currently being expressed in mammalian cells, most... more ABSTRACT The majority of biopharmaceuticals is currently being expressed in mammalian cells, mostly Chinese hamster ovary (CHO) and mouse myeloma (NS0) cells. The two most prominent challenges for mammalian cell-based systems relate to (i) product titer in the cell culture fluid at the end of cultivation and (ii) development times from final drug candidate selection to an established process to produce clinical grade material. The newest generations of mammalian cell culture production processes evolve rapidly to overcome these limitations. Boehringer Ingelheim’s current high expression (BI HEX™) cell line generation concept combines many improvements including novel or modified genetic elements to improve transcription rate, high throughput screening concepts to obtain highly productive clones reliably, and host cell lines that grow to high densities in serum-free chemically defined media. Specific productivities above 50 pg per cell and day for monoclonal antibodies have been achieved in CHO cells and were successfully translated into product titers of up to 4 g/L in an 11-day process. Furthermore, a strategy is presented for supply of material for toxicological studies produced from CHO cells in as little as 15 months starting with cloning of product-encoding genetic sequences into BI HEX vectors. At the same time it ensures that the production cell generated during this program will have the high expression potential needed to avoid a change in production cell line at later stages in development.
Genomics and Systems Biology of Mammalian Cell Culture, 2011
Development of efficient bioprocesses is essential for cost-effective manufacturing of recombinan... more Development of efficient bioprocesses is essential for cost-effective manufacturing of recombinant therapeutic proteins. To achieve further process improvement and process rationalization comprehensive data analysis of both process data and phenotypic cell-level data is essential. Here, we present a framework for advanced bioprocess data analysis consisting of multivariate data analysis (MVDA), metabolic flux analysis (MFA), and pathway analysis for mapping of large-scale gene expression data sets. This data analysis platform was applied in a process development project with an IgG-producing Chinese hamster ovary (CHO) cell line in which the maximal product titer could be increased from about 5 to 8 g/L.Principal component analysis (PCA), k-means clustering, and partial least-squares (PLS) models were applied to analyze the macroscopic bioprocess data. MFA and gene expression analysis revealed intracellular information on the characteristics of high-performance cell cultivations. By MVDA, for example, correlations between several essential amino acids and the product concentration were observed. Also, a grouping into rather cell specific productivity-driven and process control-driven processes could be unraveled. By MFA, phenotypic characteristics in glycolysis, glutaminolysis, pentose phosphate pathway, citrate cycle, coupling of amino acid metabolism to citrate cycle, and in the energy yield could be identified. By gene expression analysis 247 deregulated metabolic genes were identified which are involved, inter alia, in amino acid metabolism, transport, and protein synthesis.
Nucleic Acids Research, 2010
The arrival of next-generation sequencing (NGS) technologies has led to novel opportunities for e... more The arrival of next-generation sequencing (NGS) technologies has led to novel opportunities for expression profiling and genome analysis by utilizing vast amounts of short read sequence data. Here, we demonstrate that expression profiling in organisms lacking any genome or transcriptome sequence information is feasible by combining Illumina's mRNA-seq technology with a novel bioinformatics pipeline that integrates assembled and annotated Chinese hamster ovary (CHO) sequences with information derived from related organisms. We applied this pipeline to the analysis of CHO cells which were chosen as a model system owing to its relevance in the production of therapeutic proteins. Specifically, we analysed CHO cells undergoing butyrate treatment which is known to affect cell cycle regulation and to increase the specific productivity of recombinant proteins. By this means, we identified sequences for >13 000 CHO genes which added sequence information of $5000 novel genes to the CHO model. More than 6000 transcript sequences are predicted to be complete, as they covered >95% of the corresponding mouse orthologs. Detailed analysis of selected biological functions such as DNA replication and cell cycle control, demonstrated the potential of NGS expression profiling in organisms without extended genome sequence to improve both data quantity and quality.
Biotechnology and Bioengineering, 2010
Increase in both productivity and product yields in biopharmaceutical process development with re... more Increase in both productivity and product yields in biopharmaceutical process development with recombinant protein producing mammalian cells can be mainly attributed to the advancements in cell line development, media, and process optimization. Only recently, genome-scale technologies enable a system-level analysis to elucidate the complex biomolecular basis of protein production in mammalian cells promising an increased process understanding and the deduction of knowledge-based approaches for further process optimization. Here, the use of gene expression profiling for the analysis of a low titer (LT) and high titer (HT) fed batch process using the same IgG producing CHO cell line was investigated. We found that gene expression (i) significantly differed in HT versus LT process conditions due to differences in applied chemically defined, serum-free media, (ii) changed over the time course of the fed batch processes, and that (iii) both metabolic pathways and 14 biological functions such as cellular growth or cell death were affected. Furthermore, detailed analysis of metabolism in a standard process format revealed the potential use of transcriptomics for rational media design as is shown for the case of lipid metabolism where the product titer could be increased by about 20% based on a lipid modified basal medium. The results demonstrate that gene expression profiling can be an important tool for mammalian biopharmaceutical process analysis and optimization.
Analytical and Bioanalytical Chemistry, 2010
The "Pharmaceutical Current Good Manufacturing Practices (CGMPs) for the 21st Century-A Risk Base... more The "Pharmaceutical Current Good Manufacturing Practices (CGMPs) for the 21st Century-A Risk Based Approach" initiative announced by the FDA in August 2002 to improve and modernize pharmaceutical manufacturing facilitated adoption of process analytical technology (PAT) by the pharmaceutical industry. The potential for improved operational control and compliance resulting from continuous real-time quality assurance was highlighted as a likely benefit that would result from PAT implementation. A considerable amount of work has been done on this topic by academic and industrial contributors in the last decade. In this paper, we will start with a brief overview of evolution of PAT concepts and a review of their application in the wider pharmaceutical industry. The rest of the paper focuses on PAT applications for biotech processes with emphasis on developments in the last five years. It is our observation that while significant advances have been accomplished with regard to our ability to analyze/monitor key process and quality attributes in the biotech industry, much more needs to be done with regard to utilizing the collected data for subsequent control of the process, to achieve optimum yield and product quality. The latter is necessary to achieve the benefits that will result from PAT implementation.