Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 7 de 7
Filter
Add more filters










Database
Language
Publication year range
1.
AAPS PharmSciTech ; 21(7): 271, 2020 Oct 06.
Article in English | MEDLINE | ID: mdl-33033946

ABSTRACT

To develop a comprehensive understanding of pharmaceutical drug substance manufacturing (DSM) processes, we conducted a data mining study to examine 50 new drug applications (NDAs) approved in 2010-2016. We analyzed the prevalence of several frequently deployed in-process control (IPC) techniques and postreaction workup procedures, as well as the operational conditions specified for reactions and workups. Our findings show that crystallization and high-performance liquid chromatography (HPLC) were the most commonly used workup steps and in-process controls, respectively, in drug substance manufacturing. On average, each NDA implemented 12.6 in-process controls and 11.3 workups. Operation time for reactions and workup procedures varied from a few minutes to multiple days, though 61% of these were between 1 and 10 h.


Subject(s)
Pharmaceutical Preparations/chemical synthesis , Crystallization , Data Mining , Quality Control
2.
BMC Med Genomics ; 4: 73, 2011 Oct 14.
Article in English | MEDLINE | ID: mdl-21996057

ABSTRACT

BACKGROUND: Identifying stable gene lists for diagnosis, prognosis prediction, and treatment guidance of tumors remains a major challenge in cancer research. Microarrays measuring differential gene expression are widely used and should be versatile predictors of disease and other phenotypic data. However, gene expression profile studies and predictive biomarkers are often of low power, requiring numerous samples for a sound statistic, or vary between studies. Given the inconsistency of results across similar studies, methods that identify robust biomarkers from microarray data are needed to relay true biological information. Here we present a method to demonstrate that gene list stability and predictive power depends not only on the size of studies, but also on the clinical phenotype. RESULTS: Our method projects genomic tumor expression data to a lower dimensional space representing the main variation in the data. Some information regarding the phenotype resides in this low dimensional space, while some information resides in the residuum. We then introduce an information ratio (IR) as a metric defined by the partition between projected and residual space. Upon grouping phenotypes such as tumor tissue, histological grades, relapse, or aging, we show that higher IR values correlated with phenotypes that yield less robust biomarkers whereas lower IR values showed higher transferability across studies. Our results indicate that the IR is correlated with predictive accuracy. When tested across different published datasets, the IR can identify information-rich data characterizing clinical phenotypes and stable biomarkers. CONCLUSIONS: The IR presents a quantitative metric to estimate the information content of gene expression data with respect to particular phenotypes.


Subject(s)
Algorithms , Biomarkers, Tumor/genetics , Oligonucleotide Array Sequence Analysis/methods , Aging , Biomarkers, Tumor/metabolism , Databases, Genetic , Gene Expression Regulation, Neoplastic , Humans , Neoplasm Staging , Neoplasms/genetics , Neoplasms/pathology , Phenotype , Recurrence
3.
J Biotechnol ; 138(3-4): 103-6, 2008 Nov 25.
Article in English | MEDLINE | ID: mdl-18778741

ABSTRACT

The cell density is an inherent constraint in commercial mammalian cell cultures. Here, we describe a cell engineering strategy utilizing the overexpression of the E2F-1 cell cycle transcription factor in CHO DG44 cells that produce a monoclonal antibody in serum-free, suspension culture. Stable pools and cell lines expressing E2F-1 were isolated that attained viable cell densities 20% higher than control cell lines and continued proliferation for an additional day in batch culture. There were no significant changes in antibody production, apoptosis, and cell cycle compared to control cells, nor were the growth effects evident in fed-batch conditions. Overall, E2F-1 overexpression postponed entry into stationary phase in mammalian cells, but perhaps novel E2F-1 variants or combination cell cycle engineering strategies will be necessary to realize significant growth benefits in commercial applications.


Subject(s)
E2F1 Transcription Factor/metabolism , Gene Expression , Animals , Blotting, Western , CHO Cells , Cell Count , Cell Culture Techniques , Cell Survival , Cricetinae , Cricetulus , Humans
4.
Biotechnol Prog ; 23(2): 506-11, 2007.
Article in English | MEDLINE | ID: mdl-17381129

ABSTRACT

Mammalian cell cultures are integral to the production of therapeutic and diagnostic proteins. A common problem encountered in culturing these cell lines, however, is a loss in viability at later stages of the cell culture process. In this study the effects of three newly synthesized chemical caspase inhibitors were investigated for their capacity to inhibit cell death. Findings show that these protease inhibitors were successful in prolonging cellular viabilities among anchorage-dependent CHO-K1 and HEK-293 cells lines. Cells treated with one inhibitor, 7312, performed as well or better when compared with the commercially known caspase inhibitor, zVAD.fmk. Suspension CHO cells producing an IgG were used to investigate the capacity of 7312 to improve protein production. Treatment of cells with 7312 increased integrated cell densities by 33% with treated cells having a higher maximum cell density and higher viability. As a result, monoclonal antibody titers increased by 20% and higher in spinner flask experiments. Increasing productivity in mammalian cell cultures has key implications for the pharmaceutical and biotechnology sectors, which are presently focused on developing methods to enhance cell performance in bioreactor environments.


Subject(s)
Caspase Inhibitors , Cell Culture Techniques/methods , Enzyme Inhibitors/administration & dosage , Kidney/physiology , Protein Engineering/methods , Recombinant Proteins/metabolism , Animals , CHO Cells , Cell Line , Cell Proliferation/drug effects , Cell Survival , Cricetinae , Cricetulus , Humans , Kidney/cytology , Kidney/drug effects
5.
Biotechnol Bioeng ; 97(3): 601-14, 2007 Jun 15.
Article in English | MEDLINE | ID: mdl-17149774

ABSTRACT

The ability to regulate apoptosis in mammalian cell cultures represents one approach to developing more economical and efficient processes. Genetic modification of cells using anti-apoptotic genes is one method that may be used to improve cellular performance. This study investigates a method to inhibit upstream apoptosis pathways through the overexpression of MDM2, an E3 ubiquitin ligase for p53. Both 293 and CHO cells expressing MDM2 were examined under both batch and spent media conditions. For batch cultures, MDM2 overexpression increased viable cell densities and viabilities over control cells with the largest enhancements observed in CHO cells. When CHO cells were passaged without medium exchange, cells expressing MDM2 reached a viable cell density that was nearly double the control and survived for an extra day in culture. When exposed to spent media initially, both 293-MDM2 and CHO-MDM2 cells continued to grow for 2 days while the control cells stopped growing after the first day. DNA analysis using flow cytometry confirmed that while CHO controls were found to be undergoing DNA fragmentation, CHO-MDM2 cells exhibit DNA degradation at a much slower rate. When compared to Bcl-2-expressing cells, MDM2 expression showed greater protection against apoptosis in passaged culture, spent medium, and following transient p53 overexpression. However, expression of the RING sequence of MDM2 responsible for E3 ligase activity without the other components of the protein was found to be toxic to 293 cells in culture. These results suggest that the overexpression of heterologous MDM2 represents a promising method to delay apoptosis in mammalian cell cultures.


Subject(s)
Apoptosis/drug effects , Proto-Oncogene Proteins c-mdm2/pharmacology , Animals , Apoptosis/physiology , CHO Cells , Cell Survival/drug effects , Cell Survival/physiology , Cells, Cultured , Cricetinae , Cricetulus , Gene Expression Regulation, Neoplastic/drug effects , Mammals , Proto-Oncogene Proteins c-mdm2/chemistry
6.
Cytotechnology ; 50(1-3): 77-92, 2006 Mar.
Article in English | MEDLINE | ID: mdl-19003072

ABSTRACT

Cell culture technology has become a widely accepted method used to derive therapeutic and diagnostic protein products. Mammalian cells adapted to grow in bioreactors now play an integral role in the development of these biologicals. A major limiting factor determining the output efficiency of mammalian cell cultures however, is apoptosis or programmed cell death. Methods to delay apoptosis and increase the longevity of cell cultures can lead to more economical processes. Researchers have shown that both genetic and chemical strategies to block apoptotic signals can increase cell culture productivity. Here, we discuss various strategies which have been implemented to improve cellular viabilities and productivities in batch cultures.

7.
Trends Biotechnol ; 22(4): 174-80, 2004 Apr.
Article in English | MEDLINE | ID: mdl-15038922

ABSTRACT

Mammalian cell culture is widely used to produce valuable biotherapeutics including monoclonal antibodies, vaccines and growth factors. Industrial cell lines such as Chinese hamster ovary (CHO), mouse myeloma (NS0), baby hamster kidney (BHK) and human embryonic kidney (HEK)-293 retain many molecular components of the apoptosis cascade. Consequently, these cells often undergo programmed cell death upon exposure to stresses encountered in bioreactors. The implementation of strategies to control apoptosis and enhance culture productivities represents a major goal of biotechnologists. Fortunately, previous research has uncovered many intracellular proteins involved in activating and inhibiting apoptosis. Here, we summarize three apoptotic pathways and discuss different environmental and genetic methodologies implemented to limit cell death for biotechnology applications.


Subject(s)
Apoptosis/physiology , Cell Culture Techniques/methods , Mammals , Animals , Apoptosis/drug effects , CHO Cells , Cricetinae , Culture Media/pharmacology , Genetic Techniques , Humans
SELECTION OF CITATIONS
SEARCH DETAIL
...