Phosphate feeding improves high-cell-concentration NS0 myeloma culture performance for monoclonal antibody production

deZengotita, Vivian M.; Miller, William M.; Auniņš, John G.; Zhou, Weichang

doi:10.1002/1097-0290(20000905)69:5<566::aid-bit11>3.0.co;2-4

Cited by 48 publications

(25 citation statements)

References 34 publications

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“…PHO compensated for the complete inhibition of aerobic ATP production via glycolysis, whereas the overall metabolic activity of MG63 cells decreased when the glycolytic rate could not be enhanced further, demonstrating a connection between lactate production and metabolic activity. Maybe because of a metabolic shift the lactate production of MG63 cells declined after reaching a maximum and lactate was consumed instead of produced (15).…”

Section: Discussionmentioning

confidence: 99%

Influence on Mitochondria and Cytotoxicity of Different Antibiotics Administered in High Concentrations on Primary Human Osteoblasts and Cell Lines

Duewelhenke

Krut

Eysel

2007

Antimicrob Agents Chemother

177

138

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Osteomyelitis, osteitis, spondylodiscitis, septic arthritis, and prosthetic joint infections still represent the worst complications of orthopedic surgery and traumatology. Successful treatment requires, besides surgical débridement, long-term systemic and high-concentration local antibiotic therapy, with possible local antibiotic concentrations of 100 g/ml and more. In this study, we investigated the effect of 20 different antibiotics on primary human osteoblasts (PHO), the osteosarcoma cell line MG63, and the epithelial cell line HeLa. High concentrations of fluoroquinolones, macrolides, clindamycin, chloramphenicol, rifampin, tetracycline, and linezolid during 48 h of incubation inhibited proliferation and metabolic activity, whereas aminoglycosides and inhibitors of bacterial cell wall synthesis did not. Twenty percent inhibitory concentrations for proliferation of PHO were determined as 20 to 40 g/ml for macrolides, clindamycin, and rifampin, 60 to 80 g/ml for chloramphenicol, tetracylin, and fluoroquinolones, and 240 g/ml for linezolid. The proliferation of the cell lines was always less inhibited. We established the measurement of extracellular lactate concentration as an indicator of glycolysis using inhibitors of the respiratory chain (antimycin A, rotenone, and sodium azide) and glycolysis (iodoacetic acid) as reference compounds, whereas inhibition of the respiratory chain increased and inhibition of glycolysis decreased lactate production. The measurement of extracellular lactate concentration revealed that fluoroquinolones, macrolides, clindamycin, rifampin, tetracycline, and especially chloramphenicol and linezolid impaired mitochondrial energetics in high concentrations. This explains partly the observed inhibition of metabolic activity and proliferation in our experiments. Because of differences in the energy metabolism, PHO provided a more sensitive model for orthopedic antibiotic usage than stable cell lines.

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Section: Discussionmentioning

confidence: 99%

Influence on Mitochondria and Cytotoxicity of Different Antibiotics Administered in High Concentrations on Primary Human Osteoblasts and Cell Lines

Duewelhenke

Krut

Eysel

2007

Antimicrob Agents Chemother

177

138

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“…Previous researchers have detected lactate consumption in hybridoma, NS0 myeloma and CHO cultures (Burky et al, 2008;deZengotita et al, 2000;Pascoe et al, 2007;Zhou et al, 1995Zhou et al, , 1997. However, these cells typically exhibit lactate production in the exponential phase and transition to consumption in the stationary phases to suggest that lactate can represent both an intermediate by-product and a carbohydrate fuel source (Brooks, 1985;Burky et al, 2008).…”

Section: Comparison Of Metabolic Profiles Of Apoptotic R and Control mentioning

confidence: 99%

Expression of anti‐apoptosis genes alters lactate metabolism of Chinese Hamster Ovary cells in culture

Dorai

Kyung

Ellis

et al. 2009

Biotech & Bioengineering

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ABSTRACT:In an effort to develop robust Chinese Hamster Ovary host cell lines, a variety of anti-apoptotic genes were over-expressed, either singly or in combination, followed by screening of transfectants for improved cell growth, extended longevity, reduced caspase 3/7 activity, and enhanced mitochondrial membrane potential (MMP). Two particular cell lines, one containing two anti-apoptotic genes, E1B-19K and Aven (EA167), and another containing three, E1B-19K, Aven, and a mutant of XIAP (EAX197), exhibited a reduction in caspase 3 activity of at least 60% and a 170% enhancement in mitochondrial membrane potential compared to controls when treated with staurosporine. In batch cell growth experiments, the peak viable cell densities and viabilities were higher resulting in a 186% increase in integrated viable cell densities. Analyses of metabolite utilization and formation of waste products indicated that the apoptotic resistant cell lines depleted all the lactate when grown in commercially available CD-CHO medium while significant levels (>1.8 g/L) accumulated in the host cell lines. When the lactate level was replenished daily in the apoptotic resistant cell lines, the cell lines consumed lactate and the culture longevity was extended up to four additional days compared to control cell lines. Furthermore, the anti-apoptosis cell lines also accumulated lower levels of ammonia. The ability of the apoptotic resistant cell lines to consume lactate was exploited by cultivating them in a ''high'' glucose medium containing 15 g/L (60 mM glucose) in which apoptotic resistant cell lines exhibited lower maximum lactate (1.8 g/L) compared to control cell lines which accumulated concentrations of lactate (2.2 g/L) that appeared to be deleterious for growth. The shaker flask titer of a therapeutic antibody product expressed in an apoptotic resistant cell line in ''high'' glucose medium reached 690 mg/ L compared to 390 mg/L for a cell line derived from a control host cell line. These results represent to our knowledge the first example in the literature in which manipulation of the apoptosis pathway has altered the nutrient consumption profile of mammalian cells in culture; findings that underscore the interdependence of the apoptotic cellular machinery and metabolism and provide greater flexibility to mammalian bioreactor process development.

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“…However, for the choice of the appropriate cell line, certain key criteria have to be considered which include: the ability to provide a close to human glycosylation pattern, the capability to produce high mAb concentrations in the chosen production system, the ability to consistently produce a product of uniform characteristics (stability), and the speed with which a high yielding cell line can be obtained [1,2]. NS0, a murine myeloma cell line, has been adopted by a number of biotechnology companies for the expression of therapeutic antibodies [3][4][5][6][7]. The successful use of NS0 cells in several fusion, transfection, selection and production approaches are among the few properties of this cell line that make it a perfect candidate for the expression of the desired product [8].…”

Section: Introductionmentioning

confidence: 99%

“…Fed batch cultures have been developed with the objective of achieving maximal increase in the culture viable cell concentrations and prolonging the culture lifetime for increased product concentrations [3,5,[11][12][13][14][15] Strategies such as development of the feeding solutions, feeding rate control, maintaining low residual glucose and/or glutamine concentrations, employing dialysis membranes to remove low molecular weight components from the culture, feeding based on Oxygen UptakeHence, based on the hypothesis that none of the factors affecting the production process can really be independent, fractional factorial design using design expert software was carried out to find the interaction between the batch (Dissolved Oxygen, Temperature and Seeding Density) and fed-batch (post-feed Glucose concentration) process parameters. The usage of design of experiments also facilitates shorter development time, hence more cost effective.…”

Section: Introductionmentioning

confidence: 99%

Study on Interactions between Fed-Batch and Batch Operating Parameters for the Development of Monoclonal Antibody Fed-Batch using Design of Experiments

Raja¹,

Anuar²,

Rahman³

et al. 2014

J Bioprocess Biotech

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In conventional fed batch process development approaches, batch operating parameters (such as pH, temperature, seeding density, dissolved oxygen concentration) are kept constant and only feeding parameters such as feeding time, post-feed concentration are manipulated. The batch and fed batch operating parameters are assumed to be independent of each other. This approach to process development ignores any interactions that might exist between the batch and fed-batch operating parameters are therefore not evaluated. However in a complex bioprocess, none of the factors affecting the process can be assumed to be independent of each other and mutually exclusive. Therefore in this study an attempt was made to study the interaction between fed-batch operating parameter-post feed glucose concentration (A) and batch operating parameters-seeding density (B), temperature (C), and dissolved oxygen concentration (D) by their simultaneous manipulation, as well as the effect of these interactions on cell growth and monoclonal antibody (mAb) production. NS0 cell line producing the mAb's against carcino-embryogenic antigen (Anti-CEA) was used. The final mAb concentration, viable cell density and integral of viable cell concentration (IVCC) were the responses evaluated. Statistical analysis experimental data showed that parameter (A) and its interaction with parameter (B) were the main factors affecting both the response variables. In comparison to the batch run which yielded 5.21 mg/L mAb, the developed fed batch process increased the mAb titer by 10 fold (59.40 mg/mL), and the IVCC was increased by 7 fold. The maximum VCD value (3.46×10 6 cells/mL) of the developed fed batch process was 1.25 over fold the value for batch.

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Phosphate feeding improves high-cell-concentration NS0 myeloma culture performance for monoclonal antibody production

Cited by 48 publications

References 34 publications

Influence on Mitochondria and Cytotoxicity of Different Antibiotics Administered in High Concentrations on Primary Human Osteoblasts and Cell Lines

Influence on Mitochondria and Cytotoxicity of Different Antibiotics Administered in High Concentrations on Primary Human Osteoblasts and Cell Lines

Expression of anti‐apoptosis genes alters lactate metabolism of Chinese Hamster Ovary cells in culture

Study on Interactions between Fed-Batch and Batch Operating Parameters for the Development of Monoclonal Antibody Fed-Batch using Design of Experiments

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