scholarly journals Glycoengineering Chinese hamster ovary cells: a short history

Author(s):  
Roberto Donini ◽  
Stuart M. Haslam ◽  
Cleo Kontoravdi

Biotherapeutic glycoproteins have revolutionised the field of pharmaceuticals, with new discoveries and continuous improvements underpinning the rapid growth of this industry. N-glycosylation is a critical quality attribute of biotherapeutic glycoproteins that influences the efficacy, half-life and immunogenicity of these drugs. This review will focus on the advances and future directions of remodelling N-glycosylation in Chinese hamster ovary (CHO) cells, which are the workhorse of recombinant biotherapeutic production, with particular emphasis on antibody products, using strategies such as cell line and protein backbone engineering.

Author(s):  
Shazid Md. Sharker ◽  
Md. Atiqur Rahman

Most of clinical approved protein-based drugs or under in clinical trial have a profound impact in the treatment of critical diseases. The mammalian eukaryotic cells culture approaches, particularly the CHO (Chinese Hamster Ovary) cells are mainly used in the biopharmaceutical industry for the mass-production of therapeutic protein. Recent advances in CHO cell bioprocessing to yield recombinant proteins and monoclonal antibodies have enabled the expression of quality protein. The developments of cell lines are possible to upgrade specific productivity. As a result, it holds an interesting area for academic as well as industrial researchers around the world. This review will concentrate on the recent progress of the mammalian CHO cells culture technology and the future scope of further development for the mass-production of protein therapeutics.


1984 ◽  
Vol 4 (1) ◽  
pp. 173-180 ◽  
Author(s):  
S W Stanfield ◽  
D R Helinski

Small polydisperse circular (spc) DNA was isolated and cloned, using BglII from Chinese hamster ovary (CHO) cells. The properties of 47 clones containing at least 43 different BglII fragments are reported. The majority of the clones probably contain entire sequences from individual spcDNA molecules. Most of the clones were homologous to sequences in CHO cell chromosomal DNA, and many were also homologous to mouse LMTK- cell chromosomal sequences. The majority of homologous CHO cell chromosomal sequences were repetitive, although a few may be single copy. Only a small fraction of cloned spcDNA molecules were present in every cell; most occurred less frequently than once in 15 cells. Localization studies indicated that at least a portion of spcDNA is associated with the nucleus in CHO cells.


1977 ◽  
Vol 73 (1) ◽  
pp. 200-205 ◽  
Author(s):  
A S Weissfeld ◽  
H Rouse

When exponentially growing CHO cells were deprived of arginine (Arg), cell multiplication ceased after 12 h, but initiation of DNA synthesis continued: after 48 h of starvation with continuous [3H]thymidine exposure, 85% of the population had incorporated label, as detected autoradiographically. Consideration of the distribution of exponential cells in the various cell cycle phases leads to a calculation that most cells in G1 at the time that Arg was removed, as well as those in S, engaged in some DNA synthesis during starvation. In contrast, isoleucine (Ile)-starved cells did not initiate DNA synthesis, as has been reported by others. Experiments with cells synchronized by mitotic selection confirmed this difference in Arg- and Ile- deprived behavior, but also showed that cells which underwent the mitosis leads to G1 transition during Arg starvation remained arrested in G1 (G0?). The results suggest that Arg-deprived cells continue to maintain some proliferative function(s) while Ile-deprived cells do not.


1977 ◽  
Vol 73 (3) ◽  
pp. 601-615 ◽  
Author(s):  
RR Gould ◽  
GG Borisy

The structure and function of the centrosomes from Chinese hamster ovary (CHO) cells were investigated by electron microscopy of negatively stained wholemount preparations of cell lysates. Cells were trypsinized from culture dishes, lysed with Triton X-100, sedimented onto ionized, carbon-coated grids, and negatively stained with phosphotungstate. The centrosomes from both interphase and dividing cells consisted of pairs of centrioles, a fibrous pericentriolar material, and a group of virus-like particles which were characteristic of the CHO cells and which served as markers for the pericentriolar material. Interphase centrosomes anchored up to two dozen microtubules when cells were lysed under conditions which preserved native microtubules. When Colcemid-blocked mitotic cells, initially devoid of microtubules, were allowed to recover for 10 min, microtubules formed at the pericentriolar material, but not at the centrioles. When lysates of Colcemid-blocked cells were incubated in vitro with micotubule protein purified from porcine brain tissue, up to 250 microtubules assembled at the centrosomes, similar to the number of microtubules that would normally form at the centrosome during cell division. A few microtubules could also be assembled in vitro onto the ends of isolated centrioles from which the pericentriolar material had been removed, forming characteristic axoneme- like bundles. In addition, microtubules; were assembled onto fragments of densely staining, fibrous material which was tentatively identified as periocentriolar material by its association of CHO can initiate and anchor microtubules both in vivo and in vitro.


1996 ◽  
Vol 313 (3) ◽  
pp. 991-996 ◽  
Author(s):  
Michael R. NARKEWICZ ◽  
S. David SAULS ◽  
Susan S. TJOA ◽  
Cecilia TENG ◽  
Paul V. FENNESSEY

Serine hydroxymethyltransferase (SHMT) is the primary enzyme in the interconversion of serine and glycine. The roles of mitochondrial and cytosolic SHMT in the interconversion of serine and glycine were determined in two Chinese hamster ovary (CHO) cell lines that both contain cytosolic SHMT but either have (CHOm+) or lack (CHOm-) mitochondrial SHMT. Mitochondrial SHMT activity was significantly reduced in CHOm- (0.24±0.11 nmol/min per mg of mitochondrial protein) compared with CHOm+ (3.21±0.66 nmol/min per mg of mitochondrial protein; P = 0.02) cells, whereas cytosolic SHMT activity was similar in CHOm- and CHOm+ cells (1.09±0.31 and 1.53±0.12 nmol/min per mg of cytosolic protein respectively; P = 0.57). In CHOm+ and CHOm- cells, the relative flux of glycine to serine measured with either [1-13C]- or [2-13C]-glycine was similar (CHOm-: 538±82 nmol/24 per mg of DNA; CHOm+: 616±88 nmol/24 h per mg of DNA; P = 0.42). In contrast, the relative flux of serine to glycine measured with [1-13C]serine was low in CHOm- cells (80±28 nmol/24 h per mg of DNA) compared with CHOm+ cells (3080±320 nmol/24 h per mg of DNA; P = 0.0001). The rate of glycine production determined by UA-2[1-13C]glycine dilution was lower in CHOm- (1200±200 nmol/24 h per mg of DNA) than CHOm+ (10200±1800 nmol/24 h per mg of DNA; P = 0.03) cells, whereas glycine utilization was similar in the two cell lines. Serine production was similar in the two cell lines but serine utilization was lower in CHOm- (3800±1200 μmol/24 h per mg of DNA) than CHOm+ (6600±1000 nmol/24 h per mg of DNA; P = 0.0002) cells. Increasing the serine concentration in the medium resulted in an increase in glycine production in CHOm+ but not in CHOm- cells. Intracellular studies with [1-13C]serine confirm the findings of decreased glycine production from serine. In CHO cells there is partitioning of intracellular serine and glycine metabolism. Our data support the hypothesis that mitochondrial SHMT is the primary pathway for serine into glycine interconversion.


1991 ◽  
Vol 37 (8) ◽  
pp. 647-650 ◽  
Author(s):  
J. M. Farber ◽  
J. I. Speirs ◽  
R. Pontefract ◽  
D. E. Conner

Five strains of nonpathogenic Listeria monocytogenes were characterized for (i) hemolysin production, (ii) cytolysis of Chinese hamster ovary (CHO) cells, and (iii) ability to attach and enter intestine 407 cells. Four of the five strains produced variable hemolysis and were weakly cytolytic for Chinese hamster ovary cells, whereas the other isolate was consistently hemolytic and strongly cytolytic for CHO cells. None of the strains was able to penetrate intestine 407 cells. In addition, two of the five strains were found to be nonmotile. Key words: Listeria monocytogenes, nonpathogenic, attachment, motility, hemolysis.


1990 ◽  
Vol 95 (4) ◽  
pp. 555-561
Author(s):  
K.J. Henle ◽  
W.A. Nagle ◽  
J.S. Bedford ◽  
W.F. Harvey

Chinese hamster ovary (CHO) cells are capable of developing a high degree of thermotolerance in response to appropriate heat conditioning. In this study we examined the relationship between thermotolerance development and protein glycosylation using four sublines of CHO cells. Two of these CHO sublines are characterized by an increased heat sensitivity and impaired cellular capacity for thermotolerance development. The data show that thermotolerance development after heat conditioning in the heat-sensitive, thermotolerance-deficient mutants was accompanied by reduced labeling of a Mr 50,000 glycoprotein (GP50), in both soluble and insoluble cell fractions. Similarly, activation of UDP-N-acetylgalactosaminyltransferase (Gal-NAcT) after hyperthermia was almost completely abolished in these cell lines. Both of these endpoints have been correlated previously with thermotolerance expression. The data are consistent with the glycosylation hypothesis that attributes increased heat resistance of thermotolerant cells, at least in part, to enhanced glycosylation and accumulation of endogenous glycoproteins, such as GP50.


1978 ◽  
Vol 79 (3) ◽  
pp. 617-622 ◽  
Author(s):  
P Stanley ◽  
J P Carver

The binding of [125I]wheat germ agglutinin ([125I]WGA) of high specific activity to Chinese hamster ovary (CHO) cells has been examined over a millionfold range of WGA concentrations and correlated with the phenomena of agglutination and capping by WGA. Analysis of the binding data by the method of Scatchard gives a complex curve indicative of positive cooperativity amongst high-affinity binding sites. Binding assays performed under conditions which inhibit capping and/or agglutination, such as low temperature or glutaraldehyde fixation, give similarly complex binding curves. Thus, the gross mobility of WGA receptors in the membrane does not appear to be responsible for the cooperative binding of WGA to CHO cells.


1990 ◽  
Vol 271 (1) ◽  
pp. 11-15 ◽  
Author(s):  
C Rojas ◽  
S Enerbäck ◽  
G Bengtsson-Olivecrona

Cultured Chinese-hamster ovary cells (CHO cells) were found to produce and secrete a lipase, which was identified as a lipoprotein lipase by the following criteria. Its activity was stimulated by serum and apolipoprotein CII, and was inhibited by high salt concentration. The lipase bound to heparin-agarose and co-eluted with 125I-labelled bovine lipoprotein lipase in a salt gradient. A chicken antiserum to bovine lipoprotein lipase inhibited the activity and precipitated a labelled protein of the same apparent size as bovine lipoprotein lipase from media of CHO cells labelled with [35S]methionine. The lipase activity and secretion were similar in growing cells and in cells that had reached confluency. Hence, lipoprotein lipase appears to be expressed constitutively in CHO cells and is not linked to certain growth conditions, as in pre-adipocyte and macrophage cell lines. At 37 degrees C, but not at 4 degrees C, heparin increased the release of lipase to the medium 2-4-fold. This increased release occurred without depletion of cell-associated lipase activity, suggesting that heparin enhanced release of newly synthesized lipase.


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