Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.4.2.8 (hypoxanthine-guanine phosphoribosyltransferase)
 2,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An amino acid sequence variant is defined as an unintended amino acid sequence change and contributes to product heterogeneity. Recombinant monoclonal antibodies (MAbs) are primarily expressed from Chinese Hamster Ovary (CHO) cells using stably transfected production cell lines. Selections and amplifications with reagents such as methotrexate (MTX) are often required to achieve high producing stable cell lines. Since MTX is often used to generate high producing cell lines, we investigated the genomic mutation rates of the hypoxanthine-guanine phosphoribosyltransferase (HGPRT or HPRT) gene using a 6-thioguanine (6-TG) assay under various concentrations of MTX selection in CHO cells. Our results show that the 6-TG resistance increased as the MTX concentration increased during stable cell line development. We also investigated low levels of sequence variants observed in two stable cell lines expressing different MAbs. Our data show that the replacement of serine at position 167 by arginine (S167R) in the light chain of antibody A (MAb-A) was due to a genomic nucleotide sequence change whereas the replacement of serine at position 63 by asparagine (S63N) in the heavy chain of antibody B (MAb-B) was likely due to translational misincorporation. This mistranslation is codon specific since S63N mistranslation is not detectable when the S63 AGC codon is changed to a TCC or TCT codon. Our results demonstrate that both a genomic nucleotide change and translational misincorporation can lead to low levels of sequence variants and mistranslation of serine to asparagine can be eliminated by substituting the TCC or TCT codon for the S63 AGC codon without impacting antibody productivity.
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PMID:Mechanisms of unintended amino acid sequence changes in recombinant monoclonal antibodies expressed in Chinese Hamster Ovary (CHO) cells. 2050 32

UVB and UVC toxicity was detected in Chinese Hamster Ovary (CHO) cell lines AA8, UV5 and XEM2 (a V79-derived cell line expressing rat P450 1A1). Unlike FICZ-HPLC assay that showed induction of CYP1A1 enzyme activity after 20 minutes and 2 hour UVC exposure, the EROD assay showed no difference in cytochrome P450 1A1 (CYP1A1) activity after exposure to different doses of UVB and UVC light. Different cytotoxic and mutagenic effect of photo lesions induced by UVC and UVB light was investigated with the DRAG and HPRT assays, comparing the wild type cell line AA8 and the Nucleotide Excision Repair (NER) deficient cell line UV5. DRAG assay showed a significant difference in UV induced cytotoxicity between UVC and UVB reflecting the larger energy and toxic effect of UVC along with significant difference in UV induced toxicity between AA8 and UV5 cell lines. This was further validated through the HPRT assay, which also showed a significant difference in UVC (5 J/m(2)) induced mutagenic effect between these cell lines. In addition, HPRT assay showed the mutagenic effect of photosensitizer, acetophenone. These results show that UVB and UVC generate serious damage through photo products on DNA, and might induce the metabolic activity of CYP1A1.
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PMID:Cellular and genomic toxicity produced by UV light in Chinese hamster ovary cells. 2457 18