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Query: UNIPROT:P00492 (hypoxanthine-guanine phosphoribosyltransferase)
 2,385 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Analysis of a group of human/rodent somatic cell hybrids with nucleic acid probes prepared from cloned human variable region (VH), junctional (JH), and constant region (C epsilon) heavy chain immunoglobulin genes indicates that all of these IgH genes are localized on the subtelomeric (q32) band of chromosome 14. Somatic cell hybrids were isolated in selective medium after fusing human fibroblasts with hprt- Chinese hamster cells. The human parental cells contained two translocation chromosomes representing a reciprocal translocation between chromosomes X and 14. Only those hybrid cell lines retaining a complete human autosome 14 or the X/14 translocation chromosome (i.e. containing band 14q32) retained the human IgH genes. Retention of these genes did not correlate with the presence of the other translocation chromosome, 14/X. These results indicate that all human IgH genes (VH, JH, and CH) map to the same chromosomal band (14q32) which is commonly involved in reciprocal translocations with human chromosome 8 (8q24) in B-cell neoplasms.
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PMID:Localization of human variable and constant region immunoglobulin heavy chain genes on subtelomeric band q32 of chromosome 14. 681 44

With the goal of creating a strain of mice capable of producing human antibodies, we are cloning and reconstructing the human immunoglobulin germline repertoire in yeast artificial chromosomes (YACs). We describe the identification of YACs containing variable and constant region sequences from the human heavy chain (IgH) and kappa light chain (IgK) loci and the characterization of their integrity in yeast and in mouse embryonic stem (ES) cells. The IgH locus-derived YAC contains five variable (VH) genes, the major diversity (D) gene cluster, the joining (JH) genes, the intronic enhancer (EH), and the constant region genes, mu (C mu) and delta (C delta). Two IgK locus-derived YACs each contain three variable (V kappa) genes, the joining (J kappa) region, the intronic enhancer (E kappa), the constant gene (C kappa), and the kappa deleting element (kde). The IgH YAC was unstable in yeast, generating a variety of deletion derivatives, whereas both IgK YACs were stable. YACs encoding heavy chain and kappa light chain, retrofitted with the mammalian selectable marker, hypoxanthine phosphoribosyltransferase (HPRT), were each introduced into HPRT-deficient mouse ES cells. Analysis of YAC integrity in ES cell lines revealed that the majority of DNA inserts were integrated in substantially intact form.
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PMID:Analysis of the structural integrity of YACs comprising human immunoglobulin genes in yeast and in embryonic stem cells. 760 56

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

The potential of hypoxanthine and thymidine (H&T) to promote growth of CHO cells and production of monoclonal antibody (mAb) was explored in this study. It was demonstrated that H&T stimulated the initial cell growth and enhanced volumetric production of anti-human CD20 mAb by 22%, mainly through the elevated integrated viable cell concentration (IVCC). The moderate alteration in cell cycle distribution might partially account for the increased cell growth. Subsequent long-term stability studies indicated that H&T did not accelerate decay kinetics in mAb productivity. Specifically, cells under both nucleic acids-replete (H&T supplementation) and nucleic acids-hungry (methotrexate treatment) culture conditions showed similar stable mAb production during the first 2 months, followed by a gradual decline in the specific production rate (q(mAb)) with a 40% drop at the fourth month. In addition, the decreased transcript level of intracellular heavy chain (HC) of anti-human CD20 mAb correlated well with the decreased q(mAb). Furthermore, genomic mutation rate regarding the loss-of-function occurrence of the hypoxanthine-guanine phosphoribosyltransferase (HPRT) gene was determined, showing that H&T repressed the HPRT spontaneous mutation rate while methotrexate (MTX) provoked the mutation rate. Collectively, our data illustrated that H&T as potential medium additives promoted both initial cell growth and volumetric production of mAb, while not affecting the long-term stability of antibody-producing CHO cells.
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PMID:Insight into the roles of hypoxanthine and thymidine [corrected] on cultivating antibody-producing CHO cells: cell growth, antibody production and long-term stability. 2180 68