Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:2.4.2.8 (hypoxanthine-guanine phosphoribosyltransferase)
2,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The gene for the purine salvage enzyme hypoxanthine phosphoribosyltransferase (HPRT) is expressed at a low level in many cells. As is the case with several other "housekeeping genes," thorough studies of hprt gene regulation have been hampered by the low levels of its mRNA. We have used RNA/RNA hybridization in solution to determine the concentration of hprt-RNA in human cells. The sensitivity and specificity of the method have been validated, and it is shown that hprt-RNA can be accurately determined at a level of a few mRNA molecules per cell. As expected for a housekeeping gene, low and relatively constant hprt-RNA levels (0.3-0.8 pg/micrograms DNA) were found in primary cultures of normal amnion cells and fibroblasts, EBV-transformed lymphoblastoid cell lines, neuroblastoma, glioblastoma, and melanoma cell cultures. While resting lymphocytes were found to contain very low amounts of hprt-RNA, lymphocytes stimulated with phytohemagglutinin (PHA) showed a 10-fold increase to about 0.8-1.2 pg/microgram DNA, which corresponds to 6-10 hprt-RNA molecules per cell. The level started to increase about 20 h after PHA stimulation, 5-10 h before the onset of DNA synthesis, and a steady-state level was reached after 2-3 days in culture. In PHA-stimulated lymphocytes from two brothers with inherited HPRT deficiency (Lesch-Nyhans syndrome), the hprt-RNA level in PHA-stimulated lymphocytes was only about 25% of that in normal subjects. In T-cells selected for HPRT deficiency by growth in 6-thioguanine medium, the levels of hprt-RNA were either normal or very low, which probably reflects the different nature of the mutations involved. These results demonstrate the sensitivity of this method for determinations of low levels of RNA and clearly show induction of hprt-RNA after mitogenic stimulation of human lymphocytes.
...
PMID:Levels of hypoxanthine phosphoribosyltransferase RNA in human cells. 168 3

Some cell types within the human melanoma cell line MeWo contain homogeneously staining regions (HSRs) consisting of repetitive DNA sequences and ribosomal RNA (rRNA) genes derived from chromosome 15. To further examine the association between enhanced tumorigenicity and the presence of HSR-bearing chromosomes, hybrid cell lines were constructed by fusing X-HSR-containing MeWo cells with ouabain-resistant, HPRT-deficient Chinese hamster ovary cells and culturing in HAT medium containing ouabain. A hybrid containing the X-HSR chromosome and several MeWo chromosomes was more tumorigenic in BALB/c nude mice than derivative cells lacking the X-HSR and human chromosome 18. However, since this enhanced tumorigenicity could be due to sequences on either the X-HSR or chromosome 18, a second series of hybrids was constructed by micro-cell fusion. In this case, the tumorigenicity of hybrid cells containing 2 copies of the X-HSR as the only MeWo chromosome was similar to that of derivative cells lacking these chromosomes. Cytogenetic analysis revealed that the nucleolar organizer regions (NORs) on the HSR were inactive in the hybrid cells. Our data indicate that DNA sequences amplified on MeWo HSRs do not enhance tumorigenicity under experimental conditions in which rRNA genes are not expressed. As the only active NORs in MeWo HSR-containing cells are on the HSRs, we suggest that expression of these amplified rRNA genes is responsible for the selective growth advantage of these cell types in nude mice. Our data also indicate that the enhanced tumorigenicity of MeWo HSR-containing cells is not due to co-amplification of a dominant oncogene.
...
PMID:Relative tumorigenicities of hybrid cells with and without HSR-bearing chromosomes from a human melanoma cell line. 275 41

The hereditary dysplastic nevus syndrome (DNS) is a well-characterized disorder in which affected individuals have increased numbers of premalignant (dysplastic) nevi and a markedly increased risk of developing cutaneous melanoma. Seeking evidence of a systemic disorder in DNS, we examined the effect of ultraviolet radiation on cultured lymphoid cells. Epstein-Barr virus-transformed lymphoblastoid cell lines from patients with hereditary DNS had similar survival values following treatment with 2.3 to 9.0 J of 254-nm ultraviolet radiation per m2 as did lines from control individuals. Mutagenesis at the hypoxanthineguanine phosphoribosyltransferase locus was assessed by measuring the induction of resistance to thioguanine using a microtiter well assay. Three lymphoblastoid cell lines from patients with hereditary DNS and melanoma had a 2- to 3-fold greater frequency of induced mutants per clonable cell than three normal lines following exposure to 4.5 to 9.0 J of ultraviolet radiation per m2. Expanded clones of mutated DNS lymphoblastoid cell lines had less than 6% of normal hypoxanthine-guanine phosphoribosyltransferase activity. Inhibition and recovery of DNA synthesis following ultraviolet exposure were similar in 2 DNS and 2 normal lines. Repair by DNS lines of ultraviolet-induced DNA damage was in the normal range as measured by alkaline elution. Thus, hereditary DNS exhibits in vitro hypermutability which may reflect increased susceptibility to ultraviolet-induced somatic mutations in vivo. This abnormality may be related to the increased melanoma susceptibility of patients with hereditary DNS.
...
PMID:Hereditary dysplastic nevus syndrome: lymphoid cell ultraviolet hypermutability in association with increased melanoma susceptibility. 394 Jun 25

In studies of gene regulation using somatic cell fusion techniques, the analysis of heterokaryons circumvents several problematic aspects of the more traditional approach utilizing proliferating hybrid cells. We have analyzed the expression of muscle specific properties in heterokaryons between muscle and nonmuscle cells in order to investigate whether differentiating cells contain regulatory factors that repress the expression of alternative developmental pathways. Heterokaryons and cybrids were derived from polyethylene glycol-mediated fusion of differentiated mononucleate chicken myocytes with mouse melanoma cells, mouse melanoma cytoplasts, chicken fibroblasts, or other chicken myocytes. Our results demonstrate that fusion of a myocyte with a nonmyogenic cell generally results in extinction of muscle-specific properties in the immediate fusion product. Myocyte X melanoma heterokaryons ceased to express the skeletal muscle forms of myosin, desmin and creatine kinase, reinitiated DNA synthesis, and showed a loss of spontaneous fusion competence within 96 hr after their formation. Although chicken myocyte X mouse melanoma heterokaryons showed extinction of muscle specific properties, they continued to synthesize protein and to incorporate [3H]hypoxanthine, presumably due to the continued production of constitutive chicken HPRT. That presence of the melanoma nucleus was required for extinction to be observed was demonstrated by the continued expression of muscle proteins in cybrids between chicken myocytes and melanoma cytoplasts. Significantly, heterokaryons between chicken myocytes and chicken fibroblasts also exhibited extinction of muscle proteins, demonstrating for the first time that extinction is not restricted to fusions in which at least one parental cell type was derived from an established cell line. Our results strongly support the notion that extinction reflects cell-type specific gene regulatory mechanisms operative during development.
...
PMID:Extinction of muscle-specific properties in somatic cell heterokaryons. 669 89

Hybrids formed between HPRT- Cloudman mouse melanoma and normal cells were isolated. The parental origin of the hybrids was verified by isoenzyme and karyotype analyses. These hybrid cells differed in two major characteristics from hybrids of melanoma and established fibroblastic cells. (1) They grew as tumors when injected into mice, and (2) they expressed differentiated melanocytic functions. At least one of the differentiated functions was overexpressed. The specific activity of tyrosinase was 3-20 times higher in the hybrid cells than in the parental mouse melanoma. The overexpression of tyrosinase in these hybrid cells has been stable for more than a year, has been transmitted to subclones of the original hybrid cell lines, and has been expressed in tumors that grew after injections of hybrid cells into animals.
...
PMID:Supermelanotic hybrids derived from mouse melanomas and normal mouse cells. 676 42

Dacarbazine (DTIC) is a DNA-methylating drug used in the treatment of malignant melanoma. Among the DNA dducts induced by DTIC are N7-methylguanine (N7-meG) and O6-methylguanine (O6-meG). The latter adduct, in particular, may be important in the mutagenic as well as the cytotoxic activity of DTIC. Repair of O6-meG is carried out by the enzyme O6-alkylguanine-DNA-alkyltransferase (AGT) by a process which results in its autoinactivation. N7-meG is lost from DNA partly spontaneously and partly by enzymatic depurination followed by excision repair of the resulting apurinic site. The purpose of this study was to determine the in vivo kinetics of formation and repair of O6-meG and N7-meG and the changes in AGT in peripheral WBCs with repeated doses of DTIC, and to determine the effects on these processes of concomitant administration of hydroxyurea. In addition, we examined the induction of mutations at the HPRT gene locus. Thirty-four patients with malignant melanoma received 1.0 g/m2 DTIC i.v. every 3 weeks. Hydroxyurea was added to the second and subsequent doses of DTIC in 19 patients. The concentrations of O6-meG, N7-meG, and AGT in peripheral blood lymphocytes were determined up to 24 h after each of the first two doses of DTIC. Mutations at the HPRT gene locus were determined using the T-cell clonal assay. Peak O6-meG levels were detected 1 and 4 h after the first and second dose of DTIC, respectively. AGT concentrations declined to 56.7% (range, 40.3-76.9%) and 55.0% (range, 45.4-58.9%) of pretreatment levels 24 h after the first and second doses of DTIC, respectively, and were still approximately 25%below their initial levels just prior to administration of the second dose of DTIC. An increase in formation of O6-meG was observed at all time points after the second dose of DTIC (P = 0.0001), which was not affected by cotreatment with hydroxyurea (P > 0.5). There was a negative correlation between pretreatment AGT levels and the O6-meG concentration at 24 h after therapy (r = -0.554, P = 0.014). N7-meG levels peaked at 6 h after DTIC therapy and were not significantly influenced by the cycle number. Cotreatment with hydroxyurea tended to be associated with lower levels of N7-meG (P = 0.08). There was no correlation between either O6-meG or N7-meG levels and the grade of neutropenia. On the basis of a limited series of blood samples analyzed, there was no firm evidence that chemotherapy with DTIC resulted in induction of HPRT mutations in lymphocytes. In conclusion, repeated administrations of DTIC resulted in higher concentrations of O6-meG, probably due to reduction in cellular AGT. Hydroxyurea did not significantly influence the kinetics of O6-meG, and N7-meG adduct formation. There was no significant induction of HPRT gene mutations with DTIC. This study suggests that sequencing of DTIC doses should be evaluated using the time course of cellular AGT depletion and DNA adduct formation to achieve higher cytotoxic efficiency.
...
PMID:Methyl DNA adducts, DNA repair, and hypoxanthine-guanine phosphoribosyl transferase mutations in peripheral white blood cells from patients with malignant melanoma treated with dacarbazine and hydroxyurea. 981 73

An unusual cluster of 8 breast cancer and 8 other malignant tumor cases (ovarian, uterus, lung, colon and brain tumors and malignant melanoma) developed in a period of 12 years among 98 nurses exposed to ethylene oxide (EtOx) for 5 15 years in a unit using gas sterilizer in a hospital of the archiepiscopal city of Eger, Hungary. EtOx concentration in air samples of the working area varied from 5 to 150 mg/m3. The question was, if there was any causal relationship between the elevated incidence of breast cancer and the EtOx exposure, the other possibility was, that this cluster appeared accidentally. EtOx is a human carcinogen, however, no increased breast cancer incidence in EtOx-exposed subjects was reported in the literature. We followed up for two consecutive years the 27 non cancer patients, EtOx-exposed nurses and 11 unexposed hospital controls with the aid of a multiple genotoxicology monitor including chromosomal aberration, sister-chromatide exchange, HPRT point mutation and DNA repair studies. The results were compared with data from 30 local historical controls, 48 historical controls from Budapest, 14 hospital controls and 9 EtOx exposed nurses from Budapest. Significantly high chromosome aberration yields (especially chromosome type exchanges) were alike detected in EtOx-exposed and the two other control groups in Eger. These results could not be interpreted as a consequence of EtOx exposure only, since in the EtOx-exposed group from Budapest, beside an increased total aberration frequency, the obtained exchange type aberration yields were as low as the historical controls. A plausible explanation can be the natural low dose radioactivity (222Rn) of the local tap-water due to a specific geological situation in Eger. The spontaneous breast cancer incidence in Hungary doubled in the last 10 years compared with the previous 20 years (1960 1980), especially in Eger. The appearance of the high breast cancer incidence in the hospital of Eger indicates the combined effect of EtOx and a more common local etiologic factor, such as the naturally radioactive tap-water. However, since the reported studies did not involve the investigation either of the genetic predisposition, or the effects of other possible environmental, occupational, and/or life style confounding factors, further studies (partly in progress) are necessary to clarify the importance of these factors.
...
PMID:Is breast cancer cluster influenced by environmental and occupational factors among hospital nurses in Hungary? 1039 63

Tumor-cells have been shown to elicit MHC-restricted and antigen-specific T-cell responses. In this article, we used a new approach to study T-cell responses in tumor-bearing patients based on a global representation of the Vbeta-transcriptome, making it possible to grade CDR3-length distribution (CDR3-LD) alterations. Six patients with advanced melanoma disease, from whom blood samples were taken before and serially after tyrosinase-A peptide vaccination, were studied. The PBMC from patients displayed highly significant Vbeta transcriptome alterations as compared to healthy individuals. Similar Vbeta alterations could be detected both in PBMCs and at the tumor site. After vaccination, Vbeta alterations could also be observed by gauging individually their transcript level but not their cell-surface expression. Some Vbeta families exhibited high Vbeta/HPRT transcript ratios (e.g., Vbeta1), which represented up to 44% of the whole transcriptome, a situation that was not reflected by an increase in the percentage of T cells that expressed the corresponding protein and was not observed in normal individuals. In several instances, CDR3-LD altered T cells exhibited MHC-restricted and tumor-specific IFNgamma or GM-CSF production. Finally, we show that the presence of a tumor and probably vaccination can affect Vbeta transcriptome patterns and induce specific clones reactive to autologous tumor or vaccinating peptides. In combination with other methods, such an approach should help in identifying the clones actually involved in the response against the tumor.
...
PMID:Blood T-cell Vbeta transcriptome in melanoma patients. 1514 62

The identification of specific lymphocyte populations that mediate tumor immune responses is required for elucidating the mechanisms underlying these responses and facilitating therapeutic interventions in humans with cancer. To this end, mutant hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficient (HPRT-) T-cells were used as probes to detect T-cell clonal amplifications and trafficking in vivo in patients with advanced melanoma. Mutant T-cells from peripheral blood were obtained as clonal isolates or in mass cultures in the presence of 6-thioguanine (TG) selection and from tumor-bearing lymph nodes (LNs) or metastatic melanoma tissues by TG-selected mass cultures. Nonmutant (wild-type) cells were obtained from all sites by analogous means, but without TG selection. cDNA sequences of the T-cell receptor (TCR) beta chains (TCR-beta), determined directly (clonal isolates) or following insertion into plasmids (mass cultures), were used as unambiguous biomarkers of in vivo clonality of mature T-cell clones. Clonal amplifications, identified as repetitive TCR-beta V-region, complementarity determining region 3 (CDR3), and J-region gene sequences, were demonstrated at all sites studied, that is, peripheral blood, LNs, and metastatic tumors. Amplifications were significantly enriched among the mutant compared with the wild-type T-cell fractions. Importantly, T-cell trafficking was manifested by identical TCR-beta cDNA sequences, including the hypervariable CDR3 motifs, being found in both blood and tissues in individual patients. The findings described herein indicate that the mutant T-cell fractions from melanoma patients are enriched for proliferating T-cells that infiltrate the tumor, making them candidates for investigations of potentially protective immunological responses.
...
PMID:Clonal expansions of 6-thioguanine resistant T lymphocytes in the blood and tumor of melanoma patients. 1871 86

In vivo hypoxanthine-guanine phosphoribosyltransferase (HPRT)-deficient T cells (MT) from melanoma patients are enriched for T cells with in vivo clonal amplifications that traffic between blood and tumor tissues. Melanoma is thus a model cancer to test the hypothesis that in vivo MT from cancer patients can be used as immunological probes for immunogenic tumor antigens. MT were obtained by 6-thioguanine (TG) selection of lymphocytes from peripheral blood and tumor tissues, and wild-type T cells (WT) were obtained analogously without TG selection. cDNA sequences of the T cell receptor beta chains (TRB) were used as unambiguous biomarkers of in vivo clonality and as indicators of T cell specificity. Public TRB were identified in MT from the blood and tumor of different melanoma patients. Such public TRB were not found in normal control MT or WT. As an indicator of T cell specificity for melanoma, the >2600 MT and WT TRB, including the public TRB from melanoma patients, were compared to a literature-derived empirical database of >1270 TRB from melanoma-reactive T cells. Various degrees of similarity, ranging from 100% conservation to 3-amino acid motifs (3-mer), were found between both melanoma patient MT and WT TRBs and the empirical database. The frequency of 3-mer and 4-mer TRB matching to the empirical database was significantly higher in MT compared with WT in the tumor (p=0.0285 and p=0.006, respectively). In summary, in vivo MT from melanoma patients contain public TRB as well as T cells with specificity for characterized melanoma antigens. We conclude that in vivo MT merit study as novel probes for uncharacterized immunogenic antigens in melanoma and other malignancies.
...
PMID:In vivo 6-thioguanine-resistant T cells from melanoma patients have public TCR and share TCR beta amino acid sequences with melanoma-reactive T cells. 2118 40


1

---