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Biomedical subjects

R F Kefford

Publications and source records attributed to R F Kefford.

At least 37 records · Page 2Linked to original sources
PubMed1994

Isolation and preliminary characterisation of an X-ray-sensitive mammalian mutant cell line (WMXRS-1).

Mammalian cell lines that are sensitive to particular genotoxic agents have proved the most effective starting point for the cloning of human DNA-repair genes. After ethyl methanesulphonate mutagenesis of the parent murine fibroblast L-cell line, a new mammalian X-ray-sensitive cell line (WMXRS-1) was isolated. For selection of the mutant, a novel detection method was used: putative X-ray-sensitive clones were identified by their lack of incorporation of the DNA precursor, bromodeoxyuridine, after irradiation. The WMXRS-1 cell line was collaterally sensitive to ultraviolet radiation and some other agents known to be removed from DNA by the nucleotide excision repair pathway, but not to bleomycin or hydrogen peroxide. In relation to the wild-type strain, WMXRS-1 showed a similar pattern of induction of micronuclei up to an X-ray dose of 4 Gray and a similar DNA double-strand break (dsb) induction profile. The overall level of dsb rejoining was the same in the parent and mutant lines. However, WMXRS-1 demonstrated a reduced initial rate of dsb-rejoining, perhaps accounting for its radiosensitivity. WMXRS-1 also showed a greater G2 cell cycle phase accumulation after treatment with mitomycin-C. The cross-sensitivity profile and strand-break rejoining deficiency phenotype of WMXRS-1 is unique amongst previously characterised mammalian mutant cell lines.

Animals↗
PubMed1994

Loss of heterozygosity and homozygous deletions on 9p21-22 in melanoma.

Recent studies have implicated chromosome 9p21-22 as a location for a gene involved in cutaneous melanoma (CM). Deletion mapping in 35 matched tumour-constitutional DNA pairs from metastatic melanomas (including one melanoma cell line) and one dysplastic naevus has been performed using six short tandem repeat polymorphic (STRP) markers (D9S157-D9S162-IFNA-D9S171-DS9126-D9S10 4 ) which span approximately 19 cM across the 9p21-22 region. Both heterozygous and homozygous deletions were observed across the region in melanomas from both sporadic and familial cases. Overall 57% (20/35) of the samples displayed some form of loss. A deletion map identifies two areas of common loss either side of the interferon gene cluster. Familial CM has previously been shown to link to the more proximal of these regions. The deleted region distal to IFNA has not been previously described in melanoma. The results imply the involvement of more than one tumour suppressor gene on 9p in CM.

Chromosome Mapping↗
PubMed1993

Loss of heterozygosity in malignant melanoma at loci on chromosome 11 and 17 implicated in the pathogenesis of other cancers.

Forty-six cases of sporadic melanoma have been investigated for loss of heterozygosity at 4 loci: D11S29 (11q23), YNZ22 (17p13.3), TP53 (17p13.1); and NM23 (17q22). Each of the loci is thought to be important in the pathogenesis of other tumours. Mutations were found infrequently at the YNZ22, NM23, and TP53 loci. At D11S29, however, the frequency of mutation in the melanoma samples was high (67%) and mutations at this locus were associated with younger age at presentation. This region of chromosome 11 is also commonly mutated in breast cancers and haematological malignancies. Genetic aberrations at D11S29 may therefore represent nonspecific mutations found in several malignancies or part of a pathway common to the malignant phenotype.

Base Sequence↗
PubMed1993

Dexamethasone decreases urokinase plasminogen activator mRNA stability in MAT 13762 rat mammary carcinoma cells.

The glucocorticoid dexamethasone was observed to decrease urokinase plasminogen activator (uPA) RNA levels from within 1 h of treatment of MAT 13762 mammary adenocarcinoma cells. The drug did not alter the rate of uPA gene transcription in these cells, but decreased the stability of cytoplasmic uPA mRNA transcripts. Results from cycloheximide and actinomycin D experiments indicated that the dexamethasone-mediated reduction in uPA RNA required both new protein and RNA synthesis. Based on these results, we propose that dexamethasone induces a short-lived protein(s) which down-regulates uPA RNA levels post-transcriptionally in these metastatic tumour cells.

Animals↗
PubMed1993

Multiple levels of post-transcriptional regulation of collagenase (matrix metalloproteinase 1) in an epithelial cell line.

Multiple levels of regulation of collagenase (matrix metalloproteinase 1; MMP-1), have been demonstrated in a clonal rat epithelial cell line (A5P/B10). Secreted enzyme could not be demonstrated in culture medium from A5P/B10 cells but, using antibodies specific for collagenase, the enzyme was detected within the cytoplasm and on the surface of the cells. A probe for rat collagenase could not detect a signal for mRNA in the cytoplasm while nuclear run-on data demonstrated that the gene for collagenase was being transcribed. Incubating the cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) significantly increased cytoplasmic mRNA levels and slightly increased the intensity of staining in permeabilized cells, but collagenase activity was still not detected in the conditioned medium. This indicated that the protein was being synthesized by the TPA-treated cells but was not being secreted into the medium. These data suggest that the secretion of collagenase may be regulated both following transcription and after the completion of translation and it is suggested that multiple levels of control may be operating to determine the rate of collagenase release and hence, the rate of collagen turnover.

Animals↗
PubMed1993

Prediction of life-expectancy in hospice patients: identification of novel prognostic factors.

The prediction of life-expectancy in terminally ill patients is important both for medical and social reasons but is widely recognized as being inaccurate. In this study we prospectively collected data items which we proposed might influence survival on 148 consecutive patients at first admission to one of two hospices. Of the 19 parameters collected, four were associated with a significantly shortened survival. These were low performance status (PS), requirement for admission at first referral to the palliative care service, elevated serum bilirubin, and hypotension. Factors previously identified as predictive of shortened survival such as hyponatraemia, weight loss, confusion and tumour type were not confirmed as statistically significant independent variables. We plan to collect these data items on future patients in order to test the validity of these results.

Adult↗
PubMed1993

Interferon-alpha 2a does not improve response or survival when combined with dacarbazine in metastatic malignant melanoma: results of a multi-institutional Australian randomized trial.

Following extensive phase II trials of the combination of dacarbazine and interferon-alpha 2a we performed a prospective, randomized, controlled trial of this combination versus dacarbazine alone as systemic therapy for symptomatic, measurable metastatic malignant melanoma. The two treatment arms were well matched for age, sex, performance, status, relapse-free survival, prior therapy and sites of disease. Therapy consisted of dacarbazine given in combination in escalating doses of 200 mg/m2, 400 mg/m2 and 800 mg/m2 i.v. every 3 weeks, or alone at 800 mg/m2 i.v. every 3 weeks. Interferon was administered subcutaneously starting at 3 mU daily on days 1-3, 9 mU daily on days 4-70, then 9 mU three times per week. Therapy was continued for at least 6 months unless overt progressive disease was observed. Eighty seven patients were randomized to the combination and 83 patients to dacarbazine alone. Response rates were respectively, complete 7% and 2%, and partial 14% and 15%, for a total response rate of 21% (95% confidence limits 13-31%) and 17% (95% confidence limits 10-27%). Median duration of response was 258 and 286 days, and survival of the whole groups 229 and 269 days respectively. Toxicity was worse in the combination arm, with more patients experiencing fatigue, nausea and anorexia, flu-like symptoms and neutropenia. However quality of life was not significantly different in either group, except that fatigue, as measured at week 12 by LASA scales, and activity, as measured by the functional living index, were both improved in the combination.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1992

Loss of keratin expression in anaplastic carcinoma cells due to posttranscriptional down-regulation acting in trans.

Rat keratin K5 and vimentin complementary DNAs have been isolated, identified, and used to study keratin and vimentin expression as markers for cell differentiation. Isologous rat neoplastic epithelial cell lines used were based on a clonal benign epithelial line (A5P/B10) and a clonal anaplastic malignant derivative line (T952/F7). Stable cytoplasmic mRNA was detected for keratin but not vimentin in the benign cells. The anaplastic derivative cells expressed vimentin but showed a 1000-fold reduction in the keratin message, which nuclear run-on assays identified as being due to posttranscriptional down-regulation. An identical pattern of posttranscriptional down-regulation was found in independent malignant somatic cell hybrids of the benign and anaplastic cells. trans-acting regulatory mechanisms implicated in posttranscriptional (pretranslational) keratin down-regulation in these anaplastic malignant cells may play a role in the apparent loss of differentiation evident in tumor progression.

Animals↗
PubMed1992

Transcriptional and posttranscriptional activation of urokinase plasminogen activator gene expression in metastatic tumor cells.

Urokinase plasminogen activator (uPA) is a serine protease which has frequently been implicated in the process of tumor cell invasion and metastasis. The degree of expression and mode(s) of regulation of the uPA gene in metastatic compared with nonmetastatic tumor cells have not yet been addressed. We have cloned and sequenced a full-length rat uPA complementary DNA and utilized Northern blot analysis to report that the uPA gene is expressed at levels 3.5- to 70-fold higher in metastatic cell lines than in nonmetastatic cell lines derived from two independent rat mammary adenocarcinomas. Nuclear run-on assays and RNA half-life estimations indicated that metastatic MAT 13762 rat mammary adenocarcinoma cells expressed 3.5-fold higher levels of uPA RNA than a nonmetastatic derivative (J-clone), due to a combined increase in uPA gene transcription and cytoplasmic RNA stability. By contrast, uPA RNA (and enzyme) levels were elevated by up to 70-fold in metastatic clones of dimethylbenz(a)anthracene-induced rat mammary adenocarcinoma (DMBA-8) due to predominantly posttranscriptional mechanisms. Moreover, treatment of nonmetastatic DMBA-8 cell lines with protein synthesis inhibitors led to an increase in nuclear and cytoplasmic uPA RNA levels, without altering the rate of uPA gene transcription. These results suggest that in addition to gene transcription, posttranscriptional events localized in the nucleus and cytoplasm are key determinants of uPA gene activation in rat mammary adenocarcinomas.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1992

Post-transcriptional regulation of urokinase plasminogen activator gene expression occurs in the nucleus of BC1 rat mammary tumor cells.

The regulation of urokinase plasminogen activator (uPA) expression was investigated in 2 highly metastatic rat mammary adenocarcinoma cell lines, BC1 and MAT 13762. BC1 cells were observed to synthesize, on average, 10 times less uPA enzyme and mRNA than MAT 13762 cells; however this difference was not accounted for by differences in uPA gene copy number/structure or in the rate of uPA gene transcription in the cell lines studied. Moreover, Northern blot analysis of invasive sub-populations derived in vitro from the BC1 cell line revealed levels of uPA expression similar to those of the parent, but a 3-fold elevation in expression of the metalloprotease gene, transin. Further investigation showed that treatment of BC1 cells with either of the protein synthesis inhibitors, cycloheximide or anisomycin, increased the level of both nuclear and cytoplasmic uPA RNA 6- to 18-fold in 4 hr, whilst inducing a maximum 2.6-fold increase in the rate of uPA gene transcription. This increase in uPA gene expression may therefore reflect, in part, an increase in the stability and/or processing of nuclear uPA transcripts. These results suggest that the degree of uPA gene expression does not correlate directly with BC1 tumor-cell invasion in vitro, and that the uPA gene is down-regulated, at least in part, post-transcriptionally in the nucleus of BC1 mammary tumor cells.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1992

Amplification of HER-2/neu oncogene in human ovarian cancer.

Amplification and/or increased expression of the HER-2/neu oncogene has been reported to occur in ovarian tumors and possibly to correlate with biologic behavior and prognosis. The frequency with which amplification is reported to occur is quite variable ranging from 0-30% in different series and this variability is probably accounted for by technical and methodologic factors. The variability and lack of reproducibility has raised questions about the usefulness of assessing amplification of the HER-2/neu oncogene and in particular its clinical relevance. In this study by using strict criteria for amplification and using multiple controls we could demonstrate unequivocal amplification of the HER-2/neu oncogene by Southern blot analysis in only 11% of malignant ovarian tumors. The potential pitfalls with the techniques used to detect HER-2/neu oncogene amplification and overexpression are reviewed and possible ways to overcome some of the problems are suggested.

Journal Article↗
PubMed1992

Carboplatin-containing regimens for small cell lung cancer: implications for management in the elderly.

The efficacy and toxicity of two regimens based on etoposide/carboplatin with or without cyclophosphamide/vincristine in the management of small cell lung cancer (SCLC) were assessed by the Australian Lung Cancer Study Group. Response rates of 77% and 85% were noted for the two- and four-drug regimens, respectively, among patients with limited disease (LD). Response rates among patients with extensive disease (ED) were 58% and 79%, respectively. The profiles of nonhematologic toxicity were modest; myelosuppression was dose-limiting when colony-stimulating factors were not used. Twenty-six patients (14%) were older than 70 years of age. Although hematologic toxicity was more severe in the elderly group, there was no significant difference in nonhematologic toxicity, response rate, or overall survival between the geriatric and younger groups. When LD only was considered, 33% of those younger than 70 were alive at 2 years; no patients aged 70 years or older with LD were alive beyond 2 years. In patients with ED, there was no age-related difference in survival. Cytotoxic regimens based on etoposide/carboplatin constitute useful treatment for SCLC, with high response rates and manageable toxicity, irrespective of patient age.

Adult↗
PubMed1992

SCL gene in human tumors.

The SCL gene encodes a member of the helix-loop-helix (HLH) family of transcription factors and is reportedly involved in up to 25% of T-cell acute lymphoblastic leukemia (T-ALL). We have surveyed over 120 primary human tumors including melanomas, myeloid, and lymphoid leukemias, and other solid tumors without evidence of rearrangements involving SCL. These results are further supported by low level expression of SCL in these tumors (as assessed by a polymerase chain-reaction-based method). We conclude that rearrangement/translocation with subsequent activation of SCL occurs infrequently in myeloid leukemias and melanomas.

Base Sequence↗