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Biomedical subjects

M Clynes

Publications and source records attributed to M Clynes.

At least 55 records · Page 3Linked to original sources
PubMed1996

Multidrug resistance in ocular melanoma.

AIMS/BACKGROUND: Metastatic disease in patients with ocular melanoma is resistant to chemotherapy. One of the main mechanisms of modulating multidrug resistance is the expression of the multidrug resistance gene 1 (MDR1) product (p-glycoprotein) by tumour cells. The purpose of this study was to evaluate the frequency of expression of the MDR1 gene in ocular melanoma whose primary treatment was surgical excision or enucleation. METHODS: Twelve recent ocular melanomas were received fresh, snap frozen and cryostat sections of tumour were analysed for expression of MDR1 by immunohistochemistry using a well characterised monoclonal antibody to MDR1. Tumour explants were established in short term tissue culture from four tumours and cell blocks were examined by immunohistochemistry. RESULTS: MDR1 expression was present in five of 12 ocular melanomas. Upregulation of protein expression was found in four cell lines established in short term culture from tumour explants. A recurrent tumour, initially treated by local excision and radioactive plaque, showed overexpression of MDR1 mRNA. CONCLUSIONS: These results suggest that significant level of MDR1 may be intrinsically present in ocular melanomas before exposure to drugs involved in multidrug resistance, and indicate the possible importance of MDR1 in modulating chemoresistance in ocular melanoma. Chemosensitisation may be of potential value in planning adjuvant chemotherapy for patients with metastatic disease.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Behavior of a novel monoclonal antibody for investigation of ovarian cancer and with possible involvement in multiple drug resistance.

Monoclonal antibodies (MAbs) were raised to an ovarian cancer cell line, OAW42, derived from a patient with a histology of serous cystadenocarcinoma of the ovary, in an attempt to identify novel antigens with a possible role in cancer medicine. One antibody P1H10, subclass IgG1, with a high titer was isolated and shown to recognize an antigen of 48 kDa. Enzyme-linked immunosorbent assay and immunocytochemical studies showed the presence of the target antigen in a number of carcinoma cell lines, including lung and breast, and in two out of three frozen breast tissue specimens. The antigen was not detected in normal human lymphocytes and there was minimal binding of the antibody to normal buccal cells. The antigen was not secreted by the OAW42 or the HepG2 cell lines and was not detected in the sera of a number of ovarian cancer patients. Indirect immunofluorescence studies confirmed the localization of the antigen to be intracellular. The binding of the antibody P1H10 to a number of multidrug resistant variants of the OAW42 cell line showed that the presence and the localization of the antigen in the drug-sensitive parental line and resistant variant cell lines was distinctly different and varied with the degree of drug resistance. The relative specificity of the antibody suggests it may be a possible diagnostic agent in human cancer. A possible role of the antigen in multiple drug resistance (MDR) is also suggested.

Animals↗
PubMed1995

Microstructural musical linguistics: composers' pulses are liked most by the best musicians.

Western music is a combination of notated structure plus microstructure. Composers' pulses, as precise microstructural functions, were tested with five groups of subjects of graded musical proficiency: (1) 10 famous artists including Vladimir Ashkenazy, Yehudi Menuhin, Paul Badura-Skoda and others; (2) 14 Julliard School of Music graduate students; (3) 19 Manhattan School of Music graduate students; (4) 51 Australian conservatory students; and (5) 41 non-music students (total, 135 subjects). Composers' pulses consist of combined amplitude and timing warps specific to each composer--the composer's pulse matrix. Each subject listened to and scored 40 versions of 10 short music examples (not selected by the author) of eight bars each, composed by Beethoven, Mozart, Haydn and Schubert, performed by computer, in which composers' pulses, as developed by Clynes (1983, 1985a,b, 1986, 1987), were incorporated in a two-level hierarchic pulse configuration, and interchanged. Each example was heard with the pulses of those four composers in turn, in quasi-random sequence, one performance thus incorporating the "right" pulse and three the "wrong" ones. Results show dramatically that the greater the musical proficiency, the more marked the preference for the "right" pulse (p < .0001). Non-musicians as a group showed a similar tendency of preference, but typically to a considerably less pronounced degree. Further studies suggest that longer excerpts (32 bars) and repeated hearing would help non-musicians also to better understand composers' pulses. A theory of music as a double stream is introduced. The results suggest that the composers' pulse matrices posited by Clynes as a first approximation are in fact appropriately meaningful. Brain function aspects of the meaningful discrimination of such tiny time differences are discussed.

Adolescent↗
PubMed1993

Human cell lines as models for multidrug resistance in solid tumours.

In spite of our expanding knowledge on the molecular biology of cancer, relatively little progress has been made in improving therapy for the solid tumours which are major killers, e.g., lung, colon, breast. Significant advances over the past 10-15 years in chemotherapy of some tumours such as testicular cancer and some leukaemias indicates that, in spite of the undesirable side-effects, chemotherapy has the potential to effect cure in the majority of patients with certain types of cancer. Multidrug resistance, inherent or acquired, is one important limiting factor in extending this success to most solid tumours. In vitro studies described in this review are now uncovering a diversity of possible mechanisms of cross-resistance to different types of drug. Sensitive methods such as immunocytochemistry, RT-PCR or in situ RNA hybridisation may be necessary to identify corresponding changes in clinical material. Only by classifying individual tumours according to their specific resistance mechanisms will it be possible to define the multidrug resistance problem properly. Such rigorous definition is a prerequisite to design (and choice on an individual basis) of specific therapies suited to individual patients. Since a much larger proportion of cancer biopsies should be susceptible to accurate analysis by the immunochemical and molecular biological techniques described above than to direct assessment of drug response, it seems reasonable to hope that this approach will succeed in improving results for cancer chemotherapy of solid tumours where other approaches such as individualised in vitro chemosensitivity testing have essentially failed. Results from clinical trials using cyclosporin A or verapamil are encouraging, but these agents are far from ideal, and reverse resistance in only a subset of resistant tumours. Proper definition of the other mechanisms of MDR, and how to antagonize them, is an urgent research priority.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1993

The use of reverse transcriptase-polymerase chain reaction (RT-PCR) to investigate specific gene expression in multidrug-resistant cells.

Expression of specific genes at the level of mRNA can be studied using techniques such as Northern blot, slot/dot blot, RNase protection assay, in situ hybridisation and RT-PCR. In this article these methods of analysis are compared; RT-PCR offers higher levels of specificity and sensitivity than traditional methods of RNA analysis and as such has become the method of choice for the study of gene expression. The RT-PCR technique is described in detail with sections dealing with RNA extraction, choice of primers (including the use of cDNA sequence data bases), PCR and RT-PCR protocols in addition to the limitations of the method. The study of one particular mRNA transcript (MDR1) using RT-PCR is discussed in detail. Recently described methods for quantitation of PCR products are discussed. Quantitative PCR would appear to offer a method of studying gene expression in a more extensive way than has been possible to date.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1993

Comparison of 5 microplate colorimetric assays for in vitro cytotoxicity testing and cell proliferation assays.

This paper describes a critical comparative evaluation of 5 miniaturised colorimetric assays applicable to cytotoxicity testing of anti-tumour drugs (and other toxins) in vitro. Each assay shows a different linear range for optical density versus cell number, a different sensitivity to change in cell number and a different minimum detectable cell number; the values of these parameters vary with experimental conditions and with cell line used. All the methods gave good correlation with viable cell number (determined by colony forming efficiency) in toxicity assays after 3 or 4 days of treatment, but they underestimated cell death after 2 days. Toxicity levels for individual chemicals (in a standard 6-day assay) are similar for the different assays, irrespective of the mechanism of action of the chemical being tested. Two of the more recently developed assays (APNaOH and SRB) were found to be very sensitive under the conditions examined.

Antineoplastic Agents↗
PubMed1993

Production of autostimulatory growth factors by the human carcinoma line, RPMI 2650.

The human carcinoma line RPMI 2650 produces autocrine factors; they are detected by the ability of RPMI 2650 conditioned medium (CM) to stimulate growth in soft agar of RPMI 2650 cells plated at low density. The autocrine activity in crude CM can be fractionated by ultrafiltration into a lower molecular weight (MW) fraction (R1-30), which concentrates molecules in the 1000-30,000 Da range; and a higher MW fraction (R30) with molecules greater than 30,000 Da in a more concentrated form. R1-30 is labile to acid, base, and heat treatment, whereas R30 is stable to (and sometimes activated by) these treatments. Boiling of R30, however, renders it labile to acid, base, and trypsin treatments. CM can be separated into a weakly heparin-binding fraction (with stability properties similar, but not identical, to R1-30), and a non-heparin binding fraction (with stability properties similar to R30). RPMI 2650 cells secrete transforming growth factor (TGF)alpha- and TGF beta-like molecules, but the R1-30 fraction can be distinguished from these TGFs, and from most other known growth factors, by its unusual combination of acid lability and weak affinity for heparin. Since the R30/non-heparin binding fraction is rendered labile by boiling or acid treatment, it may represent a bound or conformationally stable form of a growth factor.

Carcinoma, Squamous Cell↗
PubMed1993

Multiple drug resistance in the human ovarian carcinoma cell line OAW42-A.

A new multidrug-resistant variant (OAW42-A) of a human ovarian carcinoma line has been selected by exposure to increasing concentrations of doxorubicin. The variant is resistant to doxorubicin, vincristine (but surprisingly not to colchicine), etoposide, tenoposide and also to cisplatin (a drug not usually involved in classical multidrug resistance), but not to 5-fluorouracil. Overexpression of P-glycoprotein in the resistant line was demonstrated by immunofluorescence and western blotting. Direct evidence for P-glycoprotein as a determinant of resistance was provided by transfection with a specific antisense oligonucleotide. Reversal was incomplete and this, along with the pattern of cross-resistance observed, suggests that additional mechanisms of resistance may also be involved. Substantial clonal variation in resistance exists within the cell line.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1992

Multiple drug-resistance in variant of a human non-small cell lung carcinoma cell line, DLKP-A.

A 300-fold adriamycin resistant variant (DLKP-A) of the human lung squamous cell carcinoma line DLKP was established by stepwise selection in increasing concentrations of adriamycin. Different levels of cross-resistance were observed towards VP-16, VM-26, colchicine, vincristine and, somewhat unexpectedly, cis-platin. Resistance was stable for at least 3 months in culture in the absence of drug. P-glycoprotein overexpression was detected by immunofluorescence and Western Blotting, and a direct causal role for P-glycoprotein overexpression in the resistant phenotype was established by transfection with an mdr1 specific antisense oligonucleotide. A modified cryopreservation procedure was necessary for the resistant variant line. The resistant population displays clonal heterogeneity with respect to resistance level. A higher frequency of double minute chromosomes was observed in DLKP-A when compared with the parental cell line.

Antineoplastic Agents↗
PubMed1992

Cytogenetic comparison of two poorly differentiated human lung squamous cell carcinoma lines.

This paper presents a cytogenetic analysis of two established but early-passage (passages 5 and 18) cell lines derived from histologically similar, poorly differentiated lymph node metastases of squamous cell carcinoma of the lung. The cell lines showed 3 shared marker chromosomes, del(1)(q11), del(2)(p11.1) and del(2)(q11.1). One of the lines (DLKP) had 8 additional markers including structural rearrangements such as translocations and isochromosomes. Five additional markers (including two deletions of chromosome 3) were found in DLRP. A notable feature of DLRP was the high incidence of telomeric association evident in the majority of metaphase plates. Over-representation of chromosome 7 was a characteristic feature of metaphases derived from DLKP, and identification of i(21q) in this cell line was an unusual finding. The results indicate significant cytogenetic heterogeneity between these early-passage cell lines derived from two apparently histologically similar tumors.

Carcinoma, Squamous Cell↗
PubMed1990

Cytological differences between normal and malignant human cell populations in culture.

Cells from early-passage cultures of normal and malignant human tissues were analyzed for the presence of multiple nucleoli and tri- or multipolar mitoses; these properties are characteristic of malignant cell populations in tissue sections. For the cell types examined the presence of tri- or multipolar mitoses was characteristic of cells of malignant origin. Within epithelial cell populations, cells containing more than 4 nucleoli were found in populations of malignant but not of normal origin; this distinction did not apply to fibroblast populations. Large numbers of cells must be analyzed quantitatively to establish the distinctions described here, since they are properties characteristic of populations, not of individual cells. This approach may facilitate identification of normal and malignant cell populations in primary and early-passage culture.

Cell Nucleus↗
PubMed1990

Establishment of two new multi-drug resistant variants of the human tumor line Hep-2.

Two multi-drug resistant variants of the human carcinoma line Hep-2 have been selected by adaptation to progressively increasing concentrations of adriamycin. In comparison to the wild-type Hep-2 cells, the variant lines both showed approximately 100-fold resistance to adriamycin, 10 to 20-fold resistance to the vinca alkaloids but only 2-3 fold resistance to VP-16 and VM-26. There was essentially no difference between wild-type and variant cells in regard to sensitivity to threosulfan and 5-fluorouracil. The drug-resistant phenotype is stable for at least 3 months in the absence of drug, and is partially reversible by concomitant treatment with Verapamil. Chromosomal abnormalities consistent with gene amplification were observed in one of the variant lines. Sensitivity of variant cells to adriamycin was enhanced following trypsin-EDTA treatment.

Antineoplastic Agents↗