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Biomedical subjects

A G Hall

Publications and source records attributed to A G Hall.

At least 37 records · Page 2Linked to original sources
PubMed1999

Expression of DNA mismatch repair proteins in acute lymphoblastic leukaemia and normal bone marrow.

Errors during normal DNA synthesis may produce mismatched base pairs. 6-Mercaptopurine (6MP), given during continuing therapy in acute lymphoblastic leukaemia (ALL), undergoes intracellular activation to give cytotoxic thioguanine nucleotides which are then incorporated into the DNA of dividing cells in place of guanine. Cell death is thought to result from futile attempts at mismatch repair. Previous work has shown that cell lines with a defect in this pathway develop tolerance to incorporated 6-thioguanine bases. In order to investigate the possible relevance of mismatch repair to the chemosensitivty of blasts to 6MP, relative to normal tissues, we have measured the expression of the mismatch repair proteins MLH1, MSH2, PMS2 and MSH6 in blasts from children and adults with ALL and in normal bone marrows, using western blotting. Fifty cases of childhood ALL, 22 cases of adult ALL and 7 normal marrows have been studied. Expression of MSH2, and of MLH1 in all but three cases, was detectable in all the blasts studied. Noticeably, expression of MLH1 was not detected in any of the normal marrow samples. MSH2 was detected in 4 of the normal marrows. Expression of PMS2 was not detected in 29 cases of ALL and, like MLH1, was absent from each of the normal marrow samples. In contrast, MSH6 was detected in all of the normal marrows and all but 16 of the cases of ALL. There was no difference in expression between adults and children. These results may help to explain the relative sensitivity of leukaemic blasts to thiopurines at presentation as compared to normal bone marrow.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Glucocorticoid resistance and the AP-1 transcription factor in leukaemia.

Glucocorticoids have been used in the treatment of acute lymphoblastic leukaemia (ALL) for many years, initially as the only agent and then as part of multiagent chemotherapy. In ALL 20% of patients are resistant to glucocorticoids at presentation but this rises to greater than 70% on relapse. It has recently been reported that the glucocorticoid receptor inhibits activity of the AP-1 transcription factor by the ligand-dependant binding of glucocorticoid receptor (GR) to the fos and jun components of AP-1. Since AP-1 is necessary for cell proliferation, the upregulation or over-expression of AP-1 may be a mechanism of resistance to glucocorticoids. The aim of the study was to investigate whether AP-1 levels correlate with in vitro glucocorticoid resistance. In vitro sensitivity to glucocorticoids was measured using the MTT assay. AP-1 levels were quantified using gel shift analysis: a consensus sequence for the AP-1 binding site was synthesised, labelled with 32P and incubated with nuclear extracts of leukaemic blasts from 14 ALL and 26 CLL patients. Leukaemic blasts were treated with prednisolone or with vehicle alone before preparation of nuclear extracts. The gels were dried and bands quantified using a phosphorimager, using an appropriate internal standard and correcting for protein loading and cytoplasmic contamination of nuclear extracts. The patient samples fell into two distinct groups with respect to their sensitivity to glucocorticoids: AP-1 levels were significantly higher (p < 0.02) in sensitive blasts than resistant ones. There was no significant change in AP-1 levels after treating blasts for 4 hours with 0.2 mM prednisolone. No change was seen in CLL samples. These data show that glucocorticoid resistance is not associated with increased AP-1. Conversely, glucocorticoid resistance in these samples was apparently associated with decreased AP-1 levels in ALL samples. Whether this has any causal relationship to glucocorticoid resistance is unknown. Clearly, further studies on the role of AP-1 and related transcription factors is essential for understanding the control of proliferation and apoptosis in ALL.

Adult↗
PubMed1999

NCL-CD10-270: a new monoclonal antibody recognizing CD10 in paraffin-embedded tissue.

CD10 (CALLA) antigen is expressed in a wide variety of epithelial and nonepithelial tissues, but its most significant application is in the diagnosis and classification of certain types of malignant lymphoma and leukemia. CD10 is expressed in a high percentage of cases of acute lymphoblastic leukemia (ALL), follicular lymphoma, Burkitt's lymphoma, and some hematopoietic tumors. Although the antigen is not lineage specific, CD10 expression is widely used to define subgroups within B-ALL and is a useful tool for detecting the presence of leukemic blasts in the bloodstream. Currently available monoclonal antibodies to CD10 have been found to be effective only in fresh-frozen tissue and for techniques such as flow cytometry. We have used a recombinant protein corresponding to the whole of CD10 to generate a monoclonal antibody that is effective in paraffin-embedded tissue sections. We have used this antibody to assay for the presence of CD10 on a range of normal and pathological tissues. Strong staining was seen in lymphoid germinal centers, renal tubules, glomeruli, syncytiotrophoblast, hepatic parenchymal canaliculi, B-lineage ALL, follicle center cell lymphoma, and a proportion of cases of large-B-cell lymphoma. We believe that this antibody will be of value in the characterization of malignant lymphoma, in particular the differential diagnosis of small-B-cell lymphoma and subtyping of lymphoblastic leukemia, as well as the investigation of the significance of expression of CD10 in other normal and pathological tissues.

Animals↗
PubMed1999

Different expression of glutathione S-transferase alpha, mu and pi in childhood acute lymphoblastic and myeloid leukaemia.

Expression of three major classes of glutathione S-transferases (GSTs), i.e. alpha, mu and pi class, P-glycoprotein (P-gp) and multidrug resistance-associated protein (MRP) were studied in childhood acute lymphoblastic leukaemia (ALL), acute myeloid leukaemia (AML) and normal peripheral blood lymphocytes by flow cytometry. In vitro cytotoxicity of 4-hydroxy-ifosfamide (IFOS), daunorubicin (DNR) and prednisolone (PRED) was assessed by the MTT assay. Expression of alpha, mu and pi class GST did not significantly differ between leukaemic cells from 100 initial and 14 unrelated relapse ALL patients (GSTalpha P=026; GSTmu P=O009; GSTpi P=0.13). The expression of GSTalpha (1.4-fold, P=0.0004), GSTpi (13-fold, P = 0001) and to a lesser extent also GSTmu (1.1-fold, P=0.03) was higher in ALL compared with normal peripheral blood lymphocytes. Expression of GSTmu and GST7pi was significantly higher in 18 AML compared with 100 ALL patients at initial diagnosis (respectively 1.3-fold, P=0.0005 and 2-fold, P<0.0001). In contrast, GSTalpha was median 2-fold lower expressed in the AML samples (P< 0.0001). Expression levels of alpha, mu and pi class GSTs were not related to the degree of resistance to IFOS, DNR and PRED nor to immunophenotype, white blood cell count or age at presentation of childhood ALL. One exception was a remarkably low expression of GSTalpha in IFOS-sensitive samples compared with a heterogenous expression in IFOS-resistant samples (P= 0.02). Expression of GSTpi, but not of GSTalpha or GSTmu, weakly correlated with the expression of MRP (Rs 0.36, P = 0.002, n = 74) but not with P-gp. However, a high expression of both GSTpi and MRP was not associated with in vitro resistance to IFOS, DNR or PRED. The present data suggest that expression of GSTs is not linked to the degree of resistance to IFOS, DNR and PRED or clinical risk factors in childhood ALL. Whether the high expression of GSTmu and GSTpi in AML cells contributes to the relative resistance to IFOS, DNR and PRED compared with ALL samples (P < or = 0.0001) warrants further study.

Acute Disease↗
PubMed1999

Analysis of thiopurine methyltransferase variant alleles in childhood acute lymphoblastic leukaemia.

The role of 6-mercaptopurine (6MP) in the treatment of childhood acute lymphoblastic leukaemia (ALL) is well established. However, the efficacy of 6MP is significantly influenced by inactivation by the polymorphic enzyme thiopurine methyltransferase (TPMT). In the general population 89-94% have high TPMT activity, 6-11% have intermediate activity, and approximately 0.3% have low activity. Individuals with low-activity experience severe or fatal toxicity with standard 6MP doses. Prospective identification of this group of patients might prevent this problem. Recent identification of the molecular basis for low TPMT activity enabled rapid assessment of altered 6MP metabolism by PCR methods. This study evaluated the frequency of mutant TPMT alleles in 147 children with ALL. One patient was homozygous mutant (0.7%), and 16 patients were heterozygous for variant TPMT alleles (10.9%). The majority of mutant alleles were TPMT*3A. Both the allele frequency and the pattern of TPMT mutations were similar to that observed in an adult British population. The number of weeks when 6MP therapy was administered at full dose was determined in patients on MRC UKALL X and XI. The 94 patients spent a median 11% of the maintenance period receiving no therapy as a result of haematological toxicity. There was no significant difference in the number of weeks when no therapy could be administered among patients with a wild-type or heterozygous genotype. However, the one patient with a homozygous mutant genotype had severe haematological toxicity and no therapy could be administered for 53% of the maintenance period. This study demonstrates that 11.6% of the children had variant TPMT alleles. Prospective identification of TPMT genotype may be a promising tool for decreasing excessive haematological toxicity in individuals with low activity.

Adult↗
PubMed1999

Review: The role of glutathione in the regulation of apoptosis.

Recent advances in the study of the regulation of cell death by apoptosis suggest that changes in mitochondrial permeability frequently precede the development of morphological features such as chromatin condensation, phosphatidylserine inversion of the outer cell membrane and the activation of endonucleases. There is evidence that this permeability transition is associated with, and may be regulated by, changes in the intracellular redox potential. The role of the tripeptide thiol glutathione in the regulation of apoptosis-associated redox changes and the control of mitochondrial membrane permeability is reviewed in this article.

Animals↗
PubMed1999

Organizational characteristics of solid-organ donor hospitals and nondonor hospitals.

CONTEXT: Efforts to increase organ donation include serious attempts in hospital settings, where unrealized donation potential exists. Research on hospital donation must include understanding organizational as well as patient-specific influences on the donation process. OBJECTIVE: To identify organizational characteristics that distinguish hospitals producing organ donations from those that do not, and to estimate the number of nondonor hospitals with donor potential. DESIGN: Data from the American Hospital Association's 1992 annual survey of hospitals were matched to Organ Procurement and Transplantation Network information from the United Network for Organ Sharing regarding the number of solid-organ donors in 1992. Hospitals with donation capability were identified, based on bed size and factors necessary to produce successful donor maintenance and organ recovery. Based on statistical analyses, organizational characteristics distinguishing donor hospitals from nondonor hospitals were identified. We also compared the number of donors and the number of donor hospitals in 1992 and 1996. SETTING: United States. RESULTS: Among all hospitals affiliated with the American Hospital Association (n = 5607), 1214 (22%) were identified as donor hospitals (> or = 1 donation in 1992). Of 2333 hospitals with procurement capability, 1268 (54%) produced no donors in 1992. Based on a multiple logistic regression model, donor hospitals differed from nondonor hospitals by hospital ownership, with municipally owned hospitals more likely and federally owned hospitals less likely to produce donation, compared with for-profit and not-for-profit hospitals. Other organizational characteristics associated with donor hospitals were level of trauma services, whether the hospital had a transplant surgery program or a hospital ethics committee, and whether it was located in the South Atlantic, Southwest Central, or Pacific regions of the United States. CONCLUSIONS: Among hospitals not currently producing organ donations, there is a sizable subgroup with donor potential. This area merits further attention.

American Hospital Association↗
PubMed1998

The relationship between thiopurine methyltransferase activity and genotype in blasts from patients with acute leukemia.

The level of expression of the enzyme thiopurine methyltransferase (TPMT) is an important determinant of the metabolism of thiopurines used in the treatment of acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML). Studies in red blood cells (RBC) have shown that TPMT expression displays genetic polymorphism with 11% of individuals having intermediate and one in 300 undetectable levels. The genetic basis for this polymorphism has now been elucidated and polymerase chain reaction (PCR)-based assays described for the most common mutations accounting for reduced activity. In previous studies, genotype has been correlated with red blood cell activity. In this report, we describe the relationship between genotype and TPMT activity measured directly in the target of drug action, the leukemic cell. We have demonstrated that the TPMT activity in lymphoblasts from 38 children and adults found by PCR to be homozygotes (*1/*1) was significantly higher than that in the five heterozygotes (*1/*3) detected (median, 0.25 v 0.08, P < .002, Mann-Whitney U). Similar results were obtained when results from children were analyzed separately. However, comparison of activity in blasts from AML and ALL showed a higher level in the former (0.35 v 0.22 nU/mg, P < .002, n = 17, 35), suggesting that factors other than genotype may also influence expression.

Acute Disease↗
PubMed1998

Acute changes in urine protein excretion may predict chronic ifosfamide nephrotoxicity: a preliminary observation.

PURPOSE: To evaluate proteinuria occurring early after ifosfamide therapy and to assess the use of changes in proteinuria in the prediction of severe chronic nephrotoxicity. METHODS: One-dimensional sodium dodecyl sulphate polyacrylamide gel electrophoresis was used to characterize urine protein excretion in 12 children with solid tumours before and after the first course of ifosfamide treatment, and in 24 healthy children. Chronic nephrotoxicity was evaluated at 6 months after ifosfamide treatment and graded as none, mild, moderate or severe. RESULTS: Urine from healthy children and from 10 of 12 patients before ifosfamide therapy showed a protein band with a molecular weight (95.4 kDa) corresponding to that of Tamm-Horsfall protein but no lower molecular weight proteins. After the first course of ifosfamide this 95.4-kDa protein was lost in six of ten patients with a concomitant appearance of a low molecular weight proteinuria (< 70 kDa) in eight. Tamm-Horsfall protein was lost in two of five patients who subsequently developed no or mild nephrotoxicity and in four of five patients who subsequently developed moderate or severe nephrotoxicity. CONCLUSIONS: Early subclinical changes in urine protein excretion after ifosfamide, manifested by a loss of Tamm-Horsfall protein excretion, may be predictive of subsequent chronic nephrotoxicity.

Adolescent↗
PubMed1998

A monofunctional derivative of melphalan: preparation, DNA alkylation products, and determination of the specificity of monoclonal antibodies that recognize melphalan-DNA adducts.

Bifunctional alkylating agents, such as those based on nitrogen mustard, form important parts of many anti-cancer chemotherapy protocols and are responsible for increased incidences of secondary tumors in successfully treated patients. These drugs generally form a majority of monofunctional DNA adducts, although the bifunctional adducts appear to be necessary for their powerful cytotoxic and antitumor effects. The relative importance of bifunctional as opposed to monofunctional adducts in the varied biological consequences of drug exposure has not been studied in detail, particularly in relation to the role and specificity of biochemical responses to therapy-related DNA damage. A simple method is described for the preparation of useful quantities of a pure monofunctional derivative of the nitrogen mustard-based drug melphalan. Monohydroxymelphalan was prepared by partial hydrolysis, purified by reversed phase chromatography, and characterized by MS, NMR, and HPLC. Contamination with melphalan was </=0.2%. The heat labile DNA base adducts formed by monohydroxymelphalan were shown to contain undetectable levels of cross-linked species. The ratio of adenine to guanine adducts was 0.62, similar to the equivalent ratio for melphalan. The sequence-dependent pattern of alkylation of purified DNA was indistinguishable from that of melphalan, but required a higher dose to achieve comparable extents of reaction. The specificities of two monoclonal antibodies that recognize melphalan-DNA adducts were investigated using DNA alkylated with [3H]monohydroxymelphalan. Adducts on this DNA showed similar immunoreactivities to adducts formed by melphalan. This shows clearly that neither antibody was specific for cross-linked adducts and that it is therefore possible to quantify adducts formed by both monohydroxymelphalan and melphalan with high sensitivities. The availability of monohydroxymelphalan in addition to melphalan, together with sensitive immunoassays for adducts on extracted DNA and in individual cells, constitutes a useful system for investigating cellular responses to the DNA modifications formed by a clinically relevant drug.

Alkylation↗
PubMed1997

The relationship between intrinsic thymidylate synthase expression and sensitivity to THYMITAQ in human leukaemia and colorectal carcinoma cell lines.

Thymidylate synthase (TS) expression has been characterized for a panel of eight human colorectal carcinoma and five human leukaemia cell lines, to relate differences in intrinsic TS activity, protein and mRNA levels to growth inhibition caused by continuous exposure to THYMITAQ, a specific non-classical antifolate TS inhibitor. Although a 20-fold variation in sensitivity to THYMITAQ was found within the colorectal cell line panel (IC50 0.12-2.7 microM), sensitivity was not related to TS activity, TS protein or TS mRNA levels. For the leukaemic cell lines, only a twofold range in sensitivity to THYMITAQ was observed (IC50 0.87-2.3 microM), and this did not correlate with TS activity, TS protein or TS mRNA levels. Across all of the cell lines, TS activity was linearly related to TS protein levels (r2 = 0.87, P < 0.0001). However, for both the colorectal and leukaemia cell line panels, no relationship was found between TS mRNA/18S rRNA ratios and either TS activity or TS protein, consistent with the importance of post-transcriptional mechanisms in regulating TS activity. Two of the colorectal cell lines (BE and HCT116) and one of the human leukaemic cell lines (HL60), were intrinsically resistant to THYMITAQ (IC50 > 2 microM) in the absence of TS overexpression, suggesting that, subsequent to TS inhibition, events such as DNA repair and tolerance to apoptotic stimuli are also important determinants of sensitivity to THYMITAQ.

Colorectal Neoplasms↗
PubMed1996

The effect of treatment with high dose melphalan, cisplatin or carboplatin on levels of glutathione in plasma, erythrocytes, mononuclear cells and urine.

Glutathione(GSH) has been implicated as an important factor in the detoxification of many electrophilic xenobiotics, including agents used in cytotoxic chemotherapy. Maintenance of high levels of GSH in normal tissues is believed to be important in the prevention of drug-induced toxicity. Previous studies have indicated that exposure of cells to some toxic electrophiles both in vitro and in vivo can cause a temporary decrease in intracellular levels of GSH. In this paper we report that in a series of 22 children and young adults treated with high dose melphalan (ten courses studied, all 200 mg/m2), cisplatin (eight courses, 80-104 mg/m2) or carboplatin (seven courses, 507-750 mg/m2) there was no significant alteration in the level of plasma, erythrocyte or urine GSH in the period immediately following drug administration. Fluctuations in the level of GSH in mononuclear cells were observed in some patients but did not follow any consistent pattern and were similar to those observed in a series of nine normal adult controls over the same time course. These results suggest that for melphalan, cisplatin and carboplatin, drug-GSH adduct formation is insufficient to cause a measurable decrease in intracellular GSH levels in normal peripheral haematopoietic cells during the course of treatment.

Adolescent↗
PubMed1996

Sensitivity of testis tumour cells to chemotherapeutic drugs: role of detoxifying pathways.

In contrast to most other types of cancer, metastatic testicular germ cell tumours (TGCT) are cured in most patients using cisplatin-based combination chemotherapy. The biochemical mechanisms underlying this sensitivity have not been defined. Drug detoxification can modulate response to chemotherapy in vivo and in vitro, and therefore we measured levels of glutathione (GSH), glutathione-S-transferase (GST) and both constitutive and cisplatin- and dexamethasone-induced levels of metallothionein (MT) in five human testis tumour cell lines. The levels were compared with those in five human bladder cancer cell lines and two cell lines with cisplatin resistance acquired in vitro. GSH levels were relatively low in the testis tumour cell lines, as might be expected in drug-sensitive cells, and there was a 77-fold increase in GSH levels in the cisplatin-resistant testis tumour cell line. GST levels were similar in the two cell types, while metallothionein levels were relatively high in the testis tumour cell lines. These data indicate that GSH may contribute to the sensitivity of TGCT to chemotherapy, and that GSH expression may be involved in the acquisition of cisplatin resistance in these tumours.

Antineoplastic Agents↗
PubMed1996

The relationship between tumour glutathione concentration, glutathione S-transferase isoenzyme expression and response to single agent carboplatin in epithelial ovarian cancer patients.

There is evidence to suggest that glutathione (GSH) and glutathione-S-transferases (GST) are important factors in determining sensitivity to cytotoxic drugs in vitro and in preclinical in vivo model systems. To define the relationship between tumour GSH concentration, GST isoenzyme expression and response to carboplatin in epithelial ovarian cancer (EOC), tumour samples from 39 patients with assessable disease after primary surgery were analyzed for GSH content and GST expression. Response was assessed after completing six courses of single agent carboplatin therapy. GSH was measured by high performance liquid chromatography (HPLC) in fresh tumour samples taken at primary laparatomy. GST isoenzyme expression was assessed by immunohistochemistry of fixed tumour material using antibodies specific for pi, alpha and mu classes. GST isoenzyme expression was defined as positive if the staining intensity was strong and more than 10% of tumour cells were involved. The mean GSH concentrations were: 8351 +/- 4496, 7211 +/- 5026, 6559 +/- 4573 and 3758 +/- 1885 (nmol g-1 tissue dry weight mean +/- s.d.) for tumours from patients who subsequently achieved a complete response (CR, n = 18), partial response (PR, n = 10) or who had static disease (SD, n = 7) or progressive disease (PD, n = 4) respectively. There was no relationship between GSH concentration and response (ANOVA, P = 0.32). There were also no relationship between GST isoenzyme expression and response (P Fisher's exact test 0.51-0.55 and chi-squared test 0.98-0.99). In conclusion, there was no association between the concentration of GSH or expression of GST isoenzymes and response to single agent carboplatin in primary previously untreated EOC.

Antineoplastic Agents↗
PubMed1996

A case of mature B-cell ALL with coexistence of t(1;19) and t(14;18) and expression of the E2A/PBX1 fusion gene.

The translocation t(1;19)(q23;p13) is found in 3-5% of all acute lymphoblastic leukaemias (ALL) and results in the expression of an E2A/PBX1 hybrid gene transcript. This translocation is very closely associated with a pre-B phenotype. t(14;18) is associated with follicular B-cell lymphoma and is characterized by over-expression of the bcl-2 oncogene. We describe a case of ALL in an adult with a mature B-cell immunophenotype and a single abnormal cell line with a complex karyotype showing both t(1;19) and t(14;18). Two reports of this phenomenon have been published previously and molecular analysis, where performed, showed the E2A gene was not rearranged, suggesting the t(1;19) was a molecular variant of the established translocation. In contrast, molecular analysis of our case demonstrated expression of the E2A/PBX1 fusion transcript typically associated with t(1;19) in pre-B ALL but showed it to be present at an extremely low level, despite the abnormal karyotype being found in the majority of metaphase cells. Analysis of bcl-2 expression showed a significant up-regulation. A down-regulation of the E2A/PBX1 hybrid gene as a consequence of the enhanced expression of bcl-2 may be a possible mechanism for this finding.

Burkitt Lymphoma↗
PubMed1996

Nurse caring behaviors: comparing five tools to define perceptions.

We asked which of five tools that measure perceptions of caring attributes (Larson's Care Q, Larson's Care/Sat, Nyberg's CAS, Duffy's CAT, Wolf's CBI) have the greatest usefulness in studies with patients and nurses. A convenience sample of 26 nurse administrators completed all five tools. Data was collected on completion time, demographic characteristics, and how results compared with earlier research using the same instruments. Larson's Care Q had similar results as previously described in the literature but problems included a forced choice format, difficulty with instructions, and it took the longest time to complete. Larson's Care/Sat was difficult to analyze, contained negatively worded items, but took the least time to complete. Nyberg's CAS was designed for use with nurses and would be difficult to use with patients. Duffy's CAT had negatively worded items. Similar results were obtained with Wolf's CBI as previously described in the literature and it had consistent language, short completion time, easy to understand instructions, and easy to analyze results which could be used in a correlational design study. We conclude that Larson's Care Q (using a simpler method of administration) or Wolf's CBI could be valuable in determining perceptions of caring in patients and nurses.

Adult↗