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Biomedical subjects

I Sturm

Publications and source records attributed to I Sturm.

17 recordsLinked to original sources
PubMed2002

Bax expression correlates with cellular drug sensitivity to doxorubicin, cyclophosphamide and chlorambucil but not fludarabine, cladribine or corticosteroids in B cell chronic lymphocytic leukemia.

In B-CLL, non-proliferating B cells accumulate due to defective apoptosis. Cytotoxic therapies trigger apoptosis and deregulation of apoptotic pathways contributes to chemoresistance. Loss of the apoptosis-promoting Bax has been implicated in resistance to cytotoxic therapy. We therefore evaluated ex vivo drug sensitivity of CLL, producing chemoresponse data which are prognostic indicators for B-CLL, in particular in the case of purine nucleoside analogs. To analyze the underlying mechanisms of drug resistance, we compared endogenous Bax and Bcl-2 expression to ex vivo response to eight drugs, and to survival in 39 B-CLL patients. We found that reduced Bax levels correlated well with ex vivo resistance to traditional B-CLL therapies - anthracyclines, alkylating agents and vincristine (all P < 0.04). Surprisingly, no such relationship was observed for the purine nucleoside analogs or corticosteroids (all P > 0.5). Mutational analysis of p53 could not explain the loss of Bax protein expression. Levels of Bcl-2 were not associated with sensitivity to any drug. In contrast to the ex vivo data, neither Bax or Bcl-2 expression nor doxorubicin sensitivity were associated with increased survival whereas sensitivity to fludarabine correlated with better overall survival (P = 0.031). These findings suggest that the resistance to purine nucleoside analogs and corticosteroids in B-CLL is due to inactivation of pathways different from those activated by anthracyclines, vinca alkaloids and alkylating agents and may be the molecular rationale for the efficacy of purine analogs in this disease.

Aged↗
PubMed2001

Bax expression in benign and malignant thyroid tumours: dysregulation of wild-type P53 is associated with a high Bax and P21 expression in thyroid carcinoma.

The purpose of our study was to determine the expression of the pro-apoptotic BAX protein in relation to the mutational status of BAX and p53 (as transcriptional activator of the BAX gene) in benign and malignant thyroid tissue. In 47 patients with thyroid tumours (14 follicular and 3 papillary carcinomas, 14 adenomas and 16 goitres), the DNA was screened for mutations of BAX (exon 1-6) and p53 (exon 5-8) by single-strand conformation polymorphism polymerase chain reaction (SSCP-PCR). Furthermore, the protein expression of BAX, p53 and p21 (which is also increased transcriptionally by p53) was investigated by immunohistochemistry. Surprisingly, we observed elevated BAX levels in patients with thyroid carcinomas compared with patients with adenomas (unpaired t-test: p<0.05) or with goitres (p<0.02). This is in clear contrast to other carcinomas where BAX is frequently inactivated which correlates to a poor prognosis (Sturm et al. J. Clin. Oncol. 1999;17:1364-74.). There were no significant differences of the BAX levels between goitres or the adenomas. In the SSCP-PCR analysis, no BAX mutations were detectable. P53 mutation analysis by SSCP-PCR did not reveal any functional p53 mutations in the patients with carcinomas, adenomas or goitres. Nevertheless, patients with carcinomas showed an overexpression (preferentially cytoplasmic) of p53 protein compared with patients with benign tumours (p<0.05). The absence of p53 mutations suggests that the overexpressed p53 is wild type. This is in line with the expression profile of BAX and p21, which showed a higher protein expression in these p53 positive tumours (p<0.05 in the carcinomas compared with the non-malignant lesions). Consequently, the overexpressed p53 might be a correlate for dysregulation without loss of function. This, in turn, might be a reason for the good outcome of some patients with thyroid cancer.

Adenocarcinoma, Follicular↗
PubMed2001

Analysis of p53/BAX/p16(ink4a/CDKN2) in esophageal squamous cell carcinoma: high BAX and p16(ink4a/CDKN2) identifies patients with good prognosis.

PURPOSE: We have previously shown that loss of BAX expression is a negative prognostic factor in metastatic colorectal cancer. In the present study, we addressed the prognostic relevance of BAX and its upstream regulator p53 in squamous cell carcinoma (SCC) of the esophagus. Analysis of p16(ink4a/CDKN2) was included because p16(ink4a/CDKN2) and p53 were shown previously to cooperate during induction of cell cycle arrest and apoptosis. PATIENTS AND METHODS: Retrospective analysis of 53 patients with curative intended R0 resection of esophageal SCC was done. Protein expression of BAX, p53, and p16(ink4a/CDKN2) was investigated by immunohistochemistry. In addition, tumor DNA was screened for BAX frameshift mutations by fragment length analysis and for p53 mutations by single-strand conformation polymorphism-polymerase chain reaction. RESULTS: Overall median survival was 13.7 months. Patients with high BAX protein expression had a median survival of 19.5 months versus 8.0 months with low BAX expression (P <.005). High p16(ink4a/CDKN2) protein expression was associated with a median survival of 23.8 months versus 9.7 months with low p16(ink4a/CDKN2) (P =.011). The best survival (median, 45.8 months) was seen in a subgroup of 12 patients whose tumors bore the combination of both favorite phenotypes (ie, high BAX and high p16(ink4a/CDKN2) protein expression). CONCLUSION: In this retrospective investigation, the combined analysis of BAX and p16(ink4a/CDKN2) shows subgroups in SCC of the esophagus with favorable (p16(ink4a/CDKN2)/BAX high expressing) or poor prognosis (loss of p16(ink4a/CDKN2)/loss of BAX). We suggest that such a multimarker analysis of apoptosis pathways could be useful for individualization of therapeutic strategies in the future, and suggest prospective studies to confirm these results.

Adult↗
PubMed2001

The kiss of death: promises and failures of death receptors and ligands in cancer therapy.

Death receptors and their ligands exert important regulatory functions in the maintenance of tissue homeostasis and the physiological regulation of programmed cell death. Currently, six different death receptors are known including tumor necrosis factor (TNF) receptor-1, CD95 (Fas/APO-1), TNF receptor-related apoptosis-mediating protein (TRAMP), TNF-related apoptosis-inducing ligand (TRAIL) receptor-1 and -2, and death receptor-6 (DR6). The signaling pathways by which these receptors induce apoptosis are similar and rely on oligomerization of the receptor by death ligand binding, recruitment of an adapter protein through homophilic interaction of cytoplasmic domains, and subsequent activation of an inducer caspase which initiates execution of the cell death programme. The ability of these receptors and their ligands to kill malignant cells was discovered early and helped to coin the term 'tumor necrosis factor' for the first identified death ligand. This review summarizes the current and rapidly expanding knowledge about the signaling pathways triggered by death receptor/ligand systems, their potency in experimental cancer therapy, and their therapeutic limitations, especially regarding their toxicity for non-malignant cells.

Adaptor Proteins, Signal Transducing↗
PubMed2001

The Bax/Bcl-2 ratio determines the susceptibility of human melanoma cells to CD95/Fas-mediated apoptosis.

Defective cytochrome c release and the resulting loss of caspase-3 activation was recently shown to be essential for the susceptibility of human melanoma cells to CD95/Fas-induced apoptosis. Cytochrome c release from mitochondria is regulated by the relative amounts of apoptosis-promoting and apoptosis-inhibiting Bcl-2 proteins in the outer membrane of these organelles. The assignment of Bax/Bcl-2 ratios by quantitative Western blotting in 11 melanoma cell populations revealed a relation to the susceptibility to CD95-mediated apoptosis. We could show that a low Bax/Bcl-2 ratio was characteristic for resistant cells and a high Bax/Bcl-2 ratio was characteristic for sensitive cells. Low Bax expression was not a consequence of mutations in the p53 coding sequence. The Bax/Bcl-2 ratio was also in clear correlation with sensitivity to another cell death inducer, N-acetylsphingosine. Furthermore, Bcl-2 overexpression abolished apoptosis triggered by both apoptotic stimuli, confirming the critical role of the Bax/Bcl-2 ratio as a rheostat that determines the susceptibility to apoptosis in melanoma cells by regulating mitochondrial function. Interestingly, some chemotherapeutics lead to the activation of death pathways by CD95L upregulation, ceramide generation, direct activation of upstream caspases, or upregulation of proapoptotic genes. Taken together, these signals enter the apoptotic pathway upstream of mitochondria, resulting in activation of this central checkpoint. We therefore assumed that apoptosis deficiency of malignant melanoma can be circumvented by drugs directly influencing mitochondrial functions. For this purpose we used betulinic acid, a cytotoxic agent selective for melanoma, straightly perturbing mitochondrial functions. In fact, betulinic acid induced mitochondrial cytochrome c release and DNA fragmentation in both CD95-resistant and CD95-sensitive melanoma cell populations, independent of the Bax/Bcl-2 ratio.

Antineoplastic Agents, Phytogenic↗
PubMed2001

Relevance of Ki-67 antigen expression and K-ras mutation in colorectal liver metastases.

AIMS: The liver is a frequent site of metastases from colorectal cancer. While these lesions are potentially amenable to surgical resection, they are usually very aggressive, and recurrence is frequent. Mutations of the proto-oncogene K- ras are thought to impart a strong growth signal to tumour cells and are closely associated with the development of malignancies of the colon and rectum. Hepatic metastases from colorectal cancer have notably elevated proliferative rates. The present study was performed to investigate the relationship between proliferation or K- ras mutation and prognosis following curative resection of colorectal liver metastases. METHODS: Colorectal liver metastases from 41 patients undergoing curative hepatic resection were examined for proliferation status and presence of K- ras mutations. The proliferative activity was assessed by Ki-67 immunohistochemistry. DNA from the same tissue samples was screened for point mutations in codon 12 of the K- ras gene using a novel microplate-based allelic-specific hybridization assay. Ki-67 scores and K- ras status were then related with patient survival as determined through retrospective analysis. RESULTS: Median survival was 40 months. Patients with high Ki-67 scores (> or = 50%) had significantly shorter median survival compared with those with low scores (30 vs 44 months, log-rank P=0.02). A high Ki-67 score was an independent negative prognostic factor by multivariate regression analysis (relative risk=3.04, P=0.036). K- ras point mutations were detected in 6/41 patients (15%), but mutational status did not correlate with Ki-67 score or survival. CONCLUSIONS: These findings suggest that the tumour proliferative index is a useful predictor of aggressive tumour behaviour and an indicator of patient survival. The presence of K- ras mutations does not appear to correlate with tumour proliferation status or patient survival.

Adult↗
PubMed2000

Impaired BAX protein expression in breast cancer: mutational analysis of the BAX and the p53 gene.

We have previously shown that the pro-apoptotic BAX protein is differentially expressed in breast cancer and in other epithelial tumors. In this line, a reduced BAX protein expression is a negative prognostic factor in various carcinomas including breast cancer. For p53, a trancriptional activator of BAX in apoptosis, mutations in the coding sequence were shown to modulate BAX protein expression in cell line models on the transcriptional level. We therefore investigated the BAX gene in 68 breast cancer specimens for the presence of mutations in the coding sequence by single-strand conformation polymorphism (SSCP)-PCR and direct sequencing. The expression of BAX protein was assessed by immunohistochemistry. In addition, we screened for mutations in the exons 5-8 of the p53 gene by SSCP-PCR to assess whether mutations in the DNA-binding domain of this upstream regulator of BAX gene transcription are responsible for differences in BAX protein expression. As previously observed, BAX was differentially expressed in the breast cancer samples, but no mutations in the coding sequence of the BAX gene were found besides a polymorphism in exon 6 at the position 552 (G->A) and additional intronic polymorphisms. In contrast, we identified 16 of 68 (23.5%) tumors to bear mutations in the p53 gene. In the subset of BAX-expressing tumors, the mutational inactivation of p53 did result in a reduced BAX protein expression (Fisher exact test, p = 0. 047). Nevertheless, we identified a subset of BAX-negative tumors lacking BAX or p53 mutations. Thus, additional, not yet identified regulators, apart from p53, appear to be involved in the regulation of BAX protein expression.

Base Sequence↗
PubMed2000

Relapse in childhood acute lymphoblastic leukemia is associated with a decrease of the Bax/Bcl-2 ratio and loss of spontaneous caspase-3 processing in vivo.

Dysfunction of the p53/Bax/caspase-3 apoptosis signaling pathway has been shown to play a role in tumorigenesis and tumor progression, ie the development of acquired drug resistance. Low expression of the apoptosis inducer Bax correlates with poor response to therapy and shorter overall survival in solid tumors. In the present study, we analyzed the p53/Bax/caspase-3 pathway in a paired and an unpaired sample series of children with acute lymphoblastic leukemia (ALL) at initial diagnosis and relapse. The data demonstrate that both Bax expression levels and the Bax/Bcl-2 ratio are significantly lower in samples at relapse as compared with samples at initial diagnosis (P=0.013, Wilcoxon signed rank test (paired samples); P=0.0039, Mann-Whitney U test (unpaired samples)). The loss of Bax protein expression was not a consequence of Bax frameshift mutations of the G8 tract and could not be attributed to mutations of the p53 coding sequence (exons 5 to 8) which were detected to a similar extent in de novo ALL samples and at relapse. Analysis of the downstream effector caspase-3 showed loss of spontaneous caspase-3 processing at relapse. Whereas nine out of 14 (64%, paired samples) or 37 out of 77 (48%, unpaired samples) ALL patients at initial diagnosis displayed spontaneous in vivo processing of caspase-3, this was completely absent in patients at relapse (paired samples) or detected in only one out of 34 patients at relapse (2.9%, unpaired samples). We therefore conclude that in ALL relapse a severe disturbance of apoptotic pathways occurs, both at the level of Bax expression and caspase-3 activation.

Adolescent↗
PubMed1999

Expression of the death gene Bik/Nbk promotes sensitivity to drug-induced apoptosis in corticosteroid-resistant T-cell lymphoma and prevents tumor growth in severe combined immunodeficient mice.

Members of the Bcl-2 gene family have been implicated in the regulation of cell death induced by cytostatic drugs. In some malignancies such as B-cell lymphoma, there is evidence that high expression of Bcl-2 is an independent negative prognostic marker and the overexpression of Bcl-2 has been shown to confer resistance to cytotoxic drugs by preventing drug-induced apoptosis. This function of Bcl-2 can be antagonized by apoptosis-promoting members of the Bcl-2 family. We previously showed that overexpression of Bax restores the chemosensitivity of Bax-deficient breast cancer cell lines. Therefore, we investigated whether the death-promoting Bcl-2 homologue Bik/Nbk can enhance cytostatic drug-induced apoptosis. As a model, we used the T-cell leukemia H9 (CD3(+) and CD4(+)CD8(-)), which is resistant to corticosteroid-induced cell death and does not express endogenous Bik/Nbk. Sensitivity for drug-induced apoptosis was increased 10- to 39-fold in cells transfected with the full-length coding sequence of Bik/Nbk. In addition, apoptosis induced via CD95/Fas or heat shock was increased to a similar extent. These data show that Bik/Nbk, which, unlike Bax, carries only a BH3 but no BH1 or BH2 domain may be a target to enhance chemosensitivity. The complete suppression of tumor growth in a severe combined immunodeficient mouse xenotransplant model suggests that, in analogy to Bax, Bik/Nbk may function as a tumor suppressor gene.

Adrenal Cortex Hormones↗
PubMed1999

Detection of genomic DNA fragmentation during apoptosis (DNA ladder) and the simultaneous isolation of RNA from low cell numbers.

In the present paper we describe a rapid and sensitive method for the simultaneous isolation of total RNA and genomic plus low-molecular-weight DNA from apoptotic cells. Using this method, we were able to detect a DNA ladder from as low as 30,000 apoptotic cells in only 45 min including gel electrophoresis. In addition, RNA can be readily obtained from the same specimen to assess gene expression during apoptosis. This method therefore appears to be advantageous when sensitivity and low amounts of sample material are a limiting factor.

Adsorption↗
PubMed1999

Analysis of the p53/BAX pathway in colorectal cancer: low BAX is a negative prognostic factor in patients with resected liver metastases.

PURPOSE: To determine the prognostic value of the central downstream apoptosis effector BAX in relation to its upstream regulator p53 in R0-resected hepatic metastases of colorectal cancer. PATIENTS AND METHODS: Retrospective analysis of 41 patients who underwent potentially curative resection of liver metastases from colarectal cancer was performed. Tumor DNA was screened for p53 mutations by single-stranded conformational polymorphism polymerase chain reaction and for BAX frameshift mutations by fragment length analysis. Protein expression of BAX, p21, and p53 was investigated by immunohistochemistry. RESULTS: Overall median survival was 40.2 months. Tumors with BAX frameshift mutations were considered microsatellite mutator phenotype-positive and were excluded from further prognostic analyses. Patients with high BAX protein expression had a median survival of 53.6 months compared with 35.4 months for patients with low BAX expression (P < .05). The negative prognostic value of low BAX expression was more evident in those patients with wild-type p53 (median survival, 54.0 v 23.3 months for BAX-negative tumors; P < .01). Low BAX expression was an independent negative prognostic marker in multivariate regression analysis for all patients independent of the p53 status (relative risk, 3.03, P = .03), especially for p53 wild-type tumors (relative risk, 8.21; P = .0095). CONCLUSION: We conclude that low BAX expression is an independent negative prognostic marker in patients with hepatic metastases of colorectal cancer. The best survival was seen in patients with an intact p53-to-BAX pathway; ie, wild-type p53- and BAX-positive tumors. Thus, analysis of apoptosis signaling pathways (here, p53 in concert with its downstream death effector, BAX) might yield more prognostic power in future studies as compared with analysis of single genes such as p53 alone.

Adult↗
PubMed1998

Immunotherapy of B-cell lymphoma with CD3x19 bispecific antibodies: costimulation via CD28 prevents "veto" apoptosis of antibody-targeted cytotoxic T cells.

Bispecific antibodies (CD3x19) against the CD3epsilon-chain of the T-cell-receptor/CD3 complex and the CD19 antigen on B cells can target polyclonal, nontumor-specific T cells to B lymphoma cells. This induces T-cell activation, and generation of cytotoxic T cells (CTLs). These polyclonal CTLs, targeted by the CD3x19 bispecific antibodies, can lyse CD19(+) B-lymphoma cells. In a xenotransplant model in severe combined immunodeficiency deficient (SCID) mice, we and others observed that CD28 triggering is required for efficient elimination of B-lymphoma cells and cure from the tumor in addition to CD3x19 administration. We also showed that the activation and targeting of CTLs to the target cell by signal one alone, ie, the CD3x19 mab, induces T-cell death by apoptosis. In blocking experiments we showed that this "veto" apoptosis is mediated by the CD95/Fas ligand. Addition of anti-CD28 (signal 2) renders the T cells resistant for veto apoptosis both in vitro and in vivo. We therefore conclude that the role of costimulation in immunotherapy with bispecific antibodies or other T-cell-based immune strategies is not only to facilitate T-cell activation but also to prevent T-cell deletion by apoptosis.

Animals↗
PubMed1997

A new method for data presentation in incremental cardiorespiratory exercise testing.

In incremental cardiopulmonary exercise testing, the averaging of data is usually performed to provide group mean data for statistical purposes. They are usually presented as averaged maximum values, or as averaged data at different exercise levels. However, during incremental exercise testing the change in metabolic status may vary between subjects, thus averaging data may not classify the metabolic status accurately. We present an averaging method using a segmented ordinal scale based on individual maximal work performance and the anaerobic threshold (AT). Individual exercise data are grouped into ten classes ranging from unloaded exercise to maximal exercise. The classes are defined in relation to the AT, resulting in an ordinal scale of four classes for exercise data below the AT, one class at the AT and five classes beyond the AT. Resting and unloaded pedalling are treated as separate classes. For evaluation, this method of classification is compared to one based on an absolute scale of oxygen uptake (Cabs) and to another based on a relative scale in 10% steps of maximal oxygen uptake (Crel). Ten healthy male subjects (mean age 23.3 years) performed a ramp cycle ergometer test. When using the Cabs classification method for mean data averaging, mean values for performance at high-intensity exercise were calculated using data from only two of the ten subjects because of variations in individual work capacity. In addition, the AT data were distributed across four classes, thus anaerobic and aerobic exercise data were mixed. Using the Crel classification method enabled data for all ten subjects to be included in the calculation of every data point, but the AT values were still distributed across three classes, resulting in the mixing of anaerobic and aerobic exercise data. However, using the segmented ordinal scale method of classification enabled data from all ten subjects to be included in the calculation of all data points, and it permitted the grouping of the AT values into one class. Thus, this latter method more accurately represents the data of the whole group under study and it allows the metabolic status of the subjects to be taken into consideration.

Adult↗