Electronic Library of Scientific Literature


Volume 45 / No. 3 / 1998

Bcl-2 family proteins and leukemia

J. Kusenda

Cancer Research Institute, Slovak Academy of Sciences, 833 91 Bratislava, Slovakia

The present status of Bcl-2 family proteins action and their role in leukemia and lymphoma is reviewed here in short. The Bcl-2 is an oncogenic protein that acts by inhibiting programmed cell death (apoptosis). In this article a timely review of the emerging mechanisms by which Bcl-2 and homologous family proteins might suppress cell death is presented. There have been reports that Bcl-2 and related anti-apoptotic proteins can function as a channel in the mitochondrial membrane and as an adaptor protein that can protect cells from cytotoxic agents. A dual function now seems likely, and interactions between Bcl-2 and other proteins are supposed.
The Bcl-2 family proteins have assumed an important role in leukemia and lymphoma research. The observations reviewed in this article suggest an important role of dysregulated Bcl-2 expression in the pathogenesis and prognosis of at least some types of leukemia and lymphoma. The Bcl-2 family proteins are important regulators of apoptosis that constitute a novel mechanism of chemoresistance in cancer.

Key words: Bcl-2 oncogene, Bcl-2 protein, leukemia, lymphoma, chemoresistance.
pp. 117-122

Oncogene amplification and expression in pediatric solid tumors

J. Mareš, V. Polanská, H. Görgens, Z. Sedláček, T. Maříková, P. Boček, R. Kodet, J. Schackert, P. Goetz

Institute of Biology and Medical Genetics, 2nd Medical School, Charles' University, 150 18 Prague, Czech Republic;
Carl Gustav Carus University Clinic, Technology University, Dresden, FRG;
Clinic of Pediatric Oncology, 2nd Medical School, Charles' University, Prague, Czech Republic;
Institute of Pathological Anatomy, 2nd Medical School, Charles' University, Prague, Czech Republic

Oncogene amplification and expression and their mutual relationship was analyzed in 92 pediatric tumors by Southern and Northern blot hybridization with N-MYC, ERB A, ERB B, N-RAS and Shb probes. Amplification and overexpression was associated with more advanced clinical stages of tumor, especially in neuroblastomas, rhabdomyosarcomas and ganglioneuroblastomas. The most frequent alteration observed was N-MYC amplification together with overexpression. N-RAS amplification was not detected, while the overexpression of this oncogene was found in 3 cases. Neither amplification nor overexpression was revealed in any specimen of hepatoblastoma or hepatocellular carcinoma. We suggest that oncogenes overexpression provides more accurate prognostic information than amplification.

Key words: Oncogenes, amplification, overexpression, pediatric solid tumors, clinical stages.
pp. 123-127

Phenotypic heterogeneity and aberrant markers expression in T-cell leukemia

O. Babušíková, M. Glasová, E. Koníková, J. Kusenda, J. Čáp, J.Gyárfáš, K. Koubek

Cancer Research Institute, Slovak Academy of Sciences, 812 32 Bratislava, Slovakia;
Department of Childhood Oncology, University Children´s Hospital, Bratislava, Slovakia;
National Cancer Institute, Bratislava, Slovakia;
Institute of Hematology and Blood Transfusion, Prague, Czech Republic

For exact determination of lineage assessment there is a need of surface membrane and intracellular (cytoplasmic and nuclear) immunophenotyping performed by flow cytometry. We evaluated in detail the results of surface and intracellular immunophenotyping of 34 T-ALL cases. The great heterogeneity of T-cell differentiation markers has been observed which did not allow relevant subclassification of T-ALL according to the existing subclassification schemes and the proposed three-stage model of physiological T-cell differentiation. Therefore, a simplified classification based on the CD3 marker expression either on cell membrane or in cytoplasm has been created with allocation of T-ALL into two main phenotypic groups. From 34 in detail examined T-ALL cases a great deal - 27 (79%) belonged to an immature phenotype (Stage I) and only 7 (21%) expressed more mature phenotype (Stage II). Simultaneously the presence of atypical/aberrant T-cell phenotypes has been studied. We showed that in T-ALL it was possible to specify some cases with leukemia-associated phenotype with coexistence of atypical markers which are absent in nonleukemic cells. In a majority of cases early B-lineage marker (CD10) and in a smaller proportion of them non-lineage associated marker (CD34) were observed. Myeloid marker CD13 was observed in one case of the immature T-ALL, together with CD10 and CD34. As these atypical markers were present through all differentiation stages of T-ALL we obtained a strong evidence that they might represent an abnormal rather than an immature phenotype. The prognostic significance of T-ALL subtypes and aberrant markers coexpression have been discussed. Simultaneously it was shown that quantitative immunofluorescence could provide an additional important diagnostic marker also in T-ALL cases.

Key words: T-cell leukemia, phenotypic heterogeneity, aberrant markers expression, membrane and intracellular immunophenotyping, flow cytometry.
pp. 128-134

Is T-cell acute lymphoblastic leukemia still a high risk leukemia in children?

J. Čáp, A. Foltinová, E. Kaiserová, O. Babušíková, M. Jamárik, M. Jasenková

Department of Childhood Oncology, University Children's Hospital, 833 40 Bratislava, Slovakia;
Cancer Research Institute, Slovak Academy of Sciences, Bratislava, Slovakia

In a group of 21 children with T cell ALL we compared the clinical picture and laboratory finding in a subgroup of 13 patients with less mature (mCD3 negative) and a subgroup of 8 patients with mature (mCD3 positive) phenotypical type and the therapeutic response in relation to the stage of thymic differentiation of the blastic cells as well. We could not find any significant differences concerning the presence or absence of mediastinal thymic mass, organomegaly, WBC count, morphology of the blasts and their acid phosphatase and PAS reaction between cases with less mature and mature type of thymic differentiation. Concerning the therapeutic response, children with mature type of T-ALL have shown at 5 years significantly higher event free survival rate, in comparison with the group of patients with less mature type of T cells. Overall survival rate was also higher in the first group, but statisticaly not significant.

Key words: T-cell ALL, childhood leukemia, high risk leukemia, immunophenotyping.
pp. 135-139

Expression of proliferating cell nuclear antigen (PCNA) and p53, bcl-2 or C-erb B-2 proteins on Reed-Sternberg cells: Prognostic significance in Hodgkin's disease

P. Smolewski, H. Niewiadomska, J. Z. Błoňski, T. Robak, E. Krykowski

Department of Hematology, Medical University of Lódż, Copernicus Hospital, 93-513 Lódż, Poland;
Department of Oncology, Medical University of Lódż, Lódż, Poland

Expression of proliferating cell nuclear antigen (PCNA) as well as p53, bcl-2 and C-erb B-2 genes protein products on Reed-Sternberg/Hodgkin's (R-S/H) cells was analyzed by immunohistochemistry in 65 patients with Hodgkin's disease (HD). Their significance as markers of clinical malignancy and prognostic factors was evaluated.
Positive reaction for PCNA was present in 57 cases (87.7%), for p53 in 42 (64.6%), for bcl-2 in 41 (63.1%), and for C-erb B-2 in 38 cases (58.5%) of HD. The high proliferate PCNA index and high expression of p53 and bcl-2 correlated with poor response to the treatment. Expression of both p53 and bcl-2 oncoproteins negatively influenced overall survival and disease free survival. Indexes of PCNA, p53 and bcl-2 were significantly higher in patients with advanced disease then in early clinical stages. In LP type of HD the lowest indexes of PCNA or bcl-2, and even lack of bcl-2 expression on R-S/H cells were observed. There was no correlation between expression of C-erb B-2 and response to the treatment, time of survival, clinical stage or histopathological types of HD.
Statistical analysis let us to the conclusion, that indexes of PCNA, p53 and bcl-2 expression can be taken into consideration as a new prognostic factors in Hodgkin's disease. C-erb B-2 protein expression seems to be of no value for prognosis in HD.

Key words: Hodgkin's disease, Reed-Sternberg cell, oncoproteins, prognostic factors.
pp. 140-147

Embolization and serum lysozyme activity in renal cancer

M. Mantur, J. Matowicka-Karna, B. Darewicz, H. Kemona, V. Dymicka-Piekarska, J. Prokopowicz, J. Darewicz

Institute of Laboratory Diagnostics, Medical University, 15-276 Białystok, Poland;
Department of Urology, Medical University, Białystok, Poland

The aim of the study was to determine the effect of renal tumor embolization on nonspecific immunity by evaluating lysozyme activity and leucocytosis in 45 patients and 40 healthy people. Lysozyme activity was assessed in the non-diluted serum (A1) and in the tenfold diluted serum (B1) prior to embolization and after embolization (A2, B2) and in control group. Prior to embolization, lysozyme activity was lower in the experimental group (A1 and B1), compared to the control groups, the differences being statistically significant (p < 0.05). After embolization, the activity became normalized (A2), reaching the control value and even exceeding it (C) in the diluted serum (B2). Leucocytosis prior to embolization (L1) resembled that of control group, increasing slightly after embolization (L2). The differences observed in the changes in lysozyme activity and leucocytosis were statistically significant (p < 0.05). Our findings indicate an inhibitory effect of the neoplastic process on nonspecific immunity. Embolization causes ischemic necrosis of tumor and products of neoplastic tissue disintegration exert a stimulating effect on granulopoiesis, by increasing the turnover of neutrophilic granulocytes. Granulocytic-monocytic infiltrations in tumor stroma are the source of lysozyme, enhancing not only local but also systemic immunity, which is manifested in the increased lysozyme activity in blood serum.

Key words: Lysozyme activity, cancer disease, embolization.
pp. 148-150

Glutathione and lipid peroxidation levels in human breast tumors

T. Çoban, A. Mabsout, B.C. Eke, D. Bülbül, U. Berberoglu, M. Iscan

Department of Toxicology, Faculty of Pharmacy, Ankara University, 061 00 Tandogan-Ankara, Turkey;
Departments of Pathology, Demetevler Oncology Hospital, Ankara, Turkey;
Department of Surgery, Demetevler Oncology Hospital, Ankara, Turkey

The levels of reduced glutathione (GSH) and lipid peroxidation (LP) of breast tumor and surrounding tumor free (normal) tissues of 39 breast cancer female patients with infiltrating ductal carcinoma and the relationship between these two parameters were investigated. Large interindividual variations in the levels of GSH and LP were found in both tumor and normal tissues. The mean GSH levels of tumors were significantly higher than those of normal tissues. This tendency did not change with the stage and grade (excluding grade 1) of the malignancy, menopausal status and chemotherapy treatment. No correlation was found between GSH level and stage or grade of malignancy (p > 0.05). However, although more than half of the tumor samples (23/39, 59%) had higher LP levels than their corresponding normal tissues, no significant difference was noted between the mean LP levels of tumor and normal tissues. This tendency did not change with the stage and grade of the malignancy, and menopausal status and chemotherapy treatment. No relationship was observed between the LP level and stage or grade of malignancy (p > 0.05). Overall, no association existed between the levels of GSH and LP in tumors (p > 0.05). These results reveal that the GSH, but not LP, could be a marker of breast malignancy and that the increase in GSH level is not sufficient to lower the LP level in human breast tumors.

Key words: Human breast tumor, glutathione, lipid peroxidation.
pp. 151-156

Photodynamic therapy for gastrointestinal tumors using three photosensitizers - ALA induced PPIX, Photofrin® and MTHPC. A pilot study

P. Mĺkvy, H. Messmann, J. Regula, M. Conio, M. Pauer, C.E. Millson, A.J. MacRobert, S.G. Bown

Department of Gastroenterology, St. Elisabeth Oncological Institute, 812 50 Bratislava, Slovakia;
Department of Internal Medicine, University of Regensburg, FRG;
Department of Gastroenterology, Postgraduate Institute, Warsaw, Poland;
Department of Internal Medicine, University of Genoa, Italy;
National Medical Laser Centre, University College London, Medical School, U.K.

Photodynamic therapy (PDT) produces localized necrosis with light after prior administration of a photosensitizing drug. As PDT lesions in the gastrointestinal tract heal well, the technique is suitable for repeated endoscopic use. In this study we used PDT to treat benign and malignant gastrointestinal tumors in esophagus, duodenum and rectum in 22 patients, who refused or were not suitable for surgery. Patients were sensitized with 0.15 mg/kg of body weight with mesotetrahydroxyphenylchlorin i.v. m-THPc (2 patients), with 2.0 mg/kg Photofrin® i.v. (4 patients) or 60 mg/kg 5-aminolevulinic acid orally ALA (which is converted in vivo to active derivate protoporphyrin IX -PPIX) in fractionated doses (16 patients). Laser treatment was performed 2 days after Photofrin®, 2 and 4 days after mTHPc and 4 hours after ALA, using a metal vapour laser (628 nm, 50-150 J/cm2 for ALA and Photofrin®, 650 nm and 10-15 j/cm2 for mTHPc). Using ALA, the necrosis was only superficial (up to 1.8 mm depth). Four patients treated with Photofrin® showed deeper necrosis, in one case of 8 mm colon cancer complete response, in three cases 1-1.5 cm adenomatous polyps involving the ampulla Vateri 50% longer term reduction in size - seen endoscopically. Two patients with rectal villous adenomas treated with mTHPc showed 60-80% reduction in size (observed endoscopically) within few days after PDT, with better effects for treatment carried out 4 rather than 2 days after the sensitization. In all patients the healing was without any complications. Photofrin® and mTHPc work better, but cause cutaneous photosensitivity lasting 12 and 5 weeks, respectively. Better results with ALA are possible when using higher drug doses or modified light dosimetry. PDT is a promising treatment for small localized tumors in patients unsuitable for surgery, but further work is required to optimize the treatment conditions.

Key words: Gastrointestinal tumors, photodynamic therapy, endoscopic therapy.
pp. 157-161

Cloning, expression, purification of rat tumor necrosis factor and demonstration of its antitumor activity in Bomirski amelanotic melanoma

P. Koszałka, H. Breza, J. Bigda

Department of Histology and Immunology, Medical University of Gdańsk, 80 211 Gdańsk, Poland

In order to analyze the effect of tumor necrosis factor (TNF) on the growth of Bomirski melanomas of hamster, we cloned, expressed and purified to homogeneity the rat tumor necrosis factor. The detailed procedure of chromatographic purification of the cytokine is reported. Polyclonal antibodies generated against the recombinant rat preparation were able to neutralize the rat and hamster TNF. We have found that the preparation of rat TNF is active in vivo and inhibits the growth of a Bomirski (Ab) amelanotic melanoma. Thus, a novel model system has been developed, which should facilitate analysis of antitumor and immunomodulatory function of TNF in cancer hosts.

Key words: TNF, rat, cloning, antitumor activity, melanoma.
pp. 162-168

Protection from pan masala induced genomic damage by beta-carotene and retinoic acid - an in vitro experience

R.K. Patel, A.H. Trivedi, R.J. Jaju M.S. Kukreti, J.M. Bhatavdekar, P.M. Shah, D.D. Patel

Cell Biology Division, Division of Research, The Gujarat Cancer Society, NCH Campus, Asaroa, Abmedabad, 380 016, India;
The Gujarat Cancer and Research Institute, NCH Campus, Asarwa, Ahmedabad, India

Cytogenetic studies in Chinese hamster ovary (CHO) cells using aqueous and organic extracts of pan masalas, as well as genomic damage observed among pan masala consumers have conclusively shown genotoxic potential of pan masala - a dry complex mixture of areca nut, lime, catechu, cardamom, unspecified flavoring agent, etc., often containing tobacco in it. Tobacco and areca nut, major ingredients of pan masala, are closely associated with oral cancer. The most widely studied group of compounds in the field of chemoprevention is retinoids which includes natural vitamin A, beta-carotene and synthetic derivatives of vitamin A. In the present study, antigenotoxic effect of beta-carotene (BC) and retinoic acid (RA) on genotoxic potential of pan masala have been evaluated in CHO cells with the help of sister chromatid exchange (SCE) frequency and chromosome aberration (CA) frequency as cytogenetic markers. The pulse treatment with pan masala plain/pan masala - tobacco (PM/PMT) extract in combination with either BC or RA yielded lower frequencies of CA and SCE in CHO cells as compared to the cultures treated with aqueous extract fo pan masalas alone. This antigenotoxic effect of BC and RA was more pronounced when treatment was given continuously for a longer duration. Thus, these results indicated possibility of using BC and RA to decrease the risk of oral cancer among pan masala chewers.

Key words: Pan masala, beta carotene, retinoic acid, chromosome aberration, sister chromatid exchange, chemoprevention.
pp. 169-175

Is high dose methotrexate without irradiation of the brain sufficiently effective in prevention of CNS disease in children with acute lymphoblastic leukemia?

J. Čáp, A. Foltinová, E. Kaiserová, A. Mojzešová, D. Sejnová, M. Jamárik

Department of Childhood Oncology University Children´s Hospital, 833 40 Bratislava, Slovakia

We present 5-year results of treatment in 93 children suffering from acute lymphoblastic leukemia using two therapeutic protocols containing multidrug chemotherapy including high dose methotrexate. We could ascertain different results in standard and high risk patients. In a group of 62 children with standard risk we observed improvement in complete remission rate being 98.9% after induction phase of therapy, only one patient died on septicemia. Relapse rate in this group was 21.2% and that 14.7% in the bone marrow and 6.5% in CNS and no testicular relapse at all. In the group of 31 children with high risk leukemia all patients achieved complete remission. Only one of them died on acute pancreatitis due to toxicity. Overall relapse rate in this group was 28.9% with l2.8% of medullary relapse and l6.1% of CNS relapse. The last one was significantly higher than in the previous study when brain irradiation was a part of therapeutic procedure. It seems that this treatment is effective mainly in the standard risk leukemia, however, in the high risk leukemias this procedure appears to be less effective in preventing CNS leukemia. In this group of patients irradiation of the brain need to be enclosed in the therapy.

Key words: Acute lymphoblastic leukemia in children, high dose methotrexate, prevention of CNS disease without irradiation of the brain.
pp. 176-179