Electronic Library of Scientific Literature


Volume 43 / No. 5 / 1996

A possible role for the alpha 1 -> 3 galactosyl epitope and the natural anti-gal antibody in oncogenesis

L. Gollogly, V. Castronovo

Metastasis Research Laboratory, Pathology B23, University of Liege, B-4000 Liege, Belgium

Glycoconjugates and their antibodies are vital components of host-tumor interaction. This review concentrates on the oncological implications of research concerning the alpha gal triad; the alpha 1 -> 3 galactosyl epitope (alphaGal), the enzyme responsible for its construction, alpha 1,3 galactosyl transferase (alpha1-3GT), and its associated antibody: anti-gal. Alpha gal epitopes, previously assumed to be absent from human tissue, have been demonstrated on several human cancer cell lines, senescent red blood cells, and Graves' disease thyrocytes. Alpha-gal presence on neoplastic lines is correlated with increased metastatic formation in animal models. The mechanisms of human response to these neoantigens are complex, as natural anti-gal antibodies exist in high titers in normal sera, thus predicting immunological recognition of cells expressing alpha gal epitopes. Hypotheses vary regarding the pathogenic contributions of metastasis-associated phenomena such as de novo expression of alpha gal and its unmasking by desialylation. The means by which alpha gal is sporadically expressed in human tissue remain unknown, as the galactosyl transferase which produces this epitope in constitutively expressive animals has undergone significant mutation at the genomic level in humans. Pathological re-expression is presumed to require permissive changes at a cellular level. Detailing these alterations is a prerequisite to the comprehension of the metastatic phenotype. In this context, the possibility of therapeutic strategies affecting alpha gal expression are also discussed.

Key words: Glycoconjugates, antibodies, oncogenesis.
pp. 285-289

Stimulation of 1-(beta-d-arabinofuranosyl)cytosine (AraC)-induced apoptosis in the multidrug resistant human promyelocytic leukemia cell lines with protein kinase inhibitors

L. Hunakova, M. Sulikova, J. Duraj, J. Sedlak, B. Chorvath

Cancer Research Institute, Slovak Academy of Sciences, 812 32 Bratislava, Slovakia

Stimulation of apoptosis induced by 1-(beta-d-arabinofuranosyl)cytosine (AraC) with protein kinase inhibitors (i.e. staurosporine, CGP 41 251 - a protein kinase C (PKC) - selective staurosporine derivative and protein tyrosine kinase (PKT) inhibitor genistein) was examined in two human multidrug-resistant promyelocytic leukemia (HL-60) cell lines with different cell membrane drug resistance-associated glycoproteins (i.e. HL-60/VCR: MDR1 gene coded Pgp/p170 and HL-60/ADR: MRP gene coded non-Pgp/p190). Staurosporine stimulatedAraC-induced apoptosis in the parental drug-sensitive HL-60 cells and both examined multidrug resistant HL-60 sublines. The stimulation of AraC-induced apoptosis by PKC selective inhibitor CGP 412 251 and PTK-inhibitor genistein was approximately equal to that of staurosporine in HL-60/ADR cell line. In both parental drug sensitive HL-60 cells and Pgp/p170 positive (MDR1) HL-60/VCR, staurosporine-stimulated AraC-induced apoptosis was higher than that stimulated by the PKC selective CGP 41 251 inhibitor, or PTK-inhibitor genistein. These data suggest that the molecular pathway(s) for AraC-induced apoptosis can be activated and stimulated by PKC- and PTK-inhibitors in both examined drug-resistant HL-60 cell lines. Furthermore, these data suggest that although both PKC- and PTK-dependent mechanisms are involved in AraC-induced apoptosis, in the drug-sensitive HL-60 cells and multidrug-resistant HL-60/VCR (Pgp/p170) cells this process is mediated at least partially, also by PKC- and PTK-independent mechanisms, activated by staurosporine.

Key words: Apoptosis, AraC, protein kinase inhibitors, staurosporine, CGP 41 251 genistein, multidrug resistant, human leukemia HL-60, flow cytometry.
pp. 291-295

Leukocyte alkaline phosphatase and serum alkaline phosphatase in patients with metastatic breast and colon cancer

N. Walach, Y. Gur

Department of Oncology, Assaf Harofeh Medical Center, Zerifin, 70300 Israel; Sackler School of Medicine, Tel Aviv University, Tel Aviv, Israel; Radiation Safety Department, Soreq Nuclear Research Center, Yavne, Israel

Periferal blood leukocyte alkaline phosphatase (LAP) scores and serum alkaline phosphatase (SAP) levels in 70 patients with metastatic breast and colorectal cancer (metastases to the liver, lung, bone and abdomen) and in 18 healthy controls were measured. The mean LAP score in the metastatic cancer patients was significantly higher than in the control group (244 vs. 61) and there was no overlap between the 95% confidence level intervals of the two groups. The mean (SAP) level in the metastatic patients was also higher than in the controls (249 u/l vs. 162 u/l) but the 95% confidence level interval of the controls was inside the 95% interval of the metastatic patients meaning that considerable percentage of the metastatic patients will have a SAP level within the normal range.
We conclude that of the two markers, the LAP is the better one for detection of metastatic tumors.

Key words: Marker, metastases, breast cancer, colon cancer.
pp. 297-300

Comparison of fluorescent in situ hybridization (FISH) and the polymerase chain reaction (PCR) for detection of residual neuroblastoma cells

T. Eckschlager, K. McClain

Department of Pediatric Oncology, University Hospital, Motol, 150 18 Prague, Czech Republic;
Texas Children's Cancer Center and Hematology Service, Baylor College of Medicine, Houston, Texas, USA

Residual neuroblastoma (NB) cells in bone marrow or peripheral progenitor hematopoietic cells harvests may be a source of relapse after autologous transplantation in patients with high stage neuroblastoma. Therefore a sensitive method for detecting minimal residual disease (MRD) in the harvested product is important so that an appropriate purging techniques can be applied and optimized. We report on the detection of NB cells by fluorescence in situ hybridization (FISH) for N-myc amplification and compare the sensitivity of FISH with a semiquantitative polymerase chain reaction (PCR) assay. As a model of MRD we used the neuroblastoma cell line IMR-32 diluted with normal peripheral blood lymphocytes. We were able to detect a single NB cell in 1000 normal mononuclear cells by FISH. The PCR method, using ethidium-bromide-stained gels, required at least 10% NB cells to be present for detection of an amplified band of the N-myc oncogene. Thus, FISH is ten to one hundred times more sensitive in detection of N-myc amplification than a differential PCR and thus it is the method of choice for detection of MRD in NB patients.

Key words: Minimal residual disease, fluorescent in situ hybridization (FISH), polymerase chain reaction (PCR), N/myc amplification, neuroblastoma cell line IMR-32.
pp. 301-303

p53 gene mutation in relation to p53 protein accumulation in male and female breast cancer

B.K. Nayak, R.N. Baral, B.R. Das

Molecular Biology Division, Institute of Life Sciences, Bhubaneswar-751 007, India

The aim of this investigation was to study the prevalence of p53 gene mutations in male and female breast cancers and to find out the relationship between this event and p53 protein expression. Genomic p53 was amplified by polymerase chain reaction (PCR). Exons 5-8 were screened for mutations using single stranded conformation polymorphism (SSCP) analysis. P53 protein expression was detected by immunohistochemistry (IHC) with the monoclonal antibody DO-1. In female breast cancer, p53 gene mutation was detected in 33% cases in either exon 5 or 6. However, in male, mutation was detected only in exon 6 in 90% cases. On the other hand, p53 protein expression was observed in all of these cases. Moreover, p53 protein immunostaining was observed in some of those cancer tissues, where no mutation was detected in exons 5-8. P53 dysfunction, as indicated by mutation or increased protein expression, common in both male and female breast cancer, but rate of occurrence or site of mutation differ from each other. Our results in male breast tumors indicate a positive correlation between p53 mutation and p53 protein overexpression, whereas the results in female breast tumors indicate an overexpression of p53 protein even without p53 gene mutation. Therefore, it may be presumed that p53 protein accumulation can result primarily from mutation. In addition, stabilization of p53 through binding to other proteins is another possible reason of p53 overexpression.

Key words: p53, breast cancer, exon, mutation, protein expression, PCR-SSCP, immunohistochemistry.
pp. 305-310

Effect of tumor excision on NK cytotoxic activity and formation of metastases of Bomirski melanoma in hamsters

A. Mysliwski, D. Sosnowska, J. Bigda

Medical University of Gdansk, Department of Histology and Immunology, 80-210 Gdansk, Poland

The growth of transplanted Bomirski melanoma in hamsters is accompanied by the decrease of natural killer cytotoxic activity and the formation of metastases. The excision of primary tumors was carried out to examine what was the effect of the growing tumor and its metastases on the host's NK activity. It was found that the excision of primary tumor caused increased NK cytotoxic activity in comparison to that of nonoperated animals although it was still lower than that of healthy hamsters. It is concluded that both, a growing tumor and metastases, exert suppressive effects on NK activity and those effects add up. The pattern of metastases in operated animals was different to that observed in nontreated hamsters.

Key words: Melanoma, natural killer cells, metastases, hamsters.
pp. 311-313

Biological consequences of E.coli RecA protein expression in the repair defective pso4-1 and rad51::URA3 mutants of S.cerevisiae after treatment with N-methyl-N'-nitro-N-nitrosoguanidine

M. Slaninova, V. Vlckova, J. Brozmanova, M.A. Morais Jr., J.A.P. Henriques

Department of Genetics, Faculty of Natural Sciences, Comenius University, 842 15 Bratislava, Slovakia;
Department of Molecular Genetics, Cancer Research Institute, Slovak Academy of Sciences, Bratislava, Slovakia;
Departemento de Biofisica e Centro de Biotecnologia, Universidade Federal do Rio Grande do Sul, Porto Alegre, Brazil

RecA protein of E.coli is a multifunctional protein participating in genetic recombination, recombinational repair and mutagenesis. We used E.coli recA gene as a probe for complementation of repair defects after treatment of N-methyl-N'-nitro--N-nitrosoguanidine in the pso4-1 and rad51::URA3 mutants of S. cerevisiae. We tried to find the role of the RecA protein in S.cerevisiae mutants defective in different repair pathways. The RecA protein had no effect on survival of haploid and diploid pso4-1 mutants, but it had a significant effect on MNNG induced mutagenesis, which was increased to the wild type level. No effect of the RecA protein on survival was observed in rad51::URA3 mutant after MNNG treatment. However, in this case the RecA protein decreased the induced mutagenesis. We can suppose that the RecA protein, with its multifunctional enzymatic activity, can bind to special intermediates and iniciate function of different repair pathways depending on repair defects of the mutants studied.

Key words: RecA protein, MNNG, DNA repair, mutagenesis.
pp. 315-319

Serum soluble IL-2 receptor as a reliable and noninvasive marker of disease activity in patients with hairy cell leukemia

L. Chrobak, K. Podzimek, L. Pliskova, Z. Kerekes, P. Zak, J. Voglova, J. Spacek, V. Palicka

Department of Clinical Hematology, University Hospital, Charles University, Faculty of Medicine, 500 05 Hradec Kralove, Czech Republic;
Department of Medicine, University Hospital, Charles University, Faculty of Medicine, Hradec Kralove, Czech Republic;
Institute of Clinical Biochemistry and Diagnostics, University Hospital, Charles University, Faculty of Medicine, Hradec Kralove, Czech Republic;
Institute of Pathology, University Hospital, Charles University, Faculty of Medicine, Hradec Kralove, Czech Republic;

Hairy cell leukemia is a chronic lymphoproliferative disorder of B-cell lineage. Malignant cells express the interleukin-2 receptor (IL-2R) which is released in vitro as well as in vivo. The sera of patients with hairy cell leukemia contain elevated levels of this soluble receptor (sIL-2R). Sera of 24 patients with hairy cell leukemia were tested for sIL-2R. In 9 patients treated with 2-chlorodeoxyadenosine an improved clinical status was associated with decreasing serum sIL-2R. The maximal rate of decrease of sIL-2R level was observed within the second and the third week after the therapy initiation. Patients with disease progression had an increase in serum sIL-2R level. Our results suggest that serial measurement of sIL-2R level can be used as a reliable, noninvasive means to assess the disease activity and its response to therapy.

Key words: Hairy cell leukemia, soluble interleukin-2 receptor (sIL-2R), 2-chlorodeoxyadenosine.
pp. 321-325

The effect of local hrecTNF-alpha administration upon the spontaneous lung metastases in rats with Morris 5123 hepatoma

S. Terlikowski, H.Fr. Nowak, M. Sulkowska, S. Sulkowski

Department of Gynecology and Septic Obstetrics, Medical School, PL-15-062 Bialystok, Poland;
Department of Pathological Anatomy, Medical School, Bialystok, Poland

The experiment used Morris hepatoma 5123 series growing in muscles of the right hind limb of Buffalo rats. The group I animals were given intratumor 4 doses of TNF-alpha and group II - 8 doses of TNF-alpha (10 microg/day). Control groups (III and IV) consisted of rats with injected Morris hepatoma, which were given PBS solution instead of TNF-alpha. A decrease in the volume of neoplastic metastases was observed in groups I and II, compared with groups III and IV. At the same time an increase was found in the volume of metastatic tumors in group II (8 x TNF-alpha), compared with group I (4 x TNF-alpha). Histological and ultrastructural analysis of the pulmonary tissue revealed intensified fibrotic reactions and inflammatory infiltrations around the metastatic tumors. The change were much more enhanced in group II, which might affect the results of neoplastic metastatic volume measurements. We concluded that multiple human recombinant TNF-alpha, hrec TNF-alpha, local injections inhibited dissemination of tumor cells and prolonged the survival time of rats up to the 76th day of the follow-up.

Key words: hrecTNF-alpha, Morris 5123 hepatoma, lung metastases.
pp. 327-333

Antitumor action of bovine seminal ribonuclease

J. Soucek, P. Pouckova, J. Matousek, P. Stockbauer, J. Dostal, M. Zadinova

Institute of Hematology and Blood Transfusion, 128 20 Prague 2, Czech Republic;
Institute of Biophysics, Medical Faculty, School of Medicine, Charles University, Prague, Czech Republic;
Institute of Animal Physiology and Genetics, Academy of Sciences of Czech Republic, Libechov, Czech Republic

Unlike the bovine pancreatic ribonuclease (RNase A), bovine seminal ribonuclease (BS RNase) displays various biological activities, including antitumor activity, immunosuppressivity, spermatogenicity and embryotoxicity. To learn more about its antitumor effect we tested BS RNase on the growth of 16 cell lines derived from patients with various hematological malignancies. The cells of lymphoid origin were generally more susceptible to BS RNase, administered in the range of concentrations from 2 to 100 microg/ml, than the myeloid ones. RNase A used at the same concentrations did not exert any inhibitory effect. The inhibitory effect of BS RNase persisted in cultured cells after three times wash in complete medium and cell recultivation in fresh medium free of BS RNase. Four cell lines were very little sensitive (KG-1 and U-937) or resistant (JOK and NAMALWA) to BS RNase regardless of their origin.
The in vivo antitumor effect of BS RNase was tested on human prostate carcinoma transplanted to athymic nude mice. The daily dose of BS RNase (0.25 mg/20 g) was administered for three weeks except weekends (15 doses) by three different ways (intraperitoneally - i.p., subcutaneously - s.c. and intratumorally - i.t.). Whereas i.p. administration was ineffective, s.c. administration significantly reduced size of the tumors and i.t. administration abolished half of the tumors in treated mice. The average weight of treated mice decreased during the experiment by 10-15%.

Key words: Bovine seminal ribonuclease, antitumor activity, cell lines, nude mice, prostate carcinoma.
pp. 335-340

Modulatory effect of tempol on toxicity and antitumor activity of 6-mercaptopurine and on P450 cytochrome level

N.P. Konovalova, R.F. Diatchkovskaya, L.M. Volkova, V.N. Varfolomeev

Institute of Chemical Physics, Russian Academy of Sciences, Chernogolovka, Moscow Region, 142 432 Russia

Low selectivity of contemporary antitumor drugs requires a search for its improvement. In this context, nitroxyl radicals are of interest as promising pharmacological agents. The introduction of nitroxyl radical into the structure of antitumor cytostatics was found to reduce considerably their general and specific toxicity. In this work, we demonstrate a detoxifying effect of tempol upon its combined injection with cytostatics at their absolute lethal dose in the intact mice as well as an improvement of sensitivity of tumor-bearing animals to 6-MP. Tempol is shown to normalize the level of oxidized form of P450 cytochrome in a liver, reduced as a result of the injection of 6-MP.

Key words: Nitroxyl radical, cytostatics, P450 cytochrome.
pp. 341-346

Phase I/II trial of carboplatin dose escalation in combination chemotherapy with etoposide, bleomycin and GM-CSF support for poor prognosis nonseminomatous germ cell tumors patients

S.A.Tjulandin, M.B. Stenina, N.J. Sidorova, F.G. Delgado, A.V. Sokolov, G.V. Molchanov, N.V. Ljubimova, A.M. Garin

Department of Clinical Pharmacology and Chemotherapy, Cancer Research Center, Moscow, 115 478, Russia;
Radioimmunology Laboratory, Cancer Research Center, Moscow, Russia;
Diagnostic Division, Cancer Research Center, Moscow, Russia;
Biochemistry Laboratory, Cancer Research Center AMS of Russia, Moscow, Russia

To determine the maximum tolerated dose (MTD), and therapeutic efficacy of carboplatin (CBDCA) in combination with etoposide and bleomycin (CEB) as initial chemotherapy for poor prognosis germ cell tumors, a CBDCA dose escalation supported with GM-CSF had been performed. Twenty four untreated patients were treated with CBDCA 400 mg/m[^2] on day 1, etoposide 100 mg/m[^2] on days 1 to 5 and bleomycin 30 mg on days 1, 3, 5. Four cycles were scheduled at 21-day interval. The first cohort of 6 patients received only initial chemotherapy regimen. In the subsequent cohorts of six patients, the CBDCA dose was increased by 100 mg/m[^2]. A fixed dose and schedule of GM-CSF at 5 µg/kg subcutaneously was given on days 6 through 15. Myelosuppression, with neutropenic fever and hemorrhages, was the dose-limiting toxicity at the 600 mg/m[^2] dose level. The recommended dose of CBDCA is 500 mg/m[^2]. Overall complete response (CR) rate was 71% and with median follow up of 25 (16-34) months, 58% of patients are alive and have no evidence of disease (NED). A higher number of CR was achieved with CBDCA dose higher than 400 mg/m[^2] compared with CBDCA dose of 400 mg/m[^2] (92 vs. 50%, p = 0.03), as well as a higher proportion of patients who are alive and with NED (75 vs. 42%, p = 0.1). Despite GM-CSF support, the MTD of CBDCA could not be increased beyond 500 mg/m[^2] (50% of the dose escalation), due to severe myelosuppression. The treatment outcomes obtained with CEB in our study are no better than the standard cisplatin-based chemotherapy. Further studies of this regimen, where CBDCA dose should be calculated according to the patients glomerular filtration rate are warranted.

Key words: Chemotherapy, granulocyte-macrophage colony stimulating factor, germ cell tumors, poor prognosis, carboplatin, testicular cancer.
pp. 347-352

Case-control study of pancreatic cancer in Serbia, Yugoslavia

N.Z. Kokic, J.B. Adanja, D.H. Vlajinac, P.J. Marinkovic, B.R. Colovic, S.M. Jarebinski

Health Center, Belgrade, Yugoslavia;
Institute of Epidemiology, School of Medicine, Belgrade University, 11000 Belgrade, Yugoslavia;
Institute of Social Medicine, Statistics and Health Research, School of Medicine, Belgrade University, Belgrade, Yugoslavia;
Institute of Gastroenterology, School of Medicine, Belgrade University, Belgrade, Yugoslavia

Case-control study comprised 100 pancreatic cancer patients and the same number of hospital controls individually matched with cases by sex, age and place of residence. According to logistic regression analysis following factors were found to be risk factors for pancreatic cancer: smoking 26 or more cigarettes per day (OR = 43.95, 95% CI = 7.69-192.53), consumption of 5 or more glasses of hard drinks per week (OR = 12.84, 95% CI = 2.13-77.29), coffee consumption during a period exceeding 35 years (OR = 2.50, 95% CI = 1.18-5.28), gall bladder disease in personal history (OR = 4.29, 95% CI = 1.08-16.99) and family history positive on peptic ulcer (OR = 11.71, 95% CI = 0.99-137.98). Two factors appeared to be protective for cancer of pancreas: appendectomy (OR = 0.22, 95% CI = 0.07-0.68) and blood type 0, Rh+ (OR = 0.25, 95% CI = 0.09-0.61).

Key words: Pancreatic cancer, risk factors, case-control study.
pp. 353-356