Forty years of the Oncological Institute at Brno.

The primary cultures of animal ascites tumor cells growth in tubes in suspension as a model in the screening of antitumor agents.

By alternating in vivo-in vitro passage of NK/Ly ascitic tumor cells, a line of tumor cells was achieved which can be used for primary in vitro screening of antitumor compounds. Example is given from testing various known and new compounds.

Effect of D-glucosamine and D-galactosamine on the uridine nucleotide concentration in mouse myeloid tumor (Graffi) and myeloma MOPC-21.

The effect of D-glucosamine and D-galactosamine on the content of uridine nucleotides in myeloid tumor (Graffi) and myeloma MOPC-21 of mice was investigated in vivo. After treatment with aminosugars a marked decrease in UTP and UDP-glucose quantity was found. The trapping of uridine phosphates by formation of UDP-sugar derivatives was different in the two tumors and depended on the aminosugar employed. D-glucosamine provoked an increase in the UDP-N-acetylglucosamine pool size in myeloid tumor (Graffi) and myeloma MOPC-21. D-galactosamine administration led to formation of UDP-galactosamine and UDP-N-acetylglucosamine in myeloma MOPC-21, while in myeloid tumor (Graffi) an increase in the content of UDP-N-acetylglucosamine was obtained only.

Macromolecular antitumor agents from Chamaenerium angustifolium.

The antineoplastic substances isolated from Chamaenerium angustifolium, Chanerol, a macromolecular polyphenol, and its complex with polysaccharide, Chanerozan, have been studied in vitro in tests of haemagglutination, tumor cell agglutination, leucoagglutination and some others. The results obtained show Chanerol as a new phytohaemagglutinin (lectin) which has no group specificity for human erythrocytes (ABO system). As distinct from other lectins (proteins) Chanerol belongs to another chemical class (macromolecular polyphenols) and has no effect on lymphocytes in leukoagglutination and blast transformation tests. Both preparations have been shown capable of agglutinating tumor cells in vitro and revealed antitumor activity in the intravenous and intraperitoneal administration to mice with transplanted tumors. The toxicity of Chanerozan and its titre of human and mouse erythrocyte agglutination are considerably lower than those of Chanerol. Thus, judging by haemagglutination test Chanerozan behaves as a typical complex of agglutinin with concurrent sugar.

Antiproliferative effect of thymosterin on KB tumor cells in vitro and h 18 R tumor development in the new-born rat.

The authors report on preliminary data concerning the antiproliferative activity of thymosterin upon the growth of the KB tumor cells in vitro and in vivo. The experiments in vitro showed an intense antiblastic action of thymosterin, which was proportional to the administered dose. The preliminary results of the in vivo experiments suggest that inhibitory effect of thymosterin on the h 18 R tumor growth in newborn rats.

Antileukemic effect of L-aspartyl-hydrazine.

Beta-L-aspartyl-hydrazine can significantly prolong survival of the A mouse strain infected with a lymphatic leukemia sensitive to L-asparaginase. By combining the above substance with L-asparaginase even better results can be obtained.

Morphology of metastatic cancer of the fallopian tube in uterine cervix carcinoma.

Morphological examination of 452 Fallopian tubes removed together with the uterus for uterine cervix cancer revealed in 221 patients at the time of operation and in 5 dead on autopsy, metastatic cancer in 8 females (3.5%) in 12 tubes (right-side involvement 2 cases, left-side 2 cases, ambi-lateral 4 instances). In 37 patients there were metastases in inner organs; of these diseased 21.6% were affected by metastatic cancer of the Fallopian tube. Macroscopically, metastatic tumors were of different form in 4 cases; in the remaining instances they were only discovered on microscopical examination. In two cases the tubes were affected in their whole length, in the other case the tumor was located in the ampullary portion. The cyto- and histological structures of the metastatic cancer were the same as those of the primary uterine cervix tumor. The predominating route of tumor dissemination has proved to be lymphogenic propagation (5 cases). In two cases lymphohematogenic dissemination, in one case propagation by implantation were observed.

Value of lymphangiography in the diagnosis and treatment of lymphangioma.

On the basis of the author's own results and literary data emphasis is laid on the significance of lymphography for confirming or establishing a diagnosis, for determining the extent of a lesion and for proper considerations on indications for surgical treatment. Relymphography following a surgical intervention is capable of showing whether the latter had been successful and of revealing foci of recurrence. The case history bears on two histologically verified lymphangioma -- in the first case of the soft parts of the right leg, in the second of the soft part of the left thoracic wall, the armpit and the mediastinum. In both cases, relapse after repeated surgical interventions was verified by direct lymphography from superficial lymph vessels of the foot and the cubital fossa.

Iliac lymph nodes as a primary lymph center draining the testicles.

The visceral radioisotopic lymphography, as a new technic, reveals new possibilities to investigate the lymph system of the testicle. The testicular lymph center is regularly visualized at the level of the para-aortic lymph nodes. But in 16.9% of the patients, respectively in 10.8% of the examined testes, iliac lymph nodes are demonstrated as a primary lymph center, draining the testicle. A preoperative scintigraphic search can be made for every patient with testicular neoplasma in the intention to obtain a more correct determination of the stage, the prognosis and the proper treatment planning.

Transplacental and direct action of benzo(a)pyrene studied in organ cultures of embryonic lung tissue.

Transplacental effect of benzo(a)pyrene was studied in organ cultures of embryonic lung tissue explanted from mouse donors injected by the carcinogen. Hyperplastic alteration of epithelium, followed by adenomatous changes were seen in embryonic lung tissue cultures from donor mice injected by benzo(a)pyrene. No alteration was seen in control cultures in which the mice were injected by a non-carcinogenic hydrocarbon (pyrene). Besides the blastomogenic action, a growth-promoting effect of the benzo(a)pyrene was observed in the particular organ cultures of the embryonic lung tissue.

Methodological approaches to the study of carbohydrate surface receptore on macrophages and tumor cells.

Ultracytochemical visualization of specific carbohydrate receptors by means of lectins can provide valuable topological information about the functional status of cell surfaces in normal and transformed cells. In addition to the concanavalin A (Con A) rection and precoupling of lectin--peroxidase (PO) a new method is suggested using precoupling of the marker enzyme (PO) with an appropriated glycoprotein (glycopeptide) containing a lectin-specific carbohydrate. Furthermore, quantitative estimations of Con A reaction by means of automatic umage analysis are performed on peritoneal macrophages which point out a cluster formation, but give no hints for a loss of glycocalyx during the preparation process contrary to tumor cells.

Different sensitivity to cytostatic drugs of primary tumor and metastasis of the Lewis carcinoma.

Differences in cytostatic sensitivity between subcutaneously transplanted Lewis lung carcinomas and metastases from them were studied in vitro. Sensitivity to vinblastine sulphate and melphalan was determined as the reduction of the incorporation of H3-thymidine into drug-treated cells compared with control cells. The main result of the present study is that tumor cells obtained from pulmonary metastases are more sensitive to cytostatic drugs in vitro than are the cells obtained from the corresponding subtaneous tumors. When cells from pulmonary metastases are transplanted subcutaneously, the sensitivity of the resulting tumor closely resembles that of other subcutaneous tumors. However, when the tumor cells have been passed through many cycles where the pulmonary metastases from subcutaneous tumors were re-transplanted subcutaneously and then allowed to metastasize again, a selection of tumor cells has apparently occurred: they have become more sensitive even when grown subcutaneously. Different explanations of this phenomenon are discussed.

Experimental study of the combined effect of leukeran, degranol and predisolone.

The effect of the combined treatment with leukeran, degranol and prednisolone on the lymphoid tissue and marrow hamopoiesis in rats is studied. The simultaneous administration of these drugs is shown to cause a more effective inhibition of the lymphoid tissue and a less suppression of the marrow, as compared with their separate administration.

In vitro incorporation of tritiated thymidine by the Sternberg-Reed cells in Hodgkin disease.

The study of the possibilities of new DNA synthesis by the Sternberg-Reed cells of Hodgkin disease was done with tritated thymidine and autoradiography. The results showed that after incubation pulses of 30 and 60 minutes cells with lobulated nucleus, binucleated and trinucleated cells, identifiable to the diagnostic Sternberg-Reed cells, had possibilities of new DNA synthesis. This points to a more dynamic interpretation of this cell that was considered without chances of cnromosome replication.

Comparative study of the amino acid composition of some tumor and normal melanosomes.

The amino acid composition of melanosomes from human and Harding-Passey mouse melanomas and from pigmented tissues of cattle eyes isolated according to BOLT was studied. The 18 current amino acids and moreover dopa were found in all the hydrolysates studied. By means of apolar/polar amino acid ratio suggested by HATCH the possible properties of melanosomal protein(s) were infered. The higher level of cysteine and lysine in mouse melanosomes as compared with tyrosinase supports the theory of special matrix protein in melanosomes. Lysine seems to have a role in regulation of the quantity of synthesized melanin.

Phosphatases XII. Isoenzymes of alkaline phosphatase and radionuclear investigation (85Sr) of patients with neoplastic affection of the skeleton.

In a group of 30 patients with neoplastic processes, 13 were found to have a significantly increased serum alkaline phosphatase activity. This finding correlated with the results of investigation with 85Sr in 7 patients, but owing to a concomitant hepatal symptomatology, it proved of differential diagnostic value in only one of them. On the other hand, the activity of bone isoenzyme of alkaline phosphatase was significantly altered in 15 patients. In 14 of them the increase in bone isoenzyme activity corresponded to an X-ray and a radionuclear finding of a tumor process in the bones. This activity was within the normal range of values in only one patient with a positive result of the 85Sr investigation. An agreement between the results of determination of bone isoenzyme activity of alkaline phosphatase and those of radionuclear investigations was found in 27 patients. In addition, a correlation was established between the increased activity of bone isoenzyme and that of intestinal isoenzyme of alkaline phosphatase. Enzyme investigation can be suitably utilized, alongside radionuclear examination, for early detection of a bone process.

Phagocytic activity of leukocytes in tumors (clinical studies).

1. A method of evaluating in vitro phagocytic activity of leukocytes, using inert latex particles has been described. 2. 363 investigations have been carried out according to the method in clinically healthy donors, in patients with and without malignant disorders. The subjects have been divide into groups according to nature of the disease (operable and inoperable tumors, non-malignant, acute anc chronic disorders). 3. A statistical evaluation of the results revealed that the phagocytic capacity of leukocytes is significantly activated in the presence of a pathological process in the organism and this to a considerably higher degree in tumorous disorders than in non-malignant affections. The degree of activation of the phagocytic capacity of leukocytes can be measured by this method and the latter can be used in every haematological laboratory.

Combination chemotherapy of Hodgkin disease in children.

MOPP chemotherapy program was employed in 20 children with advanced Hodgkin disease (stages III and IV). Complete remissions were achieved in 13 patients (65%)within an average duration rate of 17.1 months varying from 2 to 40 months. If thses only one patient developed recurrency and died 10 months after the onset of treatment. The remaining 12 children are alive and well; an average survival rate since the onset of MOPP chemotherapy being 20 months. The patients with partial remission died from further progression of the disease within the period from 2 to 15 months. The study showed that histological type and mode of previous treatment had an influence on the results of chemotherapy used. The complications of MOPP chemotherapy were insignificant and easily controlled.

Inhibition of hamster cell transformation and of benz(a)pyrene hydroxylation by antioxidants.

Addition of butylated hydroxyanisole (BHA) and butylated hydroxytoluene (bht) to benz(a)pyrene (BP) exposed cell cultures results in slight reduction of transformation. The benz(a)pyrene hydroxylation is reduced when antioxidants are added to the incubation mixture.

Antitumor effect of 2-thio-4-hydrazinouracil on transplantable tumors.

The cytostatic activity of 2-thio-4-hydrazinouracil (THU) on some transplantable tumors has been studied. A strong effect of this compound (between 60% and 100% suppression) has been found in the case of Myeloma P-8 (MOPC-21) and Sarcoma 180 (Crocker). A less pronounced effect has been observed on Yoshida sarcoma, while the development of Jensen sarcoma is not influenced.

DNA measurements on cell nuclei of normal, proliferating and neoplastic thyroid tissues in rats.

Nuclear DNA content was measured in 3 normal, 9 hyperplastic and 16 neoplastic rat thyroid glands. Thyroid hyperplasia and tumor growth were induced after treatment of the animals with X-rays and methylthiouracil. In the control animals only diploid thyroid epithelial cells were observed. At the stages of diffuse and nodular thyroid hyperplasia, the total DNA content per nucleus indicated for a diploid chromosome number and only a few cells were hyperdiploid. In the thyroid adenomas and carcinomas a scattering of the diploid region and an increase in the number of hyperdiploid cells was found. Among the various types of thyroid tumors neither difference in the number of hyperdiploid cells, nor typical pattern of distribution of these cells in the histogram was found. The increased number of hyperdiploid cells in the hyperplastic and neoplastic thyroids suggest an increase in the proportion of the cells entering the cell cycle and does not indicate for appearance of a neoplastic stemline.

Preliminary analysis of failures in the radiotherapy of head and neck cancer with a dynamic dose-fractionation in combination with breathing oxygen at ambient pressure.

Treatment failures were analysed in 43 head and neck cancer patients treated with a dynamic dose-fractionation in combination with breathing oxygen at ambient pressure. Infiltrative T3-4 lesions with necrosis showed poorer results than exophytic and necrosis-free infiltrative lesions. Possible differences in the oxygenation of the different types of lesion were considered to explain the results.

Clinical analysis of 43 surgically-treated patients with thyroid carcinoma.

The authors have analysed 43 patients with thyroid carcinoma who had been treated at the surgery department of the Oncological Institute between 1960 and 1970. The patients are distributed in four groups according to the histologic tumor structure. Considering the specific properties of each particular group, the occurrence of metastases, treatment, survival and prognosis are being discussed. An outline is given on the characteristics of the various forms of thyroid carcinoma from the histological but mainly clinical aspects. The development of regional and distant metastases, surgical treatment, postoperative irradiation or application of radioiodine and hormonal tumor dependence are presented in the paper. The best results were obtained in the papillary and the follicular form, the prognosis in the latter being dependent on the degree of vascular invasion. The solid carcinoma of thyroid shows considerable differences in survival of patients depending on regional metastases and histologic grade. The anaplastic carcinoma has the worst prognosis where survival is expressed in months, exceptions are rare.

Contributions of virology to oncology.

New potential cytotoxic and antitumor substances I. In vitro effect of bikaverin and its derivatives on cells of certain tumors.

Bikaverin and its derivatives have been found to affect precursor utilization of nucleic acid and protein synthesis in the cells of Ehrlich ascites carcinoma (EAC). Mainly the uridine incorporation into EAC cells was inhibited. This is in agreement with the known concept that anthraquinones, to which bikaverin may also be assigned, intervene into RNA synthesis. The substances followed exerted a cytotoxic effect on in vitro proliferating cells of the three studied tumors. The ED50 values found for cells of these tumors were: EAC 0.5 mug/ml; leukemia L 5178 1.4 mug/ml; sarcoma 37 4.2 mug/ml.

The effects of combined administration of cytembena and cyclophosphamide on the blood count and morphology of nucleoli in peripheral-blood lymphocytes in patients with malignant tumors.

In patients with malignant tumors, changes in the blood count and morphological changes in the nucleoli in peripheral-blood lymphocytes were studied during treatment with Cytembena, Cyclophosphamide, or combinations of both drugs. After administration of Cyclophosphamide (400 mg i.v. daily for 10 days, then 100 mg orally daily), the counts of leukocytes, especially of neutrophils, sank (by 40% maximally), and the erythrocyte count and the haemoglobin content in blood sank moderately. In peripheral-blood lymphocytes, the proportion of ring-shaped nucleoli (stimulable to synthesis of RNA) reversibly sank, and the proportion of micronucleoli (nonstimulable) rose. After administration of Cytembena (400 mg i.v. daily for 10 days, then 200 mg i.m. at intervals of 2--4 days), neither the leukocyte counts nor the relative percentages of lymphocytic nucleoli signigicantly changed; the mean corpuscular haemoglobin content (MCH) transitorily increased (by 10 percent max.). In the group receiving combined cytostatics (Cytembena 400 mg i.v. daily for 10 days, then 200 mg i.m. at intervals of 2--4 days plus Cyclophosphamide 50 mg orally daily), the erythrocyte count and the haemoglobin content in blood sank moderately. In the population of peripheral-blood lymphocytes in this group, there sank the absolute counts of lymphocytes with ring-shaped nucleoli (by 44% max.) and those with micronucleoli (by 33% max.).

Morphological changes in the nucleoli of peripheral-blood lymphocytes as prognostic criteria in the chemotherapy of malignant tumors.

By a simple cytochemical demonstration of RNA, the percentages of various nucleolar types, reflecting the activity of RNA synthesis in peripheral-blood lymphocytes, were established. The checkings were done in patients with advanced stages of malignant tumors processes with primary localization in the digestive tract, receiving cytostatic treatment with Cytembena Spofa and Cyclophosphamide Germed. Evidence has been brought that significant differences in the indicators investigated existed between a healthy population and patients with cancer before the start of the treatment. In the further course of the disease, during cytostatic treatment, peripheral lymphocytes underwent changes followed sooner of later by changes in the clinical state of the patients: a decrease in the percentage of activable (ring-shaped) nucleoli, and increase in the percentage of irreversibly inactivated nucleoli (micronucleoli) incapable of RNA synthesis, and an elevation of the nucleolar coefficient were followed by a deterioration of the patients' clinical state and a progression of the tumorous disease. With an opposite trend of the nucleolar-test findings (an increase in the percentage of nucleoli capable of RNA synthesis, a decrease in the percentage of nucleoli incapable of RNA synthesis, and a lowering of the nucleolar coefficient), the further development of the disease appeared to be favourable.

On the influence of chloramphenicol on the induction of lung adenomas by urethane in mice.

It has been shown by a number of authors that the antibiotic chloramphenicol (CP), produced in the USSE under the name of levomycetin, had considerably inhibited the growth of liver tumors, induced by various carcinogenic compounds [3,7,8,21]. Moreover, it has been found by ourselves and other authors that CP greatly reduces the toxic effect of carcinogenic polycyclic hydrocarbons on cultures of embryonic fibroblasts of mice and the adrenals of rats [5,9,20,23]. As the full extent of the antiblastomogenic activity of CP has so far not been studied, it was important to elucidate its influence on other groups of chemical carcinogens. In experiments described in the present paper we have investigated the influence of CP on the incidence of lung adenomas in mice after administration of urethane.

On the enhancement of oncolytic effect of laser radiation by means of fast electrons.

Non-inbred albino mice were inoculated subcutaneously or intraperitoneally with Ehrlich carcinoma cells pre-exposed in vitro either to different doses of laser radiation or to fast electrons alone, or to both agents. The statistical data on the latency of tumors, their weights (subcutaneous inoculation) and the survival time in group "laser + electrons" (intraperitoneal inoculation) were found to differ considerably from those for the mice inoculated with the ascitic tumor cells pre-exposed either to laser radiation or fast electrons alone. The enhancement of the oncolytic effect of laser radiation combined with fast electrons is gained through the addition of the effects of each of these agents.

A freeze-etch study of occurrence of nuclear pores in normal and tumor cells.

A freeze-etch study of nuclear pores performed on human lymphocytes, epidermal and corneal cells, on hamster fibroblasts, on rat and hamster sarcoma cells and cells from a human malignant melanoma, revealed that the frequency of pores as a part of very important biological cellular structures increases in proliferating cells, and that there is a statistically significand difference between normal and tumor cells. Once produced the pores maintain and their frequency practically does not change. The pores are randomly distributed on the nuclear envelope. Markham rotating method revealed an octa- or nonaedric outside shape of the pores and round inner margin with eight or nine granules. One bigger granule was found in the center of the pores. The granule is with a great probability filamentously attached to the margin of the pore. Fibrillar structures running to the pores on inner surface of nuclear envelope as far as the chemical composition of granules need special cytochemical examinations.