[Resistance testing and chemotherapy results in animal and human tumors].
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Biomedical subjects
Publications and source records attributed to M Volm.
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Treatment of the solid Walker carcinosarcoma of the rat for a period of one year with cyclophosphamide (40 mg/kg at each passage; tumours were transplanted weekly) resulted in the formation of a cyclophosphamide-resistant tumour cell line. Without further treatment, animals injected with non-treated or cyclophosphamide pre-treated cells survived for 10 days on an average. After therapy with cyclophosphamide (2 x 38 mg/kg), rats with non-treated tumour cells survived 22 days whereas those with pre-treated tumours survived only for 17 days. Tumour cells which were shown to be sensitive in vivo also exhibited a larger reduction in 3H-thymidine and 3H-uridine incorporation in the in vitro short term test after incubation with urine from cyclophosphamide-treated rats or with 4-hydroperoxy-cyclophosphamide. The resistance to cyclophosphamide which was detected in animal experiments with Walker carcinosarcoma can therefore also be observed using the in vitro short term test.
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The application of spores of the bacterium Clostridium oncolyticum s. butyricum (M 55) results in an increased incorporation of 131Iododeoxyuridine in tumors (amelanotic melanoma Fortner III of the golden hamster and Brown-Pearce carcinoma of the rabbit). This leads to an improved scintigraphic demonstration of the tumors.
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The effect of 1.38 X 10(-5) M adriamycin on tritiated uridine incorporation was studied after 3 hours treatment of suspensions of 25 advanced human lung carcinomas in vitro. The results were correlated with the responses to clinical therapy. All tumours which showed an inhibition of uridine incorporation in vitro of more than 40% were also sensitive to clinical treatment with adriamycin.
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Partial synchronization of the slow-growing cell of solid neurosarcoma in the rat can be achieved by blocking DNA synthesis with hydroxyurea. Possible augmentation after synchronization of the therapeutic effects achieved with various cytostatics was tested. The substances were administered to tumour-bearing rats at various times after synchronization (at the transition from G1 to S phase; at maximal DNA synthesis; at the transition from the S to the G2 phases; at mitosis) and therapeutic effects on tumour gs. Cyclophosphamide and adriamycin reduced tumour size, the effect being dose-dependent. No differences were noted between synchronized and non-synchronized cells. Hydroxyurea and vincristine had significantly less effect on the tumour cells, this minimal effect not being enhanced by synchronization. The reported results were obtained in tests on 360 rats and are statistically significant (double variance analysis).
Various human tumours have been investigated using, as previously described an in vitro method for sensitivity testing. This test provides information both on the rates of proliferation of tumours and on their resistance to cytostatic agents.
The incorporation of 131-Iodo-deoxyuridine in rat tumours can be increased by partial synchronisation with hydroxyurea. The reported investigation was carried out to determine whether the incorporation could be further increased by using a basic 131-Iodo-deoxycytidine-monophosphate-protamine complex as a depot of Iodo-deoxyuridine. It was shown that the incorporation observed after synchronisation could be increased by using this complex (non-synchronized tumours 1,46% retention, synchronized tumours 2,28% retention; measured 4 hour after the final injection of hydroxyurea) whereas in a control group injected with 131-Iodo-deoxycytidine-monophosphate alone, only a small increase was observed. The large increase in activity in the liver and in the spleen (liver: non-synchronized 2,76%, synchronized 2,79% retention; spleen: non-synchronized 3,00, synchronized 3,80% retention; measured 4 hours after final injection of hydroxyurea) which is observed after injection of the 131-Iodo-deoxycytidine-monophosphate-protamine complex renders it unsuitable for use in the scintigraphic detection of tumours. 131-Iodo-deoxyuridine is shown to be more suitable for this purpose.
The following cytostatic agents were tested for activity in vivo and in vitro inWalker carcinosarcoma 256 of the rat: cyclophosphamide, triaziquon, 5-fluorouracil, methotrexate, adriamycin, dactinomycin, daunorubicin, hydroxyurea, procarbazin and vincritine. With the exception of vincristine, the results of therapy in vivo could be predicted by using a rapid in vitro test system. This involved, for cyclosphosphamide, triaziquon, adriamycin, and daunorubicin, the measurement of 3H-uridine or 3H-thymidine incorporation. The activities of methotrexate and 5-fluorouracil could be determined from 3H-deoxyuridine incorporation and that of dactinomycin from 3H-uridine incorporation. The results of short-term tests (uring adriamycin, daunorubicin, and dactinomycin) in roughly 100 human tumors were compared with data in the literature on therapy with the same cytostatic agents. Good agreement was found between the results of in vitro tests and the literature data on clinical therapy.
With series of transplanted tumors, the activities of different cytostatic agents which directly influence the metabolism of nucleic acids (Actinomycin D, adriamycin, daunomycin, 5-fluorouracil, procarbazine, trenimon) was measured by determining 3-H-uridine incorporation in short-term (3hrs) incubations of tumor cell suspensions. Data obtained could be used to predict the response of each tumor to particular cytostatic agents in vivo. The activities of the cytostatic agents as determined using long-term tissue cultures (time of exposure of tumor cells to cytostatic agent 48 hrs were comparable to those obtained with the short-term test. In long-term cultures, determination of cell numbers gave results similar to those obtained by morphological evaluation. In SHORt-term test, differing sensitivities of tumors to cytostatics could be detected.
Certain substances which influenced nucleic acid metabolism were found to have about the same cytostatic activity on human cells when measured in tissue culture experiments (cell numbers) or in short-term cultures (3-H-uridine incorporation in cell suspensions). By treatment with a dose of cytostatics corresponding to 10 times therapeutic dose, chemosensitive tumors can be distinguished from non-responsive tumors. By using this in vitro test system to investigate the sensitivities of 100 human tumors, it is shown that 28 of these tumors were responsive to adriamycin, daunomycin and Actinomycin D. Good agreement was observed between these in vitro results and the literature data on clinical therapy using these particular substances.
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The sensitivity of tumors to cyclophosphamide was tested in vitro. For this purpose, the urine of cyclophosphamide-treated rats (90 min., 500 mg/kg) was used. This method of activating cyclophosphamid proved more advantageous than other methods; all cytostatically effective metabolites can be used in the test and the production of metabolites can easily be standardized. Consequently results with good reproducibility can be obtained. In various long-term and short-term test models with animal transplantation-tumors in vitro, the effects of the cyclophosphamide metabolites depended on the dose used. This test results in the various test models are comparable. In the tissue-culture of tumors (long-term test) sensitivity was evaluated by using morphological criteria as well as by counting cell numbers. In the short-term test the sensitivity of tumors was found out by influencing the nucleic acid syntheses (3-H-thymidine-, 3-H-uridine-incorporation). As the tissue-culture meets with difficulties as a routine clinical examination, the use of the short-term test is recommended. Using as model the fast proliferating WALKER-256-carcinosarcoma and a slowly proliferating rat adenocarcinoma, the results of the sensitivity-test in vitro were compared with the effects of therapy in animal tests. Tumor remissions corresponding with the results reached in vitro, which depended on dosis, could also be achieved in both transplantation-tumors. Human tumors, tested in in vitro studies (short-term-test), show a different sensitivity to cyclophosphamide.
On the basis of animal experiments it is shown that the results of tumor chemotherapy can be predicted with the aid of in-vitro test systems. In investigations carried out on human tumors, the correlations obtained between in-vitro test results and response to therapy are promising. However, definitive proof of a correlation can only be provided by planned, statistically supported clinical studies. Two types of information can already be obtained from the in-vitro tests: data concerning the rate of proliferation of the tumor, on which the activity of cytostatic agents depends, and data on certain types of resistance to cytostatics.