Antitumor activity of alkylphosphocholines and analogues in methylnitrosourea-induced rat mammary carcinomas.
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Animals↗
Biomedical subjects
M R Berger
Publications and source records attributed to M R Berger.
Effect of miltefosine on transplanted methylnitrosourea-induced mammary carcinoma growing in Fischer 344 rats.
Miltefosine (hexadecylphosphocholine, HPC, CAS 58066-85-6) was investigated in transplanted primary methylnitrosourea-induced PYH mammary carcinoma of F344 rats. The therapy was performed in the 5th and 10th passage. At first HPC (113 mg/kg body weight) significantly reduced the median tumor volume, but a loss of activity was observed in the 10th passage. To explain the loss of sensitivity and to obtain information on the mechanism of action histology, cytoskeleton and hormone receptor content were investigated. The most important change was observed in the histopathology of the tumor. The initial tubular papillary adenocarcinoma was transformed into a malignant adenoacanthoma with epithelial structure. Vimentin as an endothelial marker of the cytoskeleton was equally expressed in all passages. Cytokeratin was weakly expressed in the earlier passages and intensively present in the late passages. The histopathological change from tubular adenocarcinoma to malignant adenoacanthoma might be caused by an overgrowth of the primary epithelial tumor cells or by a real transformation in the morphological characteristics of the tumor, which may occur during repeated transplantation.
Adenocarcinoma↗
Antitumor activity of some ruthenium derivatives in human colon cancer cell lines in vitro.
Six ruthenium derivatives were evaluated in vitro in two human colon cancer cell lines (SW707 and SW948) utilizing the microculture tetrazolium test (MTT) and cell counting with a Coulter Counter. The ruthenium compound sodium-(tetrachloroimidazoledimethylsulfoxideruthenate)- bisdimethylsulfoxide (Na(RuDMSOimCl4)) showed the best efficacy in inhibiting cell proliferation of both colon cancer cell lines followed by the other DMSO ruthenium compound sodium-(tetrachloroindazoledimethylsulfoxideruthenate) - bisdimethylsulfoxide (Na(RuDMSOIndCl4)), as demonstrated by IC50 values (80 and 90 micrograms/ml in SW707 and SW948 cell lines for Na(RuDMSOImCl4); 155 and 165 micrograms/ml in SW707 and SW948 cell lines for Na(RuDMSOIndCl4), respectively). Out of the ruthenium derivatives without DMSO, transindazolium - [tetrachlorobis(1H - indazole)ruthenate (III,N2)] (HInd[RuInd2Cl4(N2)]), was as active as its DMSO-containing congener whereas trans-imidazolium- [tetrachlorobisimidazoleruthenate)(III)], (HIm(RuIm2Cl4)) was less active, as shown by the IC50 values: (HIm (RuIm2Cl4) = 250 and 260 micrograms/ml in cell lines SW707 and SW948; HInd[RuInd2Cl4(N2)] = 110 and greater than 200 micrograms/ml in cell lines SW707 and SW948, respectively). The other ruthenium derivatives containing pyrazole and triazole as ligands (trans - pyrazolium (tetrachlorobispyrazoleruthenate) (III), PzH(RuPz2Cl4) and triazolium(tetrachlorobistriazoleruthenate) (III), TrH(RuTr2Cl4)) were active only at high concentrations that cannot be regarded as realistic in vivo, as shown by the respective IC50 values: (PzH(RuPz2Cl4) = 1056 and 750 micrograms/ml in cell lines SW707 and SW948; TrH(RuTr2Cl4) = 350 and 300 micrograms/ml in cell lines SW707 and SW948). The promising activity of ruthenium compounds with DMSO, indazole and imidazole as ligands should be evaluated in vivo for elucidating their possible role in the treatment of colorectal cancer.
Antineoplastic Agents↗
Metabolic design of combination therapy: use of enhanced fluorodeoxyglucose uptake caused by chemotherapy.
In order to quantify effects of an experimental chemotherapy, MCF7 cells were studied with 14C-fluorodeoxyglucose (FDG) and high-performance liquid chromatography (HPLC). Uptake measurements were performed 1 and 4 hr after the end of a therapy with hexadecylphosphocholine (HPC). A dose- and time-dependent increase of the FDG uptake after therapy was observed, with a maximum at 1 hr after therapy. These data were used to develop a new metabolic design of combination treatment. Several time-dose combinations of HPC and deoxyglucose (DOG) were analyzed for their effects on growth inhibition. The combinations using DOG in the period of pronounced enhancement of FDG uptake (1 hr after HPC treatment) were found to be the most effective with an improvement of up to 520% in growth inhibition. This metabolic design of combination treatment may also be applied in vivo, and PET can be used to optimize the time and dose schedule of the modified treatment protocol.
Antineoplastic Agents↗
Effect of recombinant human erythropoietin on the growth of human tumor cell lines in vitro. Micro-titertec-tetrazolium assay.
Cells derived from 9 human tumors were tested in vitro using 5 doses ranging from 1 to 5000 units of recombinant human erythropoietin (rhEpo) per ml to assess a proliferation response to this hormone. The following tumors were used: 2 melanomas, 1 hepatoma, 3 renal cell carcinomas, 1 adenocarcinoma of the lung, and 2 mammary carcinomas (with and without sex steroid hormone receptors). Compared to untreated cells, none of the cell lines exposed to rhEpo revealed differences in proliferation extending beyond the limits of methodological fluctuation. rhEpo thus does not have any effect on the growth of human tumors in vitro under the experimental conditions applied.
Cell Division↗
On the long-term toxic risk of dinaline.
The long-term toxic risk of the cytostatic agent dinaline (4-amino-N-(2'-aminophenyl)-benzamide) was assessed in a rat bioassay. Regular administration of 9, 3 and 1 mg/kg for 106 weeks was associated with significantly increased survival of female rats receiving the median and low doses. Dinaline significantly reduced the occurrence of malignant tumors in male rats and prolonged the manifestation time of malignancies in female rats. Unlike malignant tumors, benign neoplasms were increased in male rats and were not significantly different from controls in female rats. Analysis of organ distribution of neoplastic lesions revealed a dose-dependently decreased tumor incidence in the hematopoietic and lymphatic tissue, the mammary gland (females only) and the pituitary gland and a not dose dependently reduced incidence of liver tumors. This contrasted with dose dependently increased tumor incidences in the adrenal gland, the gonads and the vagina. Considering these findings, dinaline has to be assessed as a modulator of carcinogenesis in rats. The observed decreased and increased tumor incidences suggest a hormone-related mechanism of action.
Animals↗
Effect of dietary calorie and fat restriction on mammary tumor growth and hepatic as well as tumor glutathione in rats.
Effect of dietary calorie restriction and fat reduction on growth of established mammary carcinoma in rats and on glutathione levels in liver and tumor tissue was investigated. Reduced (GSH) and oxidized (GSSG) glutathione were determined enzymatically. Female Sprague-Dawley rats were injected with 25 mg/kg methylnitrosourea (MNU) on day 50 of life for tumor induction, and subsequently fed a diet containing 50 kcal/day with 45% (energy %) fat. When tumors reached approximately 1 cm3, the diet was changed for 10 +/- 2 weeks. Four dietary groups were formed: two high calorie groups (50 kcal/day) with 45% or 25% fat and two calorie restricted groups (35 kcal/day) with 45% or 25% fat, respectively. Tumor growth was significantly inhibited by the 30% calorie restriction, and the inhibition was most effective in the calorie restricted group with low fat level. However, reduction of fat, alone, had no significant inhibitory effect. GSSG levels in both liver and tumor showed no differences among the groups. Hepatic GSH levels tended to be lower in the calorie-restricted groups, and showed no difference between isocaloric groups with different fat levels. In contrast, GSH in tumor tissue tended to be lower in the low fat groups, independently of calorie levels.
Animals↗
Evaluation of incorporation characteristics of mitoxantrone into unilamellar liposomes and analysis of their pharmacokinetic properties, acute toxicity, and antitumor efficacy.
Mitoxantrone (MTO) was incorporated into small unilamellar liposomes by formation of a complex between the anticancer drug and negatively charged lipids. The complex was formed at a 2:1 molar ratio between the lipids and MTO, with phosphatidic acid (PA) being the strongest complex-forming lipid. Weaker complexes and lower incorporation rates of MTO resulted when liposomes containing dicetylphosphate, phosphatidyl inositol, phosphatidyl serine, phosphatidyl glycerol, oleic acid, and tridecylphosphate were used. Thus, all further experiments were performed with PA-MTO liposomes that contained 0.1-3 mg MTO/ml and had mean vesicle sizes of 40-150 nm, depending on the drug concentration and the method of liposome preparation. In vitro incubations of free and liposomal MTO with human plasma showed that the drug is slowly transferred from the liposome membranes to the plasma proteins. For liposomal MTO a transfer rate of 48% was determined, whereas 75.8% of free MTO was bound to the plasma proteins. The organ distribution of the two preparations in mice showed that higher and longer-lasting concentrations of liposomal MTO were found in the liver and spleen. The terminal elimination halflives in the liver were 77 h for liposomal MTO and 14.4 h for free MTO. In the blood, slightly higher concentrations were detected for liposomal MTO, which also had slower biphasic elimination kinetics as compared with the free drug. Drug distribution in the heart was not significantly different from that in the kidneys. The LD25 of PA-MTO liposomes in mice was 19.6 mg/kg and that of free MTO was 7.7 mg/kg. The antitumor effects of PA-MTO liposomes were evaluated in murine L1210 leukemia, in various xenografted human tumors, and in methylnitrosourea-induced rat mammary carcinoma. Generally, the liposomal application form was more effective and less toxic than the free drug. The cytostatic effects were dependent on the tumor model, the application schedule, and the drug concentration. At doses that were toxic when free MTO was used, the liposomal preparation produced strong antitumor effects in some cases. In summary, the incorporation of MTO into liposomes changes the drug's plasma-binding properties, alters its organ distribution, reduces its acute toxicity, and increases its cytostatic efficiency in various tumor models. The liposomal PA-MTO complex represents a new application form of MTO that has advantageous properties.
Animals↗
Locoregional administration of 5-fluoro-2'-deoxyuridine (FdUrd) in Novikoff hepatoma in the rat: effects of dose and infusion time on tumor growth and on FdUrd metabolite levels in tumor tissue as determined by 19F-NMR spectroscopy.
The influence of infusion time and dose on the anticancer efficacy of 5-fluoro-2'-deoxyuridine (FdUrd) was investigated using a locoregional therapy model: Novikoff hepatoma transplanted i.m. into the thigh of Wistar rats and FdUrd infusion via a catheter implanted in the femoral artery. In experiment A the FdUrd dose (five daily doses of 12, 19 and 30 mg/kg) and the duration of administration (bolus, 1 h, 5 h, and 24 h) were varied. The change in tumor volume following treatment and the number of rats showing regression vs progression served as indicators of therapy response. The results showed a clear dose dependence, and for each infusion time the 30 mg/kg dose was the most effective, without any signs of general toxicity. At this dose the longest infusion time (24 h) was less effective (regression in three of six rats) compared with 1-h or 5-h treatments (four of five in regression). In experiment B either one or five daily FdUrd doses (15, 30, 60 mg/kg) were administered i.a. for the same infusion times used in experiment A. After treatment, tumors were explanted ex vivo and approximately 1-g tissues samples were immediately frozen in liquid nitrogen for storage. 19F-NMR spectroscopy at 11.7 T was used to quantify FdUrd metabolites [5-fluorouracil (FUra), alpha-fluoro-beta-alanine (F beta Ala), 5-fluorouracil nucleosides and nucleotides (F-Nuc)] in the solid tumor tissue samples (maintained at 4 degrees C) with a detection threshold of about 5 nmol/g. The metabolite signal pattern indicated that FdUrd is first converted to FUra, followed by anabolism primarily to nucleotides in the oxy form (e.g. FUTP). The total amount of fluorine detected in tumor tissue increased with dose and decreased with infusion time. For all treatments FNuc could be detected, even after 24 h infusion, and their levels showed a good linear correlation with the total F. The major catabolite F beta Ala was present in tumor at low levels that correlated poorly with total F, indicating recirculation from other organs (e.g. liver) as the main source. Thus, the NMR method can provide detailed information regarding the efficiency of locoregional treatment (catheter function, drug uptake and metabolism). Initial results of non-invasive in vivo NMR experiments are also presented.
Animals↗
Treatment of chemically induced autochthonous rat mammary and colorectal carcinomas with interleukin-2.
The antineoplastic efficacy of human interleukin-2 (IL-2) in autochthonous methylnitrosourea-induced mammary carcinoma and in acetoxymethyl-methyl-nitrosamine-induced colorectal carcinoma of Sprague Dawley rats has been investigated. Under the conditions applied, IL-2 was non-toxic. In the mammary carcinoma IL-2 was therapeutically inactive. In the colorectal carcinoma, 1200 U IL-2/day exhibited significant antitumour activity in established tumours as well as in tumours treated "prophylactically" before their manifestation (P less than 0.05). The effect of IL-2 seemed to be more pronounced when given before manifestation of colorectal tumours (T/C = 8.7% vs 17.8% in established tumours). The differential sensitivity of the autochthonous mammary and colorectal carcinoma may be explained by differences in their proliferation rates and differences in volumes at the beginning of IL-2 therapy. IL-2 seems to be preferentially active in small tumours with a low proliferation rate, a feature typical of colon tumours.
Animals↗
Relationship between dose and risk reduction: statistical evaluation of a combination experiment with three hepatocarcinogenic N-nitrosamines in rats.
Data from an experiment on the single and combination effects of very low doses of N-nitrosodiethylamine (NDEA), N-nitrosopyrrolidine (NPYR) and N-nitrosodiethanolamine (NDELA) in 1800 male Sprague-Dawley rats were analysed for age-specific incidence (time to death with liver tumour) by Cox's proportional hazards model. The model revealed a linear relationship between daily exposure to low levels of N-nitrosamine and time to death with liver tumour within the dose range investigated: an increase in individual dose resulted in a proportional decrease in liver tumour-free survival. The finding was established for the three individual carcinogens as well as for their combination. NDEA was 40 times more active than NDELA. Extrapolation to N-nitrosamine exposure levels lower than those used in the experiment revealed only a minor reduction in age-specific liver tumour incidence compared to that achieved by an equivalent reduction within the experimental dose range. In rats at advanced age a further reduction in carcinogen-induced liver tumour incidence did not contribute to longer overall survival, due to competitive, probably independent causes of death. The data thus support the idea of a quasi-threshold in terms of a 'no observed effect level'.
Age Factors↗
Strategies of reducing the carcinogenic risk of cytostatic agents on the basis of bioassay evaluation.
This article described strategies that can be used to reduce the carcinogenic risk of cytostatic chemotherapy and summarizes our recent experimental results. Reduction of neoplasms caused by the carcinogenic potency inherent in cytostatic agents can be obtained. (A) by chemical modifications such as: (1) exchanging a chlorine atom in N, N'-bis-(2-chloroethyl)-N-nitrosourea (BCNU) in the chloroethyl group at N'-position for a hydroxyl group to form the less carcinogenic analog N-(2-chloroethyl)-N'-(2-hydroxyethyl)-N-nitrosourea (HECNU); (2) linking chlorambucil to the steroid prednisolone to obtain a conjugate (prednimustine) with distinctly lower carcinogenic potential than chlorambucil; (3) progressive ring halogenation of phenyl-triazenes to generate agents with decreased long-term toxic risk; (B) by replacing cyclophosphamide within the carcinogenic drug combination of cyclophosphamide, methotrexate and 5-fluorouracil (CMF) by vincristine to form the combination VMF which has no detectable carcinogenic potential; (C) by coadministration of cyclophosphamide and mesna to achieve a dose-related reduction of cyclophosphamide-induced urinary bladder carcinomas; (D) by administration of dinaline, a compound which reduces the spontaneous incidence of malignant tumors in rats. These examples demonstrate that the carcinogenic risk of single agents and drug combinations used for antineoplastic chemotherapy has successfully been reduced, as assessed in long-term bioassays. Such strategies should be considered in the treatment of patients with long life expectancy following cytotoxic chemotherapy.
Animals↗
In vitro investigations on the antineoplastic effect of hexadecylphosphocholine.
The in vitro antineoplastic activity of hexadecylphosphocholine (HPC, CAS 58066-85-6) on methylnitrosourea (MNU)-induced mammary carcinoma in Sprague-Dawley rats and human mammary carcinoma was investigated with the modified Hamburger-Salmon-Colony-Assay (HSCA). The effect of HPC was also compared with ilmofosine (alkyllysophospholipid derivative) and N-(2-chloroethyl-N-nitroso-N'-2-hydroxyethylurea (HECNU). Moreover, the modified HSCA was performed with bone marrow cells and KB cells. In this investigation, no specific antineoplastic effect of the test compounds on MNU-induced mammary carcinoma and human mammary carcinoma was found in HSCA. Thus the antineoplastic activity of the compounds could not be compared on these cell types. In vitro, effects were only observed in bone marrow and KB cells at a very low concentration. The in vitro effect of HPC on mammary carcinoma cells, evaluated through HSCA, did not predict the effect of HPC in vivo. The reason has remained unknown but some hypotheses are discussed. Because of the contrary results of HPC in vitro and in vivo, it should be pointed out that drug development in this class of compounds mainly has to depend on in vivo experiments.
Animals↗
The role of nitrate, nitrite and N-nitrosamines in carcinogenesis of colon tumours following ureterosigmoidostomy.
Urinary diversion in both a rat model for ureterosigmoidostomy and in ureterosigmoidostomy patients result in an increased incidence of colon tumours. Bacterial and chemical investigations on feces-urine mixtures from both the rat model and ureterosigmoidostomy patients showed the presence of a complex nitrate-reducing bacterial flora in both rats and humans. This bacterial flora actively reduced urinary nitrate to nitrite in humans and increased the endogenous formation of N-nitroso compounds. No evidence of urinary nitrate reduction and increased nitrosamine formation in the rectosigmoid of rats was found. The results support the N-nitrosamine theory of carcinogenesis of the colon following ureterosigmoidostomy in humans, but not in rats. As the rat model induces colon carcinomas, factors other than the increased endogenous formation of N-nitroso compounds in the rectosigmoid may contribute to the initiation of colon carcinomas following ureterosigmoidostomy.
Animals↗
Antineoplastic efficacy of melphalan and N-(2-chloroethyl)-N-nitrosocarbamoyl-omega-lysine, in combination with diazoxide or insulin in autochthonous mammary carcinoma of the Sprague-Dawley rat.
The anticancer activity of melphalan and N-(2-chloroethyl)-N-nitrosocarbamoyl-omega-lysine (CNC-omega-Lys), was compared in the autochthonous, methylnitrosourea-induced mammary carcinoma of the Sprague-Dawley rat. In addition, the influence on the therapeutic efficacy of the combination with diazoxide, causing a mild, reversible diabetes, and with insulin was investigated. The comparison of melphalan and CNC-omega-Lys clearly showed the superiority of melphalan. Both compounds displayed a significant tumour inhibition in their medium and the highest dosages in comparison to the untreated control. The combination with diazoxide resulted for almost all groups in an increased tumour inhibition. Only the lowest dose of CNC-omega-Lys + diazoxide did not reduce the tumour volume significantly versus the control group. The combination with insulin, however, resulted in a loss of tumour inhibition compared to the effect of the cytotoxic drug alone, although in these groups, too, a significant decrease of tumour volumes versus controls could be observed. Mortality was within tolerable limits (less than 20%) through the treatment period for all experimental groups. Median lifespans were increased in all therapy groups, but no additional benefit could be observed in the combination treatment groups.
Animals↗
Differential relationship of vitamin A and E levels in methylnitrosourea-induced Sprague-Dawley rats following prolonged feeding of fatty diets enriched with the vitamins.
Prolonged supplementation of vitamins A and E (2500 IU and 11.3 IU/rat daily) in 10.3% and 21.6% fatty diets administered to methylnitrosourea-induced Sprague-Dawley rats caused alterations in plasma and liver levels of both vitamins as well as of the total lipids. Liver vitamin A levels increased steadily (more than fourfold) until termination of the experiment, whereas plasma vitamin A levels only showed a minor increase at the end of the study in comparison to rats fed a standard diet. Liver vitamin E levels initially showed no difference from those of rats treated with a standard diet, and were decreased at the end of the experiment, whereas the respective plasma levels were increased throughout the study, only showing a decrease relative to initial values at termination of the experiment. Plasma total lipids increased with age and following supplementation of the vitamins in the high-fat diet, whereas liver total lipids were influenced less by aging than by the fat and vitamin contents of the respective diets. The observed alterations were, however, not related to significant chemopreventive activity of the vitamins A and E contained in high-fat diets.
Animals↗
Androgen-linked alkylating agents: biological activity in methylnitrosourea-induced rat mammary carcinoma.
This article gives a comprehensive survey on the anticancer activity of nitrosoureas linked to steroidal androgens in methylnitrosourea (MMU)-induced rat mammary carcinoma. cis-Androsterone, testosterone, 19-nortestosterone and 5-alpha-dihydrotestosterone were used as carrier hormones and were linked to various cytotoxic N-[N'-(2-chloroethyl)-N'-nitrosocarbamoyl] (CNC)-aminoacids and to N-(2-hydroxyethyl)-N'-(2-chloroethyl)-N'-nitrosourea hemisuccinate (HECNU-hemisuccinate). In the MNU-model used esters of dihydrotestosterone (DHT) invariably were more active and less toxic than those of testosterone, nortestosterone and cis-androsterone. Within the DHT esters of CNC-aminoacids those of CNC-glycine, CNC-methionine and CNC-alanine showed the highest antineoplastic activities and superiority compared with equimolar dosages of their unlinked mixtures. Additionally, CNC-alanine-DHT ester had the highest therapeutic ratio of all agents investigated. HECNU-hemisuccinate-DHT ester, on the other hand, achieved even higher antitumor activity at the optimal dose but had a narrower therapeutic ratio. No obvious correlation between antineoplastic efficacy and receptor binding affinity could be demonstrated, but, to be active, a conjugate apparently had to have some receptor binding affinity for both androgen and progesterone receptors. The results obtained indicate that linking antineoplastic agents to transport molecules with affinity to steroid receptors is a highly promising approach to obtain drugs with specific activity in steroid receptor containing tumors.
Alkylating Agents↗
Possible role of acetaldehyde in ethanol-related rectal cocarcinogenesis in the rat.
Prospective epidemiologic studies have reported an increased risk of rectal cancer following chronic ethanol ingestion. The effect of ethanol on chemically induced colorectal carcinogenesis is controversial depending on the experimental conditions. In the present study the effect of chronic ethanol administration on acetoxymethylmethylnitrosamine-induced rectal cancer and the possible role of acetaldehyde in this process were investigated. Chronic ethanol administration resulted in an earlier occurrence of rectal tumors in this animal model. Because the concomitant administration of cyanamide, a potent acetaldehyde dehydrogenase inhibitor, showed a positive trend toward increased incidences of tumors, acetaldehyde could be involved in the ethanol-associated carcinogenesis. To measure colonic acetaldehyde, 12 chronically ethanol-fed and control rats received an acute dose of ethanol (2.5 g/kg body wt). The mucosal concentration of acetaldehyde was significantly higher in the rectum compared with the cecum (198 +/- 23 vs. 120 +/- 23 nmoles.g colon-1, p less than 0.05), but was not affected by chronic ethanol feeding. Furthermore, 6 germ-free rats had significantly lower acetaldehyde concentrations in the rectum (84 +/- 11 vs. 234 +/- 33 nmoles.g colon-1, p less than 0.01) and in the cecum (59 +/- 13 vs. 121 +/- 33 nmoles.g colon-1, p less than 0.05) compared with 6 conventional animals, and this was paralleled by the number of fecal bacteria in the 2 intestinal segments. In addition, to determine the effect of chronic ethanol feeding on colorectal cell turnover, 30 animals were pair-fed liquid diets. Using the metaphase-arrest technique, alcohol feeding induced rectal (19.1 +/- 2.0 vs. 9.1 +/- 1.8 cells.crypt-1.h-1, p less than 0.01), but not cecal (18.9 +/- 1.3 vs. 22.2 +/- 3.3 cells.crypt-1.h-1, p greater than 0.05) hyperregeneration. This was accompanied by an increase in the crypt proliferative compartment and increased mucosal ornithine decarboxylase activity (63 +/- 18 vs. 22 +/- 6 pmoles.hr-1.mg protein-1, p less than 0.05). The data show that chronic ethanol ingestion accelerates chemically induced rectal carcinogenesis and raise the possibility that acetaldehyde probably generated through bacterial ethanol oxidation may be involved in this process. The secondary hyperregeneration of the mucosa, observed after alcohol feeding, could by itself favour carcinogenesis.
Acetaldehyde↗