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Notes: Abstract The novel cisplatin analogue D-17872 was studied for its anticancer activity using in vivo and in vitro preclinical models. The compound at the sublethal dose of 215 mg/kg (ca. 50% of the approximate LD50) induced no nephrotoxic effect strong enough to increase the blood urea level in rats. It had good in vivo antitumor efficacy against murine P388 (max. ILS: D-17872 132%, cisplatin 55%) and L1210 leukemia (max. ILS: D-17872 43%, cisplatin 38%), L5222 leukemia of the rat (max. ILS: D-17872 163%, cisplatin 163%) and murine B16 melanoma. Activity against P388 leukemia substantially exceeded that of cisplatin. Moreover, the M5076 reticulum cell sarcoma implanted into the subrenal capsule and the DMBA-induced mammary tumor of the rat were inhibited by D-17872 to a greater extent than by cisplatin (min. T/C: D-17872 −3%, cisplatin 11%). Using clonogenic microassays, D-17872 was active in vitro against a variety of human and rodent tumor cell lines, albeit at higher concentrations than cisplatin (IC50 values: D-17872 2.6–12.7 μmol/l, cisplatin 0.13–0.42 μmol/l). Apart from its cytotoxic action it was able to induce in vitro differentiation of the human HL-60 and K562 and of the murine M1-T22 cell lines, while cisplatin induced differentiation only in the HL-60 cell line. Thus D-17872 exhibited a pharmacological and toxicological profile different from that of the parent compound. The results suggest that induction of differentiation contributes to the antineoplastic efficacy of this novel cisplatin derivative.

Notes: Abstract  β-D-Glucosylisophosphoramide mustard (β-DGlc-IPM) is a new, potential chemotherapeutic agent currently under investigation. Its pharmacokinetics in plasma and elimination of the parent drug and its metabolites via urine, bile, and exhaled air were studied in female Sprague-Dawley rats after bolus injection of 315 mg/kg. Typically, the drug’s disposition from plasma follows a linear two-compartment model with half-lives (t 1/2) of 1.8 (t 1/2α) and 32 min (t 1/2β). The rate of clearance is 0.0046 (range 0.0030–0.0071) l min-1 kg-1, and the steady-state volume of distribution (V ss) is 0.18 (0.08–0.042) l/kg (mean±interindividual standard deviation). In human plasma, 28.1±2.6% (mean±SD) of the drug (concentration range 0.5–5 mg/ml) is bound to plasma proteins (predominantly to albumin). Biliary excretion of the parent drug accounts for 2.9±1.7% of the dose; its elimination in the form of 14CO2 via exhaled air is less than 1%. Within 24 h, 63.5±4.9% of the 14C-labeled drug is excreted unchanged in the urine, whereas 17.5±5.1% is excreted in the urine as metabolites. In addition, β-D-Glc-[14C]-IPM was given as a bolus injection to female Sprague-Dawley rats at dose levels of 315 and 56.2 mg/kg. The distribution of radioactivity into tissue was examined qualitatively by whole-body autoradiography (WBA). Parallel experiments were carried out using the high dose of the L-derivative. After dosing with the D-compound, the highest levels of radioactivity were found in the liver, kidneys, thymus, thyroid gland, and central nervous system, including the brain. A similar distribution pattern was observed for the L-compound, except in the brain, which contained negligible levels of radioactivity. The distribution of the D-compound (high dose) was also investigated in male Copenhagen rats bearing a Dunning prostate tumor. The results were similar to those obtained in healthy Sprague-Dawley rats. Additionally, radioactivity was found in the tumor at 1 h after dosing with the drug and remained there even after 24 h. The effects of β-D-Glc-IPM on the incorporation of [methyl-3H]-thymidine into the DNA of the liver, kidneys, thymus, spleen, esophagus, and bone marrow of the rat were examined following tissue excision and liquid scintillation counting at 2, 8, and 24 h after administration of the drug. β-D-Glc-IPM showed no effect on the incorporation of [methyl-3H]-thymidine in the liver and an insignificant reduction in kidney DNA (maximal reduction: −27.3%). However, after 8 h there was a marked reduction in the incorporation rate in the thymus (−83.7%), spleen (−74.6%), and esophagus (−87.2%), with a tendency toward recovery within 24 h. In bone marrow cells a reduction of −75.5% (8 h) and −73.3% (24 h) was observed.

Notes: Summary By cloning in vitro we have obtained two sublines of the L5222 rat leukemia, one with high (L5222-S) and the other with low (L5222-R) in vivo sensitivities to non-toxic doses of mafosfamide, a stabilized derivative of 4-hydroxy-cyclophosphamide. This sensitivity in vivo was not related to the cytotoxic activity of the drug in vitro. Treatment of rats bearing the L5222-S and of mice transplanted with the MOPC-315 plasmocytoma with low doses of mafosfamide or cyclophosphamide resulted in a high percentage of surviving animals, which were resistant to a subsequent tumor challenge. Viable leukemic cells were needed to establish antitumor immunity, since it was not possible to induce resistance by injection of mitomycin-C-treated, non-viable L5222 cells. The adoptive transfer of spleen cells from animals immune against the L5222-S and the MOPC-315 resulted in resistance of the syngeneic recipients against a rechallenge with tumor cells, provided that the animals were treated with an immunosuppressive dose (100 mg/kg) of cyclophosphamide prior to the spleen cell implantation. In nude mice treatment of the L5222 with low doses of mafosfamide also resulted in surviving animals, however resistance to a second tumor challenge occurred only sporadically. The data presented confirm that therapy with cyclophosphamide or mafosfamide enhances host antitumor immunity but, contrary to previous reports, it could be demonstrated that successful tumor rejection was independent of T cells.

Notes: Summary Anticancer agents so far available and their mechanisms of action suffer from the problem of their relatively low selectivity. Their insufficient clinical efficacy against the common, slowly growing solid tumors of the lung, gastrointestinal system, kidneys, urinary bladder, and brain remains disappointing. Recently the possibility has been discussed that the limited clinical activity of current anticancer drugs could result from the screening models and methods used in their selection. The initial approach to drug discovery used by the National Cancer Institute, Bethesda, USA (NCI), the greatest oncological research unit in the world, has been empirical large-scale screening in transplantable rodent tumor models. In the past, these preclinical models have been changed periodically in line with retrospecitve analyses of preclinical predictivity for clinical efficacy. Recently, as a new strategy, a “disease-orientated” concept has been developed to screen agents against particular types of human cancer on the basis of the human tumor colony-forming assay in vitro. Each compound should now be tested directly against a spectrum of human tumor lines without passing through a rodent prescreen. Additional assays in vivo may be performed later on. This new screening concept seems to be suitable for identifying the cytotoxicity of new chemical structures and for an evaluation of sensitivity or resistance of the different tumor types. The contrasting concept of “rational drug design” is exemplified by the development of the oxazaphosphorinecytostatics. The basis of this concept was the application of the transport form/active form principle to the antiproliferative nitrogen mustard. Cyclophosphamide, the first representative of this group, had already largely reached the given objective. Generalizing conclusions from the different concepts are as follows. 1. Methods and perceptions of general pharmacotherapy must be the principal basis for the development of antitumor compounds. 2. Progress and essential new developments in cancer chemotherapy are based on experiments in intact animals. 3. An important feature of rational drug design is the stepwise or sequential procedure: the design of new drugs is based on the screening results of former drugs to achieve an optimal progress. 4. For the analysis of the activity of alkylating agents on rats and mice, the panel of test tumors for screening and pharmacological evaluation must be selected according to a different degree of chemoresistance or chemosensitivity, respectively. It should be aimed at a complete dose/activity curve with cyclophosphamide as standard at least for the most sensitive tumors of that panel. 5. The therapeutic index, i.e. LD5/CD95, has proved to be a valuable tool for chemotherapeutic usefulness, as has the danger coefficient for the quantification of organotoxic side-effects. These values provide a measure of the therapeutic range and, consequently, of the selectivity of the antitumor activity. Results from a given tumor have proved to be predictive for other tumors and turned out to be relevant also for clinical trials. 6. The different sensitivities of experimental tumors against alkylating agents is not a fundamental property but a quantitative feature. With sub- or even supra-lethal doses it is possible to overcome vitality and transplantability of even the very most resistant tumors. A new product with impressively increased selectivity is consequently expected to achieve remissions in more resistant tumors also. This evaluation system also remains applicable and useful in the context of a “disease-oriented” concept. 7. A profound knowledge of the mechanism of action of cytostatic agents and a deep insight into the metabolic patterns in the host and in tumor cells are the essential basis for a rational augmentation of cytostatic activity. 8. Promising leads, identified initially by empirical large-scale screening programs, mostly need forther optimization through the rational approach. Thus there is most often an essential and intimate interplay between the rational and large-scale screening strategies.

Notes: Abstract An experimental rat model for the study of venous pain induced by 4-hydroxy-cyclophosphamide (4-OH-CP) derivatives was developed and validated. Using various metabolites and chemical variants of 4-OH-CP it was found that pain induction was independent from the compound's alkylating activity but possibly related to the spontaneous generation of minute amounts of acrolein from the 4-OH-CP molecule. Accordingly, the pain could be prevented by the addition of thiol compounds such as mesna or N-acetyl-cysteine.

Notes: Summary The LD 50 of Z 7557 in mice was between 500 and 625 mg/kg after i.v. and around 2310 mg/kg after oral administration. The corresponding LD 10 was around 435 mg/kg (i.v.) or 1100–1250 mg/kg (p.o.), respectively. The LD 50 values in rats were in the range of 250–310 mg/kg after i.v. administration and around 1000–1250 mg/kg if given orally. With repeated daily i.v. injections only 70% of the daily dose contributed to lethality. A second administration of Z 7557 to rats after reversal of all toxic signs from the first administration induced the same symptoms and degree of toxicity as the initial injection. Reversible myelosuppression was the predominant feature of toxicity in mice and rats, but at equimolar doses this myelotoxicity was less than half that of cyclophosphamide (CP). First signs of immunosuppression were seen only at 100 mg/kg i.v. No severe urotoxicity was observed in rats with single i.v. doses up to 192 mg/kg. This might be due to the fast and complete renal excretion of the thiol moiety of Z 7557, whereas the activated oxazaphosphorine component occurred in the urine only to a much smaller amount, as could be shown by a pilot pharmacokinetic study. In conclusion, Z 7557 appeared to have an overall tolerance in rats and mice similar to cyclophosphamide but was clearly less toxic with respect to the bone marrow, the immune system and the urinary tract.