[Chemoprevention of cancer]. / Chemoprevencia rakoviny.

Chemoprevention, as one of the newest components of the fight against cancer, deals with various chemical compounds which can prevent the occurrence of malignant tumours. New experimental results achieved during the recent decades in the field of understanding of the process of carcinogenesis, have enabled a more rational approach to the prevention of normal cell malignization. It has been shown, that a number of investigated compounds are able to interact selectively with the particular phase (initiation, promotion, progression) of the multistep process of carcinogenesis. Activity of chemopreventive compounds can be evaluated by systematic analysis of various biomarkers in patients who are in high risk. The biomarkers include genetic markers (mutations, gene amplification), cellular markers (rate of proliferation), histologic markers (leukoplakia, metaplasia), or biochemical markers (blood level of various enzymes, or tumour antigens). The number of extensive clinical trials has recently increased and some positive results have been reported in the testing of various compounds preventing the appearance of some malignant tumours (breast, colon, prostate, etc.). The search for new, effective chemopreventive compounds might provide a valuable new tool aimed at the decrease in both cancer morbidity and mortality. (Ref. 37.)

The disposition of new arabinosylcytosine derivative-- 5'-chloro-5'-deoxy-arabinosylcytosine--in rats.

1. Pharmacokinetic properties of a new derivative of the widely used and very potent antileukemic agent arabinosylcytosine (araC)--5'chloro-5'-deoxy-arabinosylcytosine (5'-Cl-araC)--were investigated after intraperitoneal (i.p.) and oral routes of administration in rats and compared with the equimolar dose of araC administered orally. 2. It was found that substitution of the hydroxyl group at position 5' resulted in a change of pharmacokinetic parameters. 3. There is a large difference in average serum concentrations of 5'-Cl-araC administered by the i.p. and oral routes; the average serum concentration obtained after i.p. injection being several times higher in comparison to those after oral administration. 4. However, the latter are, at the same time, lower than the average serum concentrations of araC administered by the same route in an equimolar dose. 5. On the other hand, the apparent volume of distribution is much larger, and the area under the curve of serum concentration of 5'-Cl-araC is smaller, after oral as compared to the i.p. route of administration indicating more extensive tissue distribution together with higher tissue binding of 5'-Cl-araC when compared to the parental drug araC.

Effect of arabinosylcytosine derivative cyclocytidine on hepatal functions in rats and on Zajdela hepatoma.

1. The effectivity of the arabinosylcytosine (araC) derivative, cyclocytidine (cC), against Zajdela hepatoma was evaluated. It was established that the cC effect was dose-dependent. 2. Zajdela hepatoma-bearing rats, given a single dose of 500 mg of cC per kg of body weight, showed an increased life span of 48 and 39%. 3. cC significantly decreased the number of Zajdela hepatoma cells in ascitic fluid and affected the cytochrome content in hepatal mitochondria. 4. The overall cC effect on the hepatal function of normal and hepatectomized liver was marginal, thus making this araC derivative an interesting candidate for further evaluation of its effectivity against non-hematological tumors.

Biotransformation and in vitro activity of an arabinosylcytosine 5'-chloro-5'-deoxy analog.

5'-Chloro-5'-deoxyarabinosylcytosine (5'-chloro-araC), a lipophilic and cytidine-deaminase resistant analog of the cytotoxic agent arabinosylcytosine (araC) was evaluated in terms of bioactivation, transformation and its cytotoxic activity in vitro. 5'-Chloro-araC interferes with DNA synthesis (IC50 = 2.8 mumol/l) and inhibits the growth of L1210 cells in suspension culture (IC50 = 1.05 mumol/l) and in the soft agar assay (IC50 = 0.65 mumol/l). Being phosphorylated to the triphosphate of araC-araCTP (5'-triphosphate of araC), 5'-chloro-araC has the same mechanism of action as arabinosylcytosine. In alkaline solutions 5'-chloro-araC is transformed to another (cytostatically inactive) araC analog--2',5'-anhydroarabinosylcytosine--but at physiological pH and temperature conditions, it has sufficient stability to be phosphorylated and thus activated. A lower rate of araCTP formation from 5'-chloro-araC explains the somewhat lower cytotoxic effect of this compound against various established cell lines in vitro compared to araC. Lipophilicity that would allow an oral drug formulation and certain other superior physico-chemical and biochemical characteristics of 5'-chloro-araC make this compound an interesting candidate for further investigations.

Treatment of leukemia L1210 and P388 by arabinosylcytosine-polysaccharide conjugates.

Conjugates of 1-beta-D-arabinofuranosylcytosine (araC) with two polysaccharides such as polygalacturonic acid (PGA) and carboxymethylated yeast beta-D-glucan (CMG) were tested for their antileukemic activity in vitro on a L1210 cell line in suspension culture, in soft agar assay and in vivo on L1210, L1210/araC- and P388-leukemia-bearing mice. Both conjugates showed high activity in vitro in soft agar assay, compared with araC. Single administration of PGA-araC or CMG-araC increased the survival time 1.5 x or 1.7 x, respectively, compared with araC in vivo in L1210-leukemia-bearing mice. The conjugates were not active against araC-resistant leukemia line L1210/araC. The marked effect of both PGA-araC and CMG-araC against leukemia L1210 and P388 is probably due to the prolonged release of free araC from conjugates caused by hydrolysis.

Cross resistance studies with L1210 leukemia subline single and double resistant to cisplatin and iproplatin (CHIP).

Resistance to cisplatin (DDP) and/or iproplatin (CHIP) was induced in vitro in murine L1210 leukemia cells. Double-resistant sublines with combined resistance to both drugs were also developed. Cross resistance investigations with DDP, CHIP, oxoplatinum (OXO), carboplatin (CBDCA) and its quadrivalent derivative OXOCBDCA were performed in these resistant sublines. Lack of cross resistance between DDP and CHIP was found. A higher resistance to CHIP in the double-resistant sublines was observed. A multistep process in the development of resistance to this compound is supposed. The importance of the aminoligand and the role of different pharmacokinetics in the cross resistance are discussed.

Comparison of some biochemical parameters of arabinosylcytosine and cyclocytidine in L1210 murine leukemia cells.

The basic biochemical characteristics of cyclocytidine hydrochloride (cC.HCl) and arabinosylcytosine (araC) were compared. It was demonstrated that despite different lipophilicity and different pK (4.15 for araC and 6.60 for cC.HCl), the mechanism of inhibition of DNA synthesis by both compounds is the same (ID50 for araC was 0.048 mumol/l and for cC. HCl 0.23 mumol/l). The compounds had a different mechanism of inhibition of RNA synthesis (ID50 for araC was 2.69 mmol/l and for cC.HCl 1.08 mmol/l) and showed a marginal effect on protein synthesis. Hydrolysis of the 0(2),2'-anhydro bond in cC.HCl and formation of araC in vivo was characterized by a Km = 280 mumol/l using HPLC. Deamination of araC in vivo was studied in healthy mice (Km = 247 mumol/l), 8.6% of arabinosyluracil 15 minutes after araC administration) and in mice with sensitive and araC resistant leukemia L1210 (15.5% and 8.5% of arabinosyluracil 15 minutes after araC administration, respectively). On the basis of different physico-chemical properties of cC.HCl and different mechanisms of inhibition of RNA synthesis it can be assumed that cC.HCl, when therapeutically used, may have its own mechanism of biological effect(s) and that its application may be therapeutically advantageous in some aspects as compared to araC.

Antitumor activity and cross-resistance studies with Pt-ascorbato complexes.

Two ascorbatoplatinum complexes, cis-diammineascorbatoplatinum(II) (AMA) and cis-bis(ascorbato)-trans-diaminocyclohexaneplatinum(II) (CHA), were tested for antitumor activity in vivo on P388 leukemia and in vitro in suspension culture and soft agar assay. Sensitive line of L1210 and sublines with resistance induced against cis-diamminedichloroplatinum(II) (DDP) and two derivatives of trans-1,2-diaminocyclohexane (DACH) were used for the in vitro tests. DNA synthesis inhibition in both sensitive and resistant cells was tested. The results are compared with DDP and DACH-Pt(II)-4-carboxyphtalate (TMA). Both tested complexes proved their antitumor activity in our experimental systems. The CHA complex was more effective than AMA and its effectiveness is comparable with that of DDP and TMA. Cross-resistance was found between DDP and AMA as well as TMA and CHA. There was no cross-resistance between DDP versus CHA, and TMA versus AMA.

DNA synthesis, protein synthesis and platinum content following drug administration in cis-diamminedichloroplatinum(II) -sensitive and -resistant L1210 cells transferred from in vitro to in vivo conditions.

L1210 cells with in vitro induced drug resistance against cis-diamminedichloroplatinum(II) (cis-Pt(II] were inoculated in mice and several times transplanted. Then the effect of cis-Pt(II) on drug-sensitive and drug-resistant L1210 cells in mice was investigated. While the DNA and protein synthesis in drug-sensitive cells after in vivo cis-Pt(II) treatment was inhibited by 50%, that of drug-resistant cells remained virtually unaffected. The content of platinum in drug-sensitive cells was approximately three times higher in comparison with drug-resistant L1210 cells.

Antitumor activity of four polymer bound trans-1,2-diaminocyclohexaneplatinum(II)-4-carboxyphtalate complexes tested in different model systems.

Four polymer bound Pt-complexes have been tested in in vivo and in in vitro systems. No substantial difference in effectivity against P388 leukemia in vivo was found when free trans-1,2-diaminocyclohexaneplatinum(II)-4-carboxyphtalate (TMA) was compared with the polymer bound complexes. The compound with the highest ID50 value in soft agar assay exhibited low effectivity in in vivo testing. Polymer bound Pt-complexes with faster release of the active molecule exhibited in in vivo and in soft agar assay slightly lower activity, when compared with suspension culture test system. Cross resistance of polymer bound complexes was investigated on three cell lines with induced drug resistance against different Pt-complexes. Cross resistance was found between TMA (free and polymer bound) and trans-1,2-diaminocyclohexaneplatinum(II)citrate (PEX) as well as trans-1,2-diaminocyclohexaneplatinum(II)glucarate (PTU) but there was no cross resistance between TMA and cis-diamminedichloroplatinum(II) (cis-DDP).

The value of the short term predictive assay in the inductive treatment of patients with acute nonlymphoblastic leukemia.

Predictive capacity and clinical usefulness of the short term predictive assay (STPA) in the inductive treatment of patients with acute nonlymphoblastic leukemia (ANLL) was studied. Inductive treatment consisted of daunorubicin, arabinoside C and 6-thioguanine (TAD regimen). Leukemic cells of 20 previously untreated patients with ANLL were incubated in vitro with two doses of daunorubicin (1 microgram/ml and 10 micrograms/ml), arabinoside C (10 micrograms/ml and 100 micrograms/ml) and 6-thioguanine (10 micrograms/ml and 100 micrograms/ml). The 3H-thymidine as well as 3H-uridine uptake was measured in the treated and untreated cells. The highest predictive presence of the in vivo drug-sensitive disease was adequately reflected by the level of 3H-uridine incorporation suppression 30% of control value in the case of daunorubicin (concentration: 10 micrograms/ml) and 80% in the case of 6-thioguanine (concentration: 100 micrograms/ml). In the case of arabinoside C (concentration: 10 micrograms/ml) the limit of 3H-thymidine uptake depression was 20% of control value. It was rather difficult to define the indicative degree of precursors incorporation inhibition for prediction of the drug-resistant disease, because of low number of patients primary resistant to TAD regimen. No correlation was found between the degree of the pre-treatment DNA synthesis rates and the precursors uptake inhibition by the tested drugs.

Comparison of the effectivity of two diaminocyclohexane Pt-complexes.

A group of Pt-complexes was tested in various in vitro and in vivo systems. Murine P388 leukemia for the in vivo testing and three sublines of L1210 leukemia for the in vitro testing of two 1,2-diaminocyclohexane (DACH) derivatives of platinum were used. The effectivity of DACH-Pt(II) citrate and DACH-Pt(II) isocitrate with different ratio of trans- and/or cis-DACH was compared on the sensitive strain of L1210 and two resistant sublines: L1210 resistant to cis-diammine-dichloroplatinum(II) (cis-DDP) and L1210 resistant to trans-DACH-Pt(II) citrate. No cross-resistance was found between the DACH derivatives and cis-DDP. Slightly higher activity of citrate and trans-DACH in comparison with isocitrate and cis-DACH was found both in suspension culture and in vivo testing.

In vivo and in vitro effectivity of some platinum complexes.

The following platinum complexes have been tested: cis-DDP--cis-diamminedichloroplatinum(II), Platinex--1,2-diaminocyclohexaneplatinum(II)citrate, Platuran--1,2-diaminocyclohexaneplatinum(II)-glucarate , TMA--1,2-diaminocyclohexaneplatinum(II)-4-carboxyphthalate,o xo-PT--cis-diamminedichloro-trans-dihydroxyplatinum(IV), CHIP--cis-dichloro-bis-(isopropylamine)-trans-dihydroxyplatinum(IV ), CBDCA--cis-diammine-cyclobutane-1,1-dicarboxylatoplatinum(II ). The activity of all tested complexes againt L1210 cells was higher in soft agar colony assay when compared with suspension culture of the same target cells. Using various doses and schedules oxo-Pt, CBDCA and cis-DDP exhibited the highest in vivo activity against P388 leukemia.

Drug resistance induction and cross-resistance studies with Pt-complexes.

In vitro drug resistance was induced against cis-diamminedichloroplatinum(II) (cis-DDP), 1,2-diaminocyclohexaneplatinum-(II)citrate (PEX) and 1,2-diaminocyclohexaneplatinum(II)glucarate (PTU) in L1210 leukemia cell line. Using the resistant sublines cross-resistance was found between cis-DDP and cis-diamminecyclobutane-1,1-dicarboxylatoplatinum(II) (CBDCA) and between the three 1,2-diaminocyclohexane (DACH) derivatives tested. No (or low degree) cross-resistance was found between cis-DDP and DACH derivatives.

Effect of combined chemotherapy tested in partially hepatectomized rats bearing Zajdela hepatoma.

Ascitic Zajdela hepatoma growing in partially hepatectomized rats was used for testing cytostatics in single and two-drugs combination chemotherapy. At the optimal dosage the highest selective activity against tumor cells (hepatoma) with low inhibition of normal cells (regenerating liver cells) was seen in the combination cis-platinum + methotrexate. Synergistic effect of this combination was found when suboptimal dose of MTX was combined with low doses of cis-Pt. Dose-dependent DNA synthesis inhibition following i.p. administration of cis-Pt was documented by 3H-thymidine incorporation. Although the content of platinum expressed per DNA amount was four times higher in regenerating hepatocytes when compared with hepatoma cells, the growth inhibiting effect of cis-Pt was selectively expressed against the hepatoma cells.