Proliferation rate--drug sensitivity relation in clones isolated from heterogeneic tumor cell population.

Four separate cell clones were isolated from parental rat RBB/R cell line. Sensitivity of clones to 3-oxauracil varied, and was dependent on the proliferation rate of particular clone. Colony formation method was more appropriate than 3H-thymidine incorporation for determination of post-treatment surviving cells.

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.

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.

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.

Single versus combination chemotherapy of L1210 leukemia.

Thirteen anticancer agents, ten dual-agent combinations and five three-drug combinations were tested for treatment of L1210 leukemia in dBA/2-J mice. Data obtained form each three-drug regimen were compared with those obtained after administration of each drug alone and each two-drug combination. Cure (greater than 60 days survival) was observed in most of animals treated with VP-16 213 and VM-26. Certain regimens produced 90-100% cure rates (cyclophosphamide plus VP-16 213 or cytosine arabinoside). Inclusion of second or third agent in the treatment schedules produced improvement, deterioration or no effect on median survival time and cure rates, depending on the choice and sequences of evaluated agents. Eighty per cent of mice inoculated even with 10(7) L1210 can be cured by administration of modified schedule of VP-16 213 plus cyclophosphamide.

Combined chemoimmunotherapy of L5178Y lymphoma.

The antitumoral effect of chemotherapy and immunotherapy with BCG and/or irradiated L5178Y cells has been compared to antitumoral effect of chemotherapy alone. Significantly better results were noted in a transplantable L5178Y lymphoma grafted on DBA/2-J mice treated with methotrexate and BCG than in any other group which was not treated with BCG. The effect of irradiated lymphoma cells was similar to effectivity of methotrexate alone.

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.

Inhibition of colony-forming cells from bone marrow of leukemic patients by 3-oxauracil.

An in vitro system of colony forming cell inhibition was used for evaluation of 3-oxauracil (2,3-dihydro-1,3-6H-oxazine-2,6-dione) effectivity against bone marrow cells from 18 leukemic patients. The highest inhibitive activity of the 3-oxauracil was found in cases of acute lymphoblastic leukemia, comparing with bone marrow cells from patients with acute myeloblastic and chronic myeloid leukemia. According to the morphological characteristics of the formed colonies, lymphoblastic leukemia cells could be discerned from the myeloblastic ones in this system.

Effect of "3-oxauracil" (2,3-dihydro-1,3-6H-oxazine-dione) in L1210 leukemia.

2.3-Dihydro-1,3-6H-oxazine-dione ("3-oxauracil") was tested against L1210 leukemia. The compound exhibited significant growth-inhibiting effect against this tumor both in vitro and also in vivo following daily intraperitoneal application.

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).

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.

Mitochondrial adenosine triphosphatase of Zajdela hepatoma. III. Effect of uncouplers on the hydrolysis of intramitochondrial ATP.

Hydrolysis of extramitochondrial ATP by coupled Zajdela hepatoma mitochondria is not stimulated by uncouplers of oxidative phosphorylation. The results of the present study show that the hydrolysis of intramitochondrial ATP in these mitochondria is stimulated by DNP and CCCP. It is proposed that the uncoupler insensitivity of ATPase in coupled Zajdela hepatoma mitochondria with exogenous ATP as a substrate result from an altered functional relationship between ATPase and ADP, ATP translocase.

Stimulation of rat liver regeneration by chloramphenicol.

The administration of chloramphenicol in high doses (300 mg per kg body weight in 6 hour intervals) to partially (70%) hepatectomized rats resulted in a marked stimulation of liver regeneration.

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.

Mitochondrial ATPase of Zajdela hepatoma. V. Mitochondria of Zajdela hepatoma contain membrane sectors of ATPase complex unassociated with F1.

Relative content of membrane sectors of ATPase complex in rat liver and Zajdela hepatoma mitochondria and the ability of mitochondrial membrane of the two sources to bind isolated soluble (F1) ATPase were examined. Approximately equal concentrations of oligomycin were required for 50% inhibition of ATPase activity in submitochondrial particles of rat liver and Zajdela hepatoma indicating practically identical content of membrane sectors of ATPase complex in both types of mitochondria. As detected by the increase in oligomycin-sensitive ATPase activity of submitochondrial particles incubated with isolated F1, the submitochondrial particles of Zajdela hepatoma in contrast to those of rat liver were able to bind specifically considerable amounts of exogenously added F1. The results indicate that mitochondria of Zajdela hepatoma contain membrane sectors of ATPase complex unassociated with F1 but capable of association with this enzyme.

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.

Different inhibitory effect of nitrogen mustard (HN2) and TS-160 (HN3) on choline transport in L 5178Y lymphoblasts.

The inhibitory effect of the alkylating agents nitrogen mustard (HN2) and TS-160 (HN3) on the choline transport in L 5178Y lymphoblasts was studied. The two alkylating agents differed in the degree and in the nature of the inhibition of the process examined. Competitive inhibition of choline transport by HN2 was confirmed, whereas the noncompetitive inhibition by the less efficient inhibitor HN3 has been found. The results achieved indicate that in spite of the structural similarity between HN2 and HN3 only the former is transported into L 5178Y lymphoblasts by the choline transport carrier.

Mitochondrial ATPase of Zajdela hepatoma. II. Mitochondria of Zajdela hepatoma contain less adenosine triphosphatase than mitochondria of rat liver.

The specific activity and the content of ATPase in mitochondria of rat liver and Zajdela hepatoma were compared. The specific activity of ATPase in sonicated mitochondria and in mitochondrial membrane fraction of rat liver was almost two times higher than the specific activity in the corresponding fraction of Zajdela hepatoma. Accordingly, the autovertin binding capacity of rat liver mitochondrial membrane fraction as well as the yield of F1-ATPase from this fraction were about two times higher than those of the mitochondrial membrane fraction of Zajdela hepatoma. The results show that mitochondria of Zajdela Hapatoma possess about half amount of ATPase present in rat liver mitochondria.