Dominant expression of multidrug resistance in intraspecific murine lymphoma hybrid cells.

Cultured P388/VCR mouse lymphoma cells resistant to vincristine (VCR) and to 5-bromodeoxyuridine (BUdR) and deficient in thymidine kinase (TK-) were fused with P388/DAG cells resistant to 1.2:5,6-dianhydrogalactitol (DAG), an anticancer alkylating agent, and to 6-thioguanine (6-TG) and deficient in hypoxanthine phosphoribosyl-transferase (HPRT-). The hybrid cells expressed multidrug resistance (MDR), i.e., resistance to VCR and cross-resistance to Adriamycin (ADM) and actinomycin D (Act. D), in a dominant manner. The presence of glycoprotein p170, the MDR gene product, was detected in the hybrid cells. Resistance to DAG was also expressed dominantly, whereas cross-resistance to dibromodulcitol (DBD), a chemically related anticancer drug, was slight.

Nucleoside transport and metabolism in lymphocytes, polymorphonuclear cells and cerebral synaptosomes.

The salvage of nucleosides dominates over de novo biosynthesis in lymphocytes, polymorphonuclear cells (PMN) and in central neutral nervous system (CNS) in higher organisms. Earlier works in our laboratory have shown that the salvage of deoxycytidine (dCyd) did not correlate with DNA synthesis. The uptake and metabolism of dCyd was higher in undifferentiated germinal center lymphocytes and in follicles comparing to more differentiated cells. Recently we have compared the transport of thymidine (dThd), dCyd, uridine (Urd) and adenosine (Ado) in the three cell systems in which the salvage of nucleosides is dominating. It was found that dCyd was transported 30 times more effectively into lymphocytes than into PMN and synaptosomes, while Urd was transported about the same rate into the two cells and into synaptosomes. All transport processes could be inhibited by dipyridamole, NBRPR, papaverine and dilazep. The dCyd and dThd was phosphorylated even at 0 degrees C up to TTP and dCTP without incorporation into DNA and into liponucleotides. Our results show that the processes of transport-phosphorylation, as well as the processes of DNA-CDP-phospholipid synthesis are tightly coupled to each other in intact cells and organelles.

Changes in the rates of lipogenesis and hepatic triglyceride secretion in mice bearing Ehrlich ascites carcinoma.

The growth of Ehrlich ascites tumors (EAT) in mice induces hypertriglyceridemia and depletion of lipid stores. H-Riop: Swiss mice in early and late stages of tumor growth were examined to investigate whether an increase in liver synthesis of fatty acids (FA) and/or an increase in the liver triglyceride (TG) secretion rates (TGSR) would contribute to endogenous cancer-induced hypertriglyceridemia. Using 3H2O as tracer, FA synthesis decreased in the liver of tumorous animals. Hepatic TGSR also decreased during the development of hypertriglyceridemia. On the basis of these results, hypertriglyceridemia is probably not due to hyperproduction of lipids by the liver. In the late stage of tumor growth a considerable drop of FA synthesis also ensued in the adipose tissues, which probably participated in the loss of carcass lipids. At the early stage of tumor growth FA synthesis in the EAT cells was substantial in relation to the low lipid content of these cells, but in the late period FA synthesis slowed down, indicating that the triglyceride-rich "older" tumor cells obtained a large part of their lipids performed by the host.

Absence of cross-resistance between two alkylating agents: BCNU vs bifunctional galactitol.

Dianhydrogalactitol (DAG) increased the life span of both BCNU-sensitive and -resistant L1210 tumor-bearing mice. However, the BCNU-resistant strain showed slightly lower sensitivity against DAG, which could be overcome by an increase in drug dose of ca. 20%. The somewhat lower sensitivity was proportional to a slightly reduced DNA cross-linking formation induced by DAG in BCNU-resistant cells. The amount of DNA cross-links was determined by measurement of the 1,6-di(guaninyl)-galactitol content of DNA. The slight reduction in cross-links is not attributable to DNA repair but rather to other factors that seem to prevent the formation of DNA-drug adducts. The absence of cross-resistance is explained by different kinds of DNA damage caused by the two alkylating agents and the presumably different defense mechanisms developed by cells against these lesions.

Dominance of resistance to the alkylating agent 1,2:5,6-dianhydrogalactitol in P388 mouse lymphoma hybrid cells.

Cultured P388/S mouse lymphoma cells resistant to 5-bromodeoxyuridine (BUdR) and deficient in thymidine kinase (TK-) were fused with P388/DAG cells resistant to 1,2:5,6-dianhydrogalactitol (DAG), an anticancer alkylating agent, and to 6-thioguanine (6-TG) and deficient in hypoxanthine phosphoribosyl-transferase (HPRT-). Sensitivity to DAG in the hybrid line was very close to that in the P388/DAG line, which means that resistance to DAG was inherited in a quasi-dominant manner. Hybrid cells showed cross-resistance, similar to that of the DAG-resistant line, to two other hexitols, dibromodulcitol (DBD) and disuccinyldianhydrogalactitol (DisuDAG).

Thymidylate synthetase and thymidine kinase activities and methotrexate cytotoxicity during growth of L1210 and Ehrlich ascites tumor.

The specific activities of TS and TK determined in the cytosols of Ehrlich and L1210 ascites tumor cells changed significantly during their logarithmic growth. In both tumors, the highest activity of TK was in early log phase while that of TS occurred in late logarithmic growth. The activity curves of TS were practically the same and those of TK were similar in the two tumors; however, the absolute values of the latter were different. TK/TS activity ratios in the early and late log growth of Ehrlich tumor were 20 and 2, in those of L1210 tumor 4 and 0.15 respectively. MTX cytotoxicity was considerable throughout the logarithmic growth of L1210 tumor, significantly higher values were observed however when TS activity increased. The variable degree of MTX cytotoxicity indicated an intracellular manifestation of the changes in TS activity determined in vitro. The different patterns for the specific activities of TS and TK, as well as, the parallel changes of TS activity and MTX cytotoxicity during tumor growth may be useful information for the antimetabolite research carried out with experimental tumors.

The effect of N-formylleurosine on DNA synthesis of Ehrlich ascites tumor.

Some of the vinca alkaloids are well known and widely used in clinical practice, in spite of their numerous side-effects. For the elimination of untoward side-effects semi-synthetic alkaloids have been produced. This work reports on N-formylleurosine (N-F-Leu), one of these agents. Its antitumour activity is, in many respects, similar to that of other, presently used vinca alkaloids; it causes metaphase block, the appearance of multinucleated cells and polyploidization. In addition to these effects, N-F-Leu induces significant new phenomena, namely, changes in the activity of thymidine kinase, a key enzyme of DNA synthesis as well as in thymidine incorporation, and reduces the proportion of cells in S phase in a relatively short period of time after treatment: 24-29 hr. Based on these observations, the reevaluation of the mechanism of action of vinca alkaloids has become possible.

Incorporation of [7-3H]cholesterol into chromatin fractions of Ehrlich ascites tumour cells.

Incorporation in vivo of 3H-cholesterol into chromatin and its distribution among chromatin subfractions (DNA, histone, nonhistone proteins) of Ehrlich ascites tumour cells were studied. The majority of labeling appeared in the nonhistone fraction and only a slight incorporation or tracer amounts of 3H-labeling was found in the histone and DNA fractions. The 3H-labeling from 3H-cholesterol appeared predominantly in the non esterified cholesterol fractions. Our suggestion that neutral lipids--besides phospholipids--may also contribute to the regulatory functions of nonhistone chromosomal proteins needs further investigations.

Drug resistance studies on intraspecific hybridomas.

P388 mouse leukemia lines, one sensitive (P388/S) and the other resistance (P388/R) to vincristine (VCR), cultured in vitro, were hybridized with polyethylene glycol (PEG). A thymidine kinase-deficient mutant (TK-) was isolated from the sensitive line, and a hypoxanthine-guanine phosphoribosyltransferase-deficient mutant (HPRT-) from the resistant line. The hybrid line grows slower than the mutants. The modal chromosome numbers are: TK- = 38, HPRT- = 40, hybrid = 69 (72). The TK- cells contain a large metacentric marker which is missing from the HPRT- cells. Hybrid cells are as resistant to VCR as the P388/R and HPRT- cells.