Harumi Iida

Enhancement of vincristine- and adriamycin-induced cytotoxicity by verapamil in P388 leukemia and its sublines resistant to vincristine and adriamycin.

A calcium antagonist, verapamil, enhanced the cellular uptake and cytotoxicity of vincristine (VCR) in adriamycin-resistant P388 leukemia (P388/ADM) cells and also enhanced the cellular uptake and cytotoxicity of adriamycin (ADM) in vincristine-resistant P388 leukemia (P388/VCR) and P388/ADM cells. The enhancement of cytotoxicity and cellular uptake of VCR in P388 and P388/VCR cells has been reported previously. [1]. VCR and ADM resistance was circumvented by verapamil. A common transport mechanism for VCR and ADM, which is responsive to verapamil, seems to exist in VCR- and ADM-resistant cells. However, the enhancement of ADM cytotoxicity and cellular uptake by verapamil was not evident in P388 cells.

Tumor-induced platelet aggregation and growth promoting factors as determinants for successful tumor metastasis

Two clones isolated from a metastatic variant of mouse colon adenocarcinoma 26; the high metastatic NL-17 and the low metastatic NL-44, induced similar platelet aggregation in vitro. Heterotypic aggregates of tumor cells and platelets were injected i.v. into mice. The lung colonization potential of the NL-17 was dependent on the extent of tumor cell platelet aggregation. This clearly indicates that the interaction of tumor cells with platelets can lead to enhanced tumor metastasis possibly through more efficient intravascular arrest of the heterotypic aggregates. The interaction of tumor cells with platelets could thus be an important determinant for successful metastasis. NL-44, however, did not form pulmonary metastasis even after the tumor cells formed heterotypic aggregates with platelets, suggesting that tumor metastasis, is dependent on the intrinsic nature of tumor cells. Lung extract enhanced the growth of NL-17 more effectively than that of NL-44. These results suggest that, in addition to the interaction of metastatic cells and platelets, host factors including growth-promoting factors might also have an important role in tumor metastasis.

Potentiation of antitumor agents by calcium channel blockers with special reference to cross-resistance patterns

SummaryThe calcium channel blockers verapamil diltiazem, nicardipine, and niludipine potentiated the antitumor activities of mitotic poison antitumor agents, such as vincristine, vinblastine, vindesine, VP16-213, and taxol in P388 leukemia cells resistant to vincristine. The potentiating effect was generally dependent on the extent of cross-resistance seen in the cell line for these drugs. Calcium channel blockers also potentiate the antitumor activities of several DNA-interacting drugs, such as adriamycin, THP-adriamycin, daunomycin, aclacinomycin A, mitomycin C, actinomycin D, mitoxantrone, and nogalamycin derivatives in P388 leukemia resistant to adriamycin. Greater potentiation was observed for those antitumor agents to which the ADM-resistant cell line had become markedly cross-resistant, with the exception of the nogalamycin derivatives. Only a two-fold enhancement was observed for mitomycin C and aclacinomycin, as the cell line was only weakly cross-resistant to these agents. These results suggest the potential for therapeutic gain through the use of calcium channel blockers in combination with classic chemotherapeutic agents.

Inhibition of spontaneous and experimental tumor metastasis by the calcium antagonist verapamil

SummaryVerapamil, a calcium antagonist, inhibited both experimental (IV inoculation of tumor cells) and spontaneous metastasis (SC inoculation) of the highly metastatic B16 melanoma and colon adenocarcinoma 26 cell lines. Verapamil treatment resulted in a maximum 80% inhibition of metastases, the degree of inhibition varying among the different metastatic systems. Verapamil inhibited platelet aggregation induced by these tumor cell lines, the patterns of inhibition being different for B16 melanoma and colon adenocarcinoma. The inhibition of platelet aggregation induced by tumor cells is proposed as a mechanism by which the calcium antagonist exerts its antime-tastatic effect. These results, together with our previous findings that calcium antagonists can increase the cytotoxicity of drugs in tumor cells with induced or inherent drug resistance by inhibiting outward transport of the drug, indicate that calcium antagonists have potential as a new class of adjuvant agents in the field of cancer chemotherapy.

Growth stimulating activity of lung extract on lung-colonizing colon 26 clones and its partial characterization.

The effects of lung tissue extract on the cell growth of eight colon 26 tumor clones, four highly and four poorly lung-colonizing clones, were examinedin vitro. Addition of lung extract to serum-free medium stimulated the growth of all four of the highly lung-colonizing clones and one of the poorly lung-colonizing clones, but it had minimal effect on the other three poorly lung-colonizing clones. These results indicate that the lung extract contains a growth stimulating activity; it selectively stimulated some of the colon 26 clones including highly lung-colonizing ones. The growth stimulating activity was not dialyzable, was partially destroyed by heating at 56‡C or 80‡C for 30 min and completely destroyed by trypsin. These results suggest that the activity resides in a protein. Gel filtration chromatography of lung extract on Sephacryl S-200 revealed that the active component was eluted in a molecular weight range of 90000–120000.

Effects of cytochalasins and colchicine on the accumulation and retention of daunomycin and vincristine in drug resistant tumor cells

Cytochalasin B and D enhanced vincristine (VCR) and daunomycin (DAU) accumulation in tumor cells, especially in VCR- and DAU-resistant cell lines. The effect of cytochalasin B, and to a lesser extent cytochalasin D, was almost equivalent to that observed for verapamil, a calcium channel blocker which has been reported to enhance drug accumulation in tumor cells. Cytochalasin B was most effective in VCR- and DAU-sensitive cells; however, the effect in resistant cells was less than that observed for verapamil, suggesting a different mode of action between these drugs in sensitive and resistant cells. Enhanced accumulation of VCR and DAU by cytochalasins was mediated by the inhibition of outward transport of VCR and DAU from tumor cells. Colchicine had no effect on VCR and DAU accumulation. Cytochalasins, especially cytochalasin D is a specific inhibitor of microfilament assembly in cells. These results indicate that the cellular microfilament system plays a prominent role in drug transport of tumor cells, and that an intact microtubular system is less involved.

Prevention of vinblastine-induced cytotoxicity by ruthenium red

Abstract Ruthenium red, an inorganic dye that reacts selectively with mucopolysaccharides of the plasma membranes, prevents vinblastine-induced cytotoxicity toward cultured KB cells. At 6.7ng vinblastine/ml of the culture medium, the growth of KB cells was inhibited completely. When ruthenium red was present in the culture, however, the cytotoxicity induced by vinblastine was completely prevented at concentrations of vinblastine up to 67 ng/ml. Ruthenium red (6 μg/ml) decreased the intracellular amount of vinblastine, whereas in the absence of ruthenium red the level of cellular vinblastine was maintained for 6 hr. This rapid decrease of vinblastine can be explained by an inhibition of uptake of vinblastine through the plasma membrane, and not by an enhanced release of the drug from the cells. Ruthenium red specifically inhibited the Ca2+-ATPase activity of the plasma membrane of KB cells. The possible involvement, of this inhibition in the preventive effect of ruthenium red on vinblastine-induced cytotoxicity is discussed.

Inhibition of murine colon adenocarcinomas and lewis lung carcinoma by 1-hexylcarbamoyl-5-fluorouracil.

Summary1-Hexylcarbamoyl-5-fluorouracil (HCFU) and its parent compount 5-fluorouracil (5-FU) were tested PO for antitumor activity against mouse colon adenocarcinoma 26 (colon 26), colon adenocarcinoma 38 (colon 38), and Lewis lung carcinoma. The drugs were given orally at 2–4 days intervals for a total of ten doses. 5-FU was moderately active against colons 26 and 38 but not against Lewis lung carcinoma. In this treatment regiment the most impressive antitumor activity was obtained with HCFU against colons 26 and 38, especially colon 38 tumor. At 300 mg HCFU/kg, one out of seven mice inoculated with colon 26 and five out of ten mice inoculated with colon 38 became tumor-free. HCFU, however, was marginally effective in the prolongation of survival time of mice bearing Lewis lung carcinoma.5-FU is approximately twice as active as HCFU against cultured cell lines from colon 26, colon 38, and Lewis lung carcinoma. Lewis lung carcinoma cells were most sensitive against HCFU, which is in contrast to the results obtained in the in vivo experiment. The IC50 value of HCFU against Lewis lung carcinoma cells was approximately half that against colon 26 and colon 38 cells. This higher sensitivity of Lewis lung cells against HCFU could be explained by the higher cellular uptake of the drug.

Enhancement of vinblastine-induced cytotoxicity by lysolecithin and phosphatidylinositol☆

Vinblastine inhibited the growth of cultured KB cells 3 days after drug treatment by 55% and 67% at 2.7 ng/ml and 3.5 ng/ml of the medium, respectively. Lysolecithin and phosphatidylinositol showed only a marginal inhibitory effect on the growth of KB cells at respective concentrations of 35-125 microgram/ml and 50-150 microgram/ml of the medium. Lysolecithin, however, enhanced the cytotoxicity of vinblastine. Depending upon the concentrations of lysolecithin (35-125 microgram/ml), the growth of KB cells was inhibited by 60-91% and 86-98% at respective vinblastine concentrations of 2.7 ng/ml and 3.5 ng/ml. Enhancement of vinblastine-induced cytotoxicity also occurred similarly for phosphatidylinositol. The mechanism could be explained partly by an elevated amount of intracellular vinblastine. Other possible mechanisms can only be speculated.

Quantitative estimation of tumor metastasis by measurement of DNA polymerse activity

Metastatic tumor burden in the lung of C57BL/6 or BDF1 mice was quantitated by measuring DNA polymerase a activity in the lung of tumor-bearing animals. DNA polymerase activity in the lung increased time-dependently following the inoculation of Lewis lung carcinoma (s.c.) or B16 melanoma variant B16–B2 (i.v.). In the Lewis lung carcinoma system, the number of metastatic modules and the weight of lung also increased time-dependently. Results from the B16 melanoma showed that the increase in lung nodules occurred 10 to 20 days after i.v. inoculation of tumor cells. DNA polymerase activity increased significantly during this period. Because the lung nodules were very small there was no obvious concomitant increase in lung weight. Since no significant infiltration of host cells was observed in the lung in response to metastatic foci, the rise in DNA polymerase activity should be due to tumor cells and not to infiltrating host cells. When the metastasis of Lewis lung carcinoma was inhibited by adriamycin and cyclophosphamide, decrease in DNA polymerase activity in the lung occurred. These results indicate that the degree of tumor metastasis can be quantitated by measuring DNA polymerase activity.