Shigeru Tsukagoshi

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.

Antitumor activity of TZT-1027, a novel dolastatin 10 derivative.

Dolastatin 10, a pentapeptide isolated from the marine mollusk Dolabella auricularia, has antitumor activity. TZT‐1027, a dolastatin 10 derivative, is a newly synthesized antitumor compound. We evaluated its antitumor activity against a variety of transplantable tumors in mice. Intermittent injections of TZT‐1027 were more effective than single or repeated injections in rake with P388 leukemia and B16 melanoma. Consequently, TZT‐1027 shows schedule dependency. TZT‐1027 was effective against P388 leukemia not only when administered i.p., but also when given i.v. However, although TZT‐1027 given i.v. was active against murine solid tumors, TZT‐1027 administered i.p. was ineffective against all the tumors tested with the exception of colon 26 adenocarcinoma. The i.v. injection of TZT‐1027 at a dose of 2.0 mg/Ag remarkably inhibited the growth of three murine solid tumors; colon 26 adenocarcinoma, B16 melanoma and M5076 sarcoma, with T/C values of less than 6%. The antitumor activities of TZT‐1027 against these tumors were superior or comparable to those of the reference agents; dolastatin 10, cisplatin, vincristine, 5‐fluorouracil (5‐FU) and E7010. In experiments with drug‐resistant P388 leukemia, TZT‐1027 showed good activity against cisplatin‐resistant P388 and moderate activity against vincristine‐ and 5‐fluorouracil‐resistant P388, but no activity against adriamycin‐resistant P388. TZT‐1027 was also effective against human xenografts, that is, tumor regression was observed in mice bearing MX‐1 breast and LX‐1 lμng carcinomas. TZT‐1027 at 10 μM almost completely inhibited the assembly of porcine brain microtubules. Therefore, its mechanism of antitumor action seems to he, at least in part, ascrihable to the inhibition of microtubule assembly. Because of its good preclinical activity, TZT‐1027 has been entered into phase I clinical trials.

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.

Activation of tumor-infiltrating macrophages by a synthetic lipid A analog (ONO-4007) and its implication in antitumor effects

ONO-4007 is a novel synthetic analog of lipid A subunit and has been shown to exert antitumor activities on various experimental tumors with less toxicity than lipopolysaccharide. It remains unclear, however, what biological activities of this compound are relevant to its antitumor effects. We therefore investigated the activation of macrophages by ONO-4007 in vitro and in vivo and its implication in antitumor effects, using mouse MM46 mammary tumor as an experimental model. Intravenous injection of ONO-4007 produced significant therapeutic effects on this solid tumor. ONO-4007 could stimulate glycogen-elicited peritoneal macrophages in vitro, not only to produce tumor necrosis factor (TNF), but also to exert cytocidal activities against MM46 cells in vitro. Substantial TNF production was induced in tumor tissue by i. v. injection of ONO-4007, and its successive administration to tumor-bearing mice gave tumor-infiltrating macrophages a prominent in vitro tumoricidal activity and primed them for in vitro TNF secretion. These results suggest that activation of tumor-infiltrating macrophages to a direct tumoricidal state as well as to TNF secretion in tumor tissues may be at least some of the antitumor effects of this novel lipid A analog.

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.

Antitumor activity of MST-16, a novel derivative of bis(2,6-dioxopiperazine), in murine tumor models

SummaryWe studied the antitumor activity of newly synthesized bis(1-acyloxymethyl) derivatives of 4,4′-(1,2-ethanediyl)bis(2,6-piperazinedione) using i.p.-i.p. models of P388 leukemia and B16 melanoma. As a result, we found 4,4′-(1,2-ethanediyl)bis(1-isobutoxycarbonyloxymethyl-2,6-piperazinedione) (MST-16) to possess considerable therapeutic activity. MST-16 showed not only marked life-prolonging effects in both P388 leukemia- and B16 melanoma-bearing mice but also a greater therapeutic ratio than did its parent compounds, ICRF-154 and ICRF-159. Further studies revealed that MST-16 has considerable therapeutic activity against a number of other tumors such as ascitic forms of L1210 leukemia, colon 26 adenocarcinoma, and MH-134 hepatoma and solid forms of B16 melanoma, Lewis lung carcinoma, colon 38 adenocarcinoma, and M5076 fibrosarcoma. These results suggest that MST-16 is very promising as an antitumor agent.

Antitumor activities and schedule dependence of orally administered MST-16, a novel derivative of bis(2,6-dioxopiperazine)

SummaryWe studied bioavailability, treatment schedule dependence, and therapeutic efficacy of orally administered MST-16, a novel derivative of bis(2,6-dioxopiperazine), against murine tumors and human tumor xenografts. The rate of its intestinal absorption was about 50%, and it was immediately metabolized to its parent compound, ICRF-154. Therapeutic efficacy of MST-16 was heavily dependent on the treatment schedule: 9 daily oral administrations and treatment every 4 h on day 1 only were much more effective against s.c.-implanted L1210 leukemia than a single dose or five daily administrations giving the same total dose. Orally administered MST-16 showed potent lifeprolonging effects (196%, 219% and 148%) in mice inoculated i.p. with P388, L1210 leukemia, and C-26 colon adenocarcinoma, respectively, but had no effect on B16 melanoma inoculated in the same way. MST-16 inhibited more than 80% growth of Lewis lung carcinoma, B16 melanoma, and C-38 colon adenocarcinoma implanted s.c., but had only a minor effect on M5076 fibrosarcoma. Lung metastasis of Lewis lung carcinoma was also effectively suppressed. Furthermore, MST-16 significantly inhibited growth of human colon, lung and breast cancers implanted s.c. in nude mice. We also made a kinetic analysis of the in vitro cell-killing effect by ICRF-154, the active form of MST-16 in vivo. It demonstrated a cell cycle phase-specific and time-dependent action, providing a reasonable explanation for the schedule-dependent therapeutic effect of MST-16.

Epidermal Growth Factor Receptor‐mediated Selective Cytotoxicity of Antitumor Agents toward Human Xenografts and Murine Syngeneic Solid Tumors

Severe toxic side effects of antiproliferative agents limit their clinical usefulness as anti tumor drugs. Recently we observed that the anti tumor efficacy of various anti tumor agents (5‐fluorouracil, tegafur, adriamycin, mitomycin C, cyclophosphamide, and cisplatin) against experimental solid tumors was enhanced by prior or simultaneous administration of human epidermal growth factor (EGF). However, coadministration of EGF did not enhance the toxicity of anti tumor agents as measured by LD50 and body weight loss. The above selective potentiation of efficacy of the anti tumor agents by human EGF can be characterized as follows. In a dose‐dependent manner, human EGF enhanced the efficacy of an antitumor agent (5‐FU) treatment against human epidermoid carcinoma A431 transplanted sc in atliymic nude mice [ED50=2.9 (0.2–49.7, 95% confidence interval) μg/kg, sc]. Various degrees of enhancement were also observed against other experimental tumors transplanted sc. The degrees of enhancement were directly proportional to the numbers of human EGF binding sites present on tumor cell plasma membrane (threshold of binding site density=1.5×103 sites/cell) using 5‐FU or cisplatin as an anti tumor agent, thus suggesting that the binding of EGF to the receptors on tumor cells is an essential process in enhancing the susceptibility of tumor cells to anti tumor agents. Normal cells including intestinal epithelial and bone marrow cells are endowed with fewer EGF binding sites (less than 103 sites/cell). This may explain partially the absence of EGF‐enhanced cytotoxicity by antitumor agents toward normal cells.

Cancer chemotherapy administered by activated carbon particles and liposomes

In cancer chemotherapy, a specific drug delivery system is expected since many anticancer drugs show toxicity against not only cancer cells but also against normal tissues. The dosage form comprising anticancer drugs adsorbed on activated carbon particles or encapsulated in liposomes is developed as a drug-delivery system which enhances the therapeutic efficacy and reduces the adverse effects. The dosage forms are versatile in size and electric charge, so that large amounts of the drugs are distributed to the targeted organs or tissues and lesser amounts are distributed to the whole body. The dosage forms are designed to release the drugs slowly for a long time at local sites. Through this process, practical use of the dosage forms as an anticancer drug carrier results in an enhancement of anticancer efficacy on the local lesion and a decrease of systemic toxicity.