The purpose of this study is to examine the effects on the HSC-1 cell cycle of etretinate and anticancer drugs (BLM, 5-FU, MMC, PSK), using flow cytometry, and those of combined treatment on cell counts in HSC-1 cell culture. In the DNA histogram, when HSC-1 cells in the stationary period were exposed to etretinate, the S phase cells increased as a proportion of the cell population, mainly because of accelerated development from the G1 + 0 phase into the S phase. Combine treatment with etretinate, BLM and 5-FU decreased the number of HSC-1 cells to a greater extent than combined treatment with BLM and 5-FU. These results suggest that the stimulation of the G1 + 0 phase by etretinate enhanced the cytotoxicity of the anticancer drugs, and that a combination of etretinate with these drugs was effective in treating malignant epithelial tumors.

Seventeen patients with advanced malignancy were treated with recombinant human granulocyte colony stimulating factor rhG-CSF (KRN 8601) infused intravenously over a period of 30 minutes once daily at the dose level/25 micrograms, 50 micrograms, 100 micrograms, 200 micrograms, 400 micrograms, 800 micrograms/m2 for 14 consecutive days, and the effect was compared to the period without rhG-CSF treatment. The maximum numbers of peripheral leukocyte (granulocyte) showed a dose-related increase and the nadir of leukocyte counts escalated with shortening of the period. After stopping infusion, the neutrophil count dropped to the base line level within two or three days. RhG-CSF did not affect other components of peripheral blood such as monocyte, lymphocyte, eosinophil, and hemoglobin value and platelet counts. Transient bone pain occurred in two patients receiving a dose of 800 micrograms/m2. The biochemical changes detected were increased total alkaline phosphatase activity in serum, which appeared in parallel with the increase of neutrophil numbers, and less elevation of total uric acid values. We conclude that an optimal dose of rhG-CSF is 100 micrograms/m2 (average maximum peripheral granulocyte count, 10799/microliters; nadir granulocyte count, 3772/microliters; period of neutropenia, 2.6 days), and rhG-CSF is useful for acceleration of neutrophil recovery and prevention of infection from chemotherapy.

Nephrotoxic effects of cis-Diammine-1,1-cyclobutane dicarboxylate platinum II (CBDCA) were investigated in male 6 weeks old Wistar rats. The animals were divided into 4 groups: Group I (each received a single intraperitoneal injection of CBDCA 80, 120 mg/kg); Group II (a single intraperitoneal injection of CBDCA 80, 120 mg/kg after withdrawal of food and water for 2 days); Group III (intravenous administration of CBDCA 15, 30 mg/kg for 7 consecutive days after withdrawal of food and water for 2 days); Group IV (controls). Rats from each group were sacrificed at 3, 7, 10 and 14 days following the start of the experiment. Serum levels of BUN and creatinine were then measured and renal histopathological examination was conducted by light and electron microscopy. In addition, the total platinum concentration in the serum was measured in Group I, and X-ray microanalysis was performed after intraperitoneal administration of CBDCA (100 mg/kg) and cis-Diamminedichloroplatinum (II) (CDDP) (6 mg/kg) for 2 consecutive days (Group V). The results showed increased levels of BUN in each group due to catabolism. No significant increase in serum creatinine was observed and there appeared to be no evidence of renal dysfunction. For all groups, localized vacuolar degeneration in the epithelial cells of the tubules was predominantly apparent. Electron microscopy revealed only degeneration of the epithelium mainly in the proximal tubules and also showed reabsorption of platinum from the lumen of the tubules.(ABSTRACT TRUNCATED AT 250 WORDS)

Among the cooperative studies of cisplatin (CDDP) analogs in gynecologic malignancies in Japan, a phase III study of CBDCA in ovarian cancer was completed in 1988, while phase II studies of 254-S and DWA2114R are ongoing at the present time. Phase II preliminary data in ovarian cancer revealed a response rate of 25-38%, almost equi-effective and less toxic to that of CDDP. Approximately 20% response rate was achieved in CDDP refractory cases, in particular, responders were observed in mucinous and mesonephroid cases considered CDDP resistant. A phase III study of CBDCA in ovarian cancer, with a comparative study of CAP regimen suggested that CBDCA-containing regimen has an advantage of unnecessity of hydration, in spite of no significant differences response and toxicity. In the cancer of the uterine cervix, approximately 20% response rate was achieved. Of interest is that 254-S yielded a response of 60%, and the result suggested that the agent may have broad antitumor spectrums, different to that of CDDP.

We studied the efficiency and toxicity of three types of recently developed cisplatin analogous agents-carboplatin, 254-S and DW2114, in animal experiments and clinical examinations. In animal experiments, these three agents were injected into SD rats intraperitoneally to determine their effects on rat functions. The antitumor effect of these agents was about the same as that of CDDP, but the renal toxicity of 254-S and DW2114 was lower than that of CDDP. However, more serious myelopathy was observed with these agents than with CDDP. Although slight damage to the testis was seen in the 254-S group, damage to the testis by the other agents was negligible. It was suggested that the damaged region in the testis was spermatogonia. In clinical examinations, we administered the three analogous agents separately for testicular tumors, prostatic cancer and bladder tumors, but a complete response (5M) was recognized in only one case given 245-S, and we found these three agents less effective than CDDP. At present, CDDP is extensively used clinically as one of the combination therapy agents. However, in the first place, we should develop a new CDDP analogous agent that has an anti-tumor effect greater than that of CDDP.

Since the introduction of Cisplatin (CDDP) into clinical practice in 1972, CDDP has assumed an important role in the treatment of various tumors. But its renal toxicity has been proved to be a dose limiting factor. Thus the total number of courses which may be given is limited. For this reason, efforts have been made to develop CDDP analogues with reduced toxicities, especially renal toxicity, and more enhanced antitumor activity, and they are now reaching the clinical testing phase. Among them Carboplatin (CBDCA), 254-S, DWA 2114R and NK 121 have been well studied. These analogues were noted to be less nephrotoxic, but more myelosuppressive than CDDP in preclinical analysis. As for cytotoxicity, their inhibitory effects on tumor growth in murine experimental system were similar or more to that of CDDP. Due to these data clinical trials have been carried out. Phase I studies have shown that these analogues are relatively free of renal toxicity as evaluated in preclinical studies and that their dose limiting factor is myelosuppression. Estimation of cross resistance to CDDP and antitumor spectrum have been studied at phase II trials which are ongoing. Interim reports have not shown that enhanced tumor activity or enlarged antitumor spectrum are expected.

CDDP is an extremely active antineoplastic agent, yet has severe renal, upper GI and neurotoxicity. Therefore, extremely careful supportive care is necessary to administer this agent to patients. There are two ways to improve this agent. The first is to improve the activities of this agent, including widening the spectrum. The other is to reduce toxicity. By these two ways, the therapeutic window or efficacy of the agents will be increased. In these two factors in mind, 4 analogues, CBDCA, 254S, DWA 2114R and NK-121 are in clinical trials in Japan. These 4 agents are different from mother compound CDDP, that is the DLF is bone marrow depression with rather mild renal and upper GI toxicities with different degrees among analogues, compared with those of CDDP. Therefore, these 4 agents do not need hydration before and after the administration. The extensive studies of pharmacokinetics and dynamics are studied including plasma levels, protein bindings and urinary excretions. These above studies indicate some correlations in efficacy and toxicity, but not perfectly correlated. The experiences of the above 4 analogues are still too short to predict the possibility of neurotoxicity, however, it seems to be 4 analogues also neurotoxic. Above findings strongly suggests that the more clinical experiences are necessary to evaluate these analogues.

An equi-toxic dose of CDDP and its analogues (254-S, NK-121, CBDCA and DWA-2114 R) was administered to rabbits and S.180 bearing mice, and the pharmacokinetics were studied. Blood levels: Plasma total platinum (Pt) curves of 5 drugs decreased, showing a biphasic function. The shortest t 1/2 alpha and the longest t 1/2 beta were observed in CDDP group, which correlated with the rate of protein binding (CDDP greater than DWA-2114R greater than NK-121 greater than CBDCA greater than 254-S). Tissue distribution: The tissue levels of Pt decreased slowly, and showed a similar pattern among 5 drugs. The highest level was observed in the kidney, liver and skin, with a moderate high level in the tumor, lung, spleen and thymus, followed by the heart, pancreas, stomach, intestine, muscle and testis in that order. The lowest level was in the brain in S.180 bearing mice.

In the last decade a number of cisplatin analogs have been extensively synthesized to develop new Pt complexes with better antitumor activity and less nephrotoxicity than cisplatin. Antitumor Pt complexes consist of two carrier ligands and two or four leaving groups according to Pt valence. Our screening results of over 500 Pt complexes suggested that the carrier ligands influenced antitumor activity, especially the antitumor spectrum, while the leaving groups affected water solubility, stability, toxicity, and so forth. Recently, more than 10 cisplatin analogs have been clinically tested. In Japan four complexes have been evaluated in phase II. Most of them showed less nephrotoxicity than cisplatin. Instead, myelotoxicity was often observed. These complexes were significantly effective against various murine Their antitumor spectra, however, were somewhat different from each other. The Pt complexes such as cisplatin, carboplatin and 254-S, having the same carrier ligands, diammine, showed a quite similar spectrum. Sensitivity of cell lines derived from human tumors to these analogs were also examined in vivo and/or in vitro. So far as reported, their antitumor spectra were not so different from that of cisplatin. The cross-resistance patterns of cisplatin-resistant sublines to the complexes also depend on types of carrier ligands. Besides, L-1210/DDP showed cross-resistance only to a few carrier ligands, whereas P388/DDP did so to all of them tested. Thus, it might be a reasonable trial to search for a carrier ligand, the complexes of which would show a unique and broad antitumor spectrum including cisplatin-resistant lines and, in addition, not only against murine tumors but against human tumor cells.

Protein and RNA content in accumulated G1 cells was measured with B16 melanoma cells by flow cytometer to study the mechanism of antineoplastic action of delta 12PGJ2. DNA histogram showed that, after a 6 hr exposure to delta 12PGJ2, the percentage of tumor cells in G2M phase increased, and G1 accumulation with depletion of cell population in S phase was seen after 12-24 hr exposure to 2.0 micrograms/ml of delta 12 PGJ2. Protein content in accumulated G1 cells increased at 6 hr after exposure, and subsequently decreased. However, the protein content treated with delta 12PGJ2 was higher than that of the control. A reduction of RNA content in G1 cells was seen after delta 12PGJ2 exposure. As the result, it was found that delta 12PGJ2 made cell progression ceased in G1 phase of cell cycle, and where protein and RNA content was higher than that of control. It is suggested that some hyper-metabolite state was caused by delta 12 PGJ2 in cultured B16 melanoma cells.

In vitro and in vivo cytotoxicity tests were carried out using the YTS-1 human bladder carcinoma cell line. The lethal effects of a 1-hour treatment with 5 anti-cancer agents (Cisplatin [CDDP], Cytosine arabinoside [ara-C], Adriamycin [ADM], Etoposide [VP-16] and Carboplatin [CBDCA]) on the YTS-1 cells grown as monolayer were examined quantitatively by the regrowth assay methods. In addition, the lethal effects of 3 anti-cancer agents (CDDP, ADM, ara-C) were also examined in vivo. Moreover the effects of simultaneous combinations of CDDP and ara-C, and ADM and ara-C were assessed in vitro and in vivo. In vitro studies showed that, of these single agents, ADM and ara-C were cytotoxic judging from the human maximal plasma concentration of drugs, whereas CDDP, VP-16 and CBDCA were not. Combinations of CDDP and ara-C, and ADM and ara-C showed more than additive effect. In vivo studies on YTS-1 tumors implanted to athymic nude mice (BALB/c) showed that ADM was the cytotoxic agent. Although CDDP alone or ara-C alone was not cytotoxic to YTS-1 tumors, combination of CDDP and ara-C showed antitumor activity. These results show that combination of CDDP and ara-C is synergistic. The most effective combination was observed to be ADM and ara-C, but severe loss of body weight was also observed.

Metallothionein (MT), a metal binding protein induced by bismuth and other heavy metals, has been proved to have a potential to prevent toxic side effects of several antitumor drugs. It has also been demonstrated that the induction of MT in tumor tissues diminishes the antitumor activity of the drugs as well. These facts suggest a possibility that MT may play an important role in acquiring multi-drug-resistance in tumor cells. Thus, we were encouraged to examine the sensitivity of cultured cells overexpressing MT to various antitumor drugs which differ from each other in the mechanism of action. Cadmium-resistant cells (HeLa-R) were obtained from HeLa S3 cells (HeLa-S) by successive cultivation in a medium containing 100 microM CdCl2. MT level in HeLa-R was 180 times higher than that of HeLa-S. Comparison of the sensitivity of these two strains to the antitumor drugs, such as, cis-diamminedichloroplatinum (cis-DDP), adriamycin (ADR), peplomycin (PEP) and melphalan (MEL), revealed that HeLa-R was significantly more resistant to these drugs than HeLa-S. Further, there was no significant discrepancy between these strains, either in the level of major radical scavenging factors other than MT or in the cellular uptake of the drugs. The experimental results described above appear to support the hypothesis that MT may act as a multi-drug-resistance factor in tumor tissues.