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The present study was designed to elucidate the effect of cimetidine on CD4 and CD8 positive cells, interleukin-2 (IL-2) production and IL-2 receptor expression in peripheral blood lymphocytes (PBL) from patients with advanced ovarian carcinoma during the course of combination chemotherapy. The absolute number of CD8 (but not CD4) positive cells in PBL from patients with advanced ovarian carcinoma before surgery was significantly higher than that with benign ovarian tumor, while the IL-2 productivity was significantly lower. However, the IL-2 receptor expression was comparable to that with benign ovarian tumor. When a combination chemotherapy consisting of cisplatin, adriamycin and cyclophosphamide was given to these ovarian cancer patients, the IL-2 production was markedly depressed. If cimetidine was given with the combination chemotherapy, the inhibition of IL-2 production by chemotherapy was significantly diminished with a significant increase of CD4 positive cells. On the other hand, the IL-2 receptor expression was not affected by this treatment. When treatment with cimetidine was initiated after completion of chemotherapy, the depressed IL-2 production was restored to the level of control patients with benign ovarian tumor. The restoration seemed to result from an increase in proportion of CD4 positive cells. However, the expression of IL-2 receptor remained unchanged even if cimetidine was given.

The effect of intraperitoneal instillations of interleukin-2 (IL-2) and/or lymphokine-activated killer (LAK) cells on the ascites formation and the survival time was examined using nude mice as a model, with malignant ascites produced by intraperitoneal inoculation of human ovarian cancer cells derived from ascites of a patient with serous cystadenocarcinoma of the ovary. Twenty-eight days after tumor inoculation, all nude mice in the untreated group and in the group treated with spleen cells alone formed ascites. Two of ten nude mice treated with IL-2 alone after tumor inoculation survived without forming ascites during the experimental period. On the other hand, all nude mice treated with LAK cells alone had formed ascites 14 days after tumor inoculation. When LAK cells and IL-2 were combined, five of ten mice survived without forming ascites during the experimental period. The survival time of the group treated with IL-2 alone was significantly prolonged compared to the groups that received medium alone, spleen cells alone and LAK cells alone. When administration of LAK cells was followed by IL-2, the survival time was further prolonged.

In order to establish that the tumor burden in nude mice bearing a xenografted human ovarian tumor can be assessed by measuring plasma levels of alpha-hydroxybutyrate dehydrogenase (HBD) (E.C. 1.1.1.30), the plasma HBD and the tumor volume were measured every week after inoculation of a human ovarian carcinoma cell line designated HR. About 3 weeks after tumor inoculation, all nude mice had a palpable tumor while elevated levels of HBD were observed in a half of the mice at that time. Thereafter the HBD levels rose with increasing tumor volume. There was a quantitative linear relationship between the tumor weight and HBD activity per mouse. When 40 micrograms of cisplatin (about 2 mg/kg) was administered i.p. every week for 5 weeks 2 weeks after inoculation, plasma levels of HBD were significantly decreased on day 27 compared to those in untreated nude mice. Decreases in the HBD levels during treatment with cisplatin occurred 7 days prior to changes in tumor volumes. Further, removal of local tumor mass resulted in a marked decrease in HBD levels on day 1 after surgery and the HBD levels in nude mice which had no recurrent tumor fell into the normal range 8 days after surgery.

The present study was designed to elucidate the mechanism of resistance to cisplatin. A cisplatin-resistant cell line (KFr) was established from KF cells derived from human serous cystadenocarcinoma of the ovary. The DNA histogram revealed an increase of S-phase cells and a decrease of G1-phase cells in cultured KFr cells, compared to that in cultured KF cells. Although the cisplatin content in the KF cells incubated with cisplatin at 10 micrograms/ml increased in a time-dependent manner, that in the KFr cells remained unchanged during the experimental period. When 0.5 mg of cisplatin was administered ip to nude mice with KF or KFr tumor, the cisplatin content in the KFr tumor was significantly lower than that in the KF tumor. The KFr cells showed a cross-resistance to L-phenylalanine mustard, while no cross-resistance to vincristine or 5-fluorouracil was observed. These findings suggest that the mechanism of cisplatin resistance in the KFr cells involves a decrease of cisplatin accumulation in the tumor cells.

We studied production of, absorption of and response to interleukin 2 (IL-2) by peripheral blood mononuclear cells (PBMC) from 66 patients with gynecologic malignancies, in addition to measurement of the OKT 4/OKT 8 cell ratio. Patients with benign tumor served as controls. The OKT 4/OKT 8 cell ratio in patients with advanced (but not early) gynecologic malignancies was significantly lower than that in patients with benign tumor. PBMC from advanced cancer patients activated with phytohemagglutinin (PHA) had significantly lower IL-2 production compared to that from patients with benign tumors, while significant changes in their ability to respond to IL-2 and to absorb IL-2 were not observed. Absolute numbers of OKT 8 positive cells in PBMC of patients with good prognosis were significantly decreased after surgery and chemotherapy, while those of OKT 4 positive cells remained unchanged. Although IL-2 production in PBMC of patients with good prognosis was significantly elevated after chemotherapy, that in PBMC of patients with poor prognosis declined to about a half of pre-operative levels. The ability of PBMC to respond to IL-2 was significantly elevated not only in patients with good prognosis but also in patients with poor prognosis after termination of chemotherapy. On the other hand, the ability of PBMC to absorb IL-2 remained unchanged during the course of treatment. These findings may contribute to the understanding of tumor-induced immune suppression.

To differentiate between ovarian carcinoma and benign ovarian tumor, we compared alpha-hydroxybutyrate dehydrogenase (HBD), lactate dehydrogenase (LDH) and its isoenzyme activities in serum in 115 patients with ovarian carcinoma and benign ovarian tumor. Both serum HBD activity and total LDH activity in ovarian cancer patients were significantly (p less than 0.001) higher than that in patients with benign ovarian tumor. From analysis of serum LDH isoenzymes, a significant decrease in LDH-1, and significant increases in LDH-4 and LDH-5 in ovarian cancer patients were observed, when compared with those in patients with benign ovarian tumor. When the ovarian cancer mass could be removed surgically, a significant decrease in the HBD was observed, suggesting that the HBD might fairly reflect the tumor burden. Serum HBD activities in a half of ovarian cancer patients were higher than the mean +2 SD of those in benign control subjects.

The adjuvant effect of calmodulin antagonists (W-5 and W-7) to 5-fluorouracil (5-FU) was studied by using two human cultured cell lines derived from ovarian adenocarcinoma. Enhancement of the uptake of 5-[6-3H]fluorouracil ([3H]FU) by cancer cells was observed only when calmodulin antagonists were added in the presence of [3H]FU. Similarly, the adjuvant effect by W-5 or W-7 on cancer cell proliferation was also observed only when calmodulin antagonists were added in the presence of 5-FU.

The effect of cimetidine on the natural killer (NK) activity of peripheral blood lymphocytes (PBL) of 32 patients with ovarian carcinoma was studied before and after surgery or chemotherapy. There was no significant difference in the NK activity between normal healthy women and patients who had been disease-free for more than a year after initial surgery, while those in patients with a large residual tumor after surgery were profoundly suppressed. Cimetidine stimulated the NK activity in the disease-free patients in vivo or in vitro. In addition, cimetidine augmented the response to phytohemagglutinin of PBL from the disease-free patients. The NK activity of PBL of patients with a large residual tumor under chemotherapy consisting of cisplatinum, adriamycin and cyclophosphamide, "CAP," was about half of that before initiation of chemotherapy. The inhibition of the NK activity by chemotherapy was abrogated by in vitro exposure of the PBL to cimetidine.

Eleven patients with advanced ovarian carcinoma who had large quantities of ascites during the course of previous chemotherapy were tested with intraperitoneal injections of tranexamic acid, followed by combination chemotherapy. A 4-g dose of tranexamic acid was administered intraperitoneally every day for at least 2 weeks. Seven out of 11 patients (63.6%) showed changes from positive to negative in Papanicolaou smears of the ascitic fluid after treatment with tranexamic acid. A marked reduction of ascitic fluids was observed in 4 of 11 patients (36.4%). If 2 cases with partial reduction of ascites are included, the response rate improved to 54.5%. Median survival time after initiation of treatment with tranexamic acid was significantly longer in patients with good response to tranexamic acid than in patients with poor response. Proliferation of cells in cultures established from ovarian cancer tissue of a patient with good response to tranexamic acid was inhibited at all concentrations of tranexamic acid used in this study, in contrast to non-inhibition of similar cells cultured from a patient with poor response, at any concentration of tranexamic acid. These results suggest that treatment with tranexamic acid suppresses malignant cells in the ascites, followed by reduction of ascites themselves, and an improvement in the response rate to subsequent chemotherapy.

The effect of cimetidine on natural killer (NK) activity of peripheral blood lymphocytes (PBL) was studied in 37 post-operative patients with ovarian cancer. There was no significant difference in the NK activities between normal healthy women and patients with no residual tumor after surgery, while those in patients with large residual tumor after surgery were suppressed to less than a half of those in normal healthy women. Cimetidine augmented the NK activities in patients with no residual tumor but not in normal healthy women. Particularly, the NK activities in patients with low baseline NK status before cimetidine treatment was markedly enhanced. Furthermore, the NK boosting by cimetidine was more effective in patients with large residual tumor than in patients with no residual tumor. The NK activity in PBL from such patients was also enhanced by in vitro exposure to cimetidine. These findings suggest that cimetidine may be used as an immunorestorative-therapy for ovarian cancer patients with depressed NK activity.

Reactivity of lymphocytes from paired living patients with ovarian carcinoma on autologous cancer cells was examined using six established cell lines. Although allogeneic lymphocytes had the capacity to lyse other cultured cancer cells, autologous lymphocytes were found to be unable to lyse their own cultured cancer cells. The defective activity of autologous lymphocytes was restored by preincubation of autologous target cells but not autologous effector cells with the calmodulin inhibitors W-5 or W-7.

Effects of tranexamic acid on tumor growth were examined by using five kinds of human cultured cell lines derived from ovarian malignant tissues. An optimum inhibitory effect on cell proliferation was observed at a concentration of 10 mg/ml tranexamic acid, while replication of a clear cell carcinoma cell line (OK) was not inhibited by any concentration of tranexamic acid used in the present study. Exposure of a cystadenocarcinoma cell line (HR) to 10 mg/ml tranexamic acid for 2 hr resulted in significant growth retardation of tumors formed in nude mice. Further, tranexamic acid seemed to change the morphology of the ovarian carcinoma cells to enlarged cells with abundant cytoplasm. These results suggest that fibrinolytic factors are associated with growth of the tumor and tranexamic acid is useful as an adjuvant therapy for ovarian carcinoma.