Differing roles of protein kinase C on the signal transduction of tumour necrosis factor-alpha and -beta on PANC-1 cells: in vitro autoradiographic investigation.

We investigated alterations in protein kinase C (PKC) activity of PANC-1 cells following treatment with tumour necrosis factor (TNF)-alpha or TNF-beta by an in vitro autoradiographic method. Binding studies performed on whole cells using [3H]phorbol-12,13-dibutyrate (PDBu) as a ligand revealed strong activation of PKC by TNFs within 30 min. The effect was similar to that seen after 30 min treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA). After treatment for 24 h, TNF-beta caused a marked down-regulation of PKC similar to that seen after 24 h treatment with TPA; significant activation persisted, however, in the cells treated for 24 h with TNF-alpha. Our data suggest that PKC activation may play a more important role in the TNF-alpha signal transduction pathway than in that of TNF-beta.

Five primary cancers in one patient.

We report a 62-year-old man with five primary cancers. He underwent nephrectomy for a right renal cell carcinoma and removal of malignant meningioma and 6 years later was diagnosed as having a rectal cancer and hepatocellular carcinoma. He died of respiratory failure and a gastric cancer was found at autopsy.

Anti-proliferative effect of combined tumour necrosis factor-alpha and interferon-alpha on human pancreatic cancer in vitro and in vivo.

The anti-proliferative effects of tumour necrosis factor-alpha (TNF-alpha) and interferon-alpha (IFN-alpha), alone or in combination, on human pancreatic cancer cells lines (PANC-1, MIA PaCa-2 and BxPC-3) and human pancreatic cancer tumour (Exp-58), were investigated in vitro and in vivo. The anti-proliferative effect was determined using the dye uptake method and the subcutaneous tumour model. Combined TNF-alpha and IFN-alpha demonstrated marked synergistic and/or additive effects in comparison with their effects as single agents. These results suggest that combined cytokine therapy of TNF-alpha and IFN-alpha may make possible some improvement in the treatment of pancreatic carcinoma patients in the future.

Metachronous liver metastasis of colorectal cancer: timing of occurrence and efficacy of adjuvant portal chemotherapy.

We estimated the time of occurrence of metachronous liver metastasis in colorectal cancer patients from tumor diameter and doubling time. Micro-metastasis was present prior to operation in most patients and a few metastatic cases could have been initiated by the surgical procedure. Portal chemotherapy is more effective against liver metastasis than intravenous infusion because a higher drug concentration in the liver can be obtained. This efficacy of portal chemotherapy on survival was also observed in a rat model. Thus perioperative adjuvant treatment should be undertaken for metastasis which already existed before the operation and adjuvant chemotherapy via portal vein is the treatment of choice. The no touch isolation technique is also needed to avoid spreading of tumor cells during surgery.

Synchronous colorectal carcinomas.

Eighteen (5.0%) out of 358 patients who underwent resection of a colorectal carcinoma during the period 1978 through 1990 had synchronous colorectal carcinomas, and were 5.6 years younger on average than those with a single carcinoma. The distance between synchronous lesions was less than 10 cm in 69.6% of all the patients in the study. Among the synchronous carcinomas there was a higher incidence of ascending colon involvement, mucinous carcinoma, family history of malignant diseases, multiple malignant neoplasms associated with other organs and benign neoplastic polyps of the colorectum, and it is suggested that hereditary oncogenic factors influence these carcinomas. The synchronous lesions were detected pre-operatively in 14 of 18 patients with synchronous carcinomas, and the most common reason why synchronous lesions were missed was that the lesions on the anal side prevented the lesions on the proximal side from being examined. The prognosis in the synchronous lesion group was worse than in the solitary lesion group. Since it is difficult to predict synchronous colorectal carcinomas, careful pre-operative examination, including that of other organs, is necessary, and intra-operative colonoscopy should be carried out when pre-operative examination was insufficient.

Active enhancement of rat cardiac allografts induced by donor specific semisoluble antigens.

Active enhancement was induced in inbred rats with cardiac allografts using semisoluble antigens. The optimal time of antigen pretreatment and optimal dose of semisoluble antigens were examined. The presence of serum blocking factors in the sera of rats having had allografts for a long time was examined with a macrophage migration inhibition test and lymphocyte microcytotoxicity assay. Since the blocking factors of macrophage migration inhibition were increasing on the 7th day, that day was determined to be the optimal time of antigen pretreatment. The mean survival time (MST) of cardiac allografts in untreated rats was 17.2 +/- 7.5 days. Semisoluble antigens were administered at 2 mg/kg body weight 7 days before the graft, 4 mg/kg 7 days before the graft and 2 mg/kg divided over three days, 15, 8 and 1 day before the graft, and the MSTs of cardiac allografts of rats receiving these treatments were 71.2 +/- 39.9, 62.6 +/- 42.2 and 79.3 +/- 31.0 days, respectively. The MST in each group of the treated rats was significantly longer than that of the control group (p less than 0.01). Rejection of the allograft, however, was accelerated in a group treated with 8 mg/kg 7 days before the graft (MST: 8.4 +/- 3.2 days). Serum blocking factors were detected in the sera of approximately half of the rats having cardiac allografts which survived a long time.

Antitumor effect of natural human tumor necrosis factor-beta against Lewis lung carcinoma in mice and its synergistic potentiation by interferon.

We investigated the antitumor effect of purified natural human tumor necrosis factor-beta (nHuTNF-beta) produced by human acute lymphoblastic leukemia BALL-1 cells stimulated with HVJ on pulmonary metastatic tumors of Lewis lung carcinoma (3LL) transplanted into BDF1 mice. nHuTNF-beta showed antiproliferative effects on metastatic tumors in a dose-dependent manner when administered daily for 10 days by the intravenous route. Histological examination of the tumors treated with nHuTNF-beta revealed that the tumor size and number of metastases were much reduced. Lytic cellular changes, including cytoplasmic vacuolation, loosening of the intercellular junction and both cytoplasmic and nuclear swelling, were found, but tumor necrosis was not. These findings indicate a therapeutic effect of Grade IIa according to the histological criteria of Shimosato and Ohboshi. In addition, synergistic augmentation of the antiproliferative effects of nHuTNF-beta by natural murine interferon-alpha/beta (nMu-IFN-alpha/beta) or recombinant murine interferon-gamma (rMuIFN-gamma) was recognized by median effect plot analysis. The results suggested that nHuTNF-beta may well deserve clinical trial as a new immunotherapeutical agent for human cancer.

Comparison of the subrenal capsule assay and succinate dehydrogenase inhibition test as drug sensitivity tests for cancer.

The same chemotherapeutic agents were tested against fresh surgical explants of solid tumors obtained from 50 patients using the in vivo subrenal capsule (SRC) assay and the in vitro succinate dehydrogenase inhibition (SDI) test in comparison. Control growth adequate to meet evaluable assay criteria was obtained in 36 of the 50 tumors tested in the SRC assay (72.0%). In the SDI test, 46 of 50 tumors were evaluable (92.0%). Correlations between the two test systems were dependent upon the activity criteria established for each system. With activity criteria set at a change of less than or equal to -2.0 in the drug sensitivity score for the SRC assay and greater than or equal to 50.0% inhibition of succinate dehydrogenase activity for the SDI test, 12.5% of the drugs tested were active in the SRC assay and 22.3% were active in the SDI test. Correlations of tumor response between the two test systems were 31.7% for sensitivity (13/41) and 95.1% for resistance (98/103). In spite of the fundamental difference between the SRC assay and SDI test, meaningful correlations between the test results and clinical tumor responses in both test systems were obtained. This fact suggests that the two methods are complementary to each other.

Synergistic antiproliferative effect of the combination of natural human tumor necrosis factor-alpha and natural murine interferon-alpha/beta against colon-26 adenocarcinoma hepatic metastases in a murine model.

To prevent the development of hepatic metastases after surgery for colorectal cancer, it is important to inhibit the growth of any micrometastases which occur during the operation. We used a hepatic metastasis model in mice to investigate the effects of combination therapy with natural human tumor necrosis factor-alpha (nHuTNF-alpha) and natural murine interferon-alpha/beta (nMuIFN-alpha/beta). Decreased formation of hepatic metastases by murine colon-26 carcinoma was recognized following a single injection of nHuTNF-alpha, nMuIFN-alpha/beta, or both. These inhibitory effects were synergistic. NK activity was also measured, because notaral lerller cells not only have an anti-tumor effect but are also a representative of the host immune system. Both nHuTNF-alpha and nMuIFN-alpha/beta were able to activate NK cells, and the combination of the cytokines more significantly augmented NK activity. The in vivo elevation of NK activity induced by nHuTNF-alpha, nMuIFN-alpha/beta, or their combination may be one of the mechanisms of their antiproliferative effect on experimental hepatic metastases of murine colon-26 carcinoma.

Effect of selected splenic irradiation on growth of meth A-fibrosarcoma in mice and partial characterization of splenic effector and suppressor cell populations.

Meth A-fibrosarcoma bearing BALB/c mice were subjected to selected splenic irradiation (2.0-4.0 Gy) on days 7 and 14 of tumor growth. Tumor growth was recorded by serial measurement. Irradiation given on day 7 caused regression of tumor, but irradiation given on day 14 did not show tumor regression. Antitumor activity in the Winn assay was detected in spleen cells 3 days after irradiation, but was not detected 7 days after. The cell surface phenotypes were analyzed on days 3, 7 and 14 of splenic irradiation using monoclonal antibodies (anti-Thy1.2 antibody, anti-Lyt1 antibody, anti-Lyt2 antibody, anti-L3T4 antibody) by flow cytometry. Thy 1.2, Lyt1, and L3T4 cells were increased on day 3 of splenic irradiation, but were not on days 7 and 14. Lyt2-cells did not show increase on days 3, 7 and 14. It was possibly suggested that selected splenic irradiation induced tumor regression was caused by the ability of irradiation to preferentially eliminate suppressor T cells, thereby allowing effector T-cells to become relatively dominant.

Early phase II study of interferon-alpha and tumor necrosis factor-alpha combination in patients with advanced cancer.

Synergistic enhancement of anti-tumor effects through the combined use of natural human interferon-alpha (nHuIFN-alpha) and natural human tumor necrosis factor-alpha (nHuTNF-alpha) enabled us to decrease the effective dose of each cytokine and consequently to reduce side effects. One hundred and twenty patients with advanced or recurrent solid cancer were entered in the trial from April 1985 to January 1988, of whom 112 patients were evaluable. A mixture of nHuINF-alpha and nHuTNF-alpha was injected intravenously as the maintenance dose 1 x 10(6)U or more/day for over 8 weeks. There was no response in 40 patients injected with the maintenance dose of 1 x 10(6)U/day, but of 72 patients receiving more than 2 x 10(6)U/day (10 micrograms of nHuIFN-alpha and 3 micrograms of nHuTNF-alpha), 4 had complete responses, 10 had partial responses, and 4 had minor responses. The overall response rate was 12.5% (14/112) and the rate was 19.5% in 72 patients with more than 2 x 10(6)U/day. Positive responses were as follows: hepatoma 3/8), renal cell cancer (4/11), breast cancer (4/17), ovarian cancer (1/2), malignant thymoma (1/1) and liposarcoma (1/1). Serious adverse effects like hypotension, oliguria and severe hepatobiliary toxicity were never experienced. The effective and adequate dose of the mixed preparation was considered 2 to 4 x 10(6)U/day/body.

Early diagnosis of acute renal allograft rejection: efficacy of macrophage migration inhibition test as an immunological diagnosis.

1. Three cases of acute rejection were detected by macrophage migration inhibition tests (MIT) conducted directly on seven patients who had received renal allografts. The macrophage migration inhibitory factor (MIF) activity was positive in all cases 1-2 days before the appearance of acute rejection. 2. After the administration of a high dose of Solu-Medrol (1g/day for 3 days) to suppress the acute rejection, MIF activity recovered to its normal level 3 days later. These findings seem to indicate that MIT yields immunologically useful criteria for the early detection of an acute rejection.

The effect and distribution of a protein-bound polysaccharide preparation, PSK (Krestin), intratumorally injected prior to surgery into gastric cancer patients.

In order to improve the postoperative prognosis of gastric cancer patients we have performed preoperative endoscopic intratumoral administration of various biological response modifiers. In the present study we have investigated the kinetics and the immune response augmenting effect of intratumorally injected PSK, a protein-bound polysaccharide preparation, by immunohistochemical methods using anti-PSK antibody and various other antibodies. PSK-containing cells were located in the tumor tissues and follicular marginal zones of regional lymph nodes. Intratumorally administered PSK appeared to be phagocytized by the histiocytes and to cause them to become antigen-presenting cells. These cells may play a major role in augmenting immune responses in gastric cancer patients.

Primary non-Hodgkin's lymphoma of the rectum: a case report.

A rare gastrointestinal tract neoplasm, primary non-Hodgkin's B-cell lymphoma in a 39-year-old, asymptomatic woman is described. The tumor was originally localized in the rectum without evidence of any other lymphoma-involved organ and treated by curative surgical procedure associated with postoperative chemotherapy.

Comparative antitumor activity of 5-fluorouracil and its prodrugs in combination with hyperthermia in vitro.

We investigated the antitumor activities of 5-fluorouracil (5-FU), 5'-deoxy-5-fluorouridine (5'-DFUR), 1-hexylcarbamoyl-5-fluorouracil (HCFU) and 1-(tetrahydro-2-furanyl)-5-fluorouracil (FT-207) in combination with hyperthermia in vitro. The antitumor effect of 5-FU (10(-4) M) was slightly enhanced by combination with hyperthermia (42 degrees C) for 2h, and the effect was determined to be additive. Synergistic enhancement of antitumor activity was obtained by the concurrent use of hyperthermia (42 degrees C, 2h) and 5'-DFUR (10(-4) M) or HCFU (10(-5) M). However, the antitumor effect of FT-207 (10(-4) M) in combination with hyperthermia was comparable that of hyperthermia alone. The synergistic enhancement of antitumor activity was not obtained for all drugs when the cells were preheated at 42 degrees C for 2h. On the other hand, when cells were pretreated with drugs before heat exposure, weak interactions were obtained after 5-FU and 5'-DFUR treatment, and a synergistic interaction was obtained after HCFU treatment. It is speculated that the metabolites of 5'-DFUR and HCFU enhance the cytotoxicity of 5-FU, or might change the threshold concentration for a cytotoxic effect of 5-FU in cancer cells.

Beneficial effect of donor-specific blood transfusions (DST) on living-related kidney allograft survival.

The survival rate of 19 patients who underwent living-related kidney transplantation after donor-specific blood transfusions (DST) was compared with that of 32 historical controls receiving transplants without DST. The graft survival rate of the DST group was 82% after two and three years. The graft survival rate of the DST group was significantly better than the 53% rate after two years obtained with the 32 historical controls (p less than 0.05). We tested sera from 16 DST-treated recipients to study the beneficial effect of DST on kidney allograft survival using the mixed lymphocyte culture (MLC) serum inhibition test. The results demonstrated that MLC inhibitory factors were induced in the serum of the recipient after completion of DST. This inhibition of MLC was observed by treatment of responder lymphocytes with serum obtained three weeks after DST plus rabbit complement. The inhibitory effect was also specific for responder cells in anti-donor MLC. Regarding the correlation with rejection episodes, these MLC inhibitory factors were often observed in the non-rejection group (p less than 0.05). The data suggest that such factors may be anti-idiotypic antibodies and be associated with prolonged graft survival.

Effects of cyclosporine A on experimental hepatic metastases of mouse colon-26 tumour.

Cyclosporine A (CSA), an immunosuppressive agent with apparently selective effects on T lymphocytes and little myelotoxicity, was tested for its effects on hepatic metastases by inoculation of mouse colon-26 tumour cells into the portal vein in male CDF1 mice. CSA, given subcutaneously in daily doses of 10-50 mg/kg/day, for 22 days, significantly increased the incidence of hepatic nodules. This increase was positively correlated with the CSA dose. When 5-fluorouracil was injected at a dose of 15 mg/kg/day every other day after the inoculation of tumour cells a highly significant reduction in the incidence of metastases was seen (compared with the controls). This strong inhibitory effect of 5-fluorouracil on metastasis was almost completely suppressed when mice were treated with CSA concomitantly. The results suggest that CSA affects the host immune system, accelerating the production of hepatic metastases.

Anti-proliferative effect on human pancreatic cancer cells of natural human tumour necrosis factor-beta combined with natural human interferon-alpha or interferon-gamma.

The anti-proliferative effects of natural cytokines, human tumour necrosis factor-beta, natural human interferon-alpha and natural human interferon-gamma, on three human pancreatic cancer cell lines (PANC-1, MIA PaCa-2 and BxPC-3) were investigated in vitro. The anti-proliferative effect was determined using the dye uptake method and analysed for synergism by the median effect principle. Tumour necrosis factor-beta, as a single agent, had little anti-proliferative effect on any of the three cell lines, whereas interferon-alpha and interferon-gamma exhibited a strong anti-proliferative effect against two cell lines (MIA PaCa-2 and BxPC-3) and one cell line (BxPC-3), respectively. When tumour necrosis factor-beta and interferon-alpha were administered together (ratio 1:1), a synergistic effect was observed against PANC-1 cells. The combination of tumour necrosis factor-beta and interferon-gamma (ratio 10:1) was synergistic against both PANC-1 and MIA PaCa-2 cells. A synergistic anti-proliferative effect of tumour necrosis factor-beta and interferons was, therefore, observed even for cell lines that showed little biological response to each cytokine alone. The data suggest that some future improvement in the treatment of pancreatic cancer may be obtained by using combination cytokine therapy.

Inhibitory effect of nafamostat mesilate on metastasis into the livers of mice and on invasion of the extracellular matrix by cancer cells.

Although many agents that interfere with clotting mechanisms have been investigated for their potential to inhibit metastasis, their toxicity has prevented administration of sufficiently high doses to achieve inhibition of metastasis in clinical trials. Nafamostat mesilate (FUT-175), a synthetic serine protease inhibitor, inhibited liver metastasis in a CDF1 mice model with colon 26 adenocarcinoma cells. The apparently dose-dependent inhibitory effect was seen 21 days after all of the doses tested (0.3, 1.0, 3.0 and 10.0 mg/kg for 7 days) but the effect was only statistically significant (P less than 0.01) at the highest dose. The blood concentrations 3 min after dosing were less than 10(-6) M for all of the doses tested. At a concentration of 10(-5) M or less nafamostat mesilate was not cytotoxic towards colon 26 cells in vitro. The results indicate that it may not be difficult to achieve blood nafamostat mesilate concentrations that inhibit metastasis in mouse liver. Possible mechanisms of nafamostat mesilate are inhibition of extravasation and invasion of cancer cells, inactivation of collagenase due to inhibition of plasmin activity and inhibition of the formation of the cancer cell thrombus, and arrest in the capillaries through inhibition of thrombin activity. These preliminary results suggest that peri-operative administration of nafamostat mesilate may prevent metastasis into the liver after surgery for gastrointestinal malignancies.

[Clinical evaluation of cefmetazole for the prevention of postoperative wound infections].

The authors treated 415 patients, with injection of 4 g cefmetazole (CMZ) per day after operation of the digestive tract. In these cases, the prevention of postoperative wound infections was investigated and the following results were obtained. Out of 415 cases, 11 cases (2.7%) had postoperative wound infections; 6 cases of which were superficial wound infections and 5 cases deep wound infections. In relation to the degree of infection of the surgical field of them, 10 cases were performed with the contamination by bowel organisms and 1 case was in infected surgical field. Bacteriological examination was carried out. Twenty-four strains of bacteria were isolated and identified. The major bacterial strains identified were 6 strains of S. faecalis, 4 strains of E. cloacae and 4 strains of P. aeruginosa. These organisms were rarely sensitive to CMZ. The results suggest that the organisms causing postoperative wound infections are changing compared with the organisms of previous reports and that the use of CMZ as a postoperative medication is useful in the prevention of wound infections.