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A 38-year-old multiparous woman was referred at 19 weeks' gestation because of hydrops fetalis. Ultrasonic examination revealed severe pleural effusion, ascites and skin edema. Detailed examination of the amniotic fluid, fetal blood and intrathoracic fluid led to a diagnosis of congenital fetal chylothorax. Repeated thoracocenteses were not effective in improving the hydrops fetalis. We introduced fetal treatment for the pleural effusion by an intrapleural injection of OK-432 at 23, 24 and 25 weeks' gestation. The pleural effusion was reduced by adhesion of the intrathoracic space and resulted in the delivery of a neonate who was healthy except for right renal dysfunction. Pulmonary hypoplasia was successfully prevented by OK-432.

We present a case of pleurodesis by intrapleural injection of OK-432 for the treatment of fetal chylothorax at an early gestational age. OK-432 injection achieved rapid and effective control of pleural effusion with no adverse effects.

Background/aims: Although the therapeutic results of gastric cancer have markedly improved, it still remains the most common cancer death in Korea., Methodology: The clinicopathologic characteristics were analyzed for 11,491 consecutive patients who underwent operation for gastric cancer at the Department of Surgery, Seoul National University Hospital from 1971 to 1997. We also evaluated the survival and prognostic factors for 9,262 consecutive patients from 1981 to 1996. The prognostic significance of treatment modality [surgery alone, surgery + chemotherapy, surgery + immunotherapy + chemotherapy (immunochemo-surgery)] were evaluated in stage III gastric cancer., Results: The 5-year survival rate of overall patients was 55.8%, and that of patients who received curative resection was 64.8%. The 5-year survival rates according to TNM stage were 92.9% for Ia, 84.2% for Ib, 69.3% for II, 45.8% for IIIa, 29.6% for IIIb and 9.2% for IV. Depending on the extent of resection, the 5-year survival rates were 68.7% for STG, 45.4% for TG, 19.6% for combined resection and there was no 5-year survivor in the nonresection group. Regarding adjuvant treatment modality, significant survival difference was observed in stage III patients. The 5-year survival rates were 44.8% for the immunochemo-surgery group, 36.8% for the surgery + chemotherapy group and 27.2% for the surgery alone group., Conclusions: Curative resection, depth of invasion and lymph node metastasis were the most significant prognostic factors in gastric cancer. Consequently, early detection and curative resection with radical lymph node dissection, followed by immunochemotherapy especially in patients with stage III gastric cancer should be recommended as a standard treatment principle for patients with gastric cancer.

Hypothesis/objectives: Congenital lymphatic malformations of the head and neck (LMHN) present special challenges to the otolaryngologist-head and neck surgeon. Recently, a number of sclerotherapy trials have shown promising results. In this study, we present our experiences with picibanil (OK-432) sclerotherapy for this lesion., Study Design: Retrospectively review., Methods: We retrospectively reviewed 21 patients who have undergone sclerotherapy with picibanil for LMHN., Results: Satisfactory response with complete or nearly complete shrinkage of the lesions was observed in 15 cases after repeated sclerotherapy (average, two times). We did not observe any significant morbidity or complications in the patients treated with picibanil. Reduction in size of the mass was achieved in weeks to months. Some of the patients who had not had any other previous treatment showed remarkable reductions in size even after the first therapy. When we used picibanil sclerotherapy as a primary treatment for the LMHN, most of our patients showed satisfactory results regardless of the size or location of the lesions., Conclusion: Given with our experience and the reports that failure of picibanil sclerotherapy does not hinder subsequent surgical salvage procedures, we recommend trying picibanil sclerotherapy as a primary treatment for the LMHN and performing surgical excision as a secondary modality if the response to the sclerotherapy is not satisfactory.

The benefits of immunochemotherapy with a penicillin-treated, lyophilized preparation of Streptococcus pyogenes, OK-432 (Picibanil), were reassessed in patients with resected non-small-cell lung cancer through a meta-analysis based on data from 1,520 patients enrolled in 11 randomized clinical trials. All 11 trials were started before 1991, and the subjects had been followed up for at least 5 years after surgery and randomization. In these trials, standard chemotherapy was compared with the same therapy plus OK-432. The endpoint of interest was overall survival, and analysis was based on intent-to-treat population without patient exclusion. Data were analyzed using the Mantel-Haenszel method. The 5-year survival rate for all eligible patients in the 11 trials was 51.2% in the immunochemotherapy group versus 43.7% in the chemotherapy group. The odds ratio (OR) for overall survival was 0.70 (95% CI = 0.56-0.87, p = 0.0010). Analysis of four trials in which central randomization was performed also reconfirmed a significantly longer survival time for the immunochemotherapy group (OR = 0.66, 95% CI = 0.44-1.00, p = 0.049). Based on these results of meta-analysis, it is postulated that postoperative adjuvant immunochemotherapy using OK-432 might improve the survival of patients after resection of non-small-cell lung cancer.

There is no one common immunotherapy for the treatment of head and neck cancer (H&N cancer). A streptococcal agent, OK-432, which is classified as a biological response modifier (BRM), is occasionally used by means of local administration for recurrent H&N cancer, and the response rate is approximately 18%. In regard to specific immunotherapy, a murine monoclonal antibody (named mAb 225) against the epidermal growth factor receptor (FGFR) that is frequently overexpressed in H&N cancer has been produced in the U.S.A. Furthermore, to obviate human anti-mouse antibody responses, a chimeric human-to-murine version of mAb 225 (C225) was developed by exchanging the constant regions of mAb 225 to counterparts in human immune globulin. Phase I clinical trials of C225 in the U.S.A. demonstrated that treatment with C225 was well tolerated and that C225 given in combination with cisplatin has biologic activity. On the other hand, many tumor antigens recognized by cytotoxic T lymphocytes (CTL) have been identified from a variety of malignant tumors and some of them, including the MAGE-3 antigen, are frequently expressed in H&N cancer. We identified an MAGE-3-derived epitope recognized by HLA-A24-restricted CTL from peripheral blood mononuclear cells (PBMC). In contrast we failed to generate CTL specific for MAGE-3+/HLA-A24+ tumors from PBMC in any of 5 HLA-A24+ cancer patients whose tumors expressed the MAGE-3 gene. Therefore, we did not apply MAGE-3-derived CTL epitope in clinical uses such as peptide vaccine and peptide pulsed dendritic cell infusion for H&N cancer.

It is known that thymidine phosphorylase (dThdPase) is increased in various types of malignant tumors and is induced by cytokines. In this study, we have investigated the effects of OK-432, which induces multiple cytokines, on dTHdPase expression and angiogenesis in human gastric carcinomas. We examined 25 patients who underwent gastrectomy for gastric carcinoma. OK-432 was directly injected in tumors in 16 (OK group) of 25 patients via endoscopy before operation and the other 9 patients were not treated (control group). The dThdPase activity in carcinoma tissues of the OK group was significantly higher than that of the control group (P < 0.05). The amounts of IL-1 alpha, IFN-alpha, and IFN-gamma in carcinomas in the OK group were significantly higher than in the controls (P < 0.05), and these were significantly correlated with the dThdPase activity. Intratumoral OK-432 administration enhances the expression of dThdPase in gastric carcinoma cells by inducing various cytokines.

A giant multiseptate cystic mass covering the whole left hemitruncus of fetus was detected by ultrasonography at 23 weeks of gestation. Color and pulsed Doppler ultrasonography revealed marked blood flow in the mass. No other anomaly or chromosomal abnormality was recognized. Although the cystic lesion increased in size with advancing gestation a male infant weighing 4096 g was delivered in good condition at term. Histological examination of the lesion demonstrated hemangiolymphangioma with hyperplasia of both capillaries and lymphoducts. The infant was treated effectively with OK-432 and interferon-alfa-2a.

Malignant tumors such as brain tumors have been reported to be associated with immunosuppression caused by certain tumor-secreted cytokines. The reversion of tumor-derived immunosuppression has not been described. The use of OK-432, an immunomodulatory agent prepared from Su-strain of Streptococcus pyogenes A3, to activate peripheral blood mononuclear cells from a patient with glioblastoma multiforme has demonstrated a sharp rise in proliferative response. This proliferative response was compromised in the presence of living and irradiated autogeneic cancer cells. The conditioned media from cultured cells of glioblastoma multiforme, astrocytoma, and cholangiocarcinoma were tested for immunosuppressive ability. We found that conditioned media from 3 of 4 cases of glioblastoma, all 3 cases of astrocytoma, and 1 case of cholangiocarcinoma exhibited immunosuppressive activity toward the proliferative response of allogeneic peripheral blood mononuclear cells to phytohemagglutinin. This is the first report that cholangiocarcinoma produces soluble immunosuppressor(s). Our finding suggested that soluble substance(s) as well as direct cell-cell contact between tumor cells and mononuclear cells play roles in the observed tumor-derived immunosuppression.

Recurrence of hepatocellular carcinoma (HCC) is frequent, even after apparently curative resection. Preoperative transcatheter arterial chemoembolization (TAE) does not improve disease-free survival after hepatic resection. We previously reported the potential usefulness of transarterial immunoembolization (TIE), a newly developed arterial embolization technique using OK-432 and fibrinogen, as preoperative treatment. In this study, we further investigated the effect of TIE by histologic examination of the resected specimens and compared it with conventional TAE in a prospective nonrandomized manner. Thirty-nine patients underwent TIE (n = 17) or TAE (n = 22) before curative hepatectomy for HCC. Transarterial immunoembolization was performed according to the standard protocol using OK-432, fibrinogen, and thrombin. Histologic changes in cancerous and noncancerous liver tissues were examined at different stages after TIE. Histologic grading of cancer cell injury according to the modified Shimosato criteria (Grades 0-IV, in increasing order of severity of cell injury) and postoperative disease-free survival were compared between the two groups. Based on the results of histopathology, TAE was more effective than TIE against the main tumor. In contrast, TIE was significantly more effective than TAE against extracapsular invasion and intrahepatic metastasis. Disease-free survival after hepatectomy tended to be better in patients pretreated with TIE than TAE. Postoperative tumor recurrences in the TIE group (n = 4) occurred in the nontreatment regions, whereas tumor recurrences in TAE group developed mostly (8 of 11 patients) in treated liver regions. Based on results of histologic examination, TIE seems to be more effective than conventional TAE against extracapsular invasion and intrahepatic metastasis. Data for disease-free survival and recurrence site suggest TIE may be a useful preoperative treatment.

OK-432 has been used clinically as a biological response modifier for cancer therapy. We investigated here the protective effects of OK-432 against endotoxic shock and infectious death caused by Pseudomonas aeruginosa and Salmonella enteritidis in mice and proposed a possible mechanism. Pretreatment of OK-432 reduced the lethality of lipopolysaccharide (LPS)-induced endotoxic shock in D-(+)-galactosamine-sensitized C3H/HeN mice. OK-432 did not affect the TNFalpha production in blood, but it did decrease the susceptibility to TNFalpha. Furthermore, an acceleration of LPS clearance from blood was detected. The pretreatment of OK-432 also decreased the lethality of mice in bacterial infection caused by P. aeruginosa and S. enteritidis. The rapid decrease of the viable bacteria from the circulating blood and in spleen and liver in mice was observed in a manner similar to LPS clearance. These findings indicate that the protective effect of OK-432 against the endotoxemia and bacteremia may depend on an up-regulation of clearance of LPS and bacteria and the augmented resistance to TNFalpha.

Background: Although a few reports indicated some benefit to survival, the effect of adjuvant therapy for the patients with resected lung cancer was still controversial. The aim of our study was to evaluate survival advantage of CDDP-based adjuvant therapy compared with short-term immunochemotherapy., Experimental Design: prospective randomized trial., Patients: from 1990 through 1994, 94 patients were registered. Forty-seven patients were randomly assigned to each group, i.e., CDDP-based therapy group (CB Group, CDDP+VDS+tegafur+OK-432 or CDDP+OK-432+mediastinal irradiation) or immunochemotherapy group (IC Group, tegafur+OK-432). PATIENTS in both groups were followed at 4-month intervals with the routine follow-up program of our department., Results: No significant difference was observed between the patients' characteristics of two groups. Compliance of the regimen in each group was 79% in CB Group and 85% in IC Group. No treatment-related death was observed. Five-year survival rates of CB Group and IC Group were 49% and 51%, and 5-year disease-free survival rates were 46% and 44%, respectively. There were no statistical differences between the two groups. Furthermore, no survival differences could be found between CB Group and IC Group in any subgroup of patients., Conclusions: Both of these regimens were feasible. However, we have not observed any survival benefit in the CB Group in any subgroup, so far. Induction therapy, new chemotherapeutic agents, or anti-angiogenetic a agents may improve the survival of surgically treated lung cancer patients.

Biological response modifier antitumor effects are enhanced by the activation of the host defense mechanisms. We have investigated the antitumor effect of photodynamic therapy (PDT) and/or local administration of a biological response modifier, the streptococcal preparation OK-432, on transplanted NR-S1 mouse squamous cell carcinoma. Hematoporphyrin oligomers (20 mg/kg body weight) were used to photosensitize PDT. A pulsed Nd:YAG dye laser, tuned at 630 nm, was used as the light source. The laser power was 15 mJ cm(-2) pulse(-1), and the irradiation time was 40 min. The photosensitizer was injected intraperitoneally 48 h before laser irradiation. Where used, OK-432 was injected into the tumor either 3 h prior to PDT or immediately afterwards. The antitumor effects were evaluated 48 h after each protocol by (a) estimating the area of tumor necrosis (%) in hematoxylin/eosin-stained specimens, and (b) bromodeoxyuridine immunohistochemistry. Furthermore, the tumor sizes were evaluated 3, 7 and 10 days after each protocol, and the survival time after each protocol was evaluated as well. The antitumor effect of PDT was enhanced by administration of OK-432 3 h before PDT, whereas the administration of OK-432 immediately after PDT did not potentiate a PDT antitumor effect. Treatment with OK-432 alone had little effect on tumors. Photodynamic therapy in combination with local administration of OK-432 3 h before PDT is considered to be a useful treatment modality.

Locoregional administration using OK-432 was evaluated in treating malignant effusion. Positive clinical responses were seen in 19 (52%) of 36 gastric cancer patients, and in 9 (90%) of 10 colon cancer patients (p < 0.05), indicating its clinical benefit in treating malignant effusion of colon cancer. Fever elevation was observed in 43 (93%) patients and local pain occurred with 9 (20%) of 46 administrations. Immunological analysis for responder patients with rectal cancer revealed that OK-432 induced autologous tumor-reactive CD 3+ CD 4+ TCRV beta 20+ killer lymphocytes. The TCR gene analysis permitted us to clone a V beta 20 CDR 3 sequence, by which positive bands were shown in 3 (75%) of 4 responders and negative bands in 3 (100%) of 3 non-responders. It is suggested that cross-antigenicity exists between OK-432 and colon cancer, and that genetic analysis using the TCRCDR 3 sequence makes it possible to predict responder patients to OK-432 immunotherapy.

Modulation of interferon-gamma effect by other drug may enhance its tumor specific activity. The apoptosis inducing effect of interferon-gamma and its modulation by cyclosporin-A or tacrolimus (FK-506) were investigated in in vitro and ex vivo experiments. We found that a combination of cyclosporin-A (CsA) and recombinant interferon-gamma (rIFN-gamma) induced significant apoptosis in all four types of human gastric carcinoma cells tested but not in normal cells such as human peripheral blood mononuclear cells (PBMCs), human omentum-derived mesothelial cells, or human umbilical vein endothelial cells (HUVECs) in vitro. Apoptosis was also induced by a combination of rIFN-gamma with FK-506 but not with rapamycin. Next, the apoptosis-inducing effect of endogenous IFN-gamma combined with cyclosporin-A was examined using clinical samples. A streptococcal preparation, OK-432, was administered intraperitoneally for the management of 12 gastric cancer patients with malignant ascites. None of the gascitic fluids obtained before the OK-432 injection showed detectable IFN-gamma level. The OK-432 injection induced a detectable IFN-gamma production ranging from 6 to 89 pg/mL in ascitic fluids from 9 out of the 12 patients. A combination of CsA with ascitic fluids collected after but not before OK-432 injection induced significant apoptosis in MK-1 cells, a gastric carcinoma cell line. A positive correlation was found between the IFN-gamma level and CsA-induced apoptosis. The CsA-induced apoptosis was also blocked by a specific antibody against human IFN-gamma. These results indicated that both recombinant and endogenous IFN-gamma can induce potent tumor-apoptosis when combined with CsA.

Background: Cancer patients have often been reported to have impaired immune function, and the effect of treatment modalities, such as surgery, irradiation, and chemotherapy, in depressing patients' immunity has also been reported. In this investigation, the effect of treatment on the cellular immunity of esophageal cancer patients was evaluated., Methods: Immunological parameters, such as natural killer (NK) activity and lymphocyte subsets in peripheral blood, were measured in 32 esophageal cancer patients on 5 occasions (on the day of admission, 2 days before surgery, and 1 week, 1 month, and 2 months after surgery)., Results: NK activity was greatly impaired shortly after the operation, and the percentages of lymphocytes as a whole, and CD8+, CD16+, and CD57+ lymphocytes were significantly decreased, on the other hand, a postoperative increase in the CD4+/CD8+ ratio was observed. No significant depression of immune function by postoperative irradiation was observed., Conclusions: The results of this study suggest that cellular immunity, especially cytotoxicity, shortly after esophagectomy may be greatly impaired by the surgical stress of esophagectomy and an added effect of chemotherapy.

Background: Obstructive jaundice is frequently associated with septic complications. This study examined the influence of biliary obstruction on bacterial clearance and translocation. The study focused on the phagocytic and killing activities of Kupffer cells and the preventive effect on bacterial translocation of OK-432, which is a hemolytic streptococcal preparation developed as a biological response modifier., Methods: To study the mechanism of sepsis in obstructive jaundice, two groups of Wistar rats were examined: rats subjected to common bile duct ligation (CBDL) and rats subjected to a sham operation. Bacterial clearance, organ distribution, hepatic blood flow, and phagocytic function of Kupffer cells were examined. To evaluate the effect of OK-432 on bacterial translocation, rats were divided into three groups: sham operation + phosphate-buffered saline (PBS), CBDL + PBS, and CBDL + OK-432., Results: In this study, clearance of Escherichia coli. from the peripheral blood in CBDL rats was decreased significantly compared with that in sham-operated rats. Significant decreases in E.coli trapped in the liver and in hepatic blood flow were observed in CBDL rats compared with sham-operated rats. Phagocytic activity and superoxide production of Kupffer cells isolated from CBDL rats were significantly lower than in sham-operated rats. The incidence of bacterial translocation in CBDL rats was increased significantly, and oral administration of OK-432 prevented it., Conclusion: The results suggest that susceptibility to infection in obstructive jaundice is due to impaired phagocytic function of Kupffer cells. Furthermore, obstructive jaundice promotes bacterial translocation, and OK-432 may be useful in preventing this translocation., (Copyright 2000 Academic Press.)

Aim: To treat orbital lymphangioma with an intralesional injection of OK-432 (group A Streptococcus pyogenes of human origin)., Method: A 14 year old boy had a right orbital cystic lymphangioma. The visual acuity in the eye was 20/28. In an initial treatment, 0.02 mg of OK-432, was injected into the tumour after aspiration of the fluid contents, but no effect was seen. The second treatment was performed with 0.04 mg of OK-432., Result: 4 months later, the lesion had totally shrunk to fibrous tissue. The side effects were fever, a local inflammatory reaction lasting 3 days, and increased intraocular pressure, which was managed by draining the fluid contents. Visual acuity improved to 20/15, and the visual field defect and restriction of eye movement seen before treatment disappeared. No recurrence was noted 1 year after treatment., Conclusion: An intralesional injection of OK-432 shrunk the lymphangioma without functional disturbance and scar in the facial skin. OK-432 may be useful for orbital lymphangioma, but further studies are still warranted to determine efficacy, complications, and the optimal dose for safe treatment.

Over a period of seven years, 15 patients (aged from birth to 15 years; median 22 months) with lymphangioma were treated with OK-432; they received a mean of three injections each. Ten received OK-432 as first line treatment; five were treated after surgery (three had a residual lymphangioma after incomplete removal and two had a late recurrence). OK-432 proved to be effective for primitive as well as for residual and recurrent lymphangioma. Seven cases were macrocystic; complete regression was obtained in all. Five cases were microcystic: two had more than 50% regression, and three less than 50%. Three cases were mixed, with both large and microscopic cysts: one had more than 50% regression, and two less than 50%. These last two cases underwent surgery after the sclerosing treatment. The results obtained were excellent in 100% of macrocystic cases; a shrinkage in size was obtained in all microcystic cases. OK-432 is therefore proposed as a first line option for treatment of lymphangiomas.

The treatments previously used for lymphangiomas of the head and neck in children-surgery and intralesional injection of sclerosants-are associated with significant morbidity. A new treatment-intralesional injection of OK-432-was used for lymphangiomas of the head and neck in 11 children. The results were total shrinkage in two, marked shrinkage in two, slight shrinkage in five and no response in two. The results were not affected by previous surgery nor by whether aspiration prior to injection was possible. There were no recurrences in those children in whom shrinkage occurred and no child had subsequent surgery following injection. The results of this series support those of previous series showing that OK-432 injection is an effective and safe treatment for lymphangiomas of the head and neck in children.

Background: OK-432, a lyophilised incubation mixture of group A Streptococcus pyogenes of human origin, was used as a sclerosant for the involution of a giant cervical cystic hygroma in a newborn., Results: There were no systemic side effects. Blood tests and double immune diffusion tests showed no systemic infection or generalised inflammatory response, or antibody production. Cellular and cytokine-induced localised inflammatory reaction within the cystic hygroma, was observed on analysis of the intracystic fluid., Conclusions: The leucocytosis induced and activated by OK-432 probably increases the endothelial permeability of the lymphatics. This probably accelerated lymph drainage leading to involution of the cystic hygroma. Intralesional injection of OK-432 was safe and effective therapy for cystic hygroma in this newborn as its inflammatory reaction was localised., (Copyright 2000 S. Karger AG, Basel.)

We have experienced two cases of unresectable advanced gastric cancer effectively treated by chemo-immunotherapy. One case was of a 68-year-old male patient diagnosed as having inoperable advanced gastric cancer with liver and lung metastasis. This patient was treated by combined chemo-immunotherapy of MMC 10 mg/M, 5'-DFUR 800 mg/day and OK-432 5 KE/2W. At 6 months later, a computed tomography (CT) scan and upper gastrointestinal (GI) series revealed that the metastatic liver tumors and stomach lesion were remarkably decreased in size, and endoscopic biopsy confirmed no cancer cells in the stomach lesion. Moreover, the metastatic lung tumor had disappeared on chest X-ray. The other case was of a 68-year-old female patient with unresectable advanced gastric cancer treated by combined administration of MMC 10 mg/M, 5-FU 200 mg/day and OK-432 5 KE/2W. At 2 months after commencing the treatment, there was a reduction in the serum carcinoembryonal antigen (CEA) level. At 6 months later, the CEA had decreased to normal, the primary and metastatic sites had completely disappeared on CT, and endoscopic biopsy confirmed no cancer cells in the stomach lesion. This patient has survived to date for 5 years and 6 months after commencing the treatment. These results suggested that combined chemo-immunotherapy of MMC, antimetabolite, and OK-432 was an effective treatment for unresectable advanced gastric cancer.

A 67-year-old female patient with biopsy proven AL systemic amyloidosis developed rapidly progressive dyspnea. Chest roentgenogram and CT scan revealed a large right pleural effusion in addition to nodular lesions with bilateral hilar lymphadenopathy. The patient's serum showed IgG lambda type monoclonal gammopathy and she also had Bence Jones proteinuria. The pleural effusion was an exudate that contained many mononuclear cells and a high concentration of protein. Cardiac function was not seriously disturbed. Except for amyloidosis, no other causes for the severe pleural effusion were found. This patient was treated with chemical pleurodesis using Picibanil and a low dose of prednisolone. Eighteen months after this treatment, her right pleural effusion did not recur. Bronchopulmonary tissues are known to be frequently involved by AL systemic amyloidosis, but a nodular pattern of pulmonary amyloid deposition and a unilateral large pleural effusion are rare clinical manifestations in this disease.

Objective: In order to inhibit the occurrence of airborne metastasis of bronchioloalveolar cell carcinoma (BAC), we tried to activate alveolar macrophages by the inhalation of aerosolized OK-432, which is a heat and penicillin-treated lyophilized preparation of the Su strain of Streptococcus pyogenes., Methods and Results: Alveolar macrophages were obtained from resected specimens of lung cancer patients and cultured for 24 h in the presence of various concentrations of OK-432 (0.001-1 KE/ml). The cytotoxic activity against a lung cancer cell line was augmented in a dose dependent manner and reached a plateau level at 0.1 KE/ml of OK-432. Furthermore, the alveolar macrophages produced various cytokines, i.e., IL-1, TNF-alpha, and IL-6 after 72 h cultivation with 0.1 KE/ml of OK-432. Based on the in vitro experiments, six patients with intrapulmonary metastatic BAC were therefore treated by the inhalation of aerosolized OK-432. All 6 patients inhaled aerosolized OK-432 (0.1 KE/ml) twice a day for 5 days. The inhalation therapy regimen was repeated either weekly or monthly unless the tumor markedly progressed. No adverse events were observed in any patients. Either an augmentation of antitumor cytotoxicity or TNF-alpha production by the alveolar macrophages was observed in the two of three patients examined., Conclusion: OK-432 inhalation therapy was found to be safe and thus warrants further investigation to determine its clinical effectiveness for BAC.

Picibanil (OK-432) is a sclerosing agent derived from a low-virulence strain of Streptococcus pyogenes that induces regression of macrocystic lymphangiomas. This report describes a prospective, nonrandomized trial to evaluate the efficacy of Picibanil in the treatment of 13 affected children ranging in age from 1 to 94 months. On average, 4.1 fluoroscopically guided intracystic injections were performed per child, with an average total dose of 0.56 mg of Picibanil. As judged by physical examination and radiographic studies, 5 children (42%) showed a complete or substantial response, and 2 children (16%) showed an intermediate response. No response was seen in 5 children (42%), 2 of whom had massive craniofacial lymphangioma. Factors that contribute to failure with Picibanil sclerotherapy are the presence of a significant microcystic component to the lesion, massive craniofacial involvement, and previous surgical resection. Macrocystic lymphangiomas of the infratemporal fossa or cervical area have the best response to therapy.

New approaches to the treatment of head and neck cancer involve delivery of the therapeutic agent by direct injection into the tumor lesions. This locoregional therapy allows adaptation of conventional chemotherapy agents, such as bleomycin and cisplatin, to achieve higher drug concentrations in the tumor while avoiding severe systemic toxicities. Novel therapies administered through tumor injection, such as gene transfer, photosensitization, and biologic response modification, have been investigated in preclinical studies and are currently in different phases of clinical trial.

After approval of national health insurance for non-specific immunomodulation such as OK-432 (1978) and PSK (1980), lentinan, SPG, bestatin and dried BCG vaccine have been tried. Including cytokines such as IL-2, IL-12, IFN, TNF or monoclonal antibodies, they have been widely used as biological response modifiers (BRM). Progress in BRM therapy may be broken down into the first 10 years as development, the next 10 years as disappointment and the most recent 5 years as dream-like progress owing to molecular biological techniques. An interdisciplinary approach has been taken by the Japanese Research Society for Surgical Cancer Immunology, founded in 1980, and the Japanese Society of BRM founded in 1988. Many investigations have been performed on issues such as the clinical evaluation or criteria for responder cases, host immunocompetency, post-operative adjuvant immuno-chemotherapy, locoregional immunotherapy, cytokine therapy, adoptive immunotherapy, and tumor specific immunotherapy. Attention has also focused on malignant tumor injury, surgical stress, the advantages or disadvantages of splenectomy, and discussions of the current status and future prospects in the next century for new BRM therapy.

The current prospective randomized study was designed to compare the effects of intracavitary (i.c.) chemotherapy vs immunotherapy vs immunochemotherapy for malignant effusion. Between 1992 and 1995, a total of 42 patients with malignant effusion were registered, and 41 patients were eligible for statistical analysis. The primary diseases of the eligible patients included 27 gastric, four colorectal, four pancreatic, three lung, two liver and one oesophageal cancers. The patients with malignant effusion were randomly assigned into one of three i.c. therapeutic regimens: chemotherapy alone with weekly injection of anticancer agents (ACAs: cisplatin, mitomycin-C, adriamycin, etc.) (Group A, n = 13); immunotherapy alone with weekly injection of streptococcal preparation OK-432 (Group B, n = 14); or immunochemotherapy with ACAs and OK-432 (Group C, n = 14). The response of the effusion, patient survival and the kinetics of cytokines in the effusion were compared. There were no differences in the patients' backgrounds. The side-effects of the regimens included pain, anorexia, fever, leucopenia and anaemia and there were no differences in their incidence among the three groups. One patient died after cisplatin (CDDP) administration in Group A. Cytologic examination revealed that tumour cells in the effusion disappeared in 23% of Group A cases, 36% of Group B cases and 36% of Group C cases. The malignant effusion did not disappear in any of the Group A cases; however, the effusion disappeared in 29% of Group B cases and 43% of Group C cases (P = 0.03, Group A vs Group C). Furthermore, the 50% survival period was 1.6 months for Group A, 2.4 months for Group B and 3.5 months for Group C. The 6-month survival rate was 7% for Group A, 6% for Group B and 34% for Group C, and the 1-year survival rate was 0%, 0% and 17% respectively (P = 0.048, Group A vs Group C by the log-rank test). The analysis of the cytokine kinetics revealed a prominent increase in the level of interleukin-6 in the effusion in Group C. These results suggest that i.c. immunochemotherapy with OK-432 and ACAs may be more beneficial than i.c. chemotherapy alone or immunotherapy alone.

To determine the significance of interleukin (IL)-10 in antitumor immune response, the effect of the down-regulation of tumor-derived IL-10 on locoregional immunotherapy was investigated. C3H/HeN mice were intraperitoneally (i.p.) inoculated with IL-10-producing murine breast cancer cell line, FM3A, and treated with locoregional administration of OK-432 with or without anti-IL-10 monoclonal antibody (mAb). Anti-IL-10 mAb did not affect the in vitro growth of FM3A cells. Administration of OK-432 plus anti-IL-10 mAb remarkably delayed the retention of malignant ascites and prolonged the survival of mice compared with the administration of OK-432 alone. Spleen cells which were collected from mice treated with OK-432 plus anti-IL-10 mAb and further stimulated in vitro with inactivated FM3A cells exhibited significantly higher cytotoxicity against FM3A cells than those from mice treated with OK-432 alone or from the control mice. The expression of major histocompatibility complex (MHC) class II molecules on spleen cells was up-regulated in vitro by the addition of OK-432 and anti-IL-10 mAb. Using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR), cytokine mRNA levels of peritoneal exudate cells (PEC) and spleen cells were assessed on day 7 (before treatment) and day 14 (after treatment). In PEC, increased expression of IL-2 was observed with the administration of OK-432 plus anti-IL-10 mAb. In spleen cells, the expression of IL-2, IL-12 and IFN-gamma were strongly induced, and IL-4 expression was reduced by the administration of OK-432 plus anti-IL-10 mAb. It is suggested that down-regulation of tumor-derived IL-10 induces the up-regulation of the T helper type (Th) 1 population, resulting in an enhancement of the efficacy of locoregional immunotherapy with OK-432.

Chylothorax is an uncommon but well recognized complication of esophagectomy. We present the case of a 57 year-old man with squamous cell carcinoma of the abdominal esophagus who underwent subtotal esophagectomy by right thoracotomy. Post-operatively, the volume of pleural effusion from the right chest was increased (1600-2000 ml/day). The effusion was straw colored, not changing to milky after meals. The characteristics and composition of the pleural fluid were similar to those of chyle. We therefore treated this patient using methods for treatment of chylothorax, conservatively, by administration of OK-432 and minocycline/hydrochloride into the pleural cavity from the chest tube with success. We discuss the pathophysiology of this unusual condition and its treatment.

We report herein the case of a woman with bronchopleural fistulas treated with the endobronchial placement of vascular embolization coils. She was referred to our hospital to undergo lavage of a postoperative empyema. She had undergone an air plombage operation for pulmonary tuberculosis 9 years previously. However, bronchopleural fistulas occurred postoperatively and she had to continue the use of a chest drainage tube since then. Lavage of her empyema space with 5kE of OK-432 (Picibanil: Chugai) plus 100 mg minocycline was performed once every 2 weeks for 3 months, and the purulent discharge from the empyema remarkably decreased. Thereafter, the bronchopleural fistulas were occluded endobronchially by the placement of vascular embolization coils. Soon after the procedure, air leakage from the fistulas was stopped and the drainage tube was removed 2 days later. The patient remains well without any additional treatment at 20 months after this treatment. As treatment for empyema with bronchopleural fistulas, it would be worth trying to lavage the empyema space with OK-432 until it is cleaned out and to plug the fistulas by the endobronchial placement of embolization coils, before such radical operations as thoracoplasty and space-filling of the empyema are considered.

Bacterial endotoxin, lipopolysaccharide (LPS), is a causative agent of Gram-negative septic shock. However, if preadministered at a low dose, LPS makes mice resistant to subsequent endotoxin challenge, the phenomenon known as LPS tolerance. Here we demonstrated that the pharmaceutical preparation of Gram-positive Streptococcus pyogenes, OK-432, also induced a state analogous to LPS tolerance if administered 6-48 h prior to LPS challenge. The preadministration of OK-432 increased the lethal dose of LPS threefold in BDF1 mice, and this was accompanied by reduced gene expression of IL-6, IFN-gamma, inducible nitric oxide synthase, and IL-10 in spleen and peritoneal cells. Serum concentrations of IL-6 and IFN-gamma were also suppressed by the preadministration of OK-432. In contrast to the LPS tolerance, the levels of TNF-alpha mRNA were not suppressed in OK-432-administered mice, and their peritoneal cells produced high levels of TNF-alpha and soluble TNF receptor p75 in response to LPS in vitro. Peritoneal cells from OK-432 but not LPS-administered mice were hyporesponsive to IFN-gamma in terms of nitric oxide synthesis, and this hyporesponsiveness to IFN-gamma was abrogated by anti-IL-10 antibodies. Likewise, peritoneal cells from both OK-432- and LPS-administered mice were hyporesponsive to LPS, serum, TNF-alpha, IFN-gamma, and PMA in terms of IL-6 production. Anti-IL-10 antibodies increased IL-6 production eightfold in cells from OK-432-administered mice, but marginally in cells from LPS-administered mice. Even in peritoneal cells from OK-432-administered mice, anti-IL-10 antibodies failed to fully restore IL-6 production. Thus, the hyporesponsive state of peritoneal cells was mediated by both IL-10-dependent and -independent mechanisms. These results demonstrated that OK-432 controlled endotoxin shock by blocking the cytokine cascade from TNF-alpha., (Copyright 1998 Academic Press.)

For the past 20 years, our group has treated over 400 cases of malignant effusion by the intraperitoneal injection of streptococcal preparation OK432 (OK-432 therapy) and has investigated extensively the antitumor mechanisms of this therapy. Prospective clinical data has demonstrated that the OK-432 therapy induced a definite reduction of the effusions in around 60% (responders) of cases and significantly prolonged the survival time in patients who responded well. In addition, a definite reduction of original tumor mass volume was found in around 20% of cases. We have shown that OK432-induced neutrophils, lymphocytes, and probably macrophages may play an important role in tumor cell destruction in ascites. Tumor necrosis factor alpha (TNF-alpha)-induced CD11b/CD18 expression on leukocytes and interferon-gamma (IFN-gamma)-induced ICAM-1 expression on tumor cells may play an important role in leukocyte-mediated tumor destruction. It has also been shown that OK-432 induces various cytokines, such as TNF-alpha, TNF-beta, IFN-alpha IFN-gamma, interleukin-1 (IL-1), IL-2, IL-6, IL-12, tumor growth inhibitory factor(s) (TGIF), and possibly unknown apoptosis-inducing factor(s). Some of these cytokines have been adduced as representing the antitumor activity. These data suggest that two pathways of antitumor activity, i.e., cell-mediated and cytokine-mediated, can be induced simultaneously in the peritoneal cavity. OK-432 therapy may be valuable in the management of patients with malignant effusions. Future clinical and basic research should contribute to further progress in OK-432 therapy.

We have adopted OK-432 as a sclerosing agent in the treatment of cystic predominant thyroid nodules and have analyzed our findings to assess the efficacy of intralesional instillation of OK-432. From 1992 through 1993, 48 patients with recurrent or progressive cystic thyroid nodules after 2 or 3 aspirations alone, and whom were cytologically negative for malignancy, were used for this study. The OK-432 solution was prepared by dissolving 0.1 mg of OK-432 in 2 ml of physiologic saline and it was instilled in the amount of 1/10-to-1/5 of the aspirated cystic fluid. A repeated course of therapy was given up to 3 times when sufficient resolution was not obtained 4-to-6 weeks after treatment. The mean number of treatment sessions per patient was 1.5. Throughout the follow-up period from 30-to-45 months (mean, 38 months), 32 (66.7%) patients experienced an almost complete disappearance (< 0.05 cm in diameter) of the cystic lesion, and 12 (25%) patients responded partially by having it decrease by more than half (> 0.5 cm in diameter) of the initial cyst size, and none of these patients required further treatment. The remaining 4 (8.3%) patients showed insufficient resolution despite 3 courses of therapy and 2 of these patients underwent thyroidectomy, in which the lesion proved benign in both cases. All of the patients tolerated the sclerotherapy well. No significant local complications attributed to this treatment were observed. However, a low-grade fever was observed in 26 (54.2%) patients for 2 to 5 days after instillation, which subsided with symptomatic treatment. On the basis of our experience, OK-432 sclerotherapy appears to be safe, simple and effective, and can be a useful alternative treatment for cystic thyroid nodules.

Hyperthermia is being investigated as a cancer treatment. Many of its basic mechanisms, particularly those related to cell killing, are still poorly understood. We used a transplanted squamous cell carcinoma cell line to investigate the therapeutic effect of hyperthermia. In particular, we examined the effect of OK-432 (biological response modifier) on hyperthermia-induced apoptosis. In the hyperthermia only group the most extensive necrosis occurred on day 3 (70.3%), and the apoptosis index also was highest on that day (69.3). These results suggest that the induction of apoptosis is closely related to the cell death caused by hyperthermia. The percent of necrosis was significantly higher in the groups given hyperthermia and combined OK-432 and hyperthermia treatment than in the OK-432 group (p < 0.05). The apoptosis index was significantly lower in the combined OK-432 and hyperthermia treatment group than in the hyperthermia only group, indicative that the antitumor effect of combined hyperthermia and OK-432 therapy is not ascribable to the induction of apoptosis.

We have previously reported that intratumoral injection of a mixture of OK-432 and fibrinogen (OK/fbg) is very effective for local immunotherapy of colorectal cancer, but has little influence on breast cancer, while addition of activated macrophages to OK/fbg (OK/fbg/mo) has a marked effect on breast cancer. In this study, we analyzed the role of urokinase-type plasminogen activator (uPA) in local immunotherapy. We found that uPA levels in breast cancer tissue were lower than in colorectal cancer tissue. Injection of OK-432 into breast cancers neither increased the uPA level nor caused tumor regression. Although OK/fbg injection caused an increase of uPA, tumor regression was restricted to the area around the injection site. With OK/fbg/mo however, uPA levels increased much more markedly and extensive tumor necrosis was observed in all cases. These findings suggest that uPA plays an important role in tumor regression during local immunotherapy.

Considerable controversy surrounds the management of gastric cancer and this has largely overshadowed recent progress in our understanding of the epidemiology and molecular pathogenesis of the disease, and improvements in diagnostic and staging techniques. Differences identifiable in the molecular pathogenesis of the 'intestinal' and 'diffuse' types of gastric cancer may help to unravel the biological behaviour of variants and ultimately influence therapeutic strategies. Endoscopic ultrasound is well established as being accurate for T staging and the introduction of laparoscopy, with or without ultrasound, is obviating unnecessary laparotomy in non-bleeding, non-obstructed patients. Controversies in surgery encompass the role of laparoscopic surgery in early gastric cancer, the extent of lymphadenectomy including para-aortic nodal dissection, resection of en bloc contiguous organ involvement, pancreatosplenectomy, left upper abdominal evisceration, and modes of reconstruction (pylorus-preserving gastrectomy, pouch formation) to enhance quality of life. Whereas adjuvant chemotherapy does not impact favourably on survival, emphasis has now shifted to neoadjuvant (induction) chemotherapy to downstage the disease. Preoperative regional chemotherapy and intra-operative hyperthermic chemotherapy or irradiation may prove to be of benefit in patients with resectable disease, but some scepticism still exists as to the usefulness of biological response modifiers (e.g. OK432, PSK) for adjuvant treatment. Ethical issues relating to cultural differences in Asia sometimes mitigate against adequate trial design (e.g. a surgery-alone control group or a no adjuvant therapy treatment group may be considered inappropriate) and this has understandably hindered acceptance in Western countries of the value of current management practices in Asia. These issues and the need for ongoing well-conducted randomized trials with prospective subset analysis are now being addressed.

OK-432, a killed preparation of Streptococcus pyogenes, as well as Bacillus Calmette-Guerin (BCG) and Corynebacterium parvum are all known biological response modifiers. To examine the immunomodulatory effects of OK-432, natural killer cell activity and cytokine production by peripheral blood mononuclear cells (PBMCs) were assessed in 32 patients with gastric cancer. Skin tests for Streptococcus pyogenes A-3Su (Su-PS) and BCG were performed in all patients. Other nutritional and immunological parameters were also determined. OK-432-treated PBMCs showed a significant increase of cytotoxicity against K562 cells (p < 0.01). Increased levels of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha) and interleukin-2 (IL-2) were found in the supernatants of cultures treated with OK-432 in 29 (90.6%), 20 (62.5%), and 8 (25.0%) out of 32 patients, respectively. Natural killer cell activity, IFN-gamma production, and the Su-PS skin test were positively correlated (p < 0.01). In contrast, the BCG test and other markers were not correlated with natural killer cell activity and IFN-gamma production. These results suggest that the Su-PS skin test could predict OK-432-induced natural killer cell activity and IFN-gamma production in patients with gastric cancer, and was therefore useful to determine whether patients were responders to OK-432.

Severe osteolysis is a rare complication of lymphangioma. It can progress despite treatment and may be fatal. Four patients presented with massive osteolysis and serious related complications. Of these, two responded well to OK-432 therapy with arrest of the osteolysis. Complete recovery from life-threatening pleural effusion has been noted in one. Of the two nonresponders, one died of progressive osteolysis of the cervical and occipital bones, the other, despite therapy, still shows progressive osteolysis of the left leg from the femur to the phalanges. A boneless leg may be the outcome.

The mean survival rates of female BDF1 mice transplanted intravenously (i.v.) with murine L1210 leukemia cells were significantly prolonged by intraperitoneal (i.p.) pretreatment (before i.v. transplantation) or by i.p. pre- and post-treatment (before and after the i.v. transplantation) with heat-killed Actinobacillus suis cells ATCC 15557 (AS 15557) alone, as compared with untreated L1210-leukemia-cell-bearing control mice. However, significant prolongation of the mean survival rates was not elicited by the i.p. post-treatment with AS 15557 alone. When 5-fluorouracil (5-FU) was applied i.p. to mice receiving post-treatment with AS 15557 alone, the mean survival rates of the L1210-leukemia-cell-bearing mice were significantly prolonged. The antileukemic action of AS 15557, alone or in combination with 5-FU, against L1210 leukemia was superior to that of a streptococcal preparation (OK-432) and was almost the same as of bacillus Calmette-Guérin with or without 5-FU. The results suggest the possibility that the synergism of AS 15557 in combination with 5-FU may be dependent on the relationship between the indirect immunological function of AS 15557 and the direct cytotoxic action of 5-FU on L1210 leukemia cells.

Background: The purpose of the study was to verify whether OK-432 in combination with 5'-DFUR induced thymidine phosphorylase (TdR Pase) and cytokines in gastric cancer patients as well as in vitro., Materials and Methods: Fifty patients with invasive gastric cancer were randomly assigned, upon admission using by a closed-envelope technique, to either a group receiving 5'DFUR or OK-432 alone, to a group receiving both 5'DFUR and OK-432, or to a non- treated group up. Surgical specimens of the tumor and normal tissues were taken soon after gastrectomy to evaluate TdR Pase activity, IL-1 alpha and TNF-production., Results: TdR Pase activities were several times higher in tumor than in normal tissues. In normal tissues, TdR Pase activities in the 5'-DFUR + OK-432 group were significantly higher than in the OK-432 group. TdR Pase activity in tumors, however, showed no significant difference between treated group. In the 5'-DFUR + OK-432 group, the level of IL-1 alpha production in tumor was significantly higher compared to the control group. In the 5'-DFUR + OK-432 group, the level of TNF alpha production in tumor was significantly higher than in normal tissue. TNF alpha production in tumor showed no significant difference in each treated group compared to the control. There was a significant correlation between TdR Pase activity and IL-1 alpha production levels in tumor., Conclusions: TdR Pase was induced by IL-1 alpha in tumor tissues of gastric cancer patients. OK-432 in combination with 5'-DFUR, however, did not induce TNF alpha and IL-1 alpha, and increase TdR Pase activity in gastric cancer tumors.

Twelve patients with warts recalcitrant to various treatment, including cryotherapy, were treated with OK-432 injection therapy. Six patients received only subcutaneous injection, three received only intralesional injection, and the other three received both subcutaneous and intralesional injections. Complete clearing of the warts occurred in nine (75%) of 12 patients, while the other three patients were not cured. Grouping the patients by the method of injection, the success rate of subcutaneous injection was 5/9 (55.6%), and that of intralesional injection was 4/6 (66.7%). Although five patients complained of mild fever and malaise, these side effects gradually disappeared during the repeated injections. OK-432 injection is considered as a hopeful therapy for recalcitrant warts.

We investigated whether the combination treatment of 15 cGy total body irradiation (TBI) and a streptococcal preparation, OK-432, synergistically suppresses spontaneous lung metastasis and augments phytohemagglutinin (PHA) and concanavalin A (Con A) responses of splenocytes in WHT/Ht mice. TBI with 15 cGy was carried out 20 days after subcutaneous injection of squamous cell carcinoma cells into a hind leg. Lung colony number was counted 40 days after tumor injection. For PHA and Con A responses, mice were killed 4 hr after 15 cGy TBI. The combination treatment of 15 cGy TBI and OK-432 was most effective when OK-432 was administered 2 days before 15 cGy TBI. The combination treatment decreased the lung colony number to 29.9% of the control number. OK-432 slightly increased the PHA and Con A responses, and 15 cGy TBI did not increase them. However, when these two were combined, the PHA and Con A responses were significantly increased to 393% and 278% of the control levels, respectively. It was suggested that TBI and OK-432 acted synergistically in suppressing the lung metastasis and mitogenic response of splenocytes.

We previously reported that the antitumor effect of OK-432, a streptococcal preparation, was markedly augmented when this agent was injected into tumors together with fibrinogen. In order to elucidate the effect of this treatment on the spleen, we assessed splenic function in gastric cancer patients receiving preoperative local immunotherapy with OK-432 and fibrinogen. Immunohistochemical studies of the spleen at 7 days after intratumoral injection therapy revealed numerous macrophages phagocytizing OK-432 in the splenic sinuses. Phenotypic analysis of splenocytes by flow cytometry revealed an increase in the CD4/CD8 ratio and in the expression of HLA-DR, CD25, and Leu M3 by splenic T cells of the patients treated with OK-432 plus fibrinogen when compared to patients treated with OK-432 alone or untreated patients. Splenic T cells from patients treated with OK-432 plus fibrinogen showed significantly higher cytotoxicity against Daudi and K562 cells than T cells from control patients (p < 0.05), and culture of these splenic T cells with recombinant IL-2 induced the expansion of lymphokine-activated killer cells. These results demonstrate that local immunotherapy with a mixture of OK-432 and fibrinogen effectively augumented splenic antitumor immunity in gastric cancer patients.

Epithelioid haemangioendothelioma is an unusual type of endothelium-derived vascular tumour of borderline malignancy, which has high variability in clinical presentations, depending on the primary site of involvement. We report on a 20-year-old woman who presented with progressive abdominal fullness for 6 months. Multiple lung and liver nodules with pleural effusion and profuse ascites were found. The diagnosis of epithelioid haemangioendothelioma was made after wedge biopsy of the liver. The ascites was intractable and refractory to strong diuretic therapy and repeated paracentesis. Therefore, six courses of intraperitoneal injection of OK-432 were administered. The ascites subsided to a minimal amount after treatment and the patient remained symptom-free for approximately 8 months. The ascites recurred later and another three courses of intraperitoneal injection of OK-432 were administered. The ascites disappeared again. The patient has remained symptom-free since the end of the second period of treatment.