picibanil and Fibrosarcoma

A 72-year-old man was referred to our hospital because of bilateral pleural effusion. Although examination of pleural effusion obtained by thoracentesis did not show any specific etiology, we diagnosed yellow nail syndrome due to his yellow nails and lymphedema of both lower limbs. Diuretics were effective for the control of his pleural effusion. Subsequently, fibrosarcoma was found in his abdominal skin and was resected. The pleural effusion gradually increased after the cessation of oral diuretics. Histological examination of a pleural biopsy specimen obtained by thoracoscopy showed chronic lymphocytic inflammation, but no malignancy. His previously intractable right pleural effusion was successfully treated with pleurodesis using OK-432, suggesting that pleurodesis with OK-432 could be an effective method for the control of pleural fluid in this disease.

Topics: Aged; Fibrosarcoma; Humans; Male; Picibanil; Pleurodesis; Skin Neoplasms; Yellow Nail Syndrome

We isolated the lipoteichoic-acid-related molecule (OK-PSA) from OK-432, a streptococcal preparation, by affinity chromatography on CNBr-activated Sepharose-4B-bound monoclonal antibody TS-2, which neutralizes the interferon (IFN)-gamma-inducing activity of OK-432. We have previously reported that OK-PSA is a potent inducer of Th1-type cytokines in human peripheral blood mononuclear cells in vitro. In this study, we conducted an animal experiment to examine whether OK-PSA exhibits an anti-tumor effect in vivo by acting as a Th1 inducer in syngeneic Meth-A tumor-bearing BALB/c mice, in which the Th2 response is genetically dominant. It was found that OK-PSA induced Th1-type cytokines [IFN-gamma, tumor necrosis factor-alpha, interleukin (IL)-2, IL-12 and IL-18] in BALB/c mice bearing Meth-A tumor and caused a marked anti-tumor effect. Although it was suggested by an in vitro study. using spleen cells derived from the animals, that IL-18 plays the greatest role in the induction of the Th1-dominant state and tumor cell killing induced by OK-PSA, the in vivo experiments demonstrated that both IL-12 and IL-18 are essential in the anti-tumor effect exhibited by OK-PSA. These findings strongly suggest that OK-PSA is a major effector molecule of OK-432 and may be a useful immunotherapeutic agent, as a potent Th1 inducer, for cancer patients with a Th2-dominant state.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Monoclonal; Antineoplastic Agents; Apoptosis; Chromatography, Affinity; Drug Screening Assays, Antitumor; Fas Ligand Protein; fas Receptor; Female; Fibrosarcoma; Interleukin-12; Interleukin-18; Killer Cells, Natural; Lipopolysaccharides; Lymphokines; Lymphoma; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Moloney murine leukemia virus; Neoplasm Transplantation; Penicillin G; Perforin; Picibanil; Pore Forming Cytotoxic Proteins; Spleen; Streptococcus pyogenes; Teichoic Acids; Th1 Cells; Th2 Cells; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

It was previously demonstrated that a single injection of OK-432 (a penicillin-treated freeze-dried Streptococcus) mixed with fibrinogen into cancer tissues induces marked infiltration by eosinophils of the tumor stroma and leads to tumor necrosis. In the present study, we examined mechanisms regulating the local accumulation of eosinophils and the role of infiltrating eosinophils in tumor regression using the OK-432/fibrinogen injected Meth-A fibrosarcoma tumor. After injection of OK-432/fibrinogen into the tumor on the left flank of the BALB/c mice, eosinophil infiltration became obvious in the tumor stroma on day 3 following the accumulation of macrophages and neutrophils, was massive on day 5 and decreased by day 10. After the decrease in the infiltration of eosinophils, the tumor injected with OK-432/fibrinogen diminished markedly in size with ulceration as compared with control. Northern blot analysis revealed that expression of IL-5 mRNA in the tumor tissue was not detected on day 0, was significantly on day 3, reached the maximum on day 5, and thereafter decreased by day 10. Although intraperitoneal injection of rat anti-IL-5 monoclonal antibody in tumor bearing mice prior to OK-432 injection inhibited the infiltration of eosinophils, the antitumor effects of OK-432 persisted. In the blood, neither eosinophilia nor IL-5 activity was recognized during the course of the experiment. These results suggest that intratumoral injection of OK-432/fibrinogen induces local production of IL-5, which in turn recruits eosinophils into the tumor tissue, however, the infiltrating eosinophils do not play an important role in tumor regression.

Topics: Animals; Antibodies, Monoclonal; Antineoplastic Agents; Eosinophils; Fibrinogen; Fibrosarcoma; Interleukin-5; Male; Mice; Mice, Inbred BALB C; Picibanil; Rats; RNA, Messenger

Effective treatment of a rat transplanted ascites tumor by i.p. injection of a streptococcal biological response modifier, OK-432, was abrogated by selective in vivo depletion of neutrophils by a monoclonal antibody, RP-3. The mechanisms by which neutrophils participate in the therapeutic action of OK-432 were studied with Winn's assay using peritoneal exudate cells periodically obtained from rats i.p. injected with this biological response modifier. Intraperitoneal resident macrophages were first activated with OK-432, and within 3 h, tumor-inhibitory activity had moved to the early exuded neutrophils. However, 6 h after injection, exuded macrophages were the only cells involved in tumor inhibition. Considered together with other findings, it is likely that, in this system, neutrophils may transmit information from resident macrophages to exuded inflammatory macrophages in a series of responses induced by i.p. injection of OK-432.

Topics: Animals; Antibodies, Monoclonal; Antineoplastic Agents; Fibrosarcoma; Immunologic Factors; Macrophages, Peritoneal; Neoplasm Transplantation; Neutrophils; Picibanil; Rats; Rats, Inbred Strains; Tumor Cells, Cultured

The antitumor effects of biological response modifiers (BRM) in a new experimental mouse model, the "double grafted tumor system", were analysed. BALB/c mice received simultaneous inoculations of Meth-A fibrosarcoma cells on right flank (10(6) cells) and left flank (2 x 10(5) cells) on day 0, and BRMs were injected intratumorally into right tumor on day 3, 4 and 5. The growth of the left-flank tumor was the real target for the evaluation of a given drug after 21 days. PSK (a protein-bound polysaccharide preparation), IL-1 and Cepharanthin, cured not only the right, but also the left, non-treated tumor in a double grafted tumor system. OK-432 (a Streptococcus preparation) and BCG cured the right tumor and inhibited the growth of the left tumor. Lentinan (a polysaccharide preparation) inhibited neither the right nor the left tumor. Immunosuppressive acidic protein (IAP) in serum was increased transiently soon after intradermal injection of PSK, OK-432 and TNF in BALB/c mice. But Lentinan did not induce IAP. IAP in serum was gradually increased after intradermal inoculation of Meth-A tumor in BALB/c mice. At 21 days after tumor inoculation, IAP in serum reached a maximum level (300 micrograms/ml). The serum IAP level of Meth-A-bearing mice as well as that of normal mice increased after the intratumoral injection of PSK. At 21 days after tumor inoculation, IAP in PSK-treated mice returned to normal level. The biochemical differences between PSK-induced IAP (early, inflammatory IAP) and Meth-A-induced IAP (late, tumor-induced IAP) was investigated by crossed immunoaffino electrophoresis (CIAE). Inflammatory IAP was rich in biantennary sugar chain, and tumor-induced IAP was rich in tri-tetraantennary sugar chain.

Topics: Animals; Fibrosarcoma; Immunologic Factors; Injections, Intralesional; Macrophages; Male; Mice; Mice, Inbred BALB C; Neoplasm Proteins; Neoplasm Transplantation; Picibanil; Proteoglycans

The antitumor effect of oenothein B, a macrocyclic ellagitannin from Oenothera erythrosepala Bordas, on rodent tumors was studied. Oenothein B exhibited a strong antitumor activity against MM2 ascites tumors upon intraperitoneal administration to the mice before or after the tumor inoculation. The tannin also inhibited the growth of Meth-A solid type tumor in mice. This antitumor effect of the tannin could not be attributed to its direct cytotoxic action on tumor cells, because the cytotoxicity was very weak in the presence of serum protein. When oenothein B was injected into the peritoneal cavity of mice, peritoneal exudate cells, including cytostatic macrophages, were induced. Furthermore, in the in vitro treatment of macrophages from mice and humans, the tannin stimulated release of an interleukin 1 (IL-1)-like activity and IL-1 beta from the cells. These results suggest that oenothein B exerts its antitumor effect through potentiation of the host-immune defense via activation of macrophages.

Topics: Animals; Antineoplastic Agents, Phytogenic; Cell Survival; Female; Fibrosarcoma; Hydrolyzable Tannins; Mammary Neoplasms, Experimental; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Picibanil; Sarcoma, Experimental; Tannins; Tumor Cells, Cultured

The aim of this study was to investigate the influence of oral administration of OK-432 on the tumor growth of tumor-bearing mice. In addition, the changing pattern of the splenic lymphocyte subsets of tumor-bearing mice was evaluated by flow cytometry. OK-432 at a dose of 0.1, 1 or 10 KE was administered orally every 3 days or every other day for 30 days to subcutaneously Meth A tumor-inoculated mice. The tumor growth was significantly inhibited in the 1 KE every 3 days group, in the 1 KE every other day group and in the 10 KE every 3 days group. In the 10 KE every other day group, OK-432 inhibited the tumor growth on days 10 and 20, while the agent did not show a marked inhibitory effect on day 30. The percentages of splenic L3T4-positive cells and splenic asialo GM1-positive cells were significantly increased in the 1 KE every other day group, while the Lyt2+/Thy1.2+ ratio was decreased. On the other hand, in the 10 KE every other day group, OK-432 showed no effect on the percentages of splenic L3T4-positive cells and Lyt2+/Thy1.2+ ratio on days 20 and 30. Our results suggest that the antitumor effect of oral administration of OK-432 may be correlated with the changing pattern of L3T4-positive cells and Lyt2+/Thy1.2+ ratio.

Topics: Administration, Oral; Animals; Antigens, Ly; Antigens, Surface; Antineoplastic Agents; Fibrosarcoma; Flow Cytometry; G(M1) Ganglioside; Lymphocyte Subsets; Male; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Picibanil; Spleen; Thy-1 Antigens

The antitumor effects of biological response modifiers (BRM) in a new experimental mouse model, the "double grafted tumor system", were analysed. BALB/c mice received simultaneous inoculations of Meth-A fibrosarcoma cells on right flank (10(6) cells) and left flank (2 x 10(5) cells) on day 0, and BRMs were injected intratumorally into right tumor on day 3, 4 and 5. The growth of the left-flank tumor was the real target for the evaluation of a given drug after 21 days. PSK (a protein-bound polysaccharide preparation), IL-1 and Cepharanthin cured not only the right, but also the left, non-treated tumor in a double grafted tumor system. OK-432 (a Streptococcus preparation) and BCG cured the right tumor and inhibited the growth of the left tumor. Lentinan (a polysaccharide preparation) inhibited neither the right nor the left tumor. Spleen cells from PSK-treated tumor bearing mice produced macrophage chemotactic factor (MCF) after 48 hrs cultivation in the presence of Con A or Meth-A tumor cells. MCF producing cells were indicated to be L3T4 positive cells. On the other hand, PMN activated by PSK treatment produced MCF in the culture supernatant. Therefore, our present and previous studies on the antitumor effect of BRM in the double grafted tumor system show that intratumoral administration of BRM first induces neurophils in the right tumor via an IL-8-like factor and then cytotoxic macrophages are induced by MCF. Then Lyt-1 (L3T4)-positive cells are induced in the right regional lymph nodes and in the spleen, probably via IL-1, which might be produced from macrophages in contact with tumor cells. Subsequently, Lyt-1-positive cells reach the left tumor through the blood stream, come into contact with Meth-A tumors and then produce MCF. Intratumoral administration of PSK in the right tumor thus induces cytotoxic macrophages in the left, non-treated tumor, thereby bringing about the regression of the distant tumor.

Topics: Animals; Fibrosarcoma; Immunologic Factors; Injections, Intralesional; Male; Mice; Mice, Inbred BALB C; Neoplasm Transplantation; Picibanil; Proteoglycans

OK-432, a streptococcal preparation with potent biological response modifying activities, had the ability to cure mice bearing BAMC-1 (fibrosarcoma) ascites when it was injected intraperitoneally five times, 2, 4, 6, 8, and 10 days after the tumor inoculation. Previously, it was shown that the OK-432 injection on day 2 was indispensable since only a minimal antitumor effect was obtained when an inflammation-inducing agent such as thioglycolate instead of OK-432 was injected on day 2, followed by four OK-432 injections on days 4, 6, 8 and 10. In the present study, the injection of OK-432 on day 2 and a subsequent injection of either IL-2 or IFN-gamma on day 4 or 6 showed a significant antitumor effect, achieving a complete cure in approximately 50% of mice treated, although none of the mice could be cured by a single injection of either OK-432, IL-2, or IFN-gamma on day 2. Interestingly, however, the mice treated with an injection of a lymphokine (IL-2 or IFN-gamma) on day 2 followed by OK-432 on day 4 were not cured either. Peritoneal cells on day 12 in mice treated with OK-432 and either of the lymphokines contained pantropic killer cells, which were Thy-1+ and asialo GM1+ (AsGM1+). Moreover, the antitumor effect of the combined treatment was abolished when mice were pre-treated with anti-AsGM1. No significant antitumor effect was observed in athymic nu/nu mice. Together with our previous findings, these results indicate that lymphokines induced by OK-432 administration may account for some of its therapeutic effects and that these lymphokines may also facilitate the subsequent induction of specific killer cells. These results warrant further investigation on possible effective therapeutic protocols with the combined use of OK-432 and lymphokines.

Topics: Animals; Cytotoxicity, Immunologic; Female; Fibrosarcoma; G(M1) Ganglioside; Glycosphingolipids; In Vitro Techniques; Interferon-gamma; Interleukin-2; Killer Cells, Natural; Lymphokines; Mice; Mice, Inbred BALB C; Picibanil; T-Lymphocytes, Cytotoxic

Intratumoral induction of tumour necrosis factor (TNF) by administration of Bordetella pertussis vaccine (BPV) as compared with that by the agent OK-432 was investigated in mice. Two hours after such administration tumour tissues tested were resected from the mice, homogenised, and the TNF activities in the homogenate were assayed using a L-929 fibroblast assay. Intravenous injection of BPV into mice bearing the MM46 carcinoma resulted in a greater concentration of TNF in the tumour homogenate than in the serum. With OK-432, however, there was a greater concentration of TNF in the serum than in the tumour homogenates. A high level of intratumoral TNF induction by BPV was also observed in mice bearing Meth A fibrosarcoma or Lewis lung carcinoma. The therapeutic effect against the Meth A fibrosarcoma was in parallel with the intratumoral TNF activity. Intratumoral TNF activity is therefore believed to be a good index of therapeutic effect.

Topics: Animals; Antibodies; Drug Administration Routes; Female; Fibrosarcoma; Macrophages; Mice; Mice, Inbred Strains; Neoplasms, Experimental; Pertussis Vaccine; Picibanil; Time Factors; Tumor Necrosis Factor-alpha

Topics: Animals; Antibodies, Monoclonal; Antineoplastic Combined Chemotherapy Protocols; Drug Synergism; Female; Fibrosarcoma; Immunity; In Vitro Techniques; Interleukin-2; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Neoplasm Transplantation; Picibanil; Recombinant Proteins; Spleen; T-Lymphocytes, Cytotoxic; Thymoma

This study was carried out to determine the effect of local irradiation of the NFSa tumors on the incidence of lung metastases. The spontaneous lung metastases were found in those animals whose tumors had grown in size bigger than 10 mm in diameter. The incidence of metastases and the number of lung nodules were increased in those animals of irradiated group when compared to those of control group with the same size. This is probably because the irradiation of tumors resulted some retardation in their growth and thus, the irradiated tumors took a longer time to reach a given size than those unirradiated control tumors. The incidence of spontaneous lung metastases was significantly reduced by subcutaneous administration of OK-432 (2.5KE/mouse) locally into the surroundings of the tumor immediately after irradiation. The administration of OK-432 after the metastasis development was no longer effective. Both of intraperitoneal and subcutaneous administrations of OK-432 into opposite side of unirradiated thigh showed no suppression of metastasis. These results suggest that an appropriate immunostimulation combined with radiotherapy may be important for the suppression of tumor metastases.

Topics: Animals; Combined Modality Therapy; Fibrosarcoma; Lung Neoplasms; Male; Mice; Mice, Inbred C3H; Picibanil

The authors have investigated the effect of OK-432 on the spontaneous metastasis from a murine fibrosarcoma, either an NFSA or a SANH, that had been transplanted into the right thigh of C3H mice. When the NFSA or the SANH tumors grew to 7 mm in diameter, they were irradiated with a single dose of 30 Gy and 20 Gy, respectively. A local administration of OK-432 (2.5 KE) around each of the tumors was initiated immediately after irradiation and continued twice a week. It was found that regrowth in both tumors was not modified by the OK-432, though the mean number of metastatic lung nodules per mouse from either tumor was significantly decreased. The incidence of a lung metastasis also was reduced by the OK-432 but not significantly. Further, mice that had received a SANH tumor transplant developed a lymph node metastasis in the abdominal cavity that was depressed by the OK-432.

Topics: alpha 1-Antichymotrypsin; Animals; Biological Products; Combined Modality Therapy; Fibrosarcoma; Lung Neoplasms; Lymphatic Metastasis; Male; Mice; Mice, Inbred C3H; Picibanil; Radiotherapy Dosage

Protective effect of OK-432 on peritoneal recurrence was examined in an experimental mouse model with Meth A fibrosarcoma injected into both abdominal wall and intraperitoneal cavity. OK-432 was given through either intratumoral (i.t.), intradermal (i.d.), intraperitoneal (i.p.), or i.p. + i.t. route. A significant antitumor effect against abdominal tumor was observed in mice given i.t. or i.p. + i.t. injection of OK-432. A significant protective effect against peritoneal recurrence and prolonged survival time was obtained in an i.p. + i.t. group, although other treatment groups showed only minimal effect. We conclude that combined i.p. and i.t. administration of OK-432 may prevent peritoneal recurrence and lead to prolongation in clinical patients with advanced GI tract cancer.

Topics: Animals; Biological Products; Female; Fibrosarcoma; Injections, Intralesional; Injections, Intraperitoneal; Mice; Mice, Inbred BALB C; Peritoneal Neoplasms; Picibanil; Prognosis; Sarcoma, Experimental

The combined effects of rIL-2 and OK-432 were investigated against a Meth-A tumor, a syngeneic tumor of inbred BALB/c mice. An analysis of the effector cells was also performed. The treatment resulted in an inhibition in vivo of tumor growth and increased survival of the Meth-A tumor-bearing mice. Splenic cells obtained from Meth-A inoculated mice which received combination therapy were not only NK-sensitive YAC-1 and LAK-sensitive EL-4 cells, but also NK-resistant Meth-A cells, as shown in a 4-h 51Cr-release assay. Syngeneic killer cell activity against Meth-A cells was abolished almost completely with anti-Thy 1.2 treatment and about 70% of the activity was abolished with anti-asialo GM1 treatment in a complement-dependent cytotoxic assay. It was not changed by the removal of macrophages and B cells from the splenic cells. Mice which survived for 60 days after the start of therapy rejected Meth-A inoculation when rechallenged, suggesting the establishment of a specific immunity. Combination therapy appeared to be beneficial against Meth-A cells and T-cells appeared to play a determining role in the treated Meth-A bearing mice. It was suggested that more than two populations of killer cells exist in the spleen treated with the combined therapy and they may have the same characteristics as activated T and NK cells with or without specific killer T-cells.

Topics: Animals; Antibody-Dependent Cell Cytotoxicity; Antineoplastic Combined Chemotherapy Protocols; Female; Fibrosarcoma; Immunity; Injections, Intraperitoneal; Interleukin-2; Killer Cells, Natural; Mice; Mice, Inbred BALB C; Neoplasms, Experimental; Picibanil; Recombinant Proteins; Spleen; T-Lymphocytes

The antitumor effects of three biological response modifiers (BRMs; PSK, IFN alpha A/D and OK432) and two chemotherapeutics (Mitomycin C and Neocarzinostatin) in a new experimental mouse model, the "double grafted tumor system," were evaluated. BALB/c mice received simultaneous inoculations of Meth A fibrosarcoma cells on right flank (1 x 10(6) cells) and left flank (2 x 10(5) cells) on day 0, and drugs were given intratumorally into the right-flank tumor on day 3. The growth of the left-flank tumor was the real target for the evaluation of a given drug after 21 days. All tested five agents successfully cured the drug-injected right tumor with a pre-determined optimum dose. In addition, PSK, OK432, IFN alpha A/D and MMC among the five, inhibited the left-flank tumor, whereas no inhibition was observed when treated with NCS. To understand the mechanism by which the antitumor effect of the above four agents is able to influence the growth of tumor on the other side, tumor cells (2 x 10(5) cells) inoculated only into the left flank were treated with drugs given subcutaneously to the right flank (single tumor system). Among the four, MMC exhibited an effect similar to that obtained in the double tumor system, and IFN alpha A/D showed a less pronounced but still definite antitumor effect. However, PSK and OK432 failed to express anti-tumor effect in the single tumor system. These results obtained with PSK, OK432 and IFN alpha A/D suggest that the effect of the drug on the left-tumor may be mediated by certain effector cells, which are specifically induced by injection of the drug, in the right-tumor tissues. When effector cell analysis was conducted with spleen cells obtained after PSK treatment by means of intratumoral adoptive transfer into 3-day Meth A bearing recipients, these cells were shown to be Lyt-1+2(-)-T and L3T4(+)-T cell.

Topics: Animals; Antibodies; Antineoplastic Agents; Cells, Cultured; Complement System Proteins; Fibrosarcoma; Immunization, Passive; Immunologic Factors; Interferon Type I; Male; Mice; Mice, Inbred BALB C; Mitomycin; Mitomycins; Neoplasm Transplantation; Phenotype; Picibanil; Polysaccharides; Spleen; Zinostatin

The effects of a combination therapy of radiation and local administration of OK-432, which is one of BRMs (biological response modifiers), were studied using a radioresistant murine fibrosarcoma with weak immunogenicity. Fifty percent tumor control doses were 83.5 (79.6-87.4) Gy in animals given radiation alone and 64.3 (57.9-71.0) Gy in animals given OK-432 immediately after radiation, indicating that this biological response modifier can enhance the radiation dose effectiveness by a factor of 83.5/64.3, that is, 1.30. Histological observations of treated tumors showed that the combination therapy induced a marked infiltration with lymphocytes and prominent degeneration and necrosis of the tumor cells. Examination of subsets of the infiltrating lymphocytes using the monoclonal antibodies showed that Lyt-2 positive lymphocytes were more abundant in the group given the combination therapy than in the radiation alone group on day 5 (p greater than 0.05). Two days later, Lyt-1, Lyt-2, and L3T4 positive lymphocytes increased in the group given the combination therapy, whereas these lymphocytes disappeared in the group given radiation alone.

Topics: Animals; Antibodies, Monoclonal; Biological Products; Combined Modality Therapy; Fibrosarcoma; Injections; Lymphocytes; Mice; Mice, Inbred C3H; Picibanil; Radiotherapy Dosage

A synergistic therapeutic effect on Meth-A ascites tumor was obtained in BALB/c mice by the simultaneous i.p. injection of OK-432 and interferon alpha A/D (IFN-alpha). BALB/c mice that were cured of their Meth-A tumors showed significant tumor-specific rechallenge resistance. Combined therapeutic effect was significantly weaker in nude mice than in haired mice. The stimulation of host T-cell immunity was indicated to be crucially important for obtaining cured mice. Natural killer activity of peritoneal exudate cells was enhanced and peaked on day 1 with IFN-alpha, on day 3 with OK-432, and was sustained for up to 5 days with combination therapy. Macrophage activity assayed by in vivo resistance to the challenge of Listeria monocytogenes was stronger in order of OK-432 greater than OK-432 + IFN-alpha greater than IFN-alpha. Cytotoxic T-lymphocyte activity induced by immunization with the allogeneic tumor was enhanced most strongly by OK-432 plus IFN-alpha treatment. Each of the agents had tumor inhibitory activity on the growth of Meth-A cells in vitro, and there was a slight additive effect when two agents were used together. These findings suggest that the synergistic therapeutic effect of OK-432 plus IFN-alpha treatment on Meth-A ascites tumor was partly due to the direct antitumor activity of these two agents, but was also due in part to the beneficial antitumor immune responses that were induced. A similar synergism on Meth-A tumor was also observed by OK-432 plus interferon gamma treatment. However, this combination therapy was not effective on MH-134 ascites tumor in C3H mice.

Topics: Animals; Biological Products; Cells, Cultured; Combined Modality Therapy; Drug Synergism; Fibrosarcoma; Indomethacin; Interferon Type I; Interferon-gamma; Liver Neoplasms, Experimental; Lymphoma; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Inbred C57BL; Picibanil; T-Lymphocytes

Enough amounts of tumor necrosis factor (TNF) in mice serum for the therapy were observed by treatment with 100 units of recombinant human TNF-alpha (rHuTNF-alpha) followed by administration of OK-432 (a streptococcal preparation). Optimal time interval between rTNF and OK-432 to produce endogenous TNF was 3 h, and priming activity of rTNF persisted for at least 10 h. The same effect was observed using novel human recombinant TNF-SAM2 (rHuTNF-SAM2) developed by our group. Production of endogenous TNF using rTNF-alpha or rTNF-SAM2 as a priming reagent was almost equal among various mice strains. Induced TNF in mice serum was completely neutralized by anti-MuTNF antiserum, but not by anti-HuTNF monoclonal antibody. rMuTNF could also induce the priming state; however, the dose-response kinetics of the priming effect to produce endogenous TNF was different between rHuTNFs and rMuTNF-alpha, suggesting species specificity among rTNFs used. The therapeutic effect against Meth A and MH134 tumors in mice treated by rHuTNFs in combination with OK-432 was superior to that by single administration of either OK-432 or rHuTNFs or by successive administrations of OK-432. Especially, the antitumor effect against MH134 hepatoma was superior to that of any other treatment using known biological response modifiers so far experienced. These results suggest that such combination antitumor therapy as rTNF together with OK-432 should be applicable to cancer patients.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Dose-Response Relationship, Drug; Drug Therapy, Combination; Fibrosarcoma; Liver Neoplasms, Experimental; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Mice, Inbred Strains; Neoplasms, Experimental; Picibanil; Recombinant Proteins; Tumor Necrosis Factor-alpha

Topics: Animals; Biological Products; Fibrosarcoma; Lung Neoplasms; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Picibanil; Thigh

Murine spleen cells cultured for 3 or more days in medium with streptococcal preparation OK-432 became cytotoxic in vitro against several allogeneic and syngeneic tumor cells. These cytotoxic cells were designated OK-432-induced killer (OIK) cells. This study examined the in vivo antitumor efficacy of OIK cells in adoptive immuno- and immunochemo-therapy in mice bearing syngeneic tumors, such as EL-4 lymphoma, Meth-A fibrosarcoma, and MOPC-31C plasmacytoma. OIK cells neutralized these tumor cells, as shown by Winn-type tests, and the cell transfer prolonged the survival of mice inoculated intraperitoneally (ip) with EL-4 or Meth-A cells. Concomitant administration of OK-432 plus recombinant interleukin 2 (rIL-2) significantly improved the therapeutic efficacy of the transferred OIK cells. In mice inoculated with 1 x 10(4) EL-4 cells, chemoimmunotherapy consisting of ip administration of 200 mg/kg cyclophosphamide on day 3 followed by treatment with OIK cell (1 x 10(7)) transfer and with OK-432 (50 KE/kg) plus rIL-2 (50 units/mouse) 6 hr later and on day 6, prolonged the survival. Therefore, the immunotherapy with OIK-cell transfer followed by administration of OK-432 and rIL-2 may be clinically useful as an adjunct of cytoreductive chemotherapy for cancer.

Topics: Animals; Biological Products; Fibrosarcoma; Immunization, Passive; Interleukin-2; Killer Cells, Natural; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Neoplasm Transplantation; Neoplasms, Experimental; Picibanil; Plasmacytoma; Thymoma

Effect of combination use of radiation and multiple administration of OK-432 was studied using a radio-resistant and weakly immunogenic murine fibrosarcoma (NFSa) originated spontaneously in a C3H female mouse. Mice implanted the tumor in the leg were locally irradiated with 40 Gy of gamma rays and locally administered total 8 KE of OK-432. The first group was given 1 KE daily for 8 days, 2 KE every other day in the 2nd group, 4 KE at 1 week interval in the 3rd group and 8 KE immediately after irradiation in the 4th group. The tumor volume at day 19 after irradiation was compared. The tumor growth inhibitory effect was observed in the 3rd group (p less than 0.01) and 4th group (p less than 0.05) compared to the group given radiation alone. This effect was seen at even more smaller dose of OK-432 as 0.5 KE by 2 times administration at 1 week interval. The effect of route of OK-432 administration was studied comparing the tumor volume at day 22. OK-432 was given 2 times at 1 week interval. A significant difference was seen in the group given OK-432 locally compared to the control group given radiation alone (p less than 0.001). There were no significant differences between the groups given OK-432 by the routes of i.p., s.c. i.v. and the control group. TCD50 value was examined giving OK-432 4 KE on the day and 2 KE on day 7 after irradiation. TCD50 value in the group treated with irradiation alone was 83.5 (79.6-87.4) Gy and the group given combination therapy with OK-432 was 60.7 (55.9-65.4) Gy. Two times OK-432 administration at 1 week interval appears to potentiate the radiation response by about 23 Gy.

Topics: Animals; Biological Products; Combined Modality Therapy; Drug Administration Schedule; Fibrosarcoma; Mice; Mice, Inbred C3H; Picibanil; Radiotherapy Dosage; Sarcoma, Experimental

A murine malignant ascites model with BAMC-1 tumors was established previously, which was cured completely by five consecutive i.p. injections of OK-432. We have found that peritoneal mononuclear cells from these animals contained antitumor effector cells which could destroy nonspecifically a variety of tumor cells in vitro. They were tentatively called pantropic killer cells (PKCs). The present study was essentially designed to show the antitumor effectiveness of the PKCs in vivo by the use of an adoptive immunotherapy model. The growth of BAMC-1 tumors transplanted s.c. 5 days earlier was significantly suppressed by passive transfer of 5 x 10(6) to 2 x 10(7) PKCs induced by injection of OK-432 into BAMC-1 bearing donor mice, while more than 1 x 10(8) immune spleen cells from the same donors treated with OK-432 were required to achieve the similar effects. Furthermore, if the tumor-bearing recipients were pretreated with 180 mg/kg of cyclophosphamide 1 h before the adoptive transfer, even 5 x 10(6) PKCs could induce complete regression of the tumors transplanted 5 days earlier. This protocol made it possible even to achieve the complete regression of larger tumors (9-10 mm in diameter) in recipients transplanted 12 days earlier. The PKCs were, as expected, able to cure not only BAMC-1-bearing animals but also Meth-A-bearing mice. As effector cells for adoptive immunotherapy, therefore, the PKCs induced by OK-432 seem to be as effective as, if not better than, lymphokine-activated killer cells expanded in vitro by culturing tumor infiltrating lymphocytes with interleukin-2. Although the study on surface markers of PKCs did not unequivocally discriminate these from lymphokine-activated killer cells, the present findings are considered significant indicating that a potent biological response modifier such as OK-432 can induce pantropic killer cells which are extremely effective in destroying various tumor cells in vivo. One of the advantages of OK-432 therapy over lymphokine-activated killer cell therapy, therefore, is that the former does not require the tedious and time-consuming in vitro procedures which are essential for the latter.

Topics: Animals; Antibodies, Monoclonal; Ascitic Fluid; Female; Fibrosarcoma; Immunization, Passive; Killer Cells, Natural; Mice; Mice, Inbred BALB C; Picibanil

In vivo tumor inhibitory activity of polymorphonuclear leukocytes (PMN) treated in vitro with lymphokine(s) (LK) was investigated with Winn's assay. Culture supernatants of BALB/c mouse spleen cells incubated with a streptococcal preparation, OK-432, were used as an LK source. With the use of a [3H]uridine release assay, RL male-1 tumor cells were lysed to some extent by peritoneal BALB/c mouse PMN treated with this LK preparation. With Winn's assay, LK-treated PMN from BALB/c mice completely inhibited the growth of the admixed syngeneic tumor at a high effector to target ratio, when normal mice were used as recipients. When X-irradiated mice or nude mice were used as recipients, the tumor growth was partially inhibited by admixed LK-treated PMN, but the tumor began to grow gradually and finally killed the recipient mice, even when a high effector target ratio was used. When nude mice which had been given i.v. transfers of nylon wool column effluent spleen cells were used as recipients, the tumor inhibitory activity of LK-treated PMN was recovered to the same level as when normal mice were used as recipients. On the other hand, tumor inhibition by admixed LK-treated PMN in nude mice was not recovered by the transfer of X-irradiated nylon column effluent T-cells. As a mechanism of tumor inhibition by LK-treated PMN, a possible role of LK-treated PMN in reduction of tumor load is discussed.

Topics: 2-Chloroadenosine; Adenosine; Animals; Cells, Cultured; Female; Fibrosarcoma; Lymphokines; Male; Mast-Cell Sarcoma; Mice; Mice, Inbred BALB C; Mice, Inbred DBA; Mice, Nude; Neutrophils; Picibanil; Rats; Rats, Inbred Strains; T-Lymphocytes, Cytotoxic

In order to clarify the characteristics of interferon-alpha A/D (IFN-alpha) as a biological response modifier (BRM), several immunobiological activities were compared with OK-432. 1) Both IFN-alpha and OK-432 inhibited the multiplication of Meth-A cells in vitro. 2) IFN-alpha (2 X 10(5) IU, ip) augmented the NK activity of peritoneal exudate cells (PEC) and spleen cells, and the peak of NK activity was observed 1 day after injection. OK-432 (1 KE, ip) augmented the NK activity of PEC, but not of spleen cells, and the peak was 3 days after. 3) Macrophage activating activity was more potent with OK-432 (1 KE) than IFN-alpha (2 X 10(5) IU). 4) Induction of CTL against alloantigens was augmented by IFN-alpha and OK-432. 5) By the combination of IFN-alpha with OK-432, a synergistic antitumor effect was obtained against Meth-A ascites tumor. Immunobiological effects of IFN-alpha seemed to be somewhat different from those of OK-432. Therefore, the combination of the two agents might cause a synergistic antitumor effect.

Topics: Animals; Biological Products; Cell Line; Fibrosarcoma; Interferon Type I; Leukemia, Experimental; Lymphocyte Activation; Male; Mice; Mice, Inbred BALB C; Phytohemagglutinins; Picibanil; Sarcoma, Experimental

Optimal timing of topical administration of OK-432 and TCD50 were studied using weakly immunogenic and radioresistant C3H mouse fibrosarcoma (NFSa). The mechanism of action of this combined therapy was examined histologically and electron microscopically. Topical administration of OK-432 was performed from 2 days before irradiation to 7 days after irradiation and tumor volumes on the 20th day after irradiation were compared with a control group given radiation alone. Significant difference was observed only in the group which was given OK-432 just after irradiation. The TCD50 of the combined therapy of radiation with topical administration of OK-432, 4 KE which was given just after irradiation was 64.5 Gy and that of radiation alone was 83.5 Gy. Combined therapy shifted the TCD50 curve about 20Gy to the left. Histological examination of the tumor on the 6th day after combined therapy showed marked degeneration and necrosis of tumor cells with marked infiltration of lymphocytes. These lymphocytes were electron microscopically seen surrounding not only damaged cells, but also apparently active tumor cells. We postulate the latter cells had a tendency to be degenerative. This phenomenon suggests that lymphocytes recognize as foreign these tumor cells which are apparently active but some what damaged by radiation.

Topics: Administration, Topical; Animals; Biological Products; Combined Modality Therapy; Fibrosarcoma; Mice; Picibanil; Radiotherapy Dosage; Sarcoma, Experimental

The combined effect of radiation with local administration of OK-432 was studied using natural fibrosarcoma (NFSa) of C3H mice that are less immunogenic and resistant to radiation (TCD 50, 83, 15 Gy). When 2KE of OK-432 was administered together with 1 X 10(6) tumor cells, tumor survival was inhibited in 7/8 (87.5%) of mice. Single local administration of OK-432 (2KE, 4KE, 8KE) showed hardly any effect on tumor survival. Combined treatment of 2KE of OK-432 and 4-Gy radiation, however, not only controlled tumor growth but also eradicated 1/5 (20%) of them, compared with single application of 45-Gy radiation. OK-432 worked most effectively when administered just before or after the application of X-rays. The optimum dose of OK-432 was studied by administering 2KE, 4KE and 8KE of the agent together with the application of 55-Gy radiation. 8KE groups showed the best eradication rate but 4KE groups showed the best cure rate. Therefore, the optimum dose was considered to be between 4KE and 8KE. With regard to histological effects, both the single radiation treatment and its combined treatment with OK-432 were most effective on the 6th day. The degree of degeneration and necrosis of tumor cells together with lymphocyte exudation were much higher in the combined treatment than in single radiation application.

Topics: Animals; Biological Products; Cell Division; Combined Modality Therapy; Fibrosarcoma; Male; Mice; Mice, Inbred C3H; Picibanil; Radiotherapy Dosage; Sarcoma, Experimental

Topics: Administration, Oral; Animals; Biological Products; Cecal Neoplasms; Fibrosarcoma; In Vitro Techniques; Killer Cells, Natural; Lymphocyte Activation; Male; Mice; Mice, Inbred BALB C; Neoplasm Transplantation; Picibanil; Sarcoma, Experimental

Topics: Animals; Combined Modality Therapy; Fibrosarcoma; Immunotherapy; In Vitro Techniques; Male; Mice; Mice, Inbred BALB C; Picibanil; Spleen