ABSTRACT
Purification of cytokines was carried out while monitoring their in vivo anti-tumor activity and in vitro cytotoxic activities. As a result, purified new cytokines were obtained from culture supernatant of a histiocytic cell line from rabbit serum. Briefly, a new cytokine (HSF, histiocyte-secreted-factor) was purified from the culture of supernatant of the histiocytic cell line (TYH) which we established from the peripheral blood of a malignant-lymphoma patient. The purified samples exhibited suppressive effects on tumor growth but no necrotizing activity towards transplanted murine tumors. The substance displayed no cytotoxic activity against L cells (mouse fibroblast cells). The molecular weight of human HSF was about 42 kDa as estimated by SDS-PAGE. Amino-acid sequencing of the purified HSFD from the culture supernatant was performed, but the N-terminal was blocked. Next, a new cytokine was purified from rabbit serum stimulated with Propionibacterium acnes and elicited with lipopolysaccharide. The rabbit HSF was isolated by the same procedures as those used for the human HSF purification steps. Amino-acid sequencing was carried out after enzyme digestion. Three parts of the amino-acid sequence of the rabbit HSF were determined as LPPGLLAPMRQLRS-, NLEXFTNGMEQHYAQL-, NPAENQAHELPNQLN-. A computer-based homology search demonstrated that these sequences were novel. The molecular weight of HSF as determined using anti-peptide antibodies revealed the following values: human HSF, 41 and 46 kDa; rabbit HSF, 35, 42 and 55 kDa.
Subject(s)
Antineoplastic Agents/isolation & purification , Cytokines/isolation & purification , Amino Acid Sequence , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Cytokines/chemistry , Cytokines/pharmacology , Humans , Immunoblotting , Mice , Mice, Inbred BALB C , Molecular Sequence Data , Rabbits , Tumor Cells, CulturedABSTRACT
The antitumor activity of combination therapy with traditional Chinese medicines and OK432 (Streptococcus pyogenes) for cancer patients was investigated. Excellent antitumor activity of this treatment was achieved in one patient with hepatocellular carcinoma. The present report describes the clinical course of this patient and examines the contribution of production of tumor necrosis factor (TNF) and interferon-gamma (IFN). Endogenous production of TNF could be observed after drip intravenous injection of OK 432 in the serum of patients treated by previous oral administration of traditional Chinese medicines. The serum levels of IFN were very low and remained at almost undetectable levels under these conditions. The selective use of immunostimulants such as traditional Chinese medicines may be of value in combination with other therapies such as drip infusion of OK 432, in the treatment of advanced cancer or of aged patients because of the low toxicity. One patient out of 12 revealed a partial response as assessed by the antitumor activity. However, with this treatment, patients did become free from pain and a good performance status was supported.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Carcinoma, Hepatocellular/drug therapy , Liver Neoplasms/drug therapy , Aged , Aged, 80 and over , Carcinoma, Hepatocellular/immunology , Drugs, Chinese Herbal/administration & dosage , Female , Humans , Interferon-gamma/blood , Interleukin-1/blood , Liver Neoplasms/immunology , Lymphocyte Count , Male , Picibanil/administration & dosage , T-Lymphocyte Subsets , Tumor Necrosis Factor-alpha/metabolismABSTRACT
The antitumor activity of recombinant human tumor necrosis factor (rhTNF) against heterotransplanted human prostatic carcinoma (PC-3) and spontaneous lymphatic tumor metastasis was studied in vivo. The spontaneous lymphatic metastasis of PC-3 tumor was found in approximately 50% of cases. Significant antitumor activity was observed with repeated intratumoral administration of a large dose of rhTNF, not only on the subcutaneous tumor xenografts but also on the lymph node metastases. Strong antitumor activity could be achieved even with the intratumoral administration of a small dose of rhTNF in combination with mild hyperthermia on either the transplanted tumors or on the metastatic tumors.
Subject(s)
Hyperthermia, Induced , Prostatic Neoplasms/therapy , Tumor Necrosis Factor-alpha/therapeutic use , Acid Phosphatase/metabolism , Animals , Combined Modality Therapy , Humans , Lymphatic Metastasis , Male , Mice , Mice, Nude , Neoplasm Transplantation , Prostate/enzymology , Prostatic Neoplasms/enzymology , Prostatic Neoplasms/secondary , Recombinant Proteins/therapeutic use , Transplantation, HeterologousSubject(s)
Cell Division , Tumor Necrosis Factor-alpha/pharmacology , Animals , Cell Division/drug effects , Cell Line , Endothelium, Vascular/cytology , Endothelium, Vascular/drug effects , Hematopoietic Stem Cells/cytology , Hematopoietic Stem Cells/drug effects , Humans , Lymphocytes/cytology , Lymphocytes/drug effects , Neoplasms , Neutrophils/cytology , Neutrophils/drug effects , Neutrophils/physiology , Tumor Necrosis Factor-alpha/physiologyABSTRACT
The participation of tumor necrosis factor (TNF) and lipopolysaccharide (LPS) in Pseudomonas aeruginosa (Pa) infection was examined. The lethal challenge of Pa or TNF and LPS injection could be prevented by pretreatment with anti-TNF antibody, polymyxin B, ONO 1078, or Shosaiko-to. The combined effects of TNF and LPS may be deeply related to the lethality of Pa infection. The activities of leukotriene(LT) C4/D4/E4 or platelet activating factor (PAF) were also related to the lethality of Pa infection, probably due to the subsequently produced TNF which acts in combination with LPS. Activating the host defence mechanism with biological response modifiers like Chinese medicines was effective against Pa infection. One mechanism could involve an activity as an LT inhibitor or PAF antagonist. Following the administration of TNF and/or LPS, the serum levels of arachidonic cascade products underwent various changes. With a combination of TNF and LPS, there was a synergistic increment of prostaglandins, thromboxane, and LT. Following pretreatment with Shosaiko-to, suppression of LTs was dominant even with the combination of TNF and LPS, which might be related to the lethality of the infection or combined TNF with LPS.
Subject(s)
Aminoglycosides , Antineoplastic Combined Chemotherapy Protocols/toxicity , Leukotriene Antagonists , Platelet Activating Factor/antagonists & inhibitors , Pseudomonas Infections/prevention & control , Animals , Anti-Bacterial Agents/therapeutic use , Antibodies/therapeutic use , Antineoplastic Agents/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Arachidonic Acid , Arachidonic Acids/blood , Cephamycins/therapeutic use , Chromones/therapeutic use , Drug Synergism , Drugs, Chinese Herbal/therapeutic use , Female , Leukotriene E4 , Lipopolysaccharides/administration & dosage , Lipopolysaccharides/toxicity , Mice , Mice, Inbred Strains , Polymyxin B/therapeutic use , Pseudomonas Infections/blood , Recombinant Proteins/toxicity , SRS-A/analogs & derivatives , SRS-A/antagonists & inhibitors , SRS-A/blood , SRS-A/metabolism , Tumor Necrosis Factor-alpha/administration & dosage , Tumor Necrosis Factor-alpha/immunology , Tumor Necrosis Factor-alpha/toxicityABSTRACT
We have investigated the antitumor activity of tumor necrosis factor (TNF) against numerous. It appears that the efficacy of TNF against highly immunogenic tumor cell lines is greater than that against low immunogenic cell lines. Tumor cells express immunogenic tumor-associated antigen and sometimes shed antigens. Antibody against shedding antigens from Meth A sarcoma cells was prepared by immunizing rabbits seven times. This antibody by itself was not curative. However, in combination with TNF, the antibody was effective even against large and well-established tumors. When the antibody was injected 1 day after administration of TNF, the antitumor activity was stronger than in the case of simultaneous administration of TNF and antibody. We further examined the effect of this antibody against Ehrlich tumors in combination with TNF. The results were similar to those for Meth A sarcoma, although effects were weaker.
Subject(s)
Antibodies, Neoplasm/therapeutic use , Antigens, Neoplasm/immunology , Sarcoma, Experimental/therapy , Tumor Necrosis Factor-alpha/therapeutic use , Animals , Antibodies, Neoplasm/biosynthesis , Antigens, Neoplasm/administration & dosage , Carcinoma, Ehrlich Tumor/therapy , Combined Modality Therapy , Immunization , Immunoenzyme Techniques , Mice , Mice, Inbred BALB C , Mice, Inbred Strains , Recombinant Proteins/therapeutic useABSTRACT
It was found that the capacity for tumor necrosis factor (TNF) production by Japanese modified traditional Chinese medicines and crude drugs broadly paralleled their antitumor activity. Pretreatment with these drugs prevented the lethal and marked side effects of recombinant human TNF (rhTNF) and lipopolysaccharide (LPS) without impairing their antitumor activity. These drugs are thought to decrease the oxygen radicals and stabilize the cell membranes, with a deep relation to the arachidonic cascade. The release of prostaglandins and leukotriene B4 was suppressed by pretreatment with Shosaiko-to. Thromboxane B2 was transiently increased, followed by suppression. After pretreatment with Hochu-ekki-to or Juzen-taiho-to, suppression of leukotriene B4 could not be observed. The release of prostaglandin D2 was suppressed in mice pretreated with Shosaiko-to, Juzentaiho-to or Ogon (Scutellariae Radix) but it increased following pretreatment with Hochu-ekki-to. Chemicals that could prevent the lethality of rhTNF and LPS also revealed suppression of prostaglandins, leukotriene B4 and thromboxane B2. In general, drugs that prevented the lethality of rhTNF and LPS without impairing the antitumor activity could inhibit the release of leukotriene B4 and/or prostaglandin D2. rhTNF could activate the arachidonic cascade in combination with LPS. The lethality of rhTNF and LPS could be prevented by pretreatment with Japanese modified traditional Chinese medicines and the crude drug, Ogon.
Subject(s)
Antineoplastic Agents/therapeutic use , Arachidonic Acids/blood , Drugs, Chinese Herbal , Lipopolysaccharides/therapeutic use , Medicine, Chinese Traditional , Tumor Necrosis Factor-alpha/therapeutic use , Animals , Drug Therapy, Combination , Japan , Mice , Sarcoma/drug therapy , Sarcoma/prevention & control , Tumor Cells, CulturedSubject(s)
Antineoplastic Agents , Tumor Necrosis Factor-alpha/therapeutic use , Animals , Clinical Trials as Topic , Endothelium, Vascular/cytology , Endothelium, Vascular/drug effects , Lipid Peroxidation/drug effects , Lysosomes/enzymology , Models, Biological , Oxygen/physiology , Recombinant Proteins/therapeutic use , Tumor Necrosis Factor-alpha/adverse effects , Tumor Necrosis Factor-alpha/pharmacologyABSTRACT
Toxicity has been observed in mice receiving recombinant human tumor necrosis factor (rhTNF). In the present experiments, several chemicals were used to determine whether they could prevent the lethality of rhTNF without impairing its antitumor activity. Injection of phospholipase A2, cyclooxygenase, and lipoxygenase inhibitors at the same time as rhTNF administration could prevent the lethality of rhTNF, but the antitumor activity was also reduced. Urinastatin and reduced glutathione could prevent the lethality while reducing the activity. In contrast, by pretreatment with O2 scavengers, the lethality of rhTNF was markedly reduced without impairment of the antitumor activity of rhTNF. Antihistamines exerted no influence on the lethality of rhTNF. Histopathologic examinations have demonstrated that the capillaries of the tumor tissue show aggregation of platelets and formation of fibrin adherent to the vascular surface after TNF administration. Heparin or protamine revealed no effects against the lethality of rhTNF. These results strongly suggest that the arachidonic cascade is deeply related to the antitumor activity of TNF and its side effects. Pretreatment with O2 scavengers, especially bismuth subnitrate, could prevent the lethality of rhTNF without impairing its antitumor activity.
Subject(s)
Carcinoma, Ehrlich Tumor/drug therapy , Tumor Necrosis Factor-alpha/adverse effects , Animals , Blood , Chlorpromazine/pharmacology , Cortisone/pharmacology , Drug Therapy, Combination , Female , Glutathione/pharmacology , Mice , Mice, Inbred BALB C , Mortality , Neoplasm Recurrence, Local , Recombinant Proteins/adverse effects , Recombinant Proteins/therapeutic use , Tumor Necrosis Factor-alpha/therapeutic use , Vitamin E/pharmacologyABSTRACT
To examine the effects of combination therapy with traditional Chinese preparations (Syô-saiko-tô, Zyûzen-taiho-tô or Cinnamomum cortex) and OK432 or mitomycin C on the antitumor activities and TNF producibility, an investigation was carried out using mice transplanted with Ehrlich or Meth A tumor cells. Development of the transplanted tumors was strongly inhibited by the combination therapy, and the tumor necrosis factor (TNF) producibility was increased with the combination of a traditional Chinese preparation and OK432. A significant negative correlation was observed between the TNF activities and tumor weight, and there was a positive correlation between the TNF activities and spleen weight in the Ehrlich-bearing DDY mice receiving traditional Chinese preparations or OK432. Marked lymphocytosis, hyperplasia, and hypertrophy of Kupffer's cells in the liver were noted in the tumor-bearing DDY mice receiving traditional Chinese preparations or OK432. In two-step carcinogenesis experiments involving treatment with DMBA and TPA to induce skin papillomas, Zyûzen-taiho-tô appeared to be effective in inhibiting carcinogenesis. These results suggest that the antitumor activities and capacity for TNF production of the preparations are probably due in part to stimulation of the reticuloendothelial system, including macrophages, and induction of a host-mediated antitumor substance like TNF as one of the immunopotentiators.
Subject(s)
Carcinoma, Ehrlich Tumor/therapy , Immunotherapy , Medicine, Chinese Traditional , Plants, Medicinal/immunology , Animals , Combined Modality Therapy , Liver/anatomy & histology , Mice , Mice, Inbred BALB C , Mitomycin , Mitomycins/therapeutic use , Organ Size , Picibanil/therapeutic use , Spleen/anatomy & histology , Tumor Necrosis Factor-alpha/bloodABSTRACT
The antitumor activity of recombinant human tumor necrosis factor (rhTNF) against methylcholanthrene A-induced sarcoma (Meth A sarcoma) and human tumors in vivo was studied. On systemic administration of rhTNF to Meth A sarcoma-bearing mice, tumor regression was achieved with a large dose or with repeated administration of a small dose. Strong antitumor activity could be achieved in the Meth A sarcoma model by administration of a small dose of rhTNF in combination with moderate temperature hyperthermia (p less than 0.005). rhTNF (1,000 U/mouse) combined with moderate hyperthermia (40 degrees C for 40 min) within 2 h after the TNF administration effected 100% complete regression. In contrast, the rhTNF or moderate hyperthermia alone revealed 0% complete regression. The antitumor activity of rhTNF was decreased in combination with cyclophosphamide (0.6 or 1.2 mg/mouse) (p less than 0.005). However, in combination with mitomycin C (30 or 120 micrograms/mouse), the antitumor activity of rhTNF was enhanced (p less than 0.005). In combination with immunotherapy (lentinan or OK 432), the antitumor activity was mostly enhanced. Repeated rhTNF administration also displayed antitumor activity in heterotransplanted human tumors (p less than 0.005). The antitumor activity of TNF was enhanced by repeated administration of even small dosages, in combination with hyperthermia, or in combination with immunotherapy.
Subject(s)
Glycoproteins/therapeutic use , Neoplasms, Experimental/therapy , Sarcoma, Experimental/therapy , Animals , Antineoplastic Agents/administration & dosage , Combined Modality Therapy , Hot Temperature , Humans , Immunotherapy , Neoplasm Transplantation , Recombinant Proteins/therapeutic use , Tumor Necrosis Factor-alphaABSTRACT
The antitumor activity of murine, rabbit and recombinant human tumour necrosis factor (TNF) was examined in an experimental animal model. TNF showed an excellent curative effect against the murine and human tumours tested. Strong antitumour activity was obtained by combining a small dose of TNF with moderate hyperthermia (40 degrees C for 40 min). TNF was also active against metastatic tumours, especially after repeated administration. The necrotizing action of TNF in vivo mainly relates to capillary injury. TNF causes necrosis not only in tumour tissue but also in granulation tissue. It causes morphological changes in, growth inhibition of, and cytotoxicity against cultured vascular endothelial cells. TNF inhibits endothelial motility evoked by the tumour. The mechanism of the cytotoxic action of TNF was examined using a microspectrophotometric assay for the lysosomotropic probe, acridine orange. The results suggest that TNF exerts its effect by enhancing endogenous tumour lysosomal activity. The increment in cellular respiration paralleled the susceptibility to TNF.
Subject(s)
Cytotoxicity, Immunologic , Necrosis/etiology , Neoplasms, Experimental/drug therapy , Tumor Necrosis Factor-alpha/pharmacology , Animals , Humans , Hyperthermia, Induced , In Vitro Techniques , Mice , Mice, Inbred BALB C , Neoplasm Transplantation , Neoplasms, Experimental/pathology , Rabbits , Recombinant Proteins/pharmacology , Tumor Cells, CulturedABSTRACT
The effects of a highly purified tumor necrosis factor (TNF) on transplanted methylcholanthrene (Meth A)-induced murine tumors were compared with those of lipopolysaccharide (LPS). TNF caused immediate subepidermal edema and hyperemia followed 2 h later by fibrin thrombi in tumor blood vessels. Finally hemorrhagic necrosis with dispersal of tumor cells occurred. LPS produced similar hemorrhagic necrotizing changes. However, the necrotic action of LPS was delayed and complete tumor regression was not achieved with LPS. These findings suggest that tumor necrosis induced by TNF is due to circulatory disturbance associated with a microvascular injury in the tumor manifested by hyperemia and multiple fibrin thrombi.
Subject(s)
Fibrosarcoma/drug therapy , Glycoproteins/therapeutic use , Animals , Fibrosarcoma/blood supply , Fibrosarcoma/chemically induced , Fibrosarcoma/pathology , Lipopolysaccharides/therapeutic use , Methylcholanthrene , Mice , Mice, Inbred BALB C , Microcirculation , Microscopy, Electron , Necrosis , Neoplasm Transplantation , Thrombosis/chemically induced , Tumor Necrosis Factor-alphaABSTRACT
The effect of tumor necrosis factor (TNF) on the tumor-induced endothelial migration was evaluated with the use of a phagokinetic track assay. TNF from both ddy mice and Japanese albino rabbits (sp act, 3-5 X 10(5) U/mg, respectively) was found to inhibit the migration of bovine capillary endothelial (BCE) cells stimulated by factors from tumor cells, such as the medium conditioned with mouse sarcoma 180 cells or human meningioma extract. These TNF preparations, however, did not affect the spontaneous migration of the BCE cells. When mouse TNF was further fractionated by polyacrylamide gel electrophoresis, only TNF-positive fractions showed an inhibitory activity on the tumor-induced endothelial motility. Moreover, monoclonal antibody against rabbit TNF completely neutralized its migration-inhibitory activity. These findings indicate that the observed inhibitory effect of TNF preparations on the endothelial motility evoked by tumor is exclusively ascribed to the function of TNF. This activity presumably is involved in the suppression of tumor angiogenesis in vivo.
Subject(s)
Chemotaxis/drug effects , Endothelium/drug effects , Glycoproteins/pharmacology , Meningioma/analysis , Sarcoma 180/analysis , Animals , Capillaries/cytology , Cattle , Cells, Cultured , Culture Media/pharmacology , Depression, Chemical , Endothelium/physiology , Glycoproteins/isolation & purification , Humans , Mice , Mice, Mutant Strains , Neovascularization, Pathologic , Rabbits , Tumor Necrosis Factor-alphaABSTRACT
The action of tumor necrosis factor (TNF) was investigated histopathologically in mice using methylcholanthrene A (Meth A)-induced sarcomas and granulation tissue induced by autotransplantation of fragments of liver and spleen. Highly purified murine TNF caused hemorrhagic necrosis of both the tumors and the granulation tissue. Proliferation of tumor capillaries, demonstrated microangiographically, occurred 2 h after TNF administration and hyperemia of tumor vessels was obvious after 3-6 h. Hyperemia and capillary leakage were also observed in the granulation tissue 6 h after TNF injection and hemorrhage was noted in the epidermis after 12 h. These results strongly suggest that the in vivo necrotizing action of TNF is mainly related to capillary injury.
Subject(s)
Granulation Tissue/drug effects , Necrosis/chemically induced , Sarcoma, Experimental/pathology , Tumor Necrosis Factor-alpha/pharmacology , Animals , Capillaries/diagnostic imaging , Capillaries/drug effects , Capillaries/pathology , Granulation Tissue/pathology , Lipopolysaccharides/pharmacology , Methylcholanthrene , Mice , Polymyxin B/pharmacology , Radiography , Sarcoma, Experimental/chemically inducedSubject(s)
Glycoproteins/biosynthesis , Animals , Corynebacterium/immunology , Endotoxins/pharmacology , Kinetics , Lipopolysaccharides/pharmacology , Listeria monocytogenes/immunology , Mycobacterium bovis/immunology , Propionibacterium acnes/immunology , Rabbits , Species Specificity , Tumor Necrosis Factor-alphaABSTRACT
Cytotoxic factor was produced from murine macrophage-like cell line, J774, after stimulation with 12-o-tetradecanoyl phorbol-13-acetate (TPA) and lipopolysaccharide (LPS). J774-derived cytotoxic factor is a glycoprotein with a molecular weight of 39,000 by gel filtration and 18,000 by SDS-PAGE, and with an isoelectric point of below 4.3. J774-derived cytotoxic factor exhibited cytotoxicity to L(S) cells, not cytotoxic to L(R) cells, and a necrotizing action against transplanted Meth A sarcoma. J774-derived cytotoxic factor and murine serum tumor necrosis factor (TNF) were identical as regards properties.
Subject(s)
Glycoproteins/biosynthesis , Macrophages/metabolism , Animals , Cell Line , Cytotoxins/biosynthesis , Cytotoxins/isolation & purification , Glycoproteins/isolation & purification , Lipopolysaccharides/pharmacology , Macrophages/drug effects , Mice , Molecular Weight , Tetradecanoylphorbol Acetate/pharmacology , Tumor Necrosis Factor-alphaABSTRACT
Exposure to highly purified preparations of murine tumor necrosis factor (TNF) resulted in distinct morphologic changes and proliferation inhibition of cultured endothelial cells from the bovine aorta and capillary and of those from the human umbilical vein. The TNF preparation also inhibited the proliferation of bovine aortic smooth muscle cells but failed to inhibit the proliferation of bovine and human fibroblasts. The cytotoxic activity of the TNF preparation was prominent only in bovine capillary endothelial cells. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis (PAGE), and upon PAGE only, of the TNF preparations, the fractions representing growth inhibitory and cytotoxic activities against bovine capillary endothelial cells exactly corresponded to those containing migrated TNF. Treatments of the TNF preparations with heat and polymyxin B revealed that the effects on endothelial cells were not due to possibly contaminated endotoxin. Moreover, growth inhibitory and cytotoxic actions of rabbit TNF on bovine capillary endothelial cells were completely neutralized by the addition of an anti-rabbit TNF monoclonal antibody. These results suggest that TNF causes morphologic changes in, growth inhibition of, and cytotoxicity against, the vascular endothelial cells.