lipoteichoic-acid and Fibrosarcoma

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

Comparison was made between the immunobiological and antigenic properties of two lipoteichoic acid (LTA) fractions (LTA-1 and -2) from Enterococcus hirae ATCC 9790, their glycolipid portions, and synthetic compounds partially mimicking the above bacterial products. The more lipophilic LTA-2 fraction was capable of inducing serum tumor necrosis factor alpha and interleukin-6 in muramyldipeptide-primed mice and serum gamma interferon in those primed with Propionibacterium acnes. The LTA-2 fraction also induced tumor necrosis factor alpha, interleukin-6, and thymocyte-activating factor (essentially interleukin-1) in murine peritoneal macrophage cultures. Consecutive intravenous injections of muramyldipeptide and the LTA-2 fraction in Meth A fibrosarcoma-bearing BALB/c mice caused hemorrhagic necrosis and marked regression leading to complete regression of the tumor with no accompanying weakening or lethal effects. The LTA-2 fraction was at least 10,000-fold less pyrogenic in rabbits than a reference endotoxic lipopolysaccharide. The more hydrophilic LTA-1 fraction, on the other hand, showed at most marginal activity in the in vivo and in vitro assays. Natural glycolipids (NGL-1 and -2) which were prepared from a chloroform-methanol extract of Streptococcus pyogenes and E. hirae cells, and comparable in structure to the lipid moieties of the LTA-1 and -2 fractions, respectively, were practically inactive in all of the assays. None of the test synthetic compounds was immunobiologically active, although synthetic partial counterparts of the structure of LTA proposed by W. Fischer (Handb. Lipid Res. 6:123-234, 1990) reacted with murine monoclonal antibody TS-2, which was raised against OK-432, a penicillin-killed S. pyogenes preparation, and capable of neutralizing the cytokine-inducing activities of the LTA-2 fraction.

Topics: Acetylmuramyl-Alanyl-Isoglutamine; Animals; Antigens, Bacterial; Antineoplastic Agents; Carbohydrate Sequence; Cytokines; Enterococcus; Fibrosarcoma; Glycolipids; Interferon-gamma; Interleukin-6; Lipopolysaccharides; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Molecular Sequence Data; Propionibacterium acnes; Sarcoma, Experimental; Teichoic Acids; Tumor Necrosis Factor-alpha

Two molecular species of lipoteichoic acid (LTA 1 and LTA 2) were isolated from whole cells of Streptococcus faecalis (Enterococcus hirae) ATCC 9790 by hydrophobic chromatography on Octyl Sepharose CL-4B. Chemical analysis revealed that LTA 1 and LTA 2 contained two and four acyl lipid anchors respectively. LTA 1 was less active than LTA 2 in inducing cytokines, except interleukin-1 (IL-1), but their in vivo antitumour effects were similar. LTA 2 was a potent inducer of tumour necrosis factor (TNF) and interferon (IFN) production and had excellent antitumour activity against Meth A fibrosarcoma established in mice. Deacylation of LTA 2 by alkaline hydrolysis abolished these biological activities. The phosphatidylglycolipid fraction derived from LTA 2 after acid hydrolysis could also induce TNF, IFN, and IL-1 production, as well as having antitumour activity against Meth A fibrosarcoma. Therefore, the lipid anchor portion of S. faecalis LTA may play an important role in the manifestation of these various biological activities.

Topics: Animals; Enterococcus faecalis; Female; Fibrosarcoma; Interferons; Interleukin-1; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Inbred ICR; Structure-Activity Relationship; Teichoic Acids; Tumor Necrosis Factor-alpha

Monolayers of murine fibrosarcoma cells that had been treated either with histone-opsonized streptococci, histone-opsonized Candida globerata, or lipoteichoic acid-anti-lipoteichoic acid complexes underwent disruption when incubated with human polymorphonuclear leukocytes (PMNs). Although the architecture of the monolayers was destroyed, the target cells were not killed. The destruction of the monolayers was totally inhibited by proteinase inhibitors, suggesting that the detachment of the cells from the monolayers and aggregation in suspension were induced by proteinases releases from the activated PMNs. Monolayers of normal endothelial cells and fibroblasts were much resistant to the monolayer-disrupting effects of the PMNs than were the fibrosarcoma cells. Although the fibrosarcoma cells were resistant to killing by PMNs, killing was promoted by the addition of sodium azide (a catalase inhibitor). This suggests that the failure of the PMNs to kill the target cells was due to catalase inhibition of the hydrogen peroxide produced by the activated PMNs. Target cell killing that occurred in the presence of sodium azide was reduced by the addition of a "cocktail" containing methionine, histidine, and deferoxamine mesylate, suggesting that hydroxyl radicals but not myeloperoxidase-catalyzed products were responsible for cell killing. The relative ease with which the murine fibrosarcoma cells can be released from their substratum by the action of PMNs, coupled with their insensitivity to PMN-mediated killing, may explain why the presence of large numbers of PMNs at the site of tumors produced in experimental animals by the fibrosarcoma cells is associated with an unfavorable outcome.

Topics: Animals; Azides; Candida; Catalase; Cell Adhesion; Cell Survival; Cytotoxicity, Immunologic; Fibrosarcoma; Free Radicals; Humans; Lipopolysaccharides; Mice; Neutrophils; Opsonin Proteins; Protease Inhibitors; Sodium Azide; Streptococcus; Teichoic Acids; Tumor Cells, Cultured

The antitumour effects of lipoteichoic acids (LTA) extracted from Streptococcus pyogenes were studied in comparison with other streptococcal cellular components. LTA suppressed the tumour growth of both solid- and ascites-type Meth A fibrosarcoma as did the whole cells of S. pyogenes (OK-432). No other cellular components, such as cell wall peptidoglycan, group-specific C-carbohydrate or type-specific M protein, suppressed the growth of Meth A. LTA, but not the other cellular components, induced tumour necrosis factor (TNF) in Propionibacterium acnes-primed mice. LTA had no direct killing effects on Meth A cells. These results indicate that LTA may be an important antitumour component of OK-432 and that one of the antitumour mechanisms by this streptococcal preparation is the induction of TNF.

Topics: Animals; Antigens, Bacterial; Antineoplastic Agents; Bacterial Outer Membrane Proteins; Bacterial Proteins; Carrier Proteins; Dose-Response Relationship, Drug; Female; Fibrosarcoma; Lipopolysaccharides; Male; Mice; Mice, Inbred Strains; Peptidoglycan; Polysaccharides, Bacterial; Streptococcus pyogenes; Teichoic Acids; Tumor Necrosis Factor-alpha

Topics: Animals; Cell Line; Cell Survival; Female; Fibrosarcoma; Glycoproteins; Growth Inhibitors; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Mice, Inbred ICR; Phosphatidic Acids; Streptococcus pyogenes; Teichoic Acids; Tumor Necrosis Factor-alpha