Takuma Sakurai

Induction of an endogenous tumor necrosis factor in mice by murine recombinant interferon-γ combined with a lipid A subunit analog (GLA-60) of low toxicity

SummaryWe have investigated the endogenous production of a serum cytotoxic factor when recombinant interferon-γ (rIFN-γ) is combined with synthetic lipid A subunit analogs of low toxicity (GLA compounds). The cytotoxic activity of the serum was measured by the crystal violet staining method with L929 cells as a target. Intravenous administration of rIFN-γ followed by intravenous administration of lipopolysaccharide induced the endogenous production of a cytotoxic factor in the serum. The priming effect of rIFN-γ appeared immediately and persisted for approximately 20 h after the injection. Administration of lipopolysaccharide as a trigger enhanced the production of the cytotoxic factor in the serum maximally 2 h after the injection. The cytotoxic activity in the serum was completely inhibited by anti-(mouse tumor necrosis factor) (TNF) antibody. A synthetic lipid A subunit analog (GLA-60), which is much less toxic in its endotoxin activities than lipopolysaccharide or synthetic lipid A (compound 506), induced the endogenous production of serum TNF in rIFN-γ-primed mice. GLA-60 entrapped within liposomes induced the production of serum TNF in rIFN-γ-primed mice more effectively than GLA-60 solubilized in phosphate-buffered saline. Intravenous or intranasal administrations of rIFN-γ followed by intranasal administration of GLA-60 produced TNF in the lung washing fluid but not in the serum, indicating that TNF production can be induced locally rather than systemically by the alteration of the administration route of the primer and trigger. These results indicate that GLA-60, a lipid A subunit analog of low toxicity, is a beneficial triggering agent in the production of endogenous TNF, as well as having other immunopharmacological properties, and may provide a basis for cancer (metastases) treatment as a result of its ability to induce endogenous TNF.

Chemical synthesis and biological activities of 6,6′-di-O-mycoloyl-β,β- and -α,β-trehalose

Abstract 6,6′-Di- O -mycoloyl-β,β-trehalose (β,β-TDM) and 6,6′-di- O -mycoloyl-α,β-trehalose (α,β-TDM) were synthesized and their toxicity and ability to activate peritoneal macrophages in situ were examined in mice, in comparison with 6,6′-di- O -mycoloyl-α,α-trehalose (TDM). Both β,β-TDM and α,β-TDM caused a decrease in body weight two days after injection, however the weights reverted to a normal level. No deaths were caused by either analog. On the other hand, TDM showed potent toxicity, causing decrease in body weight and death of all animals injected. β,β-TDM and α,β-TDM were effective in the in situ activation of mouse peritoneal macrophages.

Comparative study and effects of macrophage colony-stimulating factor (M-CSF) administration on NK1.1+ cells in mouse spleen and bone marrow.

ABSTRACT We conducted a comparative study of NK1.1+ cells in spleen and bone marrow and the effects of administration of M-CSF on them. Administration of M-CSF to mice increased the number of NK1.1+ cells in spleen but not in bone marrow. The NK1.1+ cells in spleen (Spl-NK1.1) and bone marrow (BM-NK1.1) were purified by magnetic cell sorter. Their cell surface markers and functions were then examined. The percentage of Mac-1 antigen-positive cells (and F4/80 antigen-positive cells) was higher among BM-NK1.1 than Spl-NK1.1. Moreover, the administration of M-CSF increased the number of Mac-1 and F4/80 antigen-positive cells in both Spl-Nk1.1 and BM-NK1.1. The functions (cytolytic activity and IFN-γ production) of Spl-NK1.1 and BM-NK1.1 were the same and were enhanced by the administration of M-CSF. But Spl-NK1.1 produced more IFN-γ than BM-NK1.1 when M-CSF was administered. BM-NK1.1 showed a greater proliferative response to IL-2 than Spl-NK1.1. Administration of M-CSF augmented this response. BM-NK1.1 proliferated in response to IL-4 and IL-15, but Spl-NK1.1 responded only slightly. However, administration of M-CSF stimulated Spl-NK1.1 to respond to these cytokines. Both Spl-NK1.1 and BM-NK1.1 showed only a weak response to M-CSF in vitro. But the expression of c-fms antigen (M-CSFR) increased after the M-CSF injections in vivo. These results suggested that there are phenotypical and functional differences between Spl-NK1.1 and BM-NK1.1. The administration of M-CSF led to an accumulation of NK1.1+ cells which were mobilized from bone marrow in spleen.

Synthesis and immunoadjuvant activity of 2,2′-O-[2,2′-diacetamido-2,3,2′,3′-tetradeoxy-6,6′-di-O-(2-tetradecyl-hexadecanoyl)-α, α′-trahalose-3,3′-diyl]bis(N-d-lactoyl-l-alanyl-d-isoglutamine)

Abstract The disaccharide of 6- O -(2-tetradecylhexadecanoyl)muramoyl dipeptide coupled through an α-(1↔1)-α linkage, named in the title, and an analog bearing a single peptide moiety, have been synthesized from 2,2′-diazido-2,2′-dideoxy-α,α′-trehalose. The immunoadjuvant activities of the products were examined.

Chemical combination of 6-deoxy-6-mycoloylamino-α,α-trehalose and N-acetyl-6-O-(aminoacyl)muramoyl dipeptide

Abstract 6-Deoxy-6-mycoloylamino-α,α-trehalose, a biologically active derivative of 6,6′-di- O -mycoloyl-α,α-trehalose (TDM), and N -acetyl-6- O -(aminoacyl)-muramoyl dipeptide (MDP) were joined chemically by a succinic acid unit. The compounds synthesized showed activities that are characteristic of both TDM and MDP.