Tetsuya Suga

In vitro and in vivo analysis of human leukocyte binding by the antitumor polysaccharide, lentinan

Lentinan (LNT), a (1-->3)-beta-D-glucan with (1--6)-beta-D-glucopyranoside branches, has marked antitumor effects in syngeneic and autochtonous hosts. Clinically, LNT has proved effective with chemotherapeutic agents in patients with recurrent gastric and colorectal cancer. However, the mechanism that triggers subsequent immunologic reactions remains obscure. We hypothesized that LNT must first bind to the host cells. Accordingly, we analyzed LNT binding to host cells in healthy volunteers after incubating their cells under a variety of conditions as well as intravenously injecting LNT then subjecting them to flow cytometry and immunofluorescent staining using monoclonal antibody (anti-LNT mAb). LNT bound to monocytes and neutrophils, but not to lymphocytes in vitro. The most avid LNT binding was to monocytes. The percentage of LNT-bound monocytes after 60 min incubation at 4 degrees C was greater than that at 37 degrees C. The binding of LNT to monocytes was inhibited slightly by anti-iC3b receptor (anti-CR3) mAb, strongly by anti-C3b receptor (anti-CR1) mAb, and completely by anti-CR1 and anti-CR3 mAb together. The percentages of LNT-binding monocytes in the peripheral blood increased significantly 3 and 4 after 2 mg LNT-injection and returned to low levels 5 h later. However, no increase in LNT-binding neutrophils and lymphocytes was observed. We concluded that binding of LNT to human monocytes may initiate the influence of this compound on the immune system and differ between individuals. Its binding site may be similar to the C3b receptor.

Macrophage-mediated acute-phase transport protein production induced by lentinan

A new bioactive factor capable of stimulating the production of acute-phase transport proteins, haptoglobin, hemopexin and ceruloplasmin, was found in mouse serum soon after the administration of lentinan, an immunomodulatory polysaccharide. This factor (APPIF) was produced by macrophages, and may regulate the production of acute-phase transport proteins in hepatocytes. The mice given the serum obtained from donor mice 2-6 h after an injection of 10 mg/kg of lentinan showed a marked increase of the acute-phase transport proteins in their serum 4 days after the serum injection. Pretreatment with the antimacrophage agents, carrageenan and mouse Ia antiserum, before the lentinan treatment to donor mice inhibited the production of acute-phase transport proteins in the recipient mice. Thymus or T-cells had no role in the production of APPIF. As the activity of APPIF disappeared after pronase treatment of the serum, APPIF seems to be a peptide compound. Appearance of APPIF is considered to be one of the earliest manifestations of the mode of action of lentinan in addition to its augmented production of vascular dilatation and hemorrhage inducing factor and interleukin-1. The correlation between these inflammatory and immune responses in earlier stages of the host defence mechanisms is also discussed.

Synergistic Antimetastatic Effects of Lentinan and Interleukin 2 with Pre‐ and Post‐operative Treatments

The antimetastatic activity of a combination of lentinan and interleukin 2 (IL‐2) was evaluated against spontaneously metastatic 3‐methylcholanthrene‐induced DBA/2.MC.CS.T fibrosarcoma. Although pre‐operative treatment with either IL‐2 or lentinan alone exerted little effect on the reduction of lung metastasis colony numbers (7.1% or 28.4% reduction, respectively), the combination exhibited a synergistic effect (85% reduction). Furthermore, 3 of 13 mice given the pre‐operative combination treatment achieved complete cure, while no mice given saline did. Although the post‐operative combination treatment also reduced the colony number (71% reduction), it caused little prolongation of survival and no mouse achieved complete cure. Synergistic effects were observed between pre‐ and post‐operative treatments with lentinan and IL‐2: 8 of 12 mice were completely cured. The anti‐metastatic activity was abolished in mice treated simultaneously with antibodies to CD4 and CDS antigens, whereas either CD4, CDS, or NK1.1 antibody alone was ineffective. Analysis of the cellular mechanism involved in the antimetastatic activity revealed the involvement of a tumor‐associated antigen‐specific delayed‐type hypersensitivity response. These data suggest that the life‐prolonging effect of the combination of lentinan and IL‐2 is mediated by antigen‐specific T cells and that the combination of pre‐ and post‐operative therapy with lentinan and IL‐2 may be effective to prevent cancer recurrence and metastasis after surgical resection.

Antitumor activity of a new series of platinum complexes: trans(+/-)-1,2-cyclohexanediammineplatinum(II) conjugated to acid polysaccharides.

Complexes with trans (±)-1,2-cyclohexanediammineplatinum(II) conjugated to acid polysaccharides were synthesized and their antitumor activities were tested in female CDF1 mice with intraperitoneal leukemia L1210 cells. Platinum was released from the polymers under physiological conditions, with half-lives from 3.3 to 19.3 h. A hyaluronic acid-supported complex was the most effective against the tumors (all six mice survived for 60 days). The group given a chondroitin polysulfate-supported complex had five survivors, the chondroitin sulfate A group also had five, the chondroitin sulfate C group had three and the heparan sulfate group had two. Part of the antitumor activity was due to increased efficacy of the polymers. The bioavailability of these complexes is high. Therefore, acid polysaccharides should be a good system for delivering antitumor platinum complexes.

Individual diversity of IL-6 generation by human monocytes with lentinan administration.

Effect of in vivo administration of lentinan on IL-6 generation by human peripheral blood monocytes was examined using 5 healthy volunteers. Monocytes isolated from them before and 3 days after intravenous administration of 2 mg lentinan per person were cultured for 2 days and then the levels of IL-6 in the supernatants of cultured monocytes were determined. In vivo lentinan administration elicited an increase in IL-6 generation by monocytes in 3 of 5 cases. Failure to increase IL-6 generation by monocytes in two cases is characteristic of lentinan, since these monocytes generated an apparent level of IL-6 in response to lipopolysaccharide, a potent monocyte activator. Thus, it was demonstrated that lentinan was capable of augmenting IL-6 generation by human monocytes and that there was individual difference in their responsiveness to lentinan.