Comparison of imatinib, dasatinib, nilotinib and INNO-406 in imatinib-resistant cell lines.

We compared the growth-inhibitory effects and inhibition profile of the SRC family kinases (SFKs) of imatinib, dasatinib, nilotinib and INNO-406. Dasatinib exhibited the strongest potency against BCR-ABL with little selectivity over SFKs. Nilotinib exhibited a weaker affinity than the other inhibitors, but was highly specific for ABL and may be useful for the treatment of P-glycoprotein overexpressing leukemic cells. INNO-406 had an intermediate affinity for BCR-ABL between that of dasatinib and nilotinib, and inhibited only SFKs LCK and LYN among SFKs. Both nilotinib and INNO-406 were potent inhibitors of the dasatinib-resistant T315A, F317L and F317V BCR-ABL mutations.

Curcumin prevents the development of dextran sulfate Sodium (DSS)-induced experimental colitis.

Curcumin is a phenolic natural product isolated from the rhizome of Curcuma longa (turmeric). We evaluated the effects of curcumin on the development of dextran sulfate sodium (DSS)-induced experimental colitis. BALB/c mice were fed a chow containing either 3.5% (wt/wt) DSS or 3.5% DSS + 2.0% (wt/wt) curcumin. The body weight loss was more apparent in DSS-treated mice than in DSS + curcumin-treated mice. The disease activity index, histological colitis score, and MPO activity were all significantly higher in DSS-treated mice than in DSS plus curcumin-treated mice. Microscopically, mucosal edema, cellular infiltration, and epithelial disruption were much more severe in DSS-treated mice than in DSS + curcumin-treated mice. In DSS + curcumin-treated mice, NF-kappaB activation was blocked in the mucosa. In conclusion, the development of DSS-induced colitis was significantly attenuated by curcumin. Being a nontoxic natural dietary product, curcumin could be useful in treatment of IBD patients.

INNO-406, a novel BCR-ABL/Lyn dual tyrosine kinase inhibitor, suppresses the growth of Ph+ leukemia cells in the central nervous system, and cyclosporine A augments its in vivo activity.

Central nervous system (CNS) relapse accompanying the prolonged administration of imatinib mesylate has recently become apparent as an impediment to the therapy of Philadelphia chromosome-positive (Ph+) leukemia. CNS relapse may be explained by limited penetration of imatinib mesylate into the cerebrospinal fluid because of the presence of P-glycoprotein at the blood-brain barrier. To overcome imatinib mesylate-resistance mechanisms such as bcr-abl amplification, mutations within the ABL kinase domain, and activation of Lyn, we developed a dual BCR-ABL/Lyn inhibitor, INNO-406 (formerly NS-187), which is 25 to 55 times more potent than imatinib mesylate in vitro and at least 10 times more potent in vivo. The aim of this study was to investigate the efficacy of INNO-406 in treating CNS Ph+ leukemia. We found that INNO-406, like imatinib mesylate, is a substrate for P-glycoprotein. The concentrations of INNO-406 in the CNS were about 10% of those in the plasma. However, this residual concentration was enough to inhibit the growth of Ph+ leukemic cells which expressed not only wild-type but also mutated BCR-ABL in the murine CNS. Furthermore, cyclosporine A, a P-glycoprotein inhibitor, augmented the in vivo activity of INNO-406 against CNS Ph+ leukemia. These findings indicate that INNO-406 is a promising agent for the treatment of CNS Ph+ leukemia.

The S1P receptor modulator FTY720 prevents the development of experimental colitis in mice.

To evaluate the therapeutic effects of the new synthetic sphingosine-1-phosphate (S1P) receptor modulator, FTY720, we investigated how FTY720 affects the development of dextran sulfate sodium (DSS)-induced colitis and CD4+CD62L+ T cell transfer colitis. BALB/c mice were fed a chow containing 3.5% (wt/wt) DSS to induce colitis. The CD4+CD62L+ T cell transfer colitis was induced by an intraperitoneal injection of CD4+CD62L+ spleen T cells into recipient CB17 SCID mice. The FTY720 was administered by lavage at a dose of 0.3 mg/kg/day. FTY720 was effective in preventing the body weight loss in the DSS-colitis model and the CD4+CD62L+ T cell transfer model. The disease activity index, histological colitis score, and MPO activity were all significantly lower in FTY720-treated mice than in the non-treated mice. Microscopically, mucosal edema, cellular infiltration and epithelial disruption were much more moderate in the FTY720-treated mice than in the non-treated mice. In both colitis models, FTY720 prevented the infiltration of CD4+ T cells into the inflamed colonic lamina propria. In conclusion, the development of DSS-colitis and CD4+CD62L+ T cell transfer colitis were significantly attenuated by FTY720. Since FTY720 is an immunosuppressive product that does not modulate T cell functions, it could be useful in the treatment of IBD patients.

Immunohistochemical study of chymase-positive mast cells in inflammatory bowel disease.

Mast cell-derived chymase promotes inflammatory responses and tissue fibrosis. Although previous studies have reported changes in the number of mucosal mast cells in inflammatory bowel disease (IBD), the behaviour of chymase immunopositive mast cells has not been studied. In this study, we immunohistochemically investigated chymase immunopositive mast cells in the inflamed mucosa of IBD patients. Surgically-obtained or biopsy specimens from 10 patients with ulcerative colitis (UC), 10 patients with Crohn's disease (CD) and 10 normal colorectal tissue specimens were used. The chymase immunopositive cells were identified by immunohistochemical analysis using a monoclonal anti-human chymase antibody. In the normal colonic mucosa, a small number of chymase immunopositive mast cells were detected at the basal sites of the mucosa. There were no immunopositive cells in the submucosa. Chymase immunopositive mast cells were similarly observed in the inactive UC mucosa, but these cells decreased significantly in number in the active UC mucosa. In the inactive CD mucosa, the number of chymase immunopositive mast cells increased significantly (P < 0.05), and this was more clearly observed in the active CD mucosa. Furthermore, in the active CD mucosa, these cells were detected in the submucosa, propria muscularis, and surrounding fatty tissue. These observations suggest a crucial role for chymase immunopositive mast cells in the pathophysiology of CD. Since intestinal fibrotic changes such as stricture formation are a characteristic feature of CD, chymase immunopositive mast cells may act as a stimulus for the process of tissue fibrosis and tissue remodelling in the pathophysiology of CD.

Interleukin-17 augments tumor necrosis factor-alpha-induced granulocyte and granulocyte/macrophage colony-stimulating factor release from human colonic myofibroblasts.

BACKGROUND: Interleukin (IL)-17 is a newly identified T-cell-specific cytokine. In this study, we investigated the effects of IL-17 on colony-stimulating factor (CSF) release in human colonic subepithelial myofibroblasts (SEMFs). METHODS: CSF release and mRNA expression were determined by enzyme-linked immunosorbent assay (ELISA) and Northern blotting, respectively. Nuclear factor (NF)-kappaB- and activating protein (AP-1)-DNA binding activities were evaluated by electrophoretic gel mobility shift assays (EMSAs). RESULTS: Unstimulated cells secreted a small amount of granulocyte G- and granulocyte/macrophage (GM)-CSF, and a considerable amount of M-CSF. IL-17 weakly enhanced G-CSF release, but did not affect GM- and M-CSF release. IL-17 selectively enhanced tumor necrosis factor (TNF)-alpha-induced G- and GM-CSF release. The combination of IL-17 plus TNF-alpha induced a marked increase in NF-kappaB- and AP-1-DNA binding activities. The adenovirus-mediated transfer of a stable form of IkappaBalpha and/or a dominant negative mutant of c-Jun markedly inhibited the IL-17 plus TNF-alpha-induced G- and GM-CSF mRNA expression. Furthermore, a stability study showed that IL-17 plus TNF-alpha markedly enhanced the stability of G- and GM-CSF mRNA. CONCLUSIONS: IL-17 augments TNF-alpha-induced G- and GM-CSF release via transcriptional and posttranscriptional mechanisms.

Development of dextran sulfate sodium-induced colitis is aggravated in mice genetically deficient for complement C5.

The complement system is a potent effector of innate immunity. To elucidate the pathophysiological role of the complement system in inflammatory bowel disease (IBD), we evaluated the development of dextran sulfate sodium (DSS)-induced colitis in genetically complement C5-deficient mice. We used DBA2/J mice, which are genetically deficient in complement C5. DBA1/J mice have a normal complement system, and were used as controls. Experimental colitis was induced by the oral administration of 3.5% (w/v) DSS in their drinking water for 10 days. On day 10, all mice were sacrificed and their colons were collected. The development of colitis was assessed by the histological score, disease activity index, myeloperoxidase (MPO) activity, and macroscopic changes of the colon. Body weight loss was more apparent in the DBA2/J mice than in control DBA1/J mice. The colon length was shorter in the DBA2/J mice than in DBA1/J mice. The disease activity index, histological colitis score, and MPO activity were all significantly higher in the DBA2/J mice than in DBA1/J mice. Microscopically, mucosal edema, cellular infiltration and disruption of the epithelium were much more severe in the DBA2/J mice than in DBA1/J mice. The development of DSS colitis was aggravated in genetically C5-deficient DBA2/J mice. These findings suggest that the complement system might play a protective role in the development of DSS-induced experimental colitis.

Interleukin-22, a member of the IL-10 subfamily, induces inflammatory responses in colonic subepithelial myofibroblasts.

BACKGROUND & AIMS: Interleukin (IL)-22, a member of the IL-10 subfamily, is a recently identified T-cell-derived cytokine. We investigated IL-22 expression in the inflamed mucosa of patients with inflammatory bowel disease (IBD) and analyzed its biologic activities in human colonic subepithelial myofibroblasts (SEMFs). METHODS: Mucosal IL-22 expression was evaluated by immunohistochemical procedures. The effects of IL-22 on colonic SEMFs were investigated by cDNA microarrays, Northern blots, enzyme-linked immunosorbent assay, and electrophoretic gel mobility shift assays (EMSAs). RESULTS: IL-22 was not detectable in normal colonic mucosa. In IBD mucosa, IL-22 expression was detectable in CD4-positive T cells. IL-22-positive cells were increased in ulcerative colitis and even more so in Crohn's disease. IL-22 receptor expression colocalized with a marker of SEMFs. IL-22 did not modulate SEMF proliferation and collagen synthesis. cDNA microarray analyses demonstrated that, in colonic SEMFs, IL-22 increased the messenger RNA (mRNA) expression of inflammatory cytokines (IL-6, IL-8, IL-11, and leukemia inhibitory factor [LIF]), chemokines, and matrix metalloproteinases. IL-22 induced an activation of nuclear factor (NF)-kappaB and activating protein (AP)-1 within 1 hour, and a blockade of NF-kappaB and AP-1 activation markedly reduced IL-22 induction of IL-6, IL-8, IL-11, and LIF mRNA. MAP-kinase inhibitors (PD98059, U0216, and SB202190) significantly reduced IL-22 induction of cytokine secretion. The combination of either IL-17 plus IL-22 or IL-19 plus IL-22 additively up-regulated cytokine secretion. CONCLUSIONS: IL-22 derived from activated T cells acts on SEMFs to elicit expression of proinflammatory cytokines and matrix-degrading molecules indicating proinflammatory/remodeling roles in IBD.

Leukocytapheresis therapy modulates circulating t cell subsets in patients with ulcerative colitis.

The aim of this study was to elucidate the molecular mechanisms responsible for the therapeutic effects of leukocytapheresis (LCAP). We investigated the alterations in circulating T cell subsets after LCAP therapy in ulcerative colitis (UC) patients. Eighteen patients with UC were enrolled. Fourteen patients were responders, and four patients were non-responders. Peripheral venous blood was obtained within 5 min before and 5 min after LCAP therapy. Flow cytometric analysis for T cell markers and intracellular interferon (IFN)-gamma (Th1) and interleukin (IL)-4 (Th2) was then performed. The average numbers of lymphocytes, T and B cells were significantly decreased after LCAP therapy, respectively (P < 0.01). The numbers of CD4+ and CD8+ T cells were also significantly decreased, respectively (P < 0.01), but the CD4+/CD8+ ratio was not changed. The number of CD45RO+ CD4+ memory T cells was significantly decreased. The number of CD25+ CD4+ T cells tended to decrease after LCAP therapy (not significant). However, the ratio of CD25+ CD4+-cells/CD25- CD4+-cells was significantly increased (P < 0.05). The number of IFN-gamma-positive (Th1) cells was significantly decreased after LCAP therapy, but there was no significant change in the number of IL-4-positive (Th2) cells. The Th1/Th2 ratio was significantly decreased after LCAP therapy. Some of the immuno-suppressive effects of LCAP therapy may be associated with a modulation of circulating T cell subsets.

Suppression of inflammatory cytokine secretion by granulocyte/monocyte adsorptive apheresis in active ulcerative colitis.

To elucidate the molecular mechanisms involved in the therapeutic effects of granulocyte/monocyte adsorption apheresis, changes were investigated in the cytokine responses of peripheral blood mononuclear cells (PBMC) before and after granulocyte/monocyte adsorptive apheresis in ulcerative colitis (UC) patients. Four patients with active UC were enrolled. All patients responded to granulocyte/monocyte adsorptive apheresis. A total of 20 sessions of four patients were analyzed. Peripheral blood mononuclear cells were isolated from peripheral venous blood within 5 min before and after each session of granulocyte/monocyte adsorptive apheresis. The cells were stimulated with interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha for 24 h, and the secreted IL-8 and IL-6 levels were determined by enzyme-linked immunosorbent assay (ELISA). IL-1beta-induced IL-8 and IL-6 secretion was significantly decreased after granulocyte/monocyte adsorptive apheresis. TNF-alpha-induced IL-8 secretion was also significantly decreased after apheresis, but there was no significant difference in TNF-alpha-induced IL-6 secretion (P = 0.052). In conclusion, granulocyte/monocyte adsorptive apheresis down-regulates the IL-1beta- and TNF-alpha-induced inflammatory responses in PBMC. The induction of hyporesponsiveness to pro-inflammatory cytokines may be an important factor mediating the clinical effects of granulocyte/macrophage adsorptive apheresis in UC patients.

[A case of advanced gastric cancer with pulmonary carcinomatous lymphangiosis responding remarkably to combination chemotherapy of docetaxel (TXT) and TS-1].

A 51-old-female patient was admitted because of dyspnea. This case was diagnosed inoperable advanced gastric cancer and pulmonary carcinomatous lymphangiosis. She was treated by combination of docetaxel (TXT) and TS-1. TXT (40 mg/m2) was administered on day 1, and TS-1 (80 mg/body/day) was then administered for 14 days followed by a 7-day interval as one course. After two courses of chemotherapy, carcinomatous lymphangiosis declined, tumor markers decreased, and dyspnea improved. Administration of oxygen was thus discontinued. No side effects appeared (hematological or non-hematological).