ABSTRACT
AIM: Tacrolimus (TAC) is an important drug for inflammatory diseases. However, TAC has several limitations, such as variable trough concentrations among individuals and a high medication frequency. In this study, we created NK61060, a novel micellar TAC formulation, to circumvent these disadvantages. MATERIALS & METHODS: Immunosuppressive activity of NK61060 was determined in the collagen-induced arthritis rat model, mannan-induced arthritis mouse model and dextran sodium sulfate-induced colitis mouse model. The pharmacokinetics and toxicology of NK61060 were evaluated in those models. RESULTS: In arthritis and colitis models, NK61060 exhibited superior immunosuppressive activity compared with that of TAC. Pharmacokinetic and toxicological analyses indicated that NK61060 had a wider safety margin and could be administered at a reduced medication frequency. CONCLUSION: NK61060 mitigates the trough concentration variability and the medication frequency and it may be a safer and more effective option for use in clinical settings. Further studies are needed to determine its clinical usefulness.
Subject(s)
Arthritis, Experimental/drug therapy , Arthritis, Rheumatoid/drug therapy , Colitis, Ulcerative/drug therapy , Drug Carriers/chemistry , Immunosuppressive Agents/administration & dosage , Tacrolimus/administration & dosage , Animals , Area Under Curve , Arthritis, Experimental/immunology , Arthritis, Rheumatoid/immunology , Colitis, Ulcerative/chemically induced , Colitis, Ulcerative/immunology , Collagen/immunology , Dextran Sulfate/toxicity , Drug Administration Schedule , Drug Evaluation, Preclinical , Female , Humans , Mannans/immunology , Mice , Mice, Inbred ICR , Micelles , Polyethylene Glycols/chemistry , Rats , Rats, Sprague-DawleyABSTRACT
YTH domain containing 2 (YTHDC2) is a member of the DExD/H-box family of ATP-dependent RNA helicases. We previously found that YTHDC2 expression is up-regulated in several human cancer cells. In this study, we demonstrate novel roles for YTHDC2 in metastasis of colon tumor cells via translation-dependent pathway. Knockdown of YTHDC2 attenuated protein expression of metastasis-related genes, such as hypoxia-inducible factor-1alpha (HIF-1α), and inhibited metastasis of colon tumor cells in vitro and in vivo. To confirm that YTHDC2 promotes translation initiation by unwinding the 5'-untranslated region (5'UTR) of mRNA, we constructed a firefly luciferase reporter containing the 5'UTR of the HIF-1α mRNA and showed reduction in luciferase activity in YTHDC2-silenced cells. Furthermore, we examined expression levels of YTHDC2 by immunohistochemical staining in human colon cancer tissues from 72 patients and found a significantly positive correlation between YTHDC2 expression and the tumor stage, including metastasis. In conclusion, these results suggest that the RNA helicase YTHDC2 contributes to colon tumor metastasis by promoting translation of HIF-1α and that YTHDC2 is potentially a diagnostic marker and target gene for treating colon cancer patients.
Subject(s)
Adenocarcinoma/enzymology , Adenosine Triphosphatases/metabolism , Cell Movement , Colonic Neoplasms/enzymology , Hypoxia-Inducible Factor 1, alpha Subunit/biosynthesis , RNA, Messenger/metabolism , 5' Untranslated Regions , Adenocarcinoma/genetics , Adenocarcinoma/secondary , Adenosine Triphosphatases/genetics , Animals , COS Cells , Chlorocebus aethiops , Colonic Neoplasms/genetics , Colonic Neoplasms/pathology , Female , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Genes, Reporter , HCT116 Cells , HT29 Cells , Heterografts , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Mice, Inbred BALB C , Mice, Nude , Neoplasm Metastasis , Neoplasm Staging , Neoplasm Transplantation , Nuclear Proteins/metabolism , RNA Helicases , RNA Interference , RNA, Messenger/genetics , Signal Transduction , Time Factors , Transfection , Tumor Hypoxia , Twist-Related Protein 1/metabolism , Up-RegulationABSTRACT
We previously demonstrated that a cellular factor, cyclosporin A (CsA) associated helicase-like protein (CAHL) that is identical to YTH domain containing 2 (YTHDC2), forms trimer complex with cyclophilin B and NS5B of hepatitis C virus (HCV) and facilitates HCV genome replication. Gene expression of YTHDC2 was shown in tumor cell lines and tumor necrosis factor (TNF)-α-treated hepatocytes, but not in untreated. However, the function of YTHDC2 in the tumor cells and the mechanism by which the YTHDC2 gene is transcribed in these cells is largely unknown. We first evaluated that the role of YTHDC2 in the proliferation of hepatocellular carcinoma (HCC) cell line Huh7 using RNA interference and found that YTHDC2-downregulated Huh7 were significantly decreased cell growth as compared to control. We next demonstrated that the cAMP response element (CRE) site in the promoter region of the YTHDC2 gene is critical for YTHDC2 transcription. To further investigate the transcription factors bound to the CRE site, we performed chromatin immunoprecipitation assays. Our findings demonstrate that c-Jun and ATF-2 bind to the CRE site in Huh7, and that TNF-α induces the biological activity of these transcription factors in hepatocytes as well as Huh7. Moreover, treatment with the HDAC inhibitor, trichostatin A (TSA), reduces YTHDC2 expression in Huh7 and in TNF-α-stimulated hepatocytes. Collectively, these data show that YTHDC2 plays an important role in tumor cells growth and activation/recruitment of c-Jun and ATF-2 to the YTHDC2 promoter is necessary for the transcription of YTHDC2, and that HDAC activity is required for the efficient expression of YTHDC2 in both of hepatocyte and HCC cells.
