C型肝炎病毒是一個帶有正股RNA的套膜病毒。病毒套膜醣蛋白質E1和E2被證實對C型肝炎病毒的組裝是重要的，因此阻止其折疊成具有功能的蛋白質就變成了抗C型肝炎病毒的策略之一。我們建立一個以C型肝炎假病毒為主體的篩選系統，藉以篩選由影響病毒套膜醣蛋白而參與了C型肝炎病毒組裝的小分子。Iminosugars抑制位在粗糙內質網的glucosidase，藉此影響病毒的形成。在上百種的化合物中，我們篩選出五種有效化合物，化合物B6跟A6被進一步證實可以抑制病毒套膜醣蛋白質E2的醣化過程及C型肝炎病毒顆粒的組裝但不影響病毒的複製；另外也發現B6影響病毒進入宿主的能力。這些研究證實了我們建立了一個抑制C型肝炎病毒顆粒組裝的藥物篩選平台，並所篩選到的化合物有機會可發展成治療C型肝炎的先導型藥物。另一方面，Disabled-2是一個adaptor蛋白質分子，可以改變上皮細胞的cell polarity。我們建立了可以穩定抑制Dab2蛋白質表現量的HepG2細胞株，並發現Dab2影響了HepG2細胞株的cell polarity，並進一步影響了C型肝炎病毒的包裹與分泌。
HCV is an enveloped, positive-stranded RNA virus. Since envelope glycoprotein E1 and E2 of HCV are important for virus particle formation, blocking viral glycoprotein folding should be one of strategies to inhibit HCV virion assembly. For the purpose, a HCV pseudotyped virus (HCVpv)-based screen system was established to find small moleculars belonging to iminosugars derivates which inhibit the endoplasmic reticular (ER) glucosidase and may further affect the morphogenesis of viruses. The HCV-pv based screen system was validated by a well known inhibitors of the alpha-glucosidases, N-butyl-deoxynojirimycin (NB-DNJ) on both HCV cell culture viral particle (HCVcc) and HCVpv-based screen system. Among more than one hundred of compounds, five compounds were selected for advanced purification and two of the compounds B6 and A6 were further studied the activities in HCV virion assembly and secretion. These two compounds can interrupt the glycosylation processing of E2 thereby affect the viral assembly and secretion but not replication. Furthermore, B6 can also inhibit HCV viral entry. On the other side, an adapt protein Disabled-2 which is thought as an epithelial surface positioning gene. To understand the effect of loss of polarized distribution of envelope glycoprotein in HCV assembly and secretion, Dab2 knock-down stable HepG2 cell lines was established. The polarity of Dab2-knock-down cells was decreased.Furthermore, HCV assembly and secretion was increase in Dab2-knock-down HepG2 cells.
Taken together, a HCVpv-based screen system was established for drug-screening on HCV envelope protein for viral assembly/and secretion.A lead compound B6 was discovered to target on HCV envelope glycoprotein.Besides, Dab-2 disrupted hepatocyte polarity may promote HCV assembly and secretion in HepG2 cells. These two molecules may serve as potential for designing and developing drugs for treatment of HCV infection.
