$56 Journal of Clinical Virology 2006, Vol 36 (suppl 2) Abstracts, 12th ISHVLD In addition this approach might also be important for the develop- ment of novel antiviral strategies, as has been shown for gp-41- derived T 20-peptides (Fuzeon) in HIV-therapy. We have recently demonstrated that authentically myristoylated or otherwise acylated peptides encompassing the N-terminal 47 pre-S1 amino acids of the HBV L-protein block HBV infection of primary human and tupaia belangeri hepatocytes and HepaRG-cells with surprising efficacy, at already picomolar concentrations, probably by sustained inactiva- tion of a receptor on hepatocytes (J. Virol., 79:1613-1622 (2005); Gastroenterology, 129:234-245 (2005)). Methods: To test the potency of acylated HBV preS1 peptides to interfere with HBV infection also in vivo, we took advantage of the urokinase-type plasminogen activator (uPA) mouse model that supports the entire hepadnavirus replication cycle after liver repopulation with susceptible hepatocytes from humans or tupaia belangeri (J. Hepatol., 42:54-60 (2005)). Results: We here report that treatment of mice with two dif- ferent acylated preS1 peptides (HBVpreS/2-39myr (IC50-300nM) and HBVpreS/2-48stearoyl (IC50-250pM) completely inhibited the establishment of an HBV infection in vivo. HBsAg and viral DNA remained undetectable in serum of all mice (n = 6) for at least five months after inoculation (ongoing study), while establishment of chronic productive infection with high viral titers was achieved in control mice treated with a myristoylated peptide derived from the Heron Hepatitis B Virus preS domain (HHBVpreS/2-44myr). The histochemical analysis of two treated mice revealed no HBV specific antigen staining 24 weeks after virus inoculation. Using 1251 labeled H BVpreS/2-48myr we further investigated the organ distribution with respect to a specific targeting to the transplanted hepatocytes, as well as the peptide stability in serum. Conclusion: Our preclinical study proofs the principle that acy- lated preS-peptides are promising candidates for future clinical appli- cations, e.g. prevention of HBV/HDV reinfection after liver transplan- tation, post exposure prophylaxis or horizontal transfer of HBV/HDV from an infected mother to her child. It remains an open question, if efficient entry inhibition in combination with established therapies that reduce the number of infected hepatocytes can improve the therapeutic outcome in chronically infected patients. I-~-~ Drug-drug interaction between clevudine in combination with telbivudine or 3TC in human liver cells B.I. Hernandez-Santiago*, D.C. Delinsky, R.F. Schinazi. Pediatrics, Emery School of Medicine, VA Medical Center, Decatur, USA Background and Objectives: Clinical experience using approved drugs for HBV such as lamivudine and adefovir dipivoxil indicates that most subjects will require long-term therapy to maintain viral load suppression. Persons on monotherapy regimens that have inadequate viral suppression may benefit from add-on therapy or start their regimens with a combination. An understanding of inter- actions between nucleoside analogs is critical for the development of improved combinations, since this class of compound requires phosphorylation by cellular kinases. Drug~lrug interactions that decrease intracellular phosphorylation are likely to decrease their antiviral potency. The objective was to study potential drug-drug interactions at the triphosphate level between clevudine (CLV), telbivudine (LdT) and 3TC in human liver cell lines. This is important because 3TC, CLV and LdT are all substrates of 2/-deoxycytidine kinase (dCK), whereas CLV and LdT are also substrates of cytosolic thymidine kinase (TK1). Methods: Competition studies were conducted by co-incubation of either CLV with radiolabeled-3TC, radiolabeled-CLV with 3TC or LdT, or CLV with radiolabeled-LdT in HepG2 cells at equal concen- tration ratios. Samples were analyzed using reverse-phase HPLC and quantified by Perkin Elmer Beta-Counter. 2/-Deoxycytidine and thymidine were used as positive controls since they both compete with the phosphorylation of CLV and LdT. Similar studies with CLV in combination with 3TC were conducted using an LC-MS/MS approach that can simultaneously measure the nucleoside or nu- cleotide analogs. Results: No changes in the levels of nucleoside triphosphate were observed when radiolabeled-CLV was co-incubated with LdT or when CLV was co-incubated with radiolabeled-3TC. However, co- incubation of CLV with radiolabeled-LdT or radiolabeled-CLV with 3TC significantly reduced the levels of LdT triphosphate (LdT-TP) from 18.11±3.06 to 5.58±0.83 pmol/106 cells or CLV triphosphate (CLV-TP) from 32.05±6.55 to 22.49±1.69 pmol/106 cells, respec- tively (p < 0.05). Conclusion: Competition studies at equivalent concentrations showed that CLV decreased the levels of LdT-TP (3.2-fold) and 3TC reduced the levels of CLV-TP (1.4-fold) in HepG2 cells, but CLV did not change 3TC-TP concentrations and LdT did not affect CLV- TP levels. Interestingly, even when CLV-TP levels decreased in the presence 3TC, the level of the active metabolite remained well above the 50% inhibitory concentration (IC50 = 0.1 uM). This work needs to be further substantiated using an LC-MS/MS approach, virological studies, and well designed clinical trials. (BHS was supported by a supplement of NIH grant 5R37-AI-41980. This work was also supported in part by NIH 5P30-AI-50409, and the Department of Veterans Affairs). PS30 - HCV: Immunopathogenesis: cellular, humoral and innate response (Tuesday, July 4, 2006, 14:00-15:30) IO• Differential responsiveness of CD4 and CD8 T-cells during acute HCV re-infection S. Longworth 1, J. Schulze zur Wiesch 1, A.Y. Kim 1, T. Kuntzen 1, J. Timm 1, A. Jones 1, R.T. Chung 2, T.M. Allen 1, B.D. Walker 1, G.M. Lauer 1 *. 1Partners AIDS Research Center and Howard Hughes Medical Institute, Massachusetts General Hospital and Harvard Medical School, Charlestown; 2GI Unit, Massachusetts General Hospital and Harvard Medical School, Boston, USA Background and Objectives: There is clinical evidence for partially protective immunity after spontaneous resolution of HCV infection in humans. Chimpanzee experiments suggest that protection is critically mediated by cellular immune responses, but studies in humans are lacking. Methods: The current study investigates 4 subjects with acute HCV re-infection. 2 subjects had previously eliminated HCV spon- taneously and 2 subjects had cleared on therapy for acute and chronic infection, respectively. We assessed HCV-specific CD4 and CD8 responses by ELISpot, CD4 proliferative assays and tetramer staining. Virus was sequenced using standard methods. Results: In all 4 subjects we detected an HCV strain distinct from that isolated during the first infection, indicating that the patients did not experience viral recurrence but indeed re-infection. Subjects had been aviremic for between four months and three years. Three subjects cleared reinfection spontaneously and only the subject who had been treated for chronic infection earlier again required antiviral therapy. We detected a significant expansion of HCV-specific CD8+ T-cells in all subjects, including the one not controlling the virus. A majority of CD8 responses had been already detected during the first episode of HCV infection and were cross-reactive with both HCV strains. In contrast, vigorous CD4 responses were exclusively detected in the three subjects clearing reinfection spontaneously. Such CD4 responses had been detected during the first episode of infection in the two subjects clearing HCV spontaneously, but were new in the one subject treated for acute infection earlier, in whom they had been neither detected during acute HCV infection nor during successful therapy. In the subject requiring two courses of antiviral therapy CD4 responses were not detected during acute re-infection or either course of therapy. Conclusion: Acute HCV reinfection is characterized by a mas- sive expansion of memory CD8 T-cells, irrespective of outcome. In contrast, HCV-specific CD4 T-cell responses are not uniformly detected but seem to be critical for viral control. At least in the one subject reinfected after treatment of chronic HCV infection, successful antiviral treatment had not led to recovery of an effective CD4 response and virus persisted despite robust CD8 responses. Our results support the hypothesis that spontaneous resolution of