Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019163 (hepatitis B)
 38,309 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The anti-hepatitis B (anti-HBV) activities of the (-) and (+) enantiomers of cis-5-fluoro-1-[2-(hydroxymethyl)-1,3-oxathiolan-5-yl]cytosine (2'-deoxy-3'-thia-5-fluorocytosine [FTC]) were studied by using an HBV-transfected cell line (HepG2 derivative 2.2.15, subclone P5A). The (-) isomer was found to be a potent inhibitor of viral replication, with an apparent 50% inhibitory concentration of 10 nM, while the (+) isomer was found to be considerably less active. Both isomers showed minimal toxicity to HepG2 cells (50% inhibitory concentration, > 200 microM) and showed minimal toxicity in the human bone marrow progenitor cell assay. In accord with the cellular antiviral activity data, the 5'-triphosphate of (-)-FTC inhibited viral DNA synthesis in an endogenous HBV DNA polymerase assay, while the 5'-triphosphate of the (+) isomer was inactive. Unphosphorylated (-)-FTC did not inhibit product formation in the endogenous assay, suggesting that the antiviral activity of the compound is dependent on anabolism to the 5'-triphosphate. Both (-)- and (+)-FTC were anabolized to the corresponding 5'-triphosphates in chronically HBV-infected HepG2 cells. The rate of accumulation and the steady-state concentration of the 5'-triphosphate of (-)-FTC were greater. Also, (-)-FTC was not a substrate for cytidine deaminase and, therefore, is not subject to deamination and conversion to an inactive uridine analog. The (+) isomer is, however, a good substrate for cytidine deaminase.
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PMID:The anti-hepatitis B virus activities, cytotoxicities, and anabolic profiles of the (-) and (+) enantiomers of cis-5-fluoro-1-[2-(hydroxymethyl)-1,3-oxathiolan-5-yl]cytosine. 133 41

In the present study we used a HeLa whole cell extract transcription system to map the transcription start sites and the minimal promoter of the hepatitis B virus core gene. Two initiation sites located at residues 1792 +/- 5 and 1817 +/- 5 were identified. The minimal upstream region essential and sufficient for transcription was defined to a 105-base pair DNA fragment. These results are identical to the in vivo mapping of the transcription start sites and the minimal core gene promoter. When in vitro transcription elongation was carried out in the presence of the anionic detergent Sarkosyl, known to enhance premature transcription termination (attenuation), two short transcripts (as well as two run-offs) were synthesized. Kinetic studies indicated that the short transcripts resulted from a block to transcription elongation and not from RNA processing. RNA mapping showed that the short attenuated transcripts indeed initiated at the two core gene initiation sites and both prematurely terminated at nucleotide 1966 +/- 5, defined as the attenuation site. This site is located in the attenuator RNA within a uridine-rich sequence preceded by a stable hairpin structure. Attenuation at the same site occurred when transcription of the core gene was directed by the Ad2 major late promoter (MLP) and when the poly(A) signal, which precedes the attenuation site, was mutated from TATAAA to TAGAAA. We suggest that the elongation block at nt 1966 +/- 5 in vivo exerts a dual function: first, it regulates the level of RNA by attenuation during the first cycle of transcription and, second, it acts as a termination site at the end of the primary RNA transcript.
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PMID:In vitro regulation of human hepatitis B virus core gene transcription. 237 76

In contrast to hepatitis B virus infection, autoimmunity is common in chronic hepatitis D. In 1983, microsomal autoantibodies were described that differ from the previously described liver-kidney microsomal (LKM)-1 and LKM-2 antibodies. Therefore, these were named LKM-3. In addition, autoantibodies against basal layer cells of rat forestomach (basal cell layer antibodies (BCLA)) and thymic stellate epithelial cells (stellate epithelial cell antibodies (SECA)), as well as thymic reticular (thymic reticular antibodies) and perithymocytic cells (perithymocytic cell antibodies), were reported to be specifically associated with hepatitis D. Antinuclear antibodies against nuclear membrane lamin C were also found to be specifically associated with chronic hepatitis D. A molecular identification of the nonnuclear antigens has not been achieved apart from the observation that BCLA and SECA both recognize an antigen of 46 kDa and that these antibodies are immunologically cross-reactive. After the identification of cytochrome P450 2D6 as the major LKM-1 antigen and cytochrome P450 2C9 as the LKM-2 antigen, family 1 uridine diphosphate-glucuronosyltransferases have been identified as the molecules expressing the major LKM-3 autoepitope. Future studies will have to assess the possible pathogenetic and diagnostic relevance of these autoantibodies. However, chronic hepatitis D seems to be a good clinical model for the study of virus-induced autoimmunity in man.
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PMID:Molecular analysis of autoantigens in hepatitis D. 760 65

Patients with chronic hepatitis D often have liver-kidney microsomal antibodies type 3 (LKM-3). These antibodies react with several microsomal antigens that have a molecular weight of 55 KDa and an isoelectric point of about 8. We studied the molecular nature of the antigen and, by immunoscreening a human liver cDNA expression library with KM-3 sera, found that uridine diphosphate glucuronosyl transferases (UGT) appeared as candidate antigens. We confirmed the identity of UGT as an antigen by reacting the sera with recombinant rabbit liver UGT proteins. Some sera reacted with rabbit UGT-2 proteins, but UGT-1 proteins were more sensitive and specific in detecting LKM-3 autoantibodies in patient sera. Anti-UGT-1 antibodies were detected in all LKM-3 positive sera from patients with hepatitis D and 1 out of 11 patients with autoimmune hepatitis type 2. Sera from patients who had hepatitis B only did not react with UGT proteins. The UGT proteins are part of the phase II enzymes of drug metabolism and are the first such enzymes to be identified as human autoantigens.
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PMID:Recognition of uridine diphosphate glucuronosyl transferases by LKM-3 antibodies in chronic hepatitis D. 796 57

Recent reports have shown that HCV infection is not only restricted to hepatocytes. Like hepatitis B virus (HBV), which also was thought to be strictly hepatotropic in early molecular and cellular investigations, infection of lymphoid cells by HCV in vivo has been demonstrated. We showed that total peripheral blood leukocytes of chronically HCV-infected patients are infected by detection of plus- and minus-stranded HCV RNA using strand-specific oligonucleotide primers in the RT-PCR. These cells also represent extrahepatic sites for the viral replication, as demonstrated by incorporation of [3H]-uridine into nascent RNA after stimulation of the cells with a mitogen. Furthermore, total PBML from an uninfected person could be infected in vitro using an HCV-positive serum. It could be shown that replication of HCV RNA takes place in these cells. Examination of different subsets of PBML showed predominant infection of B-lymphocytes during HCV disease. Additionally, infection of T-lymphocytes was detected in about 50% of all chronically HCV-infected patients.
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PMID:B-lymphocytes are predominantly involved in viral propagation of hepatitis C virus (HCV). 803 62

The minimal substrate for human RNase P consists of the 5' leader sequence, aminoacyl acceptor stem, T-stem and T-loop of tRNA. The sequences corresponding to the D-stem, anticodon stem and loop and variable loop are replaced by a bulge which can be as small as 1 nt, but requires > 4 nt for optimal cleavage by RNase P. We found that a trans construct in which the T loop is opened between G57 and A58 (tRNA numbering system) is still processed by RNase P. The strand that is cleaved can be considered the target RNA while the other strand serves as an External Guide Sequence (EGS). We were also able to delete the nucleotides corresponding to nt 58 to 60 in the T-loop without affecting cleavage of the substrate. We propose that the sequence UUCG or UUCA (nucleotide 55 to 57 in the T-loop) positioned 3' to a double helical region of 12 to 13 basepairs containing a bulge of > 4 nt can form a structure that is recognized by human RNase P. The four nucleotides UUCR probably form a structure that resembles the uridine turn in the Tloop of tRNA. Since recognition by RNase P seems to be independent of the helical sequence, we suggest that this motif can be used for targeting RNA molecules for EGS-directed cleavage by RNase P. Based on these results, several 13-mer EGSs targeted to the 2.1 Kb surface antigen mRNA of hepatitis B virus (HBV) were designed and tested using a co-transcriptional cleavage assay with a 2.1 Kb HBV transcript. Some of these were capable of inducing cleavage of the HBV RNA by RNase P. The use of such small EGSs for the inactivation of various genes will be discussed.
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PMID:Design of short external guide sequences (EGSs) for cleavage of target molecules with RNase P. 947 94

Human RNase P recognizes a small model substrate consisting of only the 5' leader sequence, aminoacyl acceptor stem, and T stem and loop of a tRNA precursor. It was demonstrated here that a bimolecular construct in which the T loop is opened between G57 and A58 (tRNA numbering system) is still processed by RNase P. The strand that is cleaved can be considered the target RNA, whereas the other strand serves as an external guide sequence (EGS). The nucleotides corresponding to nt 58-60 in the T loop could be deleted without affecting cleavage of the substrate. Thus, the complete T loop can be replaced by the single-stranded sequence UUCG or UUCA (nt 55-57 in the T loop). The four nucleotides UUCR possibly form a structure that resembles the uridine turn in the T loop of tRNA. Because recognition by RNase P is independent of the helical sequence, this motif can be used for targeting RNA molecules for EGS-directed cleavage by human RNase P. Chemically modified EGSs with 2'-O-methyl groups also showed activity in inducing RNase P cleavage. Several 13-mer EGSs targeted to the 2.1-kb surface antigen mRNA of hepatitis B virus (HBV) were designed and tested using a co-transcriptional cleavage assay with a 2.1-kb HBV transcript. Some of the new EGSs were capable of inducing cleavage of the HBV RNA by RNase P.
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PMID:Short oligonucleotides as external guide sequences for site-specific cleavage of RNA molecules with human RNase P. 967 Oct 57

For the simultaneously visual detection of hepatitis B virus (HBV), hepatitis C virus (HCV) and human immunodeficiency virus type-1 (HIV-1), a qualitative DNA chip method, combining multiplex and nested polymerase chain reaction (PCR) with arrayed anchored primer PCR and a biotin-avidin alkaline phosphatase (Av-AP) indicator system, was developed. After pretreatment of infected blood samples and reverse transcription of the RNA virus genome, PCR was performed in a single tube by using the outer primer pairs. Second round nested multiplex PCR was performed on the DNA chip, on which the primers array had already been prepared. During the arrayed anchored multiplex PCR, 5[N-(N-biotinylaminocaproyl)-epsilon-3-aminoallyl]-2-deoxy-uridine-5-triphosphate (biotin-11-dUTP) was incorporated into the extended DNA chains in order to bind avidin alkaline phosphatase via avidin and biotin. To produce purple precipitates on the chips, the enzyme substrate 5-bromo-4-chloro-3-indolyl phosphate (BCIP) was used in conjunction with the enhancer, nitro blue tetrazolium (NBT). Blood samples containing the three viruses were tested using this DNA chip and about 1 pg of specific viral DNA fragments were detected on the chip wells after nested PCR.
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PMID:A visual DNA chip for simultaneous detection of hepatitis B virus, hepatitis C virus and human immunodeficiency virus type-1. 1470 86

Sequence-specific gene silencing by small interfering RNA (siRNA) is an intense area of focus in the development of novel therapeutic agents. Currently, there are two major hurdles to achieving clinically effective siRNA-based therapeutics: establishment of an efficient delivery system that transfers the siRNA to the correct tissue(s); and the reduction of unintended immunotoxicity associated with unmodified siRNA. We have developed a novel liver-specific delivery system of apolipoprotein A-I-decorated cationic lipids (DTC-Apo). Here, we show that intravenous injection of an unmodified hepatitis B virus (HBV)-specific siRNA encapsulated in DTC-Apo activates the innate immune response in mice. However, 2'-O-methyl (2'-OMe) modification of siRNA sense-strand uridine or uridine/adenosine residues efficiently abrogated the immunostimulatory properties of the siRNA and also silenced viral replication. In contrast, pyrimidine modification by 2'-OMe or 2'-fluoro (2'-F) substitution failed to circumvent liposome-induced immune recognition. Our findings provide useful information for the design of chemically-modified siRNAs for in vivo applications.
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PMID:Immunostimulatory properties and antiviral activity of modified HBV-specific siRNAs. 1796 21

Multiple segmental and selective isotope labeling of RNA with three segments has been demonstrated by introducing an RNA segment, selectively labeled with (13)C(9)/(15)N(2)/(2)H((1', 3', 4', 5', 5''))-labeled uridine residues, into the central position of the 20 kDa epsilon-RNA of Duck Hepatitis B Virus. The RNA molecules were produced via two efficient protocols: a two-step protocol, which uses T4 DNA ligase and T4 RNA ligase 1, and a one-pot protocol, which uses T4 RNA ligase 1 alone. With T4 RNA ligase 1 all not-to-be-ligated termini are usually protected to prevent formation of side products. We show that such labor-intensive protection of termini is not required, provided segmentation sites can be chosen such that the segments fold into the target structure or target-like structures and thus are not trapped into stable alternate structures. These sites can be reliably predicted via DINAMelt. The simplified NMR spectrum provided evidence for the presence of a U28 H(3)-imino resonance, previously obscured in the fully labeled sample, and thus of the non-canonical base pair U28:C37. The demonstrated multiple segmental labeling protocols are generally applicable to large RNA molecules and can be extended to more than three segments.
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PMID:Multiple segmental and selective isotope labeling of large RNA for NMR structural studies. 1858 61


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