[Synthesis of a novel L-nucleoside, beta-L-D4A and its inhibition on the replication of hepatitis B virus in vitro].
OBJECTIVE
Nucleoside analogues have become the most promising candidates of anti-HBV drugs. In this study, beta-L-D4A was synthesized and explored its inhibitiory action against hepatitis B virus (HBV) in 2. 2. 15 cells derived from HepG2 cells transfected with HBV genome.


METHODS
beta-L-D4A was stereo-controlled synthesized from D-glutamic acid, and the structure was identified by IR, 1H NMR and MS. 2. 2. 15 Cells were placed at a density of 5 x 10(4) per well in 12-well tissue culture plates, and treated with various concentrations of beta-L-D4A for 6 days. At the end, medium was processed to obtain virions by a polyethlene glycol precipitation method. At the same time, intracellular DNA was also extracted and digested with Hind III. Both of the above DNA were subjected to Southern blot, hybridized with a 32P-labeled HBV probe and autoradiographed. The intensity of the autoradiographic bands was quantitated by densitometric scans of computer and EC50 was calculated. 2. 2. 15 cells were also seeded in 24-well tissue culture plates, and cytotoxicity with different concentrations was examined by MTT method. IC50 was calculated.


RESULTS
The synthesized compound structure conformed with beta-L-D4A; Autoradiographic bands showed similar for supernatant and intracellular HBV DNA. Episomal HBV DNA was inhibited in a dose-dependent manner. EC50 0.2 micromol x L(-1). The experiment of cytotoxicity gained IC50 200 micromol x L(-10.


CONCLUSIONS
beta-L-D4A has been synthesized successfully. beta-L-D4A possessed potent inhibitory effect on replication of HBV in vitro with low cytotoxicity, TI value was 1 000. It is expected to be developed clinically into a new anti-HBV drug.
Wu
Jin-Ming
JM
Department of Digestive Diseases of The First Affiliated Hospital of Wenzhou Medical College, Wenzhou 325000, China.
Lin
Ju-Sheng
JS
Xie
Na
N
Qiu
Guo-Fu
GF
Hu
Xian-Ming
XM
D000998:Antiviral Agents / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004261:DNA Replication / Q000187:drug effects; D004279:DNA, Viral / Q000187:drug effects; D016048:Dideoxyadenosine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D016679:Genome, Viral; D006515:Hepatitis B virus / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology; D014162:Transfection; D014779:Virus Replication / Q000187:drug effects*
D000998:Antiviral Agents; D004279:DNA, Viral; D016048:Dideoxyadenosine; C075923:2'3'-didehydro-2'3'-dideoxyadenosine