| Backgroud/Objectives:Activation of telomerase,an enzyme that synthesizes telomeric DNA,is an essential step in cell immortalization. Telomerase is ordinarily inactive in most somatic cells,but can be detected in nearly all tumors. The activation of telomerase in malignant cancers seems to be an important step in tumorigenesis,whereby the cell gains the ability of indefinite proliferation. Due to the association between telomerase expression and malignancy,inhibition of the enzyme even particular importance inhibition the telomerase reverse transcriptase (TERT) of it is expected to be a useful a new anticancer therapeutic target. Antisense strategies,a fast-growing area,capture the imagination with their promise of rational drug design and exquisite specificity. But the design of antisense drugs is still with low efficiency and by random. It has become a difficult problem to optimize the antisense problem. We constructed an integrared bioinformatic analysis system based on WEB bio-resources. Targeted the homo sapiens TERT mRNA,we designed the antisense drug to concidering the coding region,phylogenesis of mRNA,the predicted secondary structure and the secondary structural elements of mRNA,AG between mRNA,its antisense ODNs and the phylogenetic conserved regions. It is believed that Bioinformatic targeted drug should be helpful to advence the success rate of design and to discover the role of antisense drug design.MethodsWe try to set the human telomerase reverse transcriptase mRNA as target and optimize the antisense drug design comprehensively basing on Constructing a WEB based Bioinformatic Analysis System to analyze the coding region,phylogenesis of mRNA,thepredicted secondary structure and the secondary structural elements of mRNA,AG between mRNA,its antisense ODNs and the phylogenetic conserved regions. Several excellent sequences with good activities were obtained basing on the pharmacology analysis to verify the evalution of activity of the antisense drug designed. Sequence analysis of the candidate fragment ASingl originated form a differently expressed gene showed which would be a novel one in order to character the mechanism of telomerase in cell regulation net during the period form pasttranscription to posttranslation.1 . To optimize the design of antisense drug targeting TERTmRNA,obtain better antisense drugs than TTAGGG,The basing on Constructing a WEB based Bioinformatic Analysis System was utilized to predict the optimal and suboptimal secondary structures of human TERTmRNA. Twenty-nine antisense ASODN targeting the secondary structural elements,3 partly matched ASODN and 1 -scrambled 3521 were designed. An antisense inhibitor directly against the RNA template of telomerase,TTAGGG,was already pushed into clinical trials as anti-neoplasm agent. TTAGGG was set as positive control. Mean (n = 3 - 5) 50 % inhibitory effects on proliferation of Lovo cells (IC50) of ASODNwere evaluated,AG relating to the target secondary structural elements were calculated according to the nearest neighbor model. The QSAR analysis through multiple regressions was obtained by SPSS. The results showed that three S-ODNs,ATI 755(20),AT1224(20),AP0973(20),had statistically significant lower IC50(48+7,50+4,64+2. 7 nmol. L 'respectively) than that of TTAGGG (81+25nmol. L ' P O.05). The number of bases comprising the target secondary structural elements bulge loop,internal loop,and knot,the free energy of ASODN(AG37S),and reaction (AG37R) were important parameters in QSAR equation. In the multiple regression,R was 0.68,P = 0.0193. Not tally with the equation,two ASODNs with favorable target structures and AG3?,AT 1624(20) and AT0022(20),did not behave good activities. The results indicated that computer aided antisense drug design basing on secondary structure prediction was helpful to obtain ASODNwith better in vitro effect than current positive drug. The degree of instability of secondary structural elements and A G were important factors for ASODNs activity. The WEB based Bioinformatic Analysis System is...
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