PMID: 9636294Nov 1, 1995Paper

A high capacity assay for inhibitors of human papillomavirus DNA replication

Bio/technology
M PlumptonM A Romanos

Abstract

The discovery of antiviral compounds against human papillomaviruses (HPV) has been hindered by the difficulties in culturing virus in vitro or assaying stable HPV DNA replication. However, plasmids containing the HPV replication origin replicate transiently upon co-transfection with HPV E1 and E2 expression vectors. We have adapted this assay using secreted alkaline phosphatase (SAP) as a reporter for rapid analysis of DNA copy number. Use of the SV40 early promoter in controlling SAP expression was critical in ensuring both a strong signal and copy number dependence: the stronger beta-actin promotor inhibited replication, while the weaker SV40 late promoter yielded very low levels of SAP. The precise configuration of the E1 and E2 expression vectors also was critical, most pre-existing vectors did not support efficient replication and SAP secretion. The extent of DNA replication and SAP secretion were both proportional to the amount of E1/E2 vector used in transfections; under optimal conditions SAP increased 100-fold during replication. The assay has been developed for compound screening in 96-well plates and several inhibitors have been identified. Quantitative Southern blot analysis has shown that most of these inhibit HPV ...Continue Reading

References

Jul 1, 1992·Proceedings of the National Academy of Sciences of the United States of America·C M ChiangL T Chow
Oct 1, 1992·Journal of Virology·A M Del VecchioC C Baker
Dec 1, 1990·Journal of Virology·J SterlingT Minson
Jul 1, 1987·Proceedings of the National Academy of Sciences of the United States of America·P GunningL Kedes
Aug 30, 1994·Proceedings of the National Academy of Sciences of the United States of America·P ParkI J Mohr
Apr 1, 1994·Molecular and Cellular Biology·S B HaaseM P Calos
Dec 25, 1993·Nucleic Acids Research·F J Hughes, M A Romanos
Jan 15, 1993·Proceedings of the National Academy of Sciences of the United States of America·Y S SeoJ Hurwitz
Apr 1, 1993·Proceedings of the National Academy of Sciences of the United States of America·Y S SeoJ Hurwitz
Jun 1, 1993·Proceedings of the National Academy of Sciences of the United States of America·L YangM Botchan
Jul 1, 1993·International Journal of Gynecological Cancer : Official Journal of the International Gynecological Cancer Society·J.C. LongfieldG.A. Turner

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