Chemical library screen for novel inhibitors of Kaposi's sarcoma-associated herpesvirus processive DNA synthesis.
Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of Kaposi's sarcoma and certain lymphoproliferative disorders. The role of KSHV lytic replication has been implicated in the tumor pathogenesis. A highly specific molecular complex formed by the KSHV DNA polymerase (POL8) and processivity factor (PF8) is indispensable for lytic viral DNA synthesis and may serve as an excellent molecular anti-KSHV target. The majority of conventional nucleoside-based anti-herpetic DNA synthesis inhibitors require intracellular phosphorylation/activation before they can exert inhibitory activity as competitive substrates for viral DNA polymerases. Novel and more potent inhibitors of KSHV DNA synthesis may be discovered through POL8/PF8-targeted high throughput screening (HTS) of small molecule chemical libraries. We developed a microplate-based KSHV POL8/PF8-mediated DNA synthesis inhibition assay suitable for HTS and screened the NCI Diversity Set that comprised 1992 synthetic compounds. Twenty-eight compounds exhibited greater than 50% inhibition. The inhibitory activity was confirmed for 25 of the 26 hit compounds available for further testing, with the 50% inhibitory concentrations ranging from 0.12+/-0.07 microM (mean+/-S.D.) to 10.83+/-4.19 microM. Eighteen of the confirmed active compounds efficiently blocked KSHV processive DNA synthesis in vitro. One of the hit compounds, NSC 373989, a pyrimidoquinoline analog, was shown to dose-dependently reduce the levels of KSHV virion production and KSHV DNA in lytically induced KSHV-infected BCBL-1 cells, suggesting that the compound blocked lytic KSHV DNA synthesis. HTS for KSHV POL8/PF8 inhibitors is feasible and may lead to discovery of novel non-nucleoside KSHV DNA synthesis inhibitors.
Dorjsuren
Dorjbal
D
Laboratory of Antiviral Drug Mechanisms, SAIC-Frederick, Frederick, MD, USA.
Burnette
Angela
A
Gray
Glenn N
GN
Chen
Xulin
X
Zhu
Weimin
W
Roberts
Paula E
PE
Currens
Michael J
MJ
Shoemaker
Robert H
RH
Ricciardi
Robert P
RP
Sei
Shizuko
S
D000818:Animals; D002460:Cell Line; D004279:DNA, Viral / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors; D019288:Herpesvirus 8, Human / Q000187:drug effects* / Q000378:metabolism; D011744:Pyrimidinones / Q000737:chemistry / Q000494:pharmacology; D011804:Quinolines / Q000737:chemistry / Q000494:pharmacology; D014764:Viral Proteins / Q000037:antagonists & inhibitors; D014771:Virion / Q000187:drug effects* / Q000378:metabolism
D004279:DNA, Viral; D004268:DNA-Binding Proteins; C110348:ORF59 protein, Human herpesvirus 8; D011744:Pyrimidinones; D011804:Quinolines; D014764:Viral Proteins; C037219:quinoline