SiRNA structural optimization

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Human papillomaviruses (HPVs) are a major cause of anogenital and head and neck neoplasms and malignancies. Unfortunately, no specific antiviral therapies for HPV infection or HPV-induced malignancies have been identified. This U19 proposal is submitted as project by Alexander in a consortium proposal entitled siRNA Microbicides. The goal of this research consortium is to develop native and chemically modified siRNAs as effective means for blocking transmission of HIV, HSV2 and HPV. In project by Alexander, we hypothesize that chemically modified single-stranded boranophosphate-backbone short interfering RNAs (siRNAs) are superior to native siRNAs for potent and durable gene silencing, and that anti-HPV E6/E7 single-stranded boranophosphate-backbone siRNAs (ssBP-siRNAs) can be used to effectively block HPV viral replication and proliferation of HPV-transformed cells. By combining chemical modifications shown individually to increase siRNA silencing potency and duration, we will create ssBP-siRNAs, designed for topical delivery, that block episome replication in HPV-infected cells and induce irreversible growth arrest in HPV16-transformed cells. Anti-HPV16 ssBP-siRNAs identified by us will be tested in project 3 using novel cell culture and transgenic mouse models of HPV infection. Project 3 will also supply optimized ssBP-siRNAs to projects by Ramratnam and Herold to allow comparison of ssBP-siRNAs with conventional siRNAs for targeting expression of macaque CCR5, murine focal adhesion kinase, and murine nectin-1. To allow immediate work optimizing ssBP-siRNA delivery to vaginal mucosa, we will supply existing highly active ssBP-siRNAs to Core B (Formulations). The vaginal mucosal uptake and activity of formulated ssBP-siRNAs will be assessed quantitatively in project 3. At the conclusion of this study, we will have developed anti-HPV16 E6/E7 ssBP-siRNAs as effective topical microbicides for prevention of HPV infection and HPV-induced malignancies. We will also have created high potency ssBP-siRNAs active against other consortium targets, and forwarded these ssBP-siRNAs for microbicidal testing in laboratory models of viral infection.