Use of an in vivo animal model for assessing the role of integrin α(6)ß(4) and syndecan-1 in early steps in papillomavirus infection.

Human papillomaviruses (HPV) are small DNA tumor viruses. HPV infection requires entry of virions into epithelial host cells that support the viral life cycle. Here, we used an in vivo mouse model, in which HPV pseudoviruses (PVs) are scored for their ability to transduce reporter genes, to test the role of various cellular proteins in entry. We initially investigated the role of integrin α(6)ß(4) in mediating early steps of HPV infection. Deficiency of integrin α(6)ß(4) is modestly but significantly suppressed reporter-gene transduction by PVs in conditional integrin ß(4) knockout mice. We also investigated the role of syndecan 1, a heparin sulfate proteoglycan (HSPG) for its role in HPV infection. We did not see a significant reduction in reporter-gene transduction by PVs in syndecan-1 null mice. This indicates that this HSPG is not essential for early steps in HPV infection, but does not discount a need of other HSPGs in mediating HPV infection.

Novel antivirals inhibit early steps in HPV infection.

The future incidence of cervical cancer is forecast to decline because of the remarkably effective prophylactic vaccines against human papillomaviruses. However, lack of access to these expensive vaccines in the developing countries where cervical cancer is most frequent, and the restricted genotypes these vaccines protect against, will limit their impact. Clearly, there is still a need for identifying other modalities for preventing HPV infections. Ready access to effective, inexpensive antivirals represents one potentially valuable approach to the prevention of genital HPV infections. We developed a well-validated high throughput screening (HTS) assay for identifying compounds that inhibit HPV infection and applied this assay to identify lead compounds that act by inhibiting an early step in infection. We screened over 40,000 small molecules that were available at the University of Wisconsin Small Molecule Screening Facility (UW-SMSF). The top 22 compounds were chosen for further analyses based upon the pharmacological property, scaffold diversity, strength of the inhibitory activity and lack of nonspecific cytotoxicity. Of these compounds, #13 and #14 had the most acceptable properties of low to submicromolar IC(50)'s and low cytotoxicity. Optimal antiviral activities were elicited by exposure of cells to the #13 and #14 during the initial 12 h following infection. Twenty-nine #13-like and 15 #14-like analogs were identified in silico and tested for their antiviral activities corresponded to the altered structures comparing to #13 and #14, informing on the pharmacophore structure of each compound. Studies indicate that both compounds inhibit infection post-entry.

Inhibition of gamma secretase blocks HPV infection.

Human papillomaviruses (HPV) are common sexually transmitted pathogens that predispose women to cervical and other anogenital cancers. HPV vaccines can prevent infection by some but not other sexually transmitted HPVs but are too costly for use in much of the world at greatest risk to HPV-associated cancers. Microbicides provide an inexpensive alternative to vaccines. In a high throughput screen, drugs that inhibit the cellular protein complex known as gamma secretase were identified as potential HPV microbicides. gamma Secretase inhibitors (GSIs) inhibited the infectivity of HPV pseudoviruses both in human keratinocytes and in mouse cells, with IC(50) values in the picomolar to the nanomolar range. Using a mouse model, we observed that a GSI could inhibit HPV infection to the same degree as its effectiveness in inhibiting gamma secretase activity in vivo. We conclude that gamma secretase activity is required for HPV infection and that GSIs are effective microbicides against anogenital HPVs.