Scientists at the University of Alabama at Birmingham (UAB) report that cancer drugs vorinostat, belinostat, and panobinostat might be repurposed to treat infections caused by human papillomaviruses (HPVs).
Highly efficacious vaccines against HPV infection exist, including the recently approved Gardasil 9, which immunizes against nine genotypes of HPV known to cause cervical, vulvar, vaginal and anal cancers, and genital warts. But the vaccine needs to be given before a person becomes sexually active, since it has no therapeutic efficacy against existing HPV infections.
“Safe, effective, and inexpensive therapeutic agents are urgently needed,” said N. Sanjib Banerjee, Ph.D., assistant professor of biochemistry and molecular genetics at UAB and lead author of the vorinostat study (“Vorinostat, a pan-HDAC inhibitor, abrogates productive HPV-18 DNA amplification”), which appears in PNAS.
The laboratory of Louise Chow, Ph.D., and Thomas Broker, Ph.D., at UAB discovered that the development of HPV infection depends on differentiation of the epithelium into a full-thickness, squamous epithelium. Furthermore, HPV reactivates host DNA replication in these differentiated cells, such that the replication proteins and substrates become available to support viral DNA amplification.
“HPVs cause epithelial proliferative diseases. Persistent infection of the mucosal epithelia by the high-risk genotypes can progress to high-grade dysplasia and cancers. Viral transcription and protein activities are intimately linked to regulation by histone acetyltransferases and histone deacetylases (HDACs) that remodel chromatin and regulate gene expression. HDACs are also essential to remodel and repair replicating chromatin to enable the progression of replication forks. As such, vorinostat (suberoylanilide hydroximic acid), and other pan-HDAC inhibitors, are used to treat lymphomas,” wrote the investigators
“Here, we investigated the effects of vorinostat on productive infection of the high-risk HPV-18 in organotypic cultures of primary human keratinocytes. HPV DNA amplifies in the postmitotic, differentiated cells of squamous epithelia, in which the viral oncoproteins E7 and E6 establish a permissive milieu by destabilizing major tumor suppressors, the pRB family proteins and p53, respectively. We showed that vorinostat significantly reduced these E6 and E7 activities, abrogated viral DNA amplification, and inhibited host DNA replication. The E7-induced DNA damage response, which is critical for both events, was also compromised. Consequently, vorinostat exposure led to DNA damage and triggered apoptosis in HPV-infected, differentiated cells, whereas uninfected tissues were spared. Apoptosis was attributed to highly elevated proapoptotic Bim isoforms that are known to be repressed by EZH2 in a repressor complex containing HDACs. Two other HDAC inhibitors, belinostat and panobinostat, also inhibited viral DNA amplification and cause apoptosis. We suggest that HDAC inhibitors are promising therapeutic agents to treat benign HPV infections, abrogate progeny virus production, and hence interrupt transmission.”
The Chow and Broker lab re-produced a fully differentiated human squamous epithelium by culturing primary human keratinocytes at an air-media interphase for two to three weeks, a growth they call raft culture. In 2009, their lab developed a model for a raft culture of HPV-18-infected primary human keratinocytes, allowing a robust amplification of HPV-18 DNA and production of infective viral progeny. This productive raft culture is an ideal model for preclinical investigation of potential anti-HPV agents.
Dr. Banerjee and colleagues hypothesized that inhibitors of HDACs would inhibit HPV DNA amplification because of their known mechanism of disrupting chromosomal DNA replication. Chromosomal replication requires HDAC alterations of histone proteins. Vorinostat inhibits many HDACs, so it might interrupt not only chromosomal replication but also viral DNA replication.
Using the HPV-18 model raft cultures, the researchers found that vorinostat effectively inhibited HPV-18 DNA amplification and virus production. Importantly, vorinostat also induced apoptosis in a fraction of the differentiated cells. Cell death could be attributable to DNA breakage when chromosomal DNA replication was interrupted. Similar results were obtained with two additional HDAC inhibitors, belinostat and panobinostat. In contrast, the differentiated cells of uninfected raft cultures, which do not replicate their DNA, were thus largely spared in the presence of the inhibitors.
The UAB team also examined how vorinostat affected levels and functions of viral oncoproteins, and they described the mechanisms that led to programmed cell death in HPV-18-infected cultures. “On the basis of these detailed studies,” Dr. Banerjee said, “we suggest that HDAC inhibitors are promising compounds for treating benign HPV infections, abrogating progeny production and hence interrupting infectious transmission.”
The UAB team also reported that vorinostat caused extensive cell death in raft cultures of dysplastic and cancer cell lines harboring HPV-16. HPV-16 and HPV-18 are the most prevalent, high-risk HPVs responsible for causing anogenital and oropharyngeal cancers. “But further investigation would be required to verify that these agents could also be useful in treating HPV associated dysplasias and cancers,” according to Dr. Banerjee.
