Characterization of the antiviral activity against Herpes Simplex Virus 1 (HSV-1) and 2 (HSV-2) of Oximacro®, a new cranberry extract

Herpes Simplex Virus type-1 and type-2 (HSV-1 and -2) are worldwide distributed pathogens. Both HSV-1 and -2 spread by direct contact with infected skin or mucosa, and following primary infection they establish latency in the trigeminal (HSV-1) or dorsal root ganglia (HSV-2) from which reactivation may occur throughout life of infected hosts. The spectrum of clinical manifestation of HSV diseases is wide, ranging from asymptomatic up to life-threatening infections. Although in most of the cases infections are subclinical, in several pathological contexts, such as in immunocompromised individuals, HSV infections may be associated to a high degree of morbidity and mortality. Furthermore, HSV-2 is responsible for one of the most prevalent sexually transmitted infections and it is associated to an increasing risk of HIV acquisition. In the absence of an available vaccine, acyclovir, a guanosine analogue, is the most important drug used in treatment of HSV infections, despite that its use is often associated to long term toxicity and emergence of resistant strains. All these facts highlight the urgent need to develop new antiviral molecules against HSV. In this regard, naturally occurring compounds of plant and microbial origins may represent an exploitable source of antiviral substances. Among plant-derived natural products, extracts of the fruit of American cranberry (Vaccinium macrocarpon Ait., Ericaceae) are known to be rich in compounds exerting numerous antimicrobial effects, such as prevention of urinary tract infections (UTIs), as well as anti-inflammatory activities. These effect appears to be related to the cranberry specific phytochemical profile and to its content of bioactive compounds, such as A-type proanthocyanidins (PACs). In fact, PAC-A have been reported to mask P-fimbriae of uropathogenic E. coli during adhesion to uroepithelial cells. Given these facts, the aim of this thesis work was to evaluate the antiviral properties of a new cranberry extract, Oximacro®, enriched in PAC-A content, against HSV. The results of the experimental work indicated the ability of the Oximacro® to inhibit HSV replication in vitro. Immunoblot analysis then showed that the cranberry extract blocked the expression of all time-representative IE, E and L gene products of HSV, thus suggesting that the phase of HSV replication cycle affected by Oximacro® occurs prior to the expression of IE genes. Further analysis of the pre-IE phases then revealed that the antiviral activity stemmed from inhibition of HSV attachment to host cells. Since Oximacro® was found to significantly reduce the infectivity of HSV virion particles before their interaction with host cells, we investigated its impact on those viral envelope glycoproteins, gD and gB, that are required for HSV attachment and entry, respectively. By using purified HSV particles and recombinant gD, we observed that the cranberry extract was able to interact with and alter the electrophoretic mobility of both gD and gB, thus causing their structural impairment and inhibition of virions infectivity. Finally, the anti-HSV activity of Oximacro® was retained even in the presence of human serum and low pH values, both characteristics of the vaginal environment, thus suggesting its suitability as component of vaginal microbicides. In conclusion, these results indicate Oximacro® as a promising natural candidate for the development of novel topical microbicides for the prevention of HSV infections.