Liposomal siRNA delivery against Herpes Simplex Virus-1

A large majority of the world`s population is infected with herpes simplex virus (HSV-1 and -2). These viruses lay dormant inside neuronal cells, where they successfully evade the immune system and consequently establishes lifelong infections. Hence, to date, there are no effective cures for HSV infection other than symptomatic relief. When the virus initiates reactivation from latency, it expresses viral RNA transcripts known as latency associated transcripts (LAT). LATs can potentially be silenced by siRNA, thus preventing the spread and severe side effects of the disease. Naked siRNA has a very short half-life and low cell permeability. We hypothesized that siRNA encapsulated in liposomes, would effectively be protected from degradation, thereby extending its half-life and providing efficient antiviral activity. In the present study we developed PEGylated liposomal formulations containing siRNA against LAT.

We successfully prepared two PEGylated liposomal formulations containing encapsulated siRNA, using a modified hydration technique or the ethanol injection method. Both formulations possess desirable physicochemical properties in terms of nano-size (~140 nm), low polydispersity index (PDI, 0.2), and a near-neutral charge (3mV), and high encapsulation yield (>50%). Adsorption of serum proteins to the two liposomal formulations was examined in terms of size changes following incubation in serum. Neither formulation showed any significant change of both size and PDI after 24 h incubation in serum. The encapsulated siRNA in both formulations was stable during preparation and storage. In-vitro results showed a significantly time- and concentration-dependent uptake in HaCaT and RAW264.7 cultures. Confocal imaging revealed lysosome escape after 48 h of incubation. Ongoing studies in HSV-1 ex-vivo models (plaque assay and 3D epidermis model) will evaluate efficacy and cytotoxicity.