Targeted delivery of PSC-RANTES for HIV-1 prevention using biodegradable nanoparticles

Abstract

Purpose: Nanoparticles formulated from the biodegradable co-polymer poly(lactic-co-glycolic acid) (PLGA), were investigated as a drug delivery system to enhance tissue uptake, permeation, and targeting for PSC-RANTES anti-HIV-1 activity.

Materials and methods: PSC-RANTES nanoparticles formulated via a double emulsion process and characterized in both in vitro and ex vivo systems to determine PSC-RANTES release rate, nanoparticle tissue permeation, and anti-HIV bioactivity.

Results: Spherical, monodisperse (PDI = 0.098 +/- 0.054) PSC-RANTES nanoparticles (d = 256.58 +/- 19.57 nm) with an encapsulation efficiency of 82.23 +/- 8.35% were manufactured. In vitro release studies demonstrated a controlled release profile of PSC-RANTES (71.48 +/- 5.25% release). PSC-RANTES nanoparticle maintained comparable anti-HIV activity with unformulated PSC-RANTES in a HeLa cell-based system with an IC(50) of approximately 1pM. In an ex vivo cervical tissue model, PSC-RANTES nanoparticles displayed a fivefold increase in tissue uptake, enhanced tissue permeation, and significant localization at the basal layers of the epithelium over unformulated PSC-RANTES.

Conclusions: These results indicate that PSC-RANTES can readily be encapsulated into a PLGA nanoparticle drug delivery system, retain its anti-HIV-1 activity, and deliver PSC-RANTES to the target tissue. This is crucial for the success of this drug candidate as a topical microbicide product.

Fig. 1

SEM image of PLGA nanoparticles loaded with PSC-RANTES-Biotin at a magnification of 10,000× and 10 kV (A). Averaged circular dichroism comparison of PSC-RANTES and PSC-RANTES biotin in water n=3 (B).

Fig. 2

Cumulative in vitro release profile of PSC-RANTES-biotin loaded PLGA nanoparticles for 30 days. pH=4.6. (N=3).

Fig. 3

In vitro release of PLGA nanoparticles over 10 days. pH=7.4 (triangles) and pH=4.6 (diamonds). (N=3).

Fig. 4

In vitro release of PLGA nanoparticles over 10 days with varying molar ratios of L (lactide): G (glycolide). 50:50 PLGA (closed diamonds), 65:35 PLGA (closed square), 75:25 PLGA (closed triangle), 85:15 PLGA (open circle). (N=3).

Fig. 5

Toxicity of PSC-RANTES treatment to TZM-bl cells with a 24 h exposure. In 96-well plates TZM-bl were exposed to decreasing amounts of unformulated PSC-RANTES (black) and PSC-RANTES nanoparticles (white) for 24 h at 37°C. Luminescence was measured after the addition of Cell Titer-Glo in percent viability based on positive controls. A non-toxic response was determined if the % viability of the unformulated PSC-RANTES or PSC-RANTES nanoparticles did not fall below 3 standard deviations of the control. N=3 independent experiments. *p<0.05 by Student’s t test.

Fig. 6

Luciferase luminescent readings of PSC-RANTES treated TZM-bl cells exposed to HIV-1. TZM-bl cells were treated with control empty nanoparticles (closed triangles), unformulated PSC-RANTES (filled diamond) or PSC-RANTES encapsulated nanoparticles (open squares) at a dosing range of 1 mM to 1 fM for 4 h at 37°C. Presented are data from 1 nM to 1 fM. After treatment, HIV-1_BaL was added with DEAE-dextran and allowed to culture for 48 additional hours. Results are reported on a log scale of PSC-RANTES dosing levels in percent of relative luminescent units (RLU) as compared to untreated TZM-bl cells infected with HIV-1. (*p<0.05 by Student’s t test comparing unformulated PSC-RANTES to PSC-RANTES nanoparticles).

Fig. 7

Representative H&E microscopy images of ecto-cervical tissue pre-nanoparticle exposure (A) and post nanoparticle exposure (B). Representative fluorescent microscopy images of ecto-cervical tissue with FITC (green) or DAPI (blue) staining at 40× magnification. Ecto-cervical tissue treated with unformulated PSC-RANTES for 4 hours with stained with DAPI (C) or FITC (D). Ecto-cervical tissue treated with PSC-RANTES encapsulated nanoparticles for 4 h stained with DAPI (E) or FITC (F).