A novel intravaginal ring to prevent HIV-1, HSV-2, HPV, and unintended pregnancy

Abstract

Women urgently need a self-initiated, multipurpose prevention technology (MPT) that simultaneously reduces their risk of acquiring HIV-1, HSV-2, and HPV (latter two associated with increased risk of HIV-1 acquisition) and prevents unintended pregnancy. Here, we describe a novel core-matrix intravaginal ring (IVR), the MZCL IVR, which effectively delivered the MZC combination microbicide and a contraceptive. The MZCL IVR contains four active pharmaceutical ingredients (APIs): MIV-150 (targets HIV-1), zinc acetate (ZA; targets HIV-1 and HSV-2), carrageenan (CG; targets HPV and HSV-2), and levonorgestrel (LNG; targets unintended pregnancy). The elastomeric IVR body (matrix) was produced by hot melt extrusion of the non-water swellable elastomer, ethylene vinyl acetate (EVA-28), containing the hydrophobic small molecules, MIV-150 and LNG. The solid hydrophilic core, embedded within the IVR by compression, contained the small molecule ZA and the macromolecule CG. Hydrated ZA/CG from the core was released by diffusion via a pore on the IVR while the MIV-150/LNG diffused from the matrix continuously for 94 days (d) in vitro and up to 28 d (study period) in macaques. The APIs released in vitro and in vivo were active against HIV-1ADA-M, HSV-2, and HPV16 PsV in cell-based assays. Serum LNG was at levels associated with local contraceptive effects. The results demonstrate proof-of-concept of a novel core-matrix IVR for sustained and simultaneous delivery of diverse molecules for the prevention of HIV, HSV-2 and HPV acquisition, as well as unintended pregnancy.

Keywords: Contraception; Core–matrix design; Efficacy; Intravaginal ring; Microbicides; Multipurpose prevention technology.

Figure 1. Core-matrix IVR

(a) MZCL IVR (20 mm X 4mm), macaque prototype, containing ZA/CG core (off-white ring) as seen through the translucent EVA-28 matrix containing MIV-150 and LNG and a pore to elute hydrated ZA/CG gel (scale: US dime = 17.91 mm diameter). (b and c) Cross sections depicting core and matrix compartments of the same IVR with a core side pore (b) and a drilled through pore (c) eluting ZA/CG gel.

Figure 2. In vitro release of APIs from novel core-matrix IVRs

(a) Cumulative release (mean ± SD) as a function of time of hydrophilic (core) APIs - CG and ZA (upper row) under sink conditions and hydrophobic (matrix) APIs - MIV-150 and LNG under non-sink conditions (bottom row) from the same rings. Release of APIs as seen from – 500 μm core side pore, n=5, (data pooled for MZC and MZCL IVRs), 500 μm through pore, n=5, (data pooled for MZC and MZCL IVRs) and 800 μm core side pore, n=3, (MZC IVRs). (b) Daily release as a function of time of the same rings as (a). Dotted lines represent target daily release. The data is pooled as the IVR type, MZC vs. MZCL, did not affect API release in vitro over time . (c) Unhydrated IVR core (d0) and progressive hydration of the core during 10d in vitro release. ZA/CG gel eluted from the pore by d1. The IVR dimensions are same as that in Figure 1.

Figure 2. In vitro release of APIs from novel core-matrix IVRs

(a) Cumulative release (mean ± SD) as a function of time of hydrophilic (core) APIs - CG and ZA (upper row) under sink conditions and hydrophobic (matrix) APIs - MIV-150 and LNG under non-sink conditions (bottom row) from the same rings. Release of APIs as seen from – 500 μm core side pore, n=5, (data pooled for MZC and MZCL IVRs), 500 μm through pore, n=5, (data pooled for MZC and MZCL IVRs) and 800 μm core side pore, n=3, (MZC IVRs). (b) Daily release as a function of time of the same rings as (a). Dotted lines represent target daily release. The data is pooled as the IVR type, MZC vs. MZCL, did not affect API release in vitro over time . (c) Unhydrated IVR core (d0) and progressive hydration of the core during 10d in vitro release. ZA/CG gel eluted from the pore by d1. The IVR dimensions are same as that in Figure 1.

Figure 2. In vitro release of APIs from novel core-matrix IVRs

(a) Cumulative release (mean ± SD) as a function of time of hydrophilic (core) APIs - CG and ZA (upper row) under sink conditions and hydrophobic (matrix) APIs - MIV-150 and LNG under non-sink conditions (bottom row) from the same rings. Release of APIs as seen from – 500 μm core side pore, n=5, (data pooled for MZC and MZCL IVRs), 500 μm through pore, n=5, (data pooled for MZC and MZCL IVRs) and 800 μm core side pore, n=3, (MZC IVRs). (b) Daily release as a function of time of the same rings as (a). Dotted lines represent target daily release. The data is pooled as the IVR type, MZC vs. MZCL, did not affect API release in vitro over time . (c) Unhydrated IVR core (d0) and progressive hydration of the core during 10d in vitro release. ZA/CG gel eluted from the pore by d1. The IVR dimensions are same as that in Figure 1.

Figure 3. PK profiles of APIs from MZC/ MZCL IVRs with varying pore configurations in non-DP treated macaques

(a) Schematic representation of PK study timeline for IVR insertion and removal, pH measurement and sample collection at indicated timepoints (b) CG and MIV-150 levels (upper row) in VF from MZC IVRs of 500 μm pore (core side, n=10), 500 μm pore (through, n=11), 800 μm pore (core side, n= 4) and MZCL IVRs of 500 μm pore (core side, n=3) and 500 μm pore (through, n=3). MIV-150 and LNG levels (bottom left) in blood from the same rings. Data represents mean ± SEM. ** p < 0.01 (MZC vs. MZCL) (c) Image of an MZCL IVR with 800 μm pore IVR removed post 21 days in vivo insertion (as collected from the lumen with no further post-treatment). The ZA/CG from the core released as a gel (arrow) demonstrating proof-of-concept. The IVR dimensions are same as that in Figure 1A.

Figure 4. Residual APIs in the IVRs post-PK study

(a) IVRs used in the PK study were analyzed for residual API content. The plot shows % APIs (core and matrix APIs) remaining in the IVRs. (b) Plot indicating correlation between residual core APIs, CG and ZA, using Spearman analysis. The correlation was significant indicated by p<0.0001. (c) The correlation was not significant for MIV-150 and LNG at the ‘n’s tested.

Figure 5. APIs release from MZC and MZCL IVRs were active in vitro and in vivo

(a) Eluent from the in vitro release study (days 1, 25, 46 and 94, pooled). The figure represents the mean ± SD of samples from 12 replicate MZCL IVRs with each dilution run in triplicate. (b) Vaginal fluid was collected in swabs from macaques that received MZC or MZCL IVRs (n=8) and placed in 1 mL of saline. Data is for 48h for HIV-1, 7 days for HPV and day17 HSV-2 (with exception of 1 sample which was 48h) swab sample activity. The vertical lines are the IC₅₀ values with 95% confidence intervals. All dilutions from each vaginal swab sample were run in triplicate, and the graph shows mean ± SD.