Raloxifene-/raloxifene-poly(ethylene glycol) conjugate-loaded microspheres: A novel strategy for drug delivery to bone forming cells

Abstract

Raloxifene (Ral)- or Ral-poly(ethylene glycol) (PEG) conjugate-loaded microspheres were prepared with poly(ε-caprolactone) (PCL) alone or with the blend of PCL and poly(D,L-lactide-co-glycolide) (PLGA) to provide controlled and sustained Ral release systems. Benefits of these formulations were evaluated on bone regeneration. Ral-loaded PCL microspheres had the highest encapsulation efficiency (70.7±5.0%) among all groups owing to high hydrophobic natures of both Ral and PCL. Cumulative amount of Ral released from Ral-PEG (1:2) conjugate-loaded PCL:PLGA (1:1) microspheres (26.9±8.8%) after 60days was significantly higher relative to other microsphere groups. This finding can be ascribed to two factors: i) Ral-PEG conjugation, resulting in increased water-solubility of Ral and increased degradation rates of PCL and PLGA with enhanced water penetration into the polymer matrix, and ii) usage of PLGA besides PCL in the carrier composition to benefit from less hydrophobic and faster degradable nature of PLGA in comparison to PCL. In vitro cytotoxicity studies performed using adipose-derived mesenchymal stem cells (ASCs) demonstrated that all microspheres were non-toxic. Evaluation of intensities of Alizarin red S staining conducted after 7 and 14days of incubation of ASCs in the release media of the different microsphere groups was performed with Image J analysis software. At day 7, it was observed that the matrix deposited by the cells cultivated in the release medium of Ral-PEG (1:2) conjugate-loaded PCL:PLGA (1:1) microspheres had significantly higher mineral content (26.78±6.23%) than that of the matrix deposited by the cells cultivated in the release media of the other microsphere groups except Ral-loaded PCL:PLGA (1:1) microsphere group. At day 14, Ral release from Ral-PEG (1:2) conjugate-loaded PCL:PLGA (1:1) microsphere group resulted with significantly higher mineralization of the matrix (32.31±1.85%) deposited by ASCs in comparison to all other microsphere groups. Alizarin red S staining results eventuated in parallel with the release results. Thus, it can be suggested that Ral-PEG (1:2) conjugate-loaded PCL:PLGA (1:1) microsphere formulation has a potential as an effective controlled drug delivery system for bone regeneration.

Keywords: Controlled drug release; Microsphere; Poly(D,L-lactide-co-glycolide); Poly(ε-caprolactone); Raloxifene; Raloxifene-poly(ethylene glycol) conjugate.