Development of a siRNA delivery system for the treatment of osteoporosis

Sezlev Bilecen, Deniz
Osteoporosis, the most common disease of bone, is a skeletal disorder associated with low bone mass, increase in bone fragility and in susceptibility to fractures. The high bone resorption rate is shown to be due to increased number and activity of the osteoclasts. Receptor Activator of Nuclear Factor kappa B (RANK)/ Receptor Activator of Nuclear Factor kappa B Ligand (RANKL) system plays a crucial role in osteoclast differentiation and bone remodeling. RANKL participates in differentiation and activation of osteoclasts by binding to its receptor RANK expressed on osteoclast progenitors and mature osteoclasts. The currently used drugs for osteoporosis, such as use of bisphosphonates, Denosumab and teriparatide, have low bioavailability, long-term safety concerns and cause gastric problems. This led to the need for delivering the drugs in carrier systems to increase their bioavailability and decrease their side effects. In recent years, however, the increased understanding of molecular background of bone pathology, the use of RNA interference (RNAi) gained interest in developing new treatment strategies for the disease. Small interfering RNA (siRNA) is a double stranded RNA (dsRNA) molecule which is used in this context to inhibit the translation of abnormal gene expression in cells. In this study, we developed a potentially intravenously injectable siRNA-delivery system that can target osteoclasts in bone tissue for the treatment of osteoporosis. Polyethyleneimine (PEI), a polycationic molecule, was used as a complex with RANK siRNA which was then loaded into poly(lactic acid-co-glycolic acid) (PLGA) nanocapsules. The loaded nanocapsules were coated with a genetically engineered osteoconductive polypeptide, Elastin like recombinamer (ELR) designed to attach especially to bone minerals, to specifically target the drug loaded nanocapsules to the bone tissue. The carrier system constructed was studied by Scanning Electron Microscopy (SEM), (Transmission Electron Microscopy) TEM and X-Ray Photoelectron Spectroscopy (XPS) to show the ELR coat on the capsules. Different nitrogen to phosphate ratios (N/P) (presenting PEI and siRNA, respectively) for the PEI:RANK siRNA complexes were studied to determine the least toxic and most effective complex which would lead to satisfactory RANK mRNA inhibition. As a result of that study, the N/P ratio of 20 was chosen to construct the complex to be loaded in the PLGA nanocapsules. The encapsulation efficiency of the PEI:RANK siRNA complex (N/P 20) into PLGA nanocapsules were 48% and its release kinetics followed the Higuchi kinetics for 15 days. The PEI:RANK siRNA loaded PLGA nanocapsules significantly inhibited RANK mRNA (53 %) in an osteoclast precursor cell line. The differentiation of the precursors into mature osteoclasts was also suppressed by the delivery system. Treatment of differentiating osteoclasts with the PEI:RANK siRNA loaded PLGA nanocapsules inhibited their osteoclastic activity. In conclusion, the delivery system designed has the potential to serve as an alternative treatment method for use in the treatment of osteoporosis.


Construction of a PLGA based, targeted siRNA delivery system for treatment of osteoporosis
Bilecen, Deniz Sezlev; Carlos Rodriguez-Cabello, Jose; Uludag, Hasan; Hasırcı, Vasıf Nejat (2017-01-01)
Osteoporosis, a systemic skeletal disorder, occurs when bone turnover balance is disrupted. With the identification of the genes involved in the pathogenesis of the disease, studies on development of new treatments has intensified. Shortinterfering RNA (siRNA)is used to knockdown disease related gene expressions. Targeting siRNA in vivo is challenging. The maintenance of therapeutic plasma level is hampered by clearance of siRNA from the body. Targeted systems are useful in increasing the drug concentration...
Development of PEI-RANK siRNA Complex Loaded PLGA Nanocapsules for the Treatment of Osteoporosis
Bilecen, Deniz Sezlev; Uludag, Hasan; Hasırcı, Vasıf Nejat (2018-04-30)
Osteoporosis, which is characterized by low bone mineral density and susceptibility to fracture, is caused by increased osteoclastic activity. Receptor activator of nuclear factor kappa B ligand (RANKL)/RANK signaling plays an important role in osteoclast differentiation and activation. The current treatment strategies for osteoporosis do not directly address this underlying cause and generates undesired side effects. This led to emergence of controlled delivery systems to increase drug bioavailability and ...
Identification of Three Novel FBN1 Mutations and Their Phenotypic Relationship of Marfan Syndrome
KAYHAN, GÜLSÜM; ERGÜN, MEHMET ALİ; Ergun, Sezen Guntekin; KULA, SERDAR; PERÇİN, Ferda Emriye (Mary Ann Liebert Inc, 2018-07-26)
Background: Marfan syndrome (MS), a connective tissue disorder that affects ocular, skeletal, and cardiovascular systems, is caused by heterozygous pathogenic variants in FBN1. To date, over 1800 different pathogenic variants have been reported.
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Qadir, Zakria; Ali, Muhammad; Nesimoglu, Tayfun (2018-11-02)
Bone fracture is a common problem in daily life which occurs when high pressure is applied on bone or by simple accident and also due to osteoporosis and bone cancer. Different techniques are used today to detect bone fractures such as X-Ray, Computed Tomography (CT-scan), Magnetic Resonance Imaging (MRI) and Ultrasound. Among these four modalities, X-ray diagnosis is commonly used for fracture detection. However, if the fracture is complicated, a CT scan or MRI may be needed for further diagnosis and opera...
Improvement of bioactivity with dual bioceramic incorporation to nanofibrous PCL scaffolds
Altunordu, Gercem; Tezcaner, Ayşen; Evis, Zafer; Keskin, Dilek (2023-03-01)
Bone tissue injuries, diseases or related clinical interventions require bone tissue engineering (BTE) approaches for regeneration of large bone defects, especially for compromised situations. Most BTE applications in literature focused on composites of polymers with a single type of bioceramic. However, native bone matrix has various inorganic components. Accordingly, this study aimed to investigate the use of dual bioceramics in BTE scaffolds prepared by wet-electrospinning of Poly-caprolactone (PCL) and ...
Citation Formats
D. Sezlev Bilecen, “Development of a siRNA delivery system for the treatment of osteoporosis,” Ph.D. - Doctoral Program, Middle East Technical University, 2018.