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Study on Intranasal Drug Delivery to treat Brain-associated diseases

Study on Intranasal Drug Delivery to treat Brain-associated diseases
Irfan Ullah
Issue Date
Brain is the most delicate organ of body, susceptible for many life threating disorders. Despite the major advances in neuroscience and drug development, the successful treatments for brain disorders remains limited. The major problem of restricted drug delivery to brain is presence of blood brain barriers (BBB) which limits the penetration of great number of potential therapeutics. In this study we utilize novel intranasal approaches to overcome limitation associated with local or systemic brain delivery methods. We demonstrated efficient brain specific delivery of siRNA, small therapeutics peptide and drug-loaded nanoparticles for glioblastoma and ischemic stroke models. In my first experimental setting we developed a novel intranasal approach for maximizing effective delivery of siRNA therapeutics and other drug formulation into mice brain. To investigate this, we design a device named “mouse positioning device” to maintain appropriate dosing position termed as “mecca position” during intranasal inoculation of therapeutic molecules. We incorporated applicable characteristics such as desirable height by adjusting either up or down positioning chairs to visualize mouse nostril, positioning chairs to inoculate four mice at one time and heating system to control suitable environment during experiment. A single inoculation of fluorescent labeled peptide dominantly deposits in brain, avoiding airway. Further our designed device showed outstanding delivery of functional siRNA to various part of brain. In addition, except from siRNA delivery, the intranasally delivered drug-loaded nanoparticles showed promising therapeutic efficacy in glioblastoma mouse model. We believe that, our proposed device is a step forward towards delivery of various therapeutics to central nervous system diseased models. In my second part of study, we extend our intranasal approach in rat ischemic stroke model. Ischemic stroke-induced neuronal cell death results in the permanent disabling of brain function. Apoptotic mechanisms are thought to play a prominent role in neuronal injury and ample evidence implicates Fas signaling in mediating cell death. In this study, we describe the neuroprotective effects of a Fas-blocking peptide (FBP) that by obstructing Fas signaling in cerebral ischemia inhibits apoptosis. Using an intranasal administration route in a rat model of focal cerebral ischemia, we demonstrate that nose-to-brain delivery of FBP after middle cerebral artery occlusion (MCAO) surgery results in the delivery and retention of FBP in Fas-expressing ischemic areas of the brain. A single intranasal administration of 2mg/Kg FBP resulted in significantly reduced neuronal cell death by inhibiting Fas-mediated apoptosis leading to decreased infarct volumes, reduced neurologic deficit scores and recovery from cerebral ischemia. Intranasally delivered FBP might be a promising strategy for the treatment of cerebral ischemic stroke. Next, we expend intranasal approach to treat malignant glioblastoma model. The glioblastoma multiform (GBM) is an aggressive tumor with a low survival rate and lack of effective treatment. The tumor recurrence after resection often requires chemotherapy or radiation to delay the infiltration of tumor remnants. Intracerebral chemotherapies are preferentially being used to prevent tumor regrowth although treatments remain unsuccessful because of poor drug distribution in the brain. In this study, we investigated the therapeutic efficacy of cancer-targeting tripeptide RGD-conjugated paclitaxel (PTX)-loaded nanoparticles against GBM by nose-to-brain delivery. Our results demonstrated that RGD-PTX-loaded nanoparticles showed cancer-specific delivery and enhanced anti-cancer effects both in vitro and in vivo. Further, the intranasal inoculation of RGD-modified PTX-loaded nanoparticles effectively controlled tumor burden by inducing apoptosis and/or inhibiting cancer cell proliferation without affecting the G0 stage of normal brain cells. Our data provide therapeutic evidence supporting the use of intranasally delivered cancer-targeted PTX-loaded nanoparticles for GBM therapy that could be further translated to other brain disorders. Finally, we modified our siRNA delivery platform (RVG9RC) by adding tri-leucine residues (RVG9R3LC) to improve functional effect of siRNA therapeutics. Although, the RVG9RC itself allows effective siRNA delivery to the cytoplasm by non-endocytic pathways, but a significant amount of siRNA complexes also enters the cell by ligand-induced receptor endocytosis and remain localized in endosomes. Here, we report that the incorporation of trileucine (3 Leu) residues as an endo-osmolytic moiety in the peptide improves endosomal escape and intracellular delivery of siRNA. The trileucine motif did not affect early non-endosomal mechanism of cytoplasmic siRNA delivery but enhanced target gene silencing by >20 % only beyond 24 h of transfection when siRNA delivery is mostly through the endocytic route and siRNA trapped in the endosomes at later stages were subject to release into cytoplasm. The mechanism may involve endnosomal membrane disruption as trileucine residues lysed RBCs selectively under endosomal pH conditions. Interestingly <3 Leu or >3 Leu residues were not as effective suggesting that 3 Leu residues are useful for enhancing cytoplasmic delivery of siRNA routed through endosomes. In this dissertation, we demonstrated sufficient pre-clinical evidences of efficient delivery of therapeutic drugs to mice and rat animal’s models. Our approach revealed effective delivery of siRNA, therapeutic peptide and drug-loaded nanoparticles for brain cancer and ischemic stroke disease models. In future, our developed approaches could be translated to other neurodegenerative disorders like Huntington’s, Alzheimer’s, Parkinson’s diseases.
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GRADUATE SCHOOL[S](대학원) > BIOENGINEERING(생명공학과) > Theses (Ph.D.)
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