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Targeting Mitochondrial Dysfunction and Oxidative Stress in Activated Microglia using Dendrimer-Based Therapeutics.

ABSTRACT: Mitochondrial oxidative stress is associated with many neurodegenerative diseases, such as traumatic brain injury (TBI). Targeted delivery of antioxidants to mitochondria has failed to translate into clinical success due to their nonspecific cellular localization, poor transport properties across multiple biological barriers, and associated side effects. These challenges, coupled with the complex function of the mitochondria, create the need for innovative delivery strategies. Methods: Neutral hydroxyl-terminated polyamidoamine (PAMAM) dendrimers have shown significant potential as nanocarriers in multiple brain injury models. N-acetyl cysteine (NAC) is a clinically used antioxidant and anti-inflammatory agent which has shown significant potency when delivered in a targeted manner. Here we present a mitochondrial targeting hydroxyl PAMAM dendrimer-drug construct (TPP-D-NAC) with triphenyl-phosphonium (TPP) for mitochondrial targeting and NAC for targeted delivery to mitochondria in injured glia. Co-localization and mitochondrial content of mitochondria-targeted and unmodified dendrimer were assessed in microglia and macrophages in vitro via immunohistochemistry and fluorescence quantification. Therapeutic improvements of TPP-D-NAC over dendrimer-NAC conjugate (D-NAC) and free NAC were evaluated in vitro in microglia under oxidative stress challenge. In vivo neuroinflammation targeting was confirmed in a rabbit model of TBI. Results: TPP-conjugated dendrimer co-localized significantly more with mitochondria than unmodified dendrimer without altering overall levels of cellular internalization. This targeting capability translated to significant improvements in the attenuation of oxidative stress by TPP-D-NAC compared to D-NAC and free NAC. Upon systemic administration in a rabbit TBI model, TPP-conjugated dendrimer co-localized specifically with mitochondria in activated microglia and macrophages in the white matter of the ipsilateral/injured hemisphere, confirming its BBB penetration and glial targeting capabilities. Conclusion: D-NAC has shown promising efficacy in many animal models of neurodegeneration, and this work provides evidence that modification for mitochondrial targeting can further enhance its therapeutic efficacy, particularly in diseases where oxidative stress-induced glial cell death plays a significant role in disease progression.

SUBMITTER: Sharma A

PROVIDER: S-EPMC6276292 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Project description:Brain-related disorders have outmatched cancer and cardiovascular diseases worldwide as the leading cause of morbidity and mortality. The lack of effective therapies and the relatively dry central nervous system (CNS) drug pipeline pose formidable challenge. Superior, targeted delivery of current clinically approved drugs may offer significant potential. Minocycline has shown promise for the treatment of neurological diseases owing to its ability to penetrate the blood-brain barrier (BBB) and potency. Despite its potential in the clinic and in preclinical models, the high doses needed to affect a positive therapeutic response have led to side effects. Targeted delivery of minocycline to the injured site and injured cells in the brain can be highly beneficial. Systemically administered hydroxyl poly(amidoamine) (PAMAM) generation-6 (G6) dendrimers have a longer blood circulation time and have been shown to cross the impaired BBB. We have successfully prepared and characterized the in vitro efficacy and in vivo targeting ability of hydroxyl-G6 PAMAM dendrimer-9-amino-minocycline conjugate (D-mino). Minocycline is a challenging drug to carry out chemical transformations due to its inherent instability. We used a combination of a highly efficient and mild copper catalyzed azide-alkyne click reaction (CuAAC) along with microwave energy to conjugate 9-amino-minocycline (mino) to the dendrimer surface via enzyme responsive linkages. D-mino was further evaluated for anti-inflammatory and antioxidant activity in lipopolysaccharides-activated murine microglial cells. D-mino conjugates enhanced the intracellular availability of the drug due to their rapid uptake, suppressed inflammatory cytokine tumor necrosis factor α (TNF-α) production, and reduced oxidative stress by suppressing nitric oxide production, all significantly better than the free drug. Fluorescently labeled dendrimer conjugate (Cy5-D-mino) was systematically administered (intravenous, 55 mg/kg) on postnatal day 1 to rabbit kits with a clinically relevant phenotype of cerebral palsy. The in vivo imaging study indicates that Cy5-D-mino crossed the impaired blood-brain barrier and co-localized with activated microglia at the periventricular white matter areas, including the corpus callosum and the angle of the lateral ventricle, with significant implications for positive therapeutic outcomes. The enhanced efficacy of D-mino, when combined with the inherent neuroinflammation-targeting capability of the PAMAM dendrimers, may provide new opportunities for targeted drug delivery to treat neurological disorders.

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Targeting Mitochondrial Dysfunction and Oxidative Stress in Activated Microglia using Dendrimer-Based Therapeutics.

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