Targeted Gene Therapy Approaches for Huntington's Disease: Advancing Treatment through Bioengineered Self-Assembling Peptide Hydrogels and BDNF-Encoded Adeno-Associated Viruses, Applied In Vitro and In Vivo
An aging global population's demographic and epidemiological evolution has considerably affected healthcare accessibility and cost. This phenomenon is associated with progressive age-related pathologies such as neurodegenerative disorders. Also, CNS incapacity to regenerate is an obstacle to efficiently treating neurodegenerative diseases. Huntington's disease (HD) is recognized as a fatal neurodegenerative disorder caused by an unstable expansion of a single polyglutamine repeat in the huntingtin protein. It mainly enhances the loss of striatal neurons and primary cortical neurons within the forebrain. Gene therapy has provided promising targeted therapeutic strategies to treat various neurodegenerative disorders such as HD, where the genomic DNA of a cell can be rewritten, reengineered, and manipulated to change its fate controllably. This therapeutic alternative controls genetic or epigenetic disturbances, bringing promising outcomes compared with pharmacotherapy or invasive protein delivery. BDNF is a crucial regulator of synaptic and structural plasticity, higher cognitive functions, and survival of neuronal cells within the CNS. As evidenced by research, the BDNF plays a crucial role in protecting endangered striatal neurons and primary cortical neurons from cell death in the HD brain. However, several hurdles, such as short half-life, limited diffusion and cross over the blood-brain barrier, and poor pharmacokinetics, have hindered the trophic factors' progress towards therapeutic intervention. Viral gene delivery tools have emerged as alternative protein delivery strategies to surmount these issues. Despite various successes, the stimulating potential of viral gene therapy has yet to be fully realized, with multiple clinical trials failing to deliver optimum therapeutic results. Reasons include neutralization via the host immune system and resultant poor or inconsistent transduction efficiencies, off-target toxicities, and virion dissemination. Biomaterials indicate an attractive solution candidate as they can mimic Brain's ECM, exhibit biologically relevant cues, and improve cell growth. One of the recent developments to overcome above mentioned issues is integrating engineered biomaterials with viral gene delivery holding the potential to increase the vector's residence time within the target region and provide sustained delivery, targeted expression of proteins at the site of clinical need while decreasing off-target toxicity and shielding viral vectors from humoral immune responses, achieving a safer and more efficient viral-based gene delivery. Self-assembling peptide (SAP) hydrogels are fabricated from detectable peptide sequences of tissue-specific ECM proteins to not only mimic native ECM biochemically but also provide nanofibrous structural support to enhance and control tissue renovation. These SAP hydrogels are formed based on physical interactions, allowing them to undergo a profound reversible gel/solution/gel shear thinning, which is significant for a delivery tool. Therefore, the injected low-viscosity hydrogel can freely diffuse throughout the entire irregularly shaped lesion area and initiate contact with the surrounding tissue when restored to a solid (stiffer) gel. When reformed in situ, these injected hydrogels can maintain their mechanical features and biomimetic nanoscale morphology. Therefore, to address the above-mentioned critical gaps in the development of novel treatment strategies for HD, this thesis explores the ability of Fmoc-SAP hydrogel systems to sustainably deliver a viral vector at the injured area without any side effects or immune response stimulation, as a novel treatment option for HD. Strong GFP expression proves the capability of Fmoc-SAP nanoscaffolds to deliver a functional virus and provide neural protection in vitro. Also, we found that rAAVDJ-BDNF blended with Fmoc-SAPs presented the highest neuroprotection and the lowest immune responses among all other groups in vivo.
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