SYNTHESIS OF AURONE-1,3,5-TRIAZINE CONJUGATES AND RNA-PROTEIN INTERACTION OF MUTANT HUNTINGTIN

Huntington's disease is a devastating inherited neurodegenerative disorder caused by an abnormal expansion in CAG repeats of the huntingtin gene, which leads to the production of a mutant HTT protein due to the toxic aggregate formation in neurons. It results in progressive motor dysfunction with cognitive decline and psychiatric disturbances. Currently, there is no cure for HD; thus, treatments existing are only symptomatic and have no action on the underlying molecular mechanisms. Of the many pathological processes occurring during HD, one concerns the interaction of mutant HTT RNA and RNA-binding proteins, particularly the MID1 protein complex, that encourages overproduction of the toxic HTT protein by enhancing its translation. This research involves an attempt to find a novel strategy to target pathological RNA-protein interactions by the design and synthesis of a specific aurone-135-triazine conjugate. Aurones are a class of flavonoids that are well-known to have anti-inflammatory, antioxidant, and neuroprotective properties, while triazines have been noted to bind nucleobases, making them promising candidates for targeting RNA structures. The aim of the present study was to connect the two compounds, aurones and triazines, into a hybrid molecule with selective binding to the expanded CAG-repeat RNA involved in HD and to interfere with its abnormal interaction with the MID1 protein complex. Inhibition of such an interaction would reduce overproduction of mutant HTT protein, offering a possible therapeutic strategy to slow the progression of HD. This dissertation details the multi-step synthetic pathway used to create the various intermediates towards the final aurone-1,3,5-triazine conjugate. All the intermediates have been characterized by means of NMR spectroscopy. To mitigate the lack of time that did not allow to complete the designed conjugate synthesis, a small library of thirty different aurones has been selected to undergo fluorescence spectroscopy studies in view of analysing their fluorescence properties and understanding their interaction with HuntRNA. Moreover, biological studies have also been carried out to explore the effectiveness of these new aurones regarding inhibition of the MID1-HTT RNA interaction in vitro. This involved testing the compounds on Huntington's RNA and HEK cells overexpressing the MID1 protein to determine their potential to interfere with the RNA-protein interaction central in HD pathology. These results indicate promising step towards the development of such inhibitors of mutant HTT RNA-MID1 interaction, offering potential therapeutic value for the treatment of HD. Thus, the present study paves the way for the investigation of both aurone-135-triazine conjugate and aurones as a novel series of compounds targeting the molecular mechanisms underlying Huntington's Disease, with the ultimate long-term goal of contributing to disease-modifying treatments.