Senior Scientist / Group Leader Semmelweis University Budapest, Hungary
Abstract: Huntington’s disease (HD) is an autosomal dominant neurodegenerative disorder, caused by CAG expansions in the huntingtin gene (HTT), which results in the production of mutated huntingtin protein (mHtt). HD is incurable and typically presents in mid-life and progresses to death over a 20-year period. Autophagy, a lysosomal degradation pathway ensuring cytoplasmic homeostasis is dysfunctional in HD, and thus contributes mHTT protein accumulation. Preclinically, it has been shown that felodipine can upregulate autophagy and clear protein aggregates in cells, including neural cells in HD. Thus, a phase II clinical trial was undertaken (Fell-HD) to assess the tolerability and feasibility of testing this drug in patients with early stage HD while also looking for any signal of efficacy. Given we cannot look at autophagy in the living human brain, we sought to do this using induced neurons (iN) directly reprogrammed from skin fibroblasts from Fell-HD participants. iNs keep the genetic and aging signatures of the donor bypassing any stem cell or neuroprogenitor phase during conversion. We converted 7 control and 18 Fell-HD patient derived fibroblasts to iNs with the same conversion efficiency and purity. DNA methylation array and analysis in iNs showed accelerated aging in some patients. Moreover, most HD-iNs showed a less elaborate neuronal morphology and increased HTT expression using qPCR. We used 0.1 M and 1 M Felodipine treatment for 24h to assess its effects. After 28 days of conversion followed by Felodipine treatment iNs were counterstained using neuronal and autophagy markers to determine neuronal morphology and subcellular autophagy changes using high-content automated microscopy. Additionally, HTT measurements were again performed using qPCR after treatment. Our results showed that Felodipine enhanced autophagy in only a subset of patients while having no obvious adverse effects on HD-iNs. Lastly, we compared and correlated our preclinical results with FELL-HD trial outcomes - the patient’s cognitive and motor scores - and found some correlation to clinical response. In summary, this project using an in vitro preclinical iN model offers a new approach for looking at pathways targeted by drugs that cannot be studied in the living human brain and opens a new dimension in testing agents in clinic.
