Discovering and characterising molecules that rescue mitochondrial defects

Mitochondrial disease affects approximately 14,000 people in the UK. It can be caused by a plethora of genetic mutations found in nuclear or mitochondrial DNA. Disorders can present at any stage of life and affect many tissues, although often causing muscle or neurodegenerative defects. To date, treatment is palliative at best, and there are no effective therapies for these disorders. Newcastle is the major referral Centre for patients with mitochondrial disease in the UK and leads the NHS Highly Specialised Service for Mitochondrial Disorders. Researchers at the Wellcome Centre for Mitochondrial Research (WCMR) have derived over 200 patient cell lines in our Mitochondrial Bioresource with genetically-determined mitochondrial disease. Following many years of characterising these defects at the molecular and clinical level, it is now timely to focus on identifying novel therapeutics.
One area that has particular promise is mitochondrial proliferation. Genetic manipulation of mouse models of mitochondrial disease can result in such proliferation, with resultant partial rescue of mitochondrial function. Unfortunately, no compounds have been identified that can cause similar proliferation of human mitochondria.
Whilst our Wellcome Centre has a unique resource of cell lines and expertise in mitochondrial biology, we see that identifying new therapeutic agents requires collaboration with experts who have extensive experience with running primary screens, have access to extensive libraries of compounds and can perform large dataset informatics analyses.
We believe mining natural product libraries will reveal mitochondrial proliferators. This is particularly relevant considering mitochondrial evolutionary origins and the precedent of rapamycin, a molecule produced by the Streptomycetaceae, which can modulate cell signalling pathways and mitochondrial function in higher eukaryote cells. We have developed links with Bactevo, who have pioneered a platform to screen libraries of millions of independent bacterial expressor mutants at the single clone level for the production of unique natural compounds. This ultra-high throughput method utilises independent clones trapped in microspheres that act as nanolitre reaction vessels. Initially, following the production of over 1000 libraries, the platform was established to identify novel antimicrobial agents, for which there has already been success. However, together with our unique Mitochondrial Bioresource and extensive mitochondrial expertise we have begun to refocus this approach to identify, characterise and purify novel therapeutics for mitochondrial disease.
Together, we have modified and optimised the micro-droplet assay to identify bacterial producer clones that secrete compounds that can promote mitochondrial proliferation. An initial screen with 5 libraries, assessed over 10 million droplets and identified over 850 clones that produce a significant increase in mitochondrial mass in human cell lines. DNA is currently being sequenced from these clones to identify gene clusters involved in secondary metabolite production.