Oral Presentation ASSCR, AGCTS, ISCT ANZ and Friends Joint Scientific Conference 2019

USING HUMAN EMBRYONIC STEM CELL MODELS OF MITOCHONDRIAL DISEASE TO IDENTIFY CANDIDATE DRUG TREATMENTS (#2)

Cameron L McKnight 1 , David A Stroud 2 , Andrew G Elefanty 1 , Richard J Mills 3 , James E Hudson 3 4 , Michael T Ryan 5 , David A Elliott 1 , David R Thorburn 1 6 , Ann E Frazier 1
  1. Murdoch Childrens Research Institute, Parkville, VIC, Australia
  2. Department of Biochemistry and Molecular Biology, University of Melbourne/Bio21, Melbourne, VIC, Australia
  3. QIMR Berghofer, Brisbane, QLD, Australia
  4. School of Biomedical Sciences, University of Queensland, Brisbane, QLD, Australia
  5. Department of Biochemistry and Molecular Biology, University of Monash, Melbourne, VIC, Australia
  6. Victorian Clinical Genetics Service, Royal Children's Hospital, Melbourne, VIC, Australia

This project aims to differentiate human embryonic stem cell (hESC) models of mitochondrial disease to a cardiac cell fate to facilitate preclinical treatment screens and investigation of the underlying cellular mechanisms of disease. With >320 genetic causes of oxidative phosphorylation (OXPHOS) disorders, proving therapeutic efficacy remains challenging despite a number of agents showing promise1,2.

We generated a panel of 15 knockout models of nuclear-encoded OXPHOS genes in hESCs, allowing us to study tissue-specific effects via differentiation in an isogenic background. In general, we focussed on generating cell lines for disease genes where patients have presented with biallelic Loss-of-Function mutations that cover a range of primary and secondary roles in OXPHOS biogenesis.

Our studies focussed on SURF1-/- and COA6Del80/p.W59C hESC clones that display OXPHOS complex IV (CIV) deficiency3,4. To overcome any blocks in maturation potentially arising as a result of the mutation, we also generated inducible correction lines for the SURF1 and COA6 clones. SURF1-/- clones form beating cardiomyocytes that show abnormal calcium handling and a significant decrease in contraction force in an organoid system5,6. SURF1 is a CIV assembly factor and quantitative proteomic analysis of contractile cardiomyocytes shows downregulation of CIV subunit proteins while most other mitochondrial proteins – including other CIV assembly factors – trend toward upregulation.

Although COA6 is also a CIV assembly factor, the COA6Del80/p.W59C clones fail to differentiate to cardiomyocytes under standard conditions. However, adding drugs targeting specific mitochondrial pathways in the differentiation medium has shown some capacity to support the cells getting further through the differentiation.

Data from these cell lines are promising for our other hESC models of mitochondrial disorders. The SURF1 proteomic data demonstrate compensatory mitochondrial proliferation in response to decreased CIV assembly. Currently, these hESC models are being used for preliminary investigations of disease-specific treatment options.

  1. Frazier AE, Thorburn DR, Compton AG. Mitochondrial energy generation disorders: genes, mechanisms, and clues to pathology. J Biol Chem. 2019;294(14):5386-95.
  2. Gorman GS, Chinnery PF, DiMauro S, Hirano M, Koga Y, McFarland R, et al. Mitochondrial diseases. Nature Reviews Disease Primers. 2016;2:16080.
  3. Wedatilake Y, Brown RM, McFarland R, Yaplito-Lee J, Morris AA, Champion M, et al. SURF1 deficiency: a multi-centre natural history study. Orphanet J Rare Dis. 2013;8:96.
  4. Stroud DA, Maher MJ, Lindau C, Vögtle FN, Frazier AE, Surgenor E, et al. COA6 is a mitochondrial complex IV assembly factor critical for biogenesis of mtDNA-encoded COX2. Human Molecular Genetics. 2015;24(19):5404-15.
  5. Phelan DG, Anderson DJ, Howden SE, Wong RCB, Hickey PF, Pope K, et al. ALPK3-deficient cardiomyocytes generated from patient-derived induced pluripotent stem cells and mutant human embryonic stem cells display abnormal calcium handling and establish that ALPK3 deficiency underlies familial cardiomyopathy. European Heart Journal. 2016;37(33):2586-90.
  6. Voges HK, Mills RJ, Elliott DA, Parton RG, Porrello ER, Hudson JE. Development of a human cardiac organoid injury model reveals innate regenerative potential. Development (Cambridge, England). 2017;144(6):1118-27.
  • Have you presented your abstract at another international meeting?: No