Jason Mendler, MD, PhD

2010-12-06

Two years ago, Dr. Agarwal was just starting to study mitochondrial diseases using induced pluripotent stem cells (iPSCs), but the patient samples he had were inadequate. At his hospital, a sign-out sheet on all the active patients is circulated every evening, and one patient’s clinical description suggested Pearson marrow pancreas syndrome (PS), a congenital disorder caused by mutations in mitochondrial DNA (mtDNA). “It’s a really rare disease, as you know, so this was a pretty serendipitous encounter,” he explained. He was able to obtain 300 microliters of three-week-old bone marrow from the patient. From this tiny volume of bone marrow, Dr. Agarwal was able to create iPSCs and his project was born.

The clinical hallmarks of PS include sideroblastic anemia and other cytopenias, pancreatic insufficiency, metabolic acidosis, and other systemic organ dysfunction. The cause of the hematopoietic failure in PS is unknown, and adequate cellular and animal models are lacking. At the genetic level, patients with PS and other mtDNA disorders exhibit heteroplasmy, a mix of mutant and wild-type mtDNA within each cell. To better understand the effects of mutant mtDNA on stem cells and hematopoiesis, Dr. Agarwal reprogrammed fibroblasts from the patient with PS into iPSCs. PS-iPSCs carrying the pathogenic mutation could be generated and displayed all hallmarks of pluripotency. Unexpectedly, it was demonstrated that the proportion of mutant mtDNA decreased rapidly in the PS-iPSC lines as a function of passage. Whereas the fibroblasts pre-induction had approximately 70 percent mutated mtDNA, cultured iPSCs eventually lost all detectable mutant mtDNA. This corresponded to restoration of mitochondrial function in iPSCs purged of mutant mtDNA. He also showed that disease-free hematopoietic progenitor cells could be created from these purged iPSCs. According to Dr. Agarwal, “depletion of mutant mtDNA in iPSCs in vitro mimics what appears to occur slowly in some cell types, such as hematopoietic stem cells, in patients with PS. This has been hard to study, but if we can figure out how and why it happens in iPSCs, it might give us ideas on how to accelerate the process in patients with PS and other mitochondrial diseases.”

Dr. Agarwal’s talk raised several important questions including the mechanism by which mutant mtDNA is purged from iPSCs and whether the cells generated are truly disease-free. Also raised was the question of how much “good” mitochondrial DNA is required for normal cellular function and whether heteroplasmy can be manipulated for therapeutic purposes. Although there is theoretically therapeutic potential of iPSCs, use of these cells is currently limited by their tumorigenicity and need for prolonged culture.

Thanks to Dr. Agarwal and his colleagues, we learned a great deal about PS and iPSCs, although perhaps we can learn more from the remarkable circumstances of the project’s inception. One interesting patient and a drop of bone marrow can sometimes harbor gold.

Dr. Mendler indicated no relevant conflicts of interest.