Stem Cell Reports Ar ticle Induced Pluripotent Stem Cell Modeling of Multisystemic, Hereditary Transthyretin Amyloidosis Amy Leung, 1,5 Shirley K. Nah, 1,5 Whitney Reid, 1,5 Atsushi Ebata, 2 Clarissa M. Koch, 3 Stefano Monti, 1 Joseph C. Genereux, 4 R. Luke Wiseman, 4 Benjamin Wolozin, 2 Lawreen H. Connors, 3 John L. Berk, 3 David C. Seldin, 1,3 Gustavo Mostoslavsky, 5 Darrell N. Kotton, 5 and George J. Murphy 1,5, * 1 Sections of Hematology-Oncology and Computational Biomedicine, Departments of Medicine, Pathology and Laboratory Medicine, Boston University School of Medicine, Boston, MA 02118, USA 2 Departments of Pharmacology and Neurology, Boston University School of Medicine, Boston, MA 02118, USA 3 The Amyloidosis Center, Boston University School of Medicine, Boston, MA 02118, USA 4 Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA 5 Center for Regenerative Medicine, Boston University School of Medicine and Boston Medical Center, Boston, MA 02118, USA *Correspondence: gjmurphy@bu.edu http://dx.doi.org/10.1016/j.stemcr.2013.10.003 This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-No Derivative Works License, which permits non-commercial use, distribution, and reproduction in any medium, provided the original author and source are credited. SUMMARY Familial transthyretin amyloidosis (ATTR) is an autosomal-dominant protein-folding disorder caused by over 100 distinct mutations in the transthyretin (TTR) gene. In ATTR, protein secreted from the liver aggregates and forms fibrils in target organs, chiefly the heart and peripheral nervous system, highlighting the need for a model capable of recapitulating the multisystem complexity of this clinically var- iable disease. Here, we describe the directed differentiation of ATTR patient-specific iPSCs into hepatocytes that produce mutant TTR, and the cardiomyocytes and neurons normally targeted in the disease. We demonstrate that iPSC-derived neuronal and cardiac cells display oxidative stress and an increased level of cell death when exposed to mutant TTR produced by the patient-matched iPSC-derived hepa- tocytes, recapitulating essential aspects of the disease in vitro. Furthermore, small molecule stabilizers of TTR show efficacy in this model, validating this iPSC-based, patient-specific in vitro system as a platform for testing therapeutic strategies. INTRODUCTION Amyloidosis refers to a group of diseases caused by the extracellular deposition of misfolded fibrillar proteins, leading to multiorgan failure and death (Falk et al., 1997). Familial amyloidosis (AF) occurs when inherited point mutations in the genes coding for abundant serum pro- teins such as transthyretin (TTR), fibrinogen, lysozyme, or apolipoproteins lead to clinical disease. The most com- mon form of AF arises from aggregation of mutated TTR, a 55 kDa transport protein predominantly synthesized by the liver (Connors et al., 2003; Falk et al., 1997; Plante ´- Bordeneuve and Said, 2011). Familial transthyretin amyloidosis (ATTR) is a lethal, autosomal-dominant dis- ease caused by single amino acid substitutions arising from 1 of more than 100 described mutations in the TTR gene (Connors et al., 2003; online amyloidosis mutation database in Rowczenio and Wechalekar [2010]). These sin- gle amino acid substitutions destabilize the native homote- trameric structure of circulating TTR, promoting release of amyloidogenic TTR monomers, fibril formation, and depo- sition as amyloid in target end organs (Ando et al., 2005; Falk et al., 1997). Most amyloidogenic TTR variants target the nervous system and heart, inducing neuropathy and cardiomyopathy (Jacobson et al., 1992, 1997; Plante ´- Bordeneuve and Said, 2011). The organ distribution and age of onset can vary across families and endemic areas, even with identical TTR mutations. Although transgenic mouse models and immortalized human cell lines have provided some insights into disease pathogenesis (Araki et al., 1994; Kohno et al., 1997; Reixach et al., 2004; Sousa et al., 2002; Tagoe et al., 2007), these systems are indepen- dent of the genetic context of the patient. ATTR represents an important unmet medical need with significant morbidity and early mortality in affected fam- ilies. Survival after ATTR disease onset is usually only 5–15 years (Benson et al., 2011; Plante ´-Bordeneuve and Said, 2011). Orthotopic liver transplantation is currently the only US-approved treatment for ATTR. However, only about one-third of patients are candidates for surgery (Monteiro et al., 2004), and cardiomyopathy and neu- ropathy may continue to worsen after transplantation. Recently, small molecule stabilizers of variant TTR (Tafami- dis and diflunisal) shown to inhibit amyloid fibril forma- tion in vitro are being studied in clinical trials (Berk et al., 2012; Coelho et al., 2012). Patient-specific, cell-based models are needed to facilitate the study of the genetic and epigenetic factors in this disease, permitting pharma- cogenomic assessments of novel therapeutics. The generation of induced pluripotent stem cells (iPSCs) through the reprogramming of somatic cells from patients with inherited diseases provides an unprecedented oppor- tunity to study the effects of genetic abnormalities and dis- ease progression. The derivation of unlimited quantities of Stem Cell Reports j Vol. 1 j 451–463 j November 19, 2013 j ª2013 The Authors 451