replacement therapy must be explored, considering the benefit of the non-invasive nature of these evaluations. doi:10.1016/j.ymgme.2016.11.049 41 Messenger RNA (mRNA) delivery to the liver corrects ornithine transcarbamylase deficiency in a mouse disease model Gordon Brandt, Mary Prieve, Eric Bell, Teri Blevins, Anna Galperin, Pierrot Harvie, Allen Li, Jean-Rene Ella Menye, Sean Monahan, Amber Paschal, Debashish Roy, Matt Waldheim, Michael Houston, PhaseRx, Inc., Seattle, WA, United States Introduction: Messenger RNA (mRNA) replacement is a promising approach for treatment of single gene inherited metabolic diseases such as ornithine transcarbamylase deficiency (OTCD). We report on the development of a liver-targeted mRNA delivery platform which has shown efficacy in multiple disease indications and the preclinical proof of concept in a model of OTCD. Methods: The definitive preclinical model for OTCD is the OTC spf ash mouse model in which the mouse OTC mRNA is ablated resulting in urea cycle dysfunction, rapid development of hyperammonemia, and death. PRX-OTC, the liver-targeted mRNA drug product, replaces the missing mouse OTC mRNA with the human OTC mRNA. PRX-OTC was administered over 5 weeks at a dose of 3 mg/kg. Analyses included daily body weights, urinary orotic acid, plasma ammonia, survival, and human OTC protein expression in liver. Negative controls included buffer and an mRNA drug product identical to PRX-OTC except for a missing start codon. Results: Animals treated with buffer or negative control rapidly developed hyperammonemia and elevated urinary orotic acid, lost body weight and had a median survival of 20 days. PRX-OTC administration resulted in intra hepatic production of the human OTC enzyme which persisted for more than 21 days after the final dose. PRX- OTC treated animals showed normalization of plasma ammonia and urinary orotic acid, gain of body weight, and complete survival through the end of dosing (day 35) and for a further three weeks after dosing (day 56). No elevations in cytokines were observed in groups receiving PRX-OTC. Conclusions: PRX-OTC administration corrects OTCD in the spf ash model by replacing the human enzyme in the liver, thus normalizing plasma ammonia and extending survival for weeks after dosing. Clinical evaluation appears warranted. doi:10.1016/j.ymgme.2016.11.050 42 Investigation of newborns screened in a pilot program for four lysosomal diseases in Brazil Heydy V Bravo-Villalta a , Eurico C Neto b , Jaqueline Schulte b , Jamile Pereira b , Cláudio Sampaio-Filho c , Maira G Burin d , Fernanda H Bitencourt d , Fernanda M Sebastião d , Régis R Guidobono d , Ana C Brusius-Facchin d , Gabriela Pasqualim a , Diana E Rojas-Málaga a , Ursula S Matte a , Roberto Giugliani e , a UFRGS, Porto Alegre, - RS, Brazil, b Centro de Triagem Neonatal, Porto Alegre, - RS, Brazil, c Intercientífica, São José dos Campos, - SP, Brazil, d Hospital de Clinicas de Porto Alegre, Porto Alegre, - RS, Brazil, e Hospital de Clinicas de Porto Alegre, Porto Alegre, Brazil Newborn screening for lysosomal diseases has been gaining considerable interest, as several of these conditions are now treatable and as early treatment seems to improve patient outcomes. Also important, a number of different high-throughput platforms are now available for this screening. We present the preliminary results obtained with a fluorometric digital microfluidic platform to perform multiplexed enzymatic analysis of α-L-iduronidase (IDUA, to screen for MPS I), acid α-glucosidase (GAA, to screen for Pompe disease), acid β-glucosidase (GBA, to screen for Gaucher disease) and acid α-galactosidase (GLA, to screen for Fabry disease). The analyses were performed in dried blood spot (DBS) samples from 10,527 newborns randomly selected among the cards routinely received by the Neonatal Screening Center, based in Porto Alegre, Brazil. Time from sample preparation to enzyme activity result on each cartridge was less than 3 hours. Overall coefficient of variation (CV) values between cartridges, days, instruments, and operators ranged from 4 to 20%; linearity correlation coefficients were ≥ 0.98 for all assays. The method correctly discriminated samples of 9 affected patients (3 MPS I, 2 Pompe, 2 Gaucher and 2 Fabry diseases) from samples of normal babies. The analyses of all samples with activities below cutoff were repeated. After retesting, 4 showed consistently low enzyme activities (2 for IDUA, 1 for GLA and 1 for GBA). Further investigation of these 4 cases, performed at Hospital de Clinicas, Porto Alegre, Brazil, indicated 1 case of pseudodeficiency for MPS I, 1 case of pseudodeficiency for Pompe, 1 carrier for MPS I and 1 carrier for Gaucher. In conclusion, digital microfluidic technology shows potential for the routine testing for the 4 lysosomal diseases selected in a standard newborn screening laboratory. As in the screening for other conditions, well-established algorithms should be in place to quickly clarify any suspicion raised by the screening results. doi:10.1016/j.ymgme.2016.11.051 43 Sulfpraphane induces autophagy and reduces the level of mutated huntingtin in human fibroblasts Joanna Brokowska, Aleksandra Hac, Grzegorz Wegrzyn, Anna Herman-Antosiewicz, University of Gdansk, Gdansk, Poland Autophagy is a process of degradation of damaged organelles and aggregates of proteins by lysosomal acid hydrolases, therefore, lysosomes play a key role in this process. Autophagy can promotes survival, whereas in other cases it appears to promote programmed cell death. Disorders of autophagy may accompany many diseases such as Huntington disease, Parkinson disease, Crohn disease, obesity and mucopolysaccharidoses. It was demonstrated that in cancer cells autophagy can be induced by sulforaphane (SFN), a natural agent present in cruciferous plants. In our study, we determined whether sulforaphane is able to induce autophagy in normal, non-cancerous cells as well as to modulate the amount of mutated huntingtin in human fibroblasts expressing the mutated protein. In cells treated with sulforaphane, we observed reduction in the mutant huntingtin level as well as a decrease in the amount of its aggregates comparing to control cells. Simultaneously, we observed autophagy induction. Our study revealed that SFN inhibited a major negative regulator of autophagy, mTORC1. Autophagy induction and mTORC1 inhibi- tion was preceded by activation of AMPK kinase, a known inhibitor of mTORC1 and thus autophagy activator. The autophagy induction by SFN coincided with a block in protein synthesis which might be, together with the induction of autophagy, the molecular mecha- nism leading to reduction of the mutant huntingtin amount in cells by sulforaphane. Summarizing, our study shows that SFN induces autophagy as well as inhibits protein synthesis and suggests that sulforaphane can be used as potential therapeutic in diseases Abstracts / Molecular Genetics and Metabolism 120 (2016) S17–S145 S31