biostudies-literatureDiep TNational Institute of Neurological Disorders and StrokeSwiss National Science FoundationNINDS NIH HHS102470https://www.ebi.ac.uk/biostudies/studies/S-EPMC11905892biostudies-literatureUnknown36(1)Carbamoyl phosphate synthetase 1 (CPS1) deficiency, a urea-cycle disorder, results in hyperammonemia initiating a sequence of adverse events that can lead to coma and death if not treated rapidly. There is a high unmet need for an effective therapeutic for this disorder, especially in early neonatal patients where mortality is excessive. However, development of an adeno-associated virus (AAV)-based approach is hampered by large cDNA size and high protein requirement. We developed an oversized AAV vector as a gene therapy to treat <i>CPS1</i> deficiency. In order to constrain genome size, we utilized small liver-specific promoter/enhancers and a minimal polyadenylation signal. Long-term survival (9 months, end of study) with ammonia control was achieved in AAV8.CPS1-administered Cps1<sup>flMolecular therapy. Nucleic acidsUse of an oversized AAV8 vector for CPS1 deficiency results in long-term survival and ammonia control.PMC11905892320030R61 NS121348207965Barshop BADiep TMakris GZhou WReyes RENitzahn MDay ILLueptow LLi YZhuravka IGangoiti JLipshutz GSBakshi SPadaon HHaberle JfalseUse of an oversized AAV8 vector for CPS1 deficiency results in long-term survival and ammonia control.Carbamoyl phosphate synthetase 1 (CPS1) deficiency, a urea-cycle disorder, results in hyperammonemia initiating a sequence of adverse events that can lead to coma and death if not treated rapidly. There is a high unmet need for an effective therapeutic for this disorder, especially in early neonatal patients where mortality is excessive. However, development of an adeno-associated virus (AAV)-based approach is hampered by large cDNA size and high protein requirement. We developed an oversized AAV vector as a gene therapy to treat <i>CPS1</i> deficiency. In order to constrain genome size, we utilized small liver-specific promoter/enhancers and a minimal polyadenylation signal. Long-term survival (9 months, end of study) with ammonia control was achieved in AAV8.CPS1-administered Cps1<sup>fl2025-01-01T00:00:00Z2025 Mar2026-06-01T22:56:55.722Z2025-04-06T10:51:12.238ZS-EPMC119058924008364610.1016/j.omtn.2025.102470