A significant challenge of mRNA-based protein replacement therapies is the diminishing efficacy and escalating toxicity associated with repeated dosing of lipid nanoparticles (LNPs). Many existing lipid formulations were originally designed for vaccine and are not optimized for therapeutic applications. We developed two libraries of ionizable lipids—one based on piperazine and the other on a newly introduced cyclohexane structure—with variations in linker and tail groups to enhance molecular diversity. GC Biopharma’s cyclohexane- and piperazine-based LNPs (GCP LNPs) supported stable and high-level expression without inducing liver toxicity under repeated dosing regimens. These LNPs effectively corrected disease markers in mouse models of phenylketonuria (PKU) and succinic semialdehyde dehydrogenase (SSADH) deficiency. Specially, we observed that the rigid structure and chemical stability of cyclohexane-based lipids contributed to sustained delivery performance. These findings offer a promising direction for the development of LNPs suitable for chronic mRNA-based therapies.