Novel mRNA-Based Therapy Reduces Toxic Galactose Metabolites and Overcomes Galactose Sensitivity in a Mouse Model of Classic Galactosemia

Abstract

Classic galactosemia (CG) is a potentially lethal inborn error of galactose metabolism that results from deleterious mutations in the human galactose-1 phosphate uridylyltransferase (GALT) gene. Previously, we constructed a GalT À/À (GalT-deficient) mouse model that exhibits galactose sensitivity in the newborn mutant pups, reduced fertility in adult females, impaired motor functions, and growth restriction in both sexes. In this study, we tested whether restoration of hepatic GALT activity alone could decrease galactose-1 phosphate (gal-1P) and plasma galactose in the mouse model. The administration of different doses of mouse GalT (mGalT) mRNA resulted in a dose-dependent increase in mGalT protein expression and enzyme activity in the liver of GalT-deficient mice. Single intravenous (i.v.) dose of human GALT (hGALT) mRNA decreased gal-1P in mutant mouse liver and red blood cells (RBCs) within 24 h with low levels maintained for over a week. Repeated i.v. injections increased hepatic GalT expression, nearly normalized gal-1P levels in liver, and decreased gal-1P levels in RBCs and peripheral tissues throughout all doses. Moreover, repeated dosing reduced plasma galactose by 60% or more throughout all four doses. Additionally, a single intraperitoneal dose of hGALT mRNA overcame the galactose sensitivity and promoted the growth in a GalT À/À newborn pup.

Current and Future Treatments for Classic Galactosemia

“…The required repeated dosing of this therapeutic modality tends to raise concerns about immunogenicity and therapy burden for the patients. As with gene therapy targeting, potential long-term adverse effects therapeutic index and dosing interval of mRNA remain important challenges for the future [ 60 , 68 , 70 ]. The transient nature of the mRNA requiring repeated dosing to remain effective also bears an advantage, by allowing individual dose titration and cessation of therapy if necessary [ 60 , 68 , 70 ].…”

“…Several preclinical studies evaluating the potential of systemically administered mRNA in metabolic disorders have shown that mRNA provides transient, half-life dependent protein expression, while bearing a low risk of insertional mutagenesis and maintaining dose-responsiveness [ 67 , 68 , 69 , 71 , 72 , 73 , 74 , 75 ]. This therapeutic approach is being explored in classic galactosemia using lipid nanoparticles as delivery system.…”

“…This therapeutic approach is being explored in classic galactosemia using lipid nanoparticles as delivery system. Balakrishnan et al recently showed in adult Galt knockout mice that systemic administration of LNP-packaged mouse and hGALT mRNA resulted in hepatic expression of active, long-lasting GALT enzyme and effectively eliminated galactose 1-phosphate in liver and other peripheral tissues [ 68 ]. Using the galt knockout zebrafish model, Haskovic et al also showed successful delivery and translation of functional hGALT after single dose injections with LNP-packaged hGALT mRNA ( n = 3) delivered at day 0 and measured on day 5 post fertilization (dpf) [ 76 ].…”

“…Exogenous, intracellular delivery of mRNA is emerging as a new class of medicine [59][60][61][62]. Recent advances in mRNA technology have led to improvements in stability and translation by codon optimization and encapsulation within biodegradable delivery systems, preserving mRNA integrity and allowing targeted intracellular delivery [59][60][61][62][63][64][65][66][67][68][69][70]. The required repeated dosing of this therapeutic modality tends to raise concerns about immunogenicity and therapy burden for the patients.…”

Gene therapy for inherited metabolic diseases

“…Lipid nanoparticles are composed of phospholipids, sterols and polyethylene glycol ( 80 )-conjugated lipids, which protect the mRNA from nuclease degradation and immune responses and help in their cellular uptake ( 81 ). mRNAs encapsulated in LNPs have been trialled in animal models of methylmalonic acidaemia ( 82 ), acute intermittent porphyria ( 83 ), ornithine transcarbamylase ( 84 ) deficiency ( 85 ), arginase deficiency ( 86 ) and galactosaemia ( 87 ). Phase I/II clinical trials have been initiated in OTC patients to test the safety and efficacy of OTC mRNA formulated in lipid nanoparticles ( 76 ).…”