Novel Corrector for Variants of SLC6A8: A Therapeutic Opportunity for Creatine Transporter Deficiency

ACS Chem Biol. 2024 Nov 15;19(11):2372-2382. doi: 10.1021/acschembio.4c00571. Epub 2024 Oct 17.

Abstract

Mutations in creatine transporter SLC6A8 cause creatine transporter deficiency (CTD), which is responsible for 2% of all cases of X-linked intellectual disability. CTD has no current treatments and has a high unmet medical need. Inspired by the transformational therapeutic impact of small molecule "correctors" for the treatment of cystic fibrosis, which bind to mutated versions of the CFTR ion channel to promote its trafficking to the cell surface, we sought to identify small molecules that could stabilize SLC6A8 as a potential treatment for CTD. We leveraged a novel chemoproteomic technology for ligand discovery, reactive affinity probe interaction discovery, to identify small-molecule fragments with photoaffinity handles that bind to SLC6A8 in a cellular environment. We synthesized a library of irreversible covalent analogs of these molecules to characterize in functional assays, which revealed molecules that could promote the trafficking of mutant SLC6A8 variants to the cell surface. Further medicinal chemistry was able to identify reversible drug-like small molecules that both promoted trafficking of the transporter and also rescued creatine uptake. When profiled across the 27 most prevalent SLC6A8 missense variants, we found that 10-20% of patient mutations were amenable to correction by our molecules. These results were verified in an endogenous setting using the CRISPR knock-in of selected missense alleles. We established in vivo proof-of-mechanism for correctors in a novel CTD mouse model with the P544L patient-defined variant knocked in to the SLC6A8 locus, where treatment with our orally bioavailable and brain penetrant tool corrector increased brain creatine levels in heterozygous female mice, validating correctors as a potential therapeutic approach for CTD.

MeSH terms

  • Animals
  • Brain Diseases, Metabolic, Inborn / drug therapy
  • Brain Diseases, Metabolic, Inborn / genetics
  • Brain Diseases, Metabolic, Inborn / metabolism
  • Creatine / deficiency
  • Creatine / metabolism
  • HEK293 Cells
  • Humans
  • Language Development Disorders / drug therapy
  • Language Development Disorders / genetics
  • Membrane Transport Proteins
  • Mental Retardation, X-Linked* / drug therapy
  • Mental Retardation, X-Linked* / genetics
  • Mental Retardation, X-Linked* / metabolism
  • Mice
  • Muscle Hypotonia / drug therapy
  • Muscle Hypotonia / genetics
  • Mutation
  • Nerve Tissue Proteins / genetics
  • Nerve Tissue Proteins / metabolism
  • Plasma Membrane Neurotransmitter Transport Proteins* / deficiency
  • Plasma Membrane Neurotransmitter Transport Proteins* / genetics
  • Plasma Membrane Neurotransmitter Transport Proteins* / metabolism
  • Small Molecule Libraries / chemistry
  • Small Molecule Libraries / pharmacology

Substances

  • Plasma Membrane Neurotransmitter Transport Proteins
  • SLC6A8 protein, human
  • Creatine
  • Small Molecule Libraries
  • Nerve Tissue Proteins
  • creatine transporter
  • Membrane Transport Proteins

Supplementary concepts

  • Creatine deficiency, X-linked