Mutations in the ABCD1 gene that encodes peroxisomal
ABCD1 protein cause
X-linked adrenoleukodystrophy (
X-ALD), a rare
neurodegenerative disorder. More than 70% of the patient fibroblasts with this missense mutation display either a lack or reduction of the
ABCD1 protein because of posttranslational degradation. In this study, we analyzed the stability of the missense mutant ABCD1
proteins (p.A616T, p.R617H, and p.R660W) in
X-ALD fibroblasts and found that the mutant
ABCD1 protein p.A616T has the capacity to recover its function by incubating at low temperature. In the case of such a mutation, chemical compounds that stabilize mutant ABCD1
proteins could be therapeutic candidates. Here, we prepared CHO cell lines stably expressing ABCD1
proteins with a missense mutation in fusion with
green fluorescent protein (GFP) at the C-terminal. The stability of each mutant ABCD1-GFP in CHO cells was similar to the corresponding mutant
ABCD1 protein in
X-ALD fibroblasts. Furthermore, it is of interest that the GFP at the C-terminal was degraded together with the mutant
ABCD1 protein. These findings prompted us to use CHO cells expressing mutant ABCD1-GFP for a screening of chemical compounds that can stabilize the mutant
ABCD1 protein. We established a fluorescence-based assay method for the screening of
chemical libraries in an effort to find compounds that stabilize mutant ABCD1
proteins. The work presented here provides a novel approach to finding therapeutic compounds for
X-ALD patients with missense mutations.