Introduction
Adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP) is an autosomal dominant neurological disorder that predominantly affects the cerebral white matter [
1]. ALSP encompasses two similar entities previously known as hereditary diffuse leukoencephalopathy with spheroids (HDLS) and pigmentary orthochromatic leukodystrophy (POLD) [
1]. ALSP is clinically characterized by executive dysfunction, memory decline, personality changes, motor impairments, and seizures [
2]. Frontal lobe syndrome in ALSP is characterized by loss of judgment, lack of social inhibitors, lack of insight, and motor persistence, which usually appears early in the disease course. The mean age at onset is 43 years ranging from 18 to 78 years [
3]. Patients with ALSP eventually become bedridden with a mean disease duration of 6.8 years from onset to death [
3].
Mutation in the colony stimulating factor 1 receptor (
CSF1R) was identified as the cause of ALSP [
4]. To date, 58 pathological mutations in
CSF1R have been identified [
3]. All reported mutations were found in exons 12−22 including the coding sequence of the tyrosine kinase domain (TKD) of
CSF1R [
1‐
3]. We previously reported that
CSF1R mutation-mediated pathogenesis may be explained by haploinsufficiency or the loss of CSF1R-mediated signals [
5].
In this study, we attempted to identify the CSF1R mutations in patients who were referred to our institute for genetic analysis. By this analysis, we found novel and previously reported CSF1R mutations including a novel frameshift in exon 4 outside of TKD and examined the pathogenicity of novel CSF1R variants.
Discussion
In this study, we identified seven novel and two previously reported mutations in
CSF1R among patients who fulfilled the diagnostic criteria of ALSP [
6]. This study provides several noteworthy insights relevant to the role of
CSF1R mutations in ALSP.
First, we identified a novel frameshift mutation caused by single-nucleotide deletion (c.310delC) in exon 4 resulting in p.Pro104LeufsTer8 located outside of TKD. All the previously reported mutations in ALSP were found within exons 12−22 including the coding sequence of TKD [
3‐
5]. The frameshift mutation p.Pro104LeufsTer8 generates a premature stop codon, which was predicted to cause NMD [
8]. Our expression analysis revealed that the level of mRNA expression derived from the mutant allele was substantially decreased as predicted (Fig.
2a). These findings suggest that the p.Pro104LeufsTer8 mutation causes ALSP owing to the haploinsufficiency of
CSF1R.
Second, we identified another novel frameshift mutation caused by two-nucleotide deletion (c.2655_2656delAT) in exon 21 resulting in p.Tyr886GlnfsTer55. This frameshift mutation does not fulfill the criterion of NMD because the premature stop codon is generated within exon 22, the last exon of
CSF1R. Our expression analysis revealed that the mRNA expression level of the mutant allele was comparable to that of the wild-type allele (Fig.
2b). These findings suggest that this frameshift mutation does not cause
CSF1R haploinsufficiency. Because this mutation is located within TKD, we examined whether the mutant CSF1R of p.Tyr886GlnfsTer55 affects autophosphorylation upon ligand stimulation. Our examination revealed that the mutant CSF1R of p.Tyr886GlnfsTer55 did not show autophosphorylation (Fig.
3a, b, d). Thus, the pathogenic mechanism caused by p.Tyr886GlnfsTer55 mutation appears to be the loss of CSF1R-mediated signals.
Third, we showed that the autophosphorylation of novel missense variants (p.Ile662Thr, p.Asp778Glu, p.Ile794Phe, p.Pro878Ser, and p.Pro878Ala) within TKD was impaired. These findings suggest that these mutants within TKD are the causal factors [
4,
5]. As previously reported [
3], pathogenic mutations identified in this study occur more frequently in the distal part of TKD than in the proximal part. There is apparently no significant difference in the degree of autophosphorylation impairment between the p.Ile662Thr mutant within the proximal part of TKD and other mutants within the distal part of TKD.
Finally, we identified the homozygous
CSF1R variant of p.His362Arg located in the extracellular domain in patient 2 who fulfilled the possible criteria of ALSP. We considered that the p.His362Arg missense variant is not the causative factor for the following reasons. First, the degree of autophosphorylation of this variant was comparable to that of the wild type. Second, this variant was reported in the ExAC database to have a frequency of 0.049 in the general population and 0.368 in the East Asian population. Thus, the cause of leukoencephalopathy in this patient is not likely to be this
CSF1R variant. Recently, mutations in
AARS2 have been reported in ALSP patients who lack
CSF1R mutations [
9]. We performed genetic analysis of
AARS2 and found no causative mutation in this patient.
The clinical and neuropathological findings with
CSF1R mutations in this study were consistent with those reported previously [
1‐
3,
7]. Indeed, all the patients in this study fulfilled possible or probable diagnostic criteria for ALSP [
6]. The MRI findings of the patients were similar to those reported previously (Supplementary Table 2) [
10]. Attention should be paid to patient 1 in whom signal changes detected by MRI were observed with marked left-side predominance (Fig.
4). Brain calcifications were observed in all the patients with
CSF1R mutation, supporting the diagnostic value of this finding for ALSP [
11].
In conclusion, the detection of the CSF1R mutation outside of exons 12−22 may extend the genetic spectrum of ALSP with CSF1R mutations. Mutational analysis of all the exons of CSF1R should be considered for patients clinically suspected of having ALSP.
Acknowledgements
We would like to thank the patients and their relatives for participating in this study and Ms. Reiko Kawai for her technical assistance. Dr. Konno is supported by JSPS Overseas Research Fellowships and the gift from Carl Edward Bolch, Jr., and Susan Bass Bolch. Dr. Wszolek is partially supported by the NIH/NINDS P50 NS072187, NIH/NIA (primary) and NIH/NINDS (secondary) 1U01AG045390-01A1, Mayo Clinic Center for Regenerative Medicine, Mayo Clinic Neuroscience Focused Research Team (Cecilia and Dan Carmichael Family Foundation, and the James C. and Sarah K. Kennedy Fund for Neurodegenerative Disease Research at Mayo Clinic in Florida), the gift from The Sol Goldman Charitable Trust, and Donald G. and Jodi P. Heeringa. Dr. Ikeuchi is supported by JSPS KAKENHI Grant number JP16H01331 and 26117506, AMED under Grant number JP18kk0205009, and MHLW Grant number 18062640.