Non-coding repeat analyses in patients with Parkinson's disease

Abstract

Introduction: The genetic etiology of Parkinson's disease (PD) is complex; approximately 10% of patients with PD have various gene mutations that lead to familial forms of the disease. Recent analyses of non-coding repeat regions revealed that many neurodegenerative diseases are associated with pathological expansions. We evaluated the genetic background of non-coding repeat expansions in Japanese patients with PD.

Methods: We collected blood samples from 203 Japanese patients with PD and analyzed various non-coding repeat genes, including ATXN8OS, RFC1, C9ORF72, NOTCH2NLC, BEAN1/TK2, and NOP56, using PCR-Sanger sequencing, repeat-primed PCR assay, and long-read sequencing.

Results: Three patients with PD (1.5%) were found to have heterozygous repeat expansions in ATXN8OS, the gene causative of spinocerebellar ataxia type 8 and is associated with long non-coding RNA. One (0.5%) patient had compound heterozygous repeat expansions (AAGGG and ACAGG) in RFC1, the gene causative of cerebellar ataxia, neuropathy, and vestibular areflexia syndrome, which encodes a DNA repair protein. No patient had repeat expansions in C9ORF72, NOTCH2NLC, BEAN1/TK2, or NOP56. All patients with ATXN8OS repeat expansions exhibited typical parkinsonism with relatively rare subjective dysphagia, which was confirmed by videofluoroscopic results. Functional imaging, such as dopamine-transporter single photon emission computed tomography, showed abnormal findings in patients with non-coding repeat expansions.

Discussion: Our findings revealed the importance of non-coding repeat expansions in Japanese patients with PD. This is the first study to show the positive result of non-coding repeat expansions in many patients with PD in Japan.

Keywords: Canvas; RFC1; dysphagia; parkinsonism; repeat disease; spinocerebellar ataxia type 8; videofluoroscopic analysis.

Figure 1

Agarose gel electrophoretic analysis of the ATXN8OS gene in patients with Parkinson’s disease (Pt 1–3). Three patients had expansions of the CTA/CTG repeat as indicated, whereas controls (C) had a normal repeat size. M, 100 base pair (bp) marker.

Figure 2

(A) Repeat-primed PCR results for RFC1. Pathological repeats of AAGGG or ACAGG were expanded in a patient with Parkinson’s disease (PD). (B) Long-read sequencing revealed 212 repeats of AAGGG and 651 repeats of ACAGG in the patient with PD.

Figure 3

Imaging results of patients with ATXN8OS repeat expansions. (A) MRI of Patient 1 with 92 repeats (92r) showed no apparent atrophy of the cerebellum (upper panel) or cerebrum (lower panel). (B) MRI of Patient 2 with 101 repeats (101r) showed asymptomatic infarction of the cerebellum (arrow) and slight non-specific atrophy of the cerebellum or mild atrophy of the cerebrum with chronic ischemic lesions. (C) Computed tomography (CT) of the head in Patient 3 with 311 repeats (311r) showed no apparent atrophy of the cerebellum (left panel) or cerebrum (right panel). Dopamine-transporter single photon emission computed tomography showed marked reduction of the uptakes in the striatum. The specific binding ratio was 1.26 on the right striatum and 1.62 on the left.

Figure 4

Videofluoroscopic results in patients with ATXN8OS repeat expansions. (A) A patient with 92 repeats underwent aspiration (arrowhead). (B) A patient with 101 repeats had laryngeal penetration. (C) A patient with 311 repeats also had laryngeal penetration.

Figure 5

Imaging results of patients with RFC1 repeat expansions. (A) MRI of the patient with AAGGG and ACAGG repeat expansions. (B) ¹²³I-metaiodobenzylguanidine myocardial scintigraphy performed on a patient about 1 year after onset showed normal values of heart/mediastinum (H/M) ratio. The H/M ratios were standardized under medium-energy collimator conditions (normal ≥ 2.2). Additionally, the washout ratio was found to have increased (normal ≤ 22%).