Optimized testing strategy for the diagnosis of GAA-FGF14 ataxia/spinocerebellar ataxia 27B

Abstract

Dominantly inherited GAA repeat expansions in FGF14 are a common cause of spinocerebellar ataxia (GAA-FGF14 ataxia; spinocerebellar ataxia 27B). Molecular confirmation of FGF14 GAA repeat expansions has thus far mostly relied on long-read sequencing, a technology that is not yet widely available in clinical laboratories. We developed and validated a strategy to detect FGF14 GAA repeat expansions using long-range PCR, bidirectional repeat-primed PCRs, and Sanger sequencing. We compared this strategy to targeted nanopore sequencing in a cohort of 22 French Canadian patients and next validated it in a cohort of 53 French index patients with unsolved ataxia. Method comparison showed that capillary electrophoresis of long-range PCR amplification products significantly underestimated expansion sizes compared to nanopore sequencing (slope, 0.87 [95% CI, 0.81 to 0.93]; intercept, 14.58 [95% CI, − 2.48 to 31.12]) and gel electrophoresis (slope, 0.84 [95% CI, 0.78 to 0.97]; intercept, 21.34 [95% CI, − 27.66 to 40.22]). The latter techniques yielded similar size estimates. Following calibration with internal controls, expansion size estimates were similar between capillary electrophoresis and nanopore sequencing (slope: 0.98 [95% CI, 0.92 to 1.04]; intercept: 10.62 [95% CI, − 7.49 to 27.71]), and gel electrophoresis (slope: 0.94 [95% CI, 0.88 to 1.09]; intercept: 18.81 [95% CI, − 41.93 to 39.15]). Diagnosis was accurately confirmed for all 22 French Canadian patients using this strategy. We also identified 9 French patients (9/53; 17%) and 2 of their relatives who carried an FGF14 (GAA)≥250 expansion. This novel strategy reliably detected and sized FGF14 GAA expansions, and compared favorably to long-read sequencing.

Spinocerebellar ataxia 27B: A novel, frequent and potentially treatable ataxia

“…23 This observation is in line with other studies that have shown SCA27B to have a frequency of approximately 15−30% in European cohorts of patients with unsolved adult-onset ataxia. 21,22,30,35,41,42 Such high prevalence may be reflective of a population structure in individuals of European descent who appear to more commonly carry larger FGF14 GAA alleles compared to other populations. 21,22 Future studies are needed to define the regional prevalence of SCA27B, although ongoing screening efforts suggest that SCA27B is also a relatively common cause of late-onset ataxia in South Asia (10%) 21 and Brazil (9%).…”

“…In comparison with the common polyglutamine SCAs which typically have a disease onset in the third to fifth decade, 48,49 SCA27B consistently presents in the fifth to seventh decade. [21][22][23]30,31,41,42,50 Some -but not allcases carrying biallelic FGF14 GAA repeat expansions may manifest before the age of 30 and/or have a more severe phenotype. 28,29 The GAA repeat length only appears to correlate weakly with the age at onset, 21,22,30,31,42 in contrast to polyglutamine SCAs.…”

“…21,30,50 The frequency and duration of episodes of ataxia are highly variable and may last from minutes to days and occur daily to monthly, respectively. Episodic symptoms, which may antedate permanent ataxia by several years, 21,31,41 may fail to be recognised by patients and providers alike to be related to an underlying cerebellar disorder. This may partly account for the significant discrepancy in the frequency of episodic symptoms reported in studies published thus far -varying from 13 to 80% 21,30,41,50,51 -and highlights the need to establish an accepted clinical definition of these symptoms.…”

“…Episodic symptoms, which may antedate permanent ataxia by several years, 21,31,41 may fail to be recognised by patients and providers alike to be related to an underlying cerebellar disorder. This may partly account for the significant discrepancy in the frequency of episodic symptoms reported in studies published thus far -varying from 13 to 80% 21,30,41,50,51 -and highlights the need to establish an accepted clinical definition of these symptoms. We had previously proposed the following definition: 'a recognizable constellation of symptoms, which are recurrent, intermittent and discrete with clear onset and offset from the patient's established baseline, and can appear unprovoked or be induced, for example, by alcohol or physical activity; patients must have episodic cerebellar symptoms (gait ataxia, dysarthria, diplopia, oscillopsia, vertigo and/or dizziness or appendicular ataxia), but can also have other episodic symptoms'.…”

Non‐GAARepeat Expansions inFGF14Are Likely Not Pathogenic—Reply to: “Shaking Up Ataxia:FGF14andRFC1Repeat Expansions in Affected and Unaffected Members of a Chilean Family”

“…This last point highlights the importance of adopting standardized protocols to diagnose SCA27B. 9 Moreover, segregation studies in affected families, tests of association in large case-control series, and, ultimately, functional studies will be needed in the future to establish the pathogenicity of alternative motifs at the FGF14-SCA27B repeat locus.…”