Phenotype Presentation and Molecular Diagnostic Yield in Non-5q Spinal Muscular Atrophy

Clinical Description of Patients and Ancillary Testing

Seventy-one patients from 65 different families were included. Thirty-four patients (47.9%) were female, and most patients had no family history of neuromuscular disease or consanguinity and were therefore classified as sporadic (60.6%). Onset of the disease ranged from 0 to 58 years, and 36 patients (50.7%) had an early onset (younger than 18 years). At presentation (38.2 ± 14.9 years), 21 patients (29.6%) showed exclusive proximal weakness (P-SMA), 35 (49.3%) associated distal weakness (PD-SMA), and 15 patients (21.1%) a scapuloperoneal phenotype (SP-SMA). Clinical and ancillary data are shown in Table 1. Differences in clinical variables between the P-SMA, PD-SMA, and SP-SMA groups are shown in eTable 2 (links.lww.com/NXG/A615).

On ENMG, all patients had neurogenic motor unit potentials as per inclusion criteria, with 27 (38%) showing reduction of CMAP amplitude and 6 (8.5%) a minimal reduction of SNAPs associated with the peripheral motor neuron involvement.

For 60 patients, follow-up data were available and 5 patients (8.3%) had lost ambulation at a median of 5 years (range 4–54 years) from symptom's onset.

Genetic Diagnosis and Predictive Variables

In 32 patients (45.1%), a genetic diagnosis was achieved. The molecular diagnostic yield was lowest in the P-SMA (6/21, 28.6%) vs the PD-SMA (16/35, 45.7%) and SP-SMA (10/15, 66.7%) clinical groups. The different genes found in each non-5q SMA clinical group can be seen in Figure 1.

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Figure 1 Distribution of Patients With a Positive Genetic Diagnosis According to the Spinal Muscular Atrophy (SMA) Subtype

Thirty-two patients with a genetic diagnosis achieved through neuropathy gene panel (n = 19: DYNC1H1 x6, BICD2 x4, TRPV4 x4, HSPB1, VCP, VRK1, DNAJB2, MORC2), individual gene sequencing (n = 8: BICD2 x3, AR x2, HSPB1, HEXB, and COL6A3), and other neuromuscular NGS (n = 5: BICD2 x2, DYNC1H1, VCP, and ASAH1). Neg: negative; NGS: next-generation sequencing; P-SMA: proximal SMA; PD-SMA; proximo-distal SMA; SP-SMA: scapuloperoneal SMA.

Fifty-nine neuropathy NGS gene panels were performed, with a positive genetic diagnosis in 19/59 (32.2%). There was no significant difference in the performance of the neuropathy gene panel from Hospice Civils de Lyon (n = 48) vs other neuropathy gene panels in this series (n = 11) (eTable 3, links.lww.com/NXG/A616). In 8 additional patients, a genetic diagnosis was achieved through individual gene sequencing: BICD2 (3 patients from a previously reported family with 6 affected individuals),¹⁶ AR (n = 2), HSPB1, HEXB, and COL6A3. Reasons for this were genetic testing before routine gene panel sequencing in family members were an index case was found to carry a variant in the gene through exome sequencing (BICD2),¹⁶ genetic testing before routine gene panel sequencing and a plausible clinical picture (HSPB1), a particular or noncorresponding pattern on muscle MRI (HEXB and COL6A3), gynecomastia and elevated CK levels (patient with Kennedy disease), and a negative neuropathy gene panel and elevated CK levels (second patient with Kennedy disease). Finally, 5 patients were diagnosed through an alternative neuromuscular NGS: BICD2 (n = 2), DYNC1H1, VCP, and ASAH1. Furthermore, 20 patients underwent WES, which did not confirm any additional pathogenic or likely pathogenic variants in a known neurologic disease-causing gene.

Patients who received a definitive genetic diagnosis were more likely to have an earlier age at onset, a family history of the disease or consanguinity, and contractures and/or arthrogryposis. In a multivariate logistic regression, an earlier onset and a family history of the disease or consanguinity retained statistical significance (Table 2).

Clinical Features of the Most Common Non-5q SMA–Associated Genes

The most common genes in non-5q SMA patients were BICD2 (n = 9) and DYNC1H1 (n = 7), associated with SMA with lower extremity predominance (SMA-LED), and TRPV4 (n = 4), implicated in SP-SMA. These 3 genes are dominantly inherited; however, only 13/20 patients had a clear AD family history of the disease, the rest were therefore categorized as sporadic cases. We summarize below the features of these 3 most common genes.

1. BICD2 (9 patients from 7 different families). Variants: c.1502G>C (n = 3) from the previously reported family,¹⁶ c.2042C>T (n = 2), c.2108C>T, c.2080C>T, c.1617C>G, and c.380A>G. The latter 2 had never been reported: The c.1617C>G variant segregated correctly within the family and predicted as likely pathogenic in silico; the c.380A>G variant was predicted as a likely pathogenic in silico (because it is absent from population databases, well conserved, and situated in a mutational hotspot, with a clinical and muscle MRI phenotype compatible with BICD2 SMA-LED).¹⁷ However, given that SMA-LED is not a disease with a single genetic etiology and that segregation was not performed, we retain this variant as a variant of unknown significance (VUS) for now. Globally, although all types of phenotypical presentations were possible between and within families [PD-SMA (n = 5), SP-SMA (n = 3) or P-SMA (n = 1)], BICD2 patients frequently showed some degree of distal weakness on presentation (7/9) and a disease onset younger than 10 years (8/9). Normal or brisk deep tendon reflexes (DTR) (6/9) with 1 patient presenting some degree of spasticity, scapula alata (4/9), and axial weakness (3/9) were relatively frequent. Two patients had mild respiratory insufficiency, 1 needing overnight NIV. No patient had lost ambulation at follow-up. All patients had a motor neuronopathy on ENMG, one with mild sensory involvement.

2. DYNC1H1 (7 patients from 7 families). Variants: c.2327C>T (n = 2), c.752G> A, c.1792C>T, c.752G>T, c.1427T>C, c.596A>C. The latter 2 variants have never been reported and were predicted as likely pathogenic in silico. Segregation for the c.596A>C confirmed a de novo origin; however, segregation for the c.1427T>C variant could not be performed because the patient was adopted. Clinically, DYNC1H1 patients presented most frequently a PD-SMA (5/7). Disease onset was always in younger than 5 years, 3/7 patients had neonatal arthrogryposis, and 3/7 learning difficulties since childhood. DTRs were mostly abolished (6/7). Only 1 patient had moderate respiratory insufficiency in the context of obesity. Of the 4 patients who were followed up, none had lost ambulation. All patients had a pure motor neuronopathy on ENMG.

3. TRPV4 (4 patients from 3 families). Variants: c.557G>A (n = 2) and c.694C>T (n = 2), all previously reported. These patients showed predominantly a SP-SMA (3/4), although 1 patient presented with a PD-SMA without scapula alata. Although onset of the first progressive symptom was in adult age (18–53 years), 2 patients had a personal history of arthrogryposis and one of scoliosis and pes cavus. DTRs were always abolished. Three patients had some degree of respiratory insufficiency, none requiring ventilation. Three of these patients also had vocal cord paralysis with hypophonia. One patient showed a minimal reduction of normal SNAP on ENMG. All but 1 patient remain ambulant, the latter losing ambulation at age 5 years (before the onset of weakness) because of arthrogryposis.

Other Non-5q SMA–Associated Genes

Other variants in genes implicated in HMNs were found. In 2 SP-SMA patients with lower-limb distal involvement, scapula alata, and a disease onset in adult life, HSPB1 variants were found (AD transmission). Two patients (SP-SMA and P-SMA) carried variants in the VCP and one (SP-SMA) in the VRK1 gene (both AD), all 3 with an adult disease onset. A previously reported patient with an adult-onset PD-SMA carried a variant in the MORC2 gene.¹⁸ One patient with an infantile-onset PD-SMA carried a homozygous pathogenic variant in the DNAJB2 gene (AR). The specific genetic variants can be seen in eTable 1 (links.lww.com/NXG/A614).

It is important that 2 patients with a PD-SMA phenotype without clinical nor ENMG sensory involvement and an onset of symptoms in their late 20s carried CAG repeat expansions in the AR gene (AD transmission). None of these patients had bulbar symptoms, and only one had very high CK levels (2000U/L) and gynecomastia.

Furthermore, 2 patients carried biallelic variants (AR transmission) in genes implicated in lysosomal storage disorders (LSD). One patient carries 2 new compound heterozygous pathogenic variants in the HEXB gene (c.881A>G and c.1082+2T>C), compatible with a diagnosis of Sandhoff disease. This patient presented with a pure P-SMA, an onset of symptoms at 11 years, and no other neurologic manifestation of the disease (cerebellar, extrapyramidal, cognitive, or psychiatric); DTRs were normal, and there was no upper limb tremor. ENMG was compatible with a motor neuronopathy. Importantly, the patient was found to carry a VUS in the DYNC1H1 gene, but the muscle MRI pattern did not correspond to the one previously described in DYNC1H1-associated SMA (Figure 2),¹⁹ so a leukocyte hexosaminidase analysis was performed, finding deficient A and B enzymatic activities. A second patient which we have previously reported²⁰ carries compound heterozygous variants in the ASAH1 gene (c.77C>G and c.125+1G>A) and has a deficient leukocyte ceramidase activity. This patient presented with a pure P-SMA, an onset of symptoms at 19 years and again no other neurologic manifestation of the disease (epilepsy). The patient had scoliosis and club feet, upper limb tremor, and retained DTRs. ENMG was compatible with a motor neuronopathy.

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Figure 2 Differing MRI Muscle Involvement in Non-5q Spinal Muscular Atrophy (SMA) Patients

T1-weighted MRI sequences of 5 patients with variants in the 3 most frequent non-5q SMA genes (DYNC1H1, BICD2, and TRPV4) as well as in Sandhoff disease (HEXB) and COL6A3. Muscles with yellow arrows had marked fat replacement; muscles with white arrows were completely or partially spared. (A) Quadriceps, semitendinosus, and adductor magnus muscle degeneration in a patient with Sandhoff disease presenting as P-SMA. (B) The previously described quadriceps muscle degeneration with adductor longus and semitendinosus sparing in a patient with DYNC1H1 PD-SMA.¹⁹ In this case, sartorius and gracillis muscle fat replacement is also observed. (C) Quadriceps and sartorious muscle degeneration with more important lower limb involvement in a patient with BICD2 PD-SMA. Notice some degree of paraspinal muscle atrophy and important scoliosis. (D) Tibialis anterior and gastrocnemius medialis involvement with sparing of muscles of the thigh and paraspinal and deltoid muscle degeneration in a patient with TRPV4 SP-SMA. (E) Finally, the characteristic muscle fat replacement in a “sandwich patter” of soleus and quadriceps muscles (yellow arrowheads) in a patient with COL6A3 mutations and a pure SMA phenotype. AL = adductor longus; AM = adductor magnus; De = deltoid; GM = gastrocnemius medialis; Gr = gracillis; PS = paraspinal; Quad = quadriceps; Sa = sartorious; ST = semitendinosus; TA = tibialis anterior.

Finally, in 1 patient with a PD-SMA, retained DTRs, contractures in all 4 limbs since early childhood, and a neurogenic ENMG and muscle biopsy with no evidence of myopathy, who had no genetic diagnosis (after a 102 gene neuropathy panel, SMN1 deletion, and hexosaminidase analysis); a characteristic “sandwich” sign on muscle MRI, typical of collagenopathies, was observed (Figure 2).²¹ Direct sequencing of the COL6A3 gene found a previously unreported likely pathogenic homozygous c.5867A>G variant as well as a previously described heterozygous missense variant (c.7447A>G), which probably acts as a modulator of the disease.²² A sibling, who presents a similar clinical phenotype and a predominant neurogenic ENMG and muscle biopsy, carried the same variants. Their mother was heterozygous for the c.5867A>G variant, and the father was deceased.