A Novel SPAST Variant Associated with Isolated Spastic Paraplegia

Genetic variants in SPAST are the most common cause of hereditary spastic paraplegia (HSP), entitled spastic paraplegia type 4 (SPG4). Inheritance is autosomal dominant, and age of onset can vary from childhood to adulthood. Pathogenic SPAST variants are often observed in isolated cases, likely due to reduced penetrance and clinical variability. We report an isolated case of SPG4 associated with a novel likely pathogenic variant in SPAST. A 38-year-old woman presented with an eight-year history of progressive difficulty walking. Neurological examination revealed spastic paraparesis in the absence of upper motor neuron dysfunction, sensory deficits, or intellectual disability. Magnetic resonance imaging (MRI) of the brain and spinal cord was normal. No family members had similar complaints. Genetic analysis revealed a novel heterozygous sequence variant in SPAST, c.1751A > G p.(Asp584Gly) (NM_014946.4). The affected amino acid is highly conserved among orthologue and paralogue species. Four other nucleotide substitutions predicted to affect the same amino acid, a “hot spot”, have been reported previously in adult-onset HSP. This report describes a novel SPAST variant in a female with HSP without a known family history of the disorder.


Introduction
Hereditary spastic paraplegia (HSP) consists of a group of neurological disorders characterised by length-dependent corticospinal tract and dorsal column degeneration, typically resulting in bilateral lower limb spasticity, hyperrefexia, and extensor plantar responses [1].In a populationbased study in Norway, the overall prevalence of HSP was 7.4/100,000.Te prevalence of cases considered to be autosomal dominant or autosomal recessive based on family history was 5.5/100, 000 and 0.6/100,000, respectively.Te prevalence of isolated cases, i.e., cases without a known family history, was 1.3/100 000 [2].A German study has more recently reported that 40% of cases are isolated [3].
Pathogenic and likely pathogenic variants in over 80 genes can cause spastic paraplegia [4].Several hundred variants in SPAST can underlie spastic paraplegia type 4 (SPG4).Pathogenic variants in SPAST account for approximately 25% of autosomal dominant HSP, and the SPAST gene is the most frequently mutated gene in isolated cases [2,5].Wide interand intrafamilial clinical variability exists, and the age of onset ranges from the frst to the eighth decade.Penetrance is almost complete after the seventh decade [6].We report an isolated case with a novel likely pathogenic variant in SPAST.

Case Presentation
Te patient is a 38-year-old woman who works full-time as a secretary.She underwent bilateral femoral osteotomies at age ten years and developed low back pain without sciatica from age 18. Magnetic resonance imaging (MRI) revealed spondylolisthesis at L4/5 with bilateral compression of L5 nerve roots.At age 36 years, she was referred to a neurologist with a six-year history of difculty walking in the absence of other complaints.She was able to walk approximately 100 meters without assistance.Her gait disturbance was least pronounced in the morning and after rest.She had no upper limb, visual, neuropsychiatric, or urinary symptoms.
Neurological examination was normal except for spastic paraparesis with bilateral ankle clonus, patellar subclonus, and positive Babinski sign.She did not have nystagmus, dysarthria, dysphagia, muscle wasting, extrapyramidal signs, or sensory loss.Her HSP rating scale (HSPRS) score was 17 of 52 [7].She had low levels of serum vitamin B9 (4 nmol/l) and D (29 nmol/l), but there was no evidence of metabolic or infammatory disease.MRI of the brain and the spinal cord was normal.
Introduction of baclofen 25 mg three times a day had some efect on her spasticity.However, her HSPRS score remained unchanged at 18-month follow-up.
Her mother who is 59 years old and her mother's halfsister have no difculty walking, according to the proband.Her father died at the age of 57 years; he had low back pain and no difculty walking.Our patient reports that her two 33-year-old half-siblings (shared father) and her 12-year-old son have no difculty walking.Te pedigree is shown in Figure 1.It has not been possible to interview, examine, or collect blood samples from the patient's relatives.

Discussion/Conclusions
SPG4 is caused by both missense and loss of function variants altering SPAST and is typically associated with slowly progressive lower extremity weakness and corticospinal tract signs, with or without sphincter disturbances and deep sensory loss, but without other neurological symptoms or signs [12].Tis is in keeping with the phenotype in our patient.
Spastin has a key role in neuronal axons where it modulates microtubule number, length, and motility [13].Parodi et al. analysed a cohort of 842 patients with SPG4 to assess genotype-phenotype correlations.Age of onset in individuals carrying missense variants tended to be in the frst to second decade, whereas individuals with loss of function variants often experienced symptoms starting in the second to sixth decade.Tere was no diference in clinical severity, but intellectual disability was more common in individuals with missense variants [6].Our patient who carried a missense variant had the onset in her early thirties and no intellectual disability.Interestingly, other patients carrying missense variants afecting the same codon also have relatively late onset ranging from 21 to 40 years [6,[8][9][10].An individual heterozygous for p.(Asp584His) had not developed symptoms at the age of 70 [10].Variation afecting p.Asp584 may predispose to a phenotype often seen in patients with loss of function variants.Te patient presented in our study reported that no other family members had symptoms similar to hers.However, since it was not possible to confrm her negative family history, other family members may carry the variant.Family history for SPG4   Case Reports in Genetics may appear to be negative due to age-related/reduced penetrance or death prior to the onset of the disorder [4].
Te proportion of cases caused by a de novo variant is low [4].
In conclusion, we report a novel heterozygous likely pathogenic missense variant in SPAST in an apparently isolated case of pure HSP.

Figure 2 :
Figure 2: Sequencing data, conservation, and in silico modelling.(a) Sequencing data shown in integrative genomics viewer (Broad Institute) for the proband (upper panel) and a control (lower panel).Te nucleotide substitution from adenine (A) to guanine (G) is shown in orange.(b) Evolutionary conservation of the amino acid afected by the spastin p.(Asp584Gly) variant across diferent species.Te afected amino acid is shown in red.Blue indicates amino acids that are similar to the human sequence.Data from Alamut Visual Software (Sophia Genetics) and protein BLAST (https://blast.ncbi.nlm.nih.gov/Blast.cgi).(c) Spastin domain structure and paralog conservation of amino acids 571-597 among 14 related orthologues identifed from the protein data bank (PDB).Alignment was created by VarSite (EMBL-EBI).Te amino acids in the paralogue alignment are coloured by residue type.Shown below the alignment is the number of disease-causing variants at each position based on the p.(Asp584Gly) substitution reported in our case and reports in the human gene mutation database (HGMD) professional version 2022.2 (QiaGen).N-term � N-terminal sequence, MIT �microtubule interacting and trafcking domain, MTBD � microtubule-binding domain, and AAA � ATPase associated with various cellular activities.Te asparagine (D) residue at position 584 is marked with a box.(d) Cartoon modelling of the human spastin hexamonomer.Tere is one colour for each monomer.Te Asp584 residue is shown in red in each of the six monomers.(e) Localization of the p.Asp584 residue (dark red) and its interacting amino acids (orange).Hydrogen bonds (light blue) predicted with p.Arg578, p.Arg581, and p.Ser588.(f ) Localization of the p.(Asp584Gly) substitution (dark red) and interacting amino acids (orange).Hydrogen bonds (light blue) are predicted to be formed with p.Ser588, but not with p.Arg578 and p.Arg581.Te position of the new amino acid change within the structure was estimated using the Rotamers tool with standard parameters.Te modelling in (d), (e), and (f ) was performed using ChimeraX-1.5 on the spastin (PDB ID: 6PEN) crystal structure.
Figure 1: Pedigree.Arrow � proband, black symbol � afected by SPG4, white symbols � not reported by proband to have difculty walking, and white symbols with ?� unknown status.d � died, y � years.