A Case Report of a Pedigree with Distal Hereditary Motor Neuropathy Caused by a Homozygous c.1124G>A Variant an the /*9Vaccinia-Related Kinase 1 Gene

Summary

This study reports a Chinese consanguineous pedigree with adult-onset distal hereditary motor neuropathy (dHMN) caused by a homozygous c.1124G>A variant in the vaccinia-related kinase 1 (VRK1) gene. The findings expand the clinical phenotypic and genetic spectrum of VRK1-related dHMN in the Chinese population.

Abstract

This study aimed to analyze the clinical phenotypes, neurophysiological characteristics, and pathogenicity of gene variants in a pedigree with distal hereditary motor neuropathy (dHMN) caused by VRK1 variants, and to provide evidence to support clinical diagnosis and genetic counseling for this disease. We report a Chinese consanguineous family with dHMN caused by a homozygous c.1124G>A variant in the VRK1 gene. Clinical and electrophysiological data of the proband were collected. Whole-exome sequencing (WES) and validation by Sanger sequencing were performed to identify the variant site, and pathogenicity interpretation was conducted in accordance with American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines. The proband was a 24-year-old male who presented with 2 years of progressive weakness and atrophy of the distal lower limbs, accompanied by slender upper limbs and no sensory disturbance. Electrophysiological examination showed decreased compound muscle action potential (CMAP) amplitude in motor nerves of both upper and lower limbs, indicating peripheral neurogenic damage, while sensory nerve conduction was normal. Genetic testing detected a homozygous c.1124G>A (p.Trp375Ter) variant in the VRK1 gene. His parents and elder sisters were heterozygous carriers, and the pedigree conformed to autosomal recessive inheritance. According to ACMG guidelines, this variant was classified as pathogenic (evidence: PVS1, PM2, PP1). The homozygous VRK1 c.1124G>A variant causes adult-onset dHMN, rather than pontocerebellar hypoplasia type 1A (PCH1A) as annotated in some genetic databases. This pedigree presents distinctive phenotypes, including slender upper limbs and diffusely decreased CMAP amplitudes in both upper and lower limbs, thereby expanding the clinical and genetic spectrum of VRK1-related dHMN in the Chinese population.

Introduction

Distal hereditary motor neuropathy (dHMN) constitutes a genetically heterogeneous group of inherited peripheral nervous system disorders, defined pathologically by length-dependent degeneration of lower motor neurons and their axons. Clinically, patients present with progressive weakness and atrophy of the distal limb muscles, while sensory function remains largely intact in most cases¹. Initial manifestations typically affect the distal lower limbs, including toe flexion weakness and atrophy of the gastrocnemius and intrinsic foot muscles, with gradual progression to involve the interosseous muscles of the hand and thenar/hypothenar muscles as the disease advances². Inheritance patterns vary across subtypes, with both autosomal dominant and autosomal recessive forms described. To date, over 50 pathogenic genes have been linked to dHMN, yet the underlying genetic cause remains unidentified in more than half of affected individuals³.

Given the marked genetic heterogeneity and extensive phenotypic overlap with other neuromuscular conditions, diagnosis of dHMN based solely on clinical evaluation is often unreliable⁴. Conventional stepwise diagnostic approaches, which proceed from clinical phenotyping to candidate gene sequencing, are inefficient and suffer from low diagnostic yield, particularly in sporadic or phenotypically atypical cases^1,2. Targeted gene panels offer a more cost-efficient alternative but are constrained by predefined gene lists and may miss rare or novel pathogenic variants. Whole-genome sequencing offers the most comprehensive genomic coverage but is limited by high cost, prolonged analytical turnaround, and high volumes of variants of uncertain significance in routine clinical settings⁵. For the present study, we employed an integrated diagnostic framework combining systematic clinical assessment, neurophysiological examination, whole-exome sequencing (WES), and familial segregation analysis. Neurophysiological testing helps confirm pure motor peripheral nerve involvement and delineate the distribution of nerve injury, thereby narrowing the differential diagnosis. WES allows simultaneous interrogation of all known dHMN-associated genes, detection of potential novel pathogenic variants, and subsequent familial segregation analysis, thereby further filtering out benign incidental variants and strengthening the evidence for the pathogenicity of candidate variants, striking a pragmatic balance among diagnostic yield, cost, and interpretability.

The vaccinia-related kinase 1 (VRK1) gene encodes a serine/threonine kinase that participates in nuclear signal transduction and DNA damage repair pathways, and its physiological function is essential for maintaining neuronal homeostasis and viability⁶. Pathogenic variants in VRK1 give rise to a broad clinical spectrum: infantile-onset pontocerebellar hypoplasia type 1A (PCH1A), characterized by microcephaly, severe hypotonia, and profound motor developmental delay; adult-onset dHMN with predominant distal limb involvement; and rarer presentations including hereditary spastic paraplegia (HSP) and amyotrophic lateral sclerosis (ALS)-like phenotypes^7,8,9. Specifically, the homozygous c.1124G>A (p.Trp375Ter) nonsense variant in VRK1 has been documented in multiple independent dHMN pedigrees with consistent autosomal recessive inheritance, supporting its established causal association with dHMN¹⁰. This variant introduces a premature termination codon in the C-terminal coding region of VRK1, and is predicted to cause loss of protein function either through production of a truncated protein or activation of nonsense-mediated mRNA decay. While the precise pathological mechanism has not been functionally validated for this specific variant, it is hypothesized that impaired VRK1 kinase activity may disrupt neuronal DNA repair and intracellular signaling cascades, thereby contributing to motor neuron degeneration and the development of dHMN.

In clinical practice, VRK1 genetic testing should be prioritized in the following scenarios: patients with clinically suspected dHMN manifesting length-dependent distal motor weakness and atrophy with preserved sensation, particularly those with autosomal recessive family history or adult disease onset; individuals from consanguineous families presenting with unexplained distal motor neuropathy; patients with overlapping central and peripheral nervous system manifestations not attributable to other known genetic disorders; and cases of dHMN or Charcot-Marie-Tooth disease type 2 (CMT2) that are negative for common pathogenic gene variants^11,12. Several important limitations of this diagnostic approach should be recognized. First, VRK1-related dHMN shares substantial phenotypic overlap with other neuromuscular disorders, including spinal muscular atrophy (SMA), CMT2, HSP, and early-stage ALS, potentially leading to diagnostic misclassification. Second, the interpretation of rare VRK1 variants remains challenging, as most lack functional validation and rely heavily on in silico prediction tools, resulting in uncertain pathogenicity classifications for a subset of variants. Third, as a single-pedigree study, the generalizability of our findings is limited, and genotype-phenotype correlations require further confirmation in larger patient cohorts.

Currently, reports of VRK1-associated dHMN in Chinese populations remain scarce, and the full phenotypic heterogeneity and clinical features of this condition are yet to be fully delineated. In this study, we report a Chinese consanguineous family with dHMN caused by a homozygous c.1124G>A variant in the VRK1 gene. By integrating clinical, electrophysiological, and genetic findings, we characterize the phenotypic and genotypic features of this pedigree, providing reference data to support accurate clinical diagnosis and genetic counseling for VRK1-related dHMN.

CASE PRESENTATION:

The proband (III₁) was a 24-year-old male from Bijie, Guizhou Province, born to first-cousin parents. The three-generation family pedigree is shown in Figure 1 and is consistent with an autosomal recessive inheritance pattern. II₁ and II₂ (the proband’s parents) were both heterozygous carriers of the VRK1 c.1124G>A variant and were asymptomatic. The proband (III₁) was homozygous for the variant and presented with clinical symptoms. III₂ and III₄ (the proband’s elder sisters) were heterozygous carriers with no clinical phenotype. III₃ (the proband’s third elder sister) was adopted in childhood and reported similar lower limb weakness and muscle atrophy, but genetic confirmation was not available.

Clinical manifestations

Chief complaint: Progressive weakness and atrophy of both lower limbs for 2 years.

Present illness history: Two years prior, the patient developed painless weakness of the distal lower limbs without obvious precipitating factors. He reported easy falling when walking on level ground, but was able to squat and stand unassisted. Gradual thinning of the legs below the knees was noted by the patient and his family. He reported poor exercise tolerance since childhood, with difficulty in running and jumping. There was no numbness, pain, or temperature disturbance in the limbs, no obvious upper limb weakness, and no bulbar symptoms. His intelligence was normal, and he worked independently as a hairdresser.

Neurological examination

Muscle strength (MRC scale): Hip flexion 5/5, knee extension 4+/5, ankle dorsiflexion 4/5, ankle plantarflexion 3+/5 bilaterally; upper limb proximal muscles 5/5, wrist extension 4+/5, finger flexion 4/5, intrinsic hand muscles 4/5 bilaterally.

Muscle tone: Normal in all four limbs; no increased or decreased tone.

Muscle bulk: Obvious atrophy of distal lower limb muscles (gastrocnemius, tibialis anterior, and intrinsic foot muscles) was observed (Figure 2A). The upper limbs appeared slender, with mild atrophy of the intrinsic hand muscles (Figure 2B).

Fasciculations: No visible fasciculations were observed in limb or trunk muscles.

Tendon reflexes: Upper limb reflexes were diminished; lower limb tendon reflexes (knee and ankle jerks) were absent bilaterally.

Pathological signs: Bilateral Babinski sign and Chaddock sign were negative.

Sensory examination: Superficial sensation (pain, touch, temperature) and deep sensation (vibration, position) were intact in all limbs.

Coordination: Finger-nose test and heel-knee-shin test were normal bilaterally.

Cranial nerves: Pupils equal and reactive to light; extraocular movements full; facial symmetry; normal palate elevation; normal tongue movement without atrophy or fasciculation; no dysphagia or dysarthria.

Gait: Steppage gait bilaterally.

Auxiliary laboratory examinations: Serum creatine kinase (CK), vitamin B₁₂ level, and thyroid function tests (TSH, FT3, FT4) were all within normal reference ranges.

Neuroimaging: Cranial and spinal magnetic resonance imaging (MRI) was not performed in this patient.

Differential diagnosis

PCH1A: Excluded. The patient had adult-onset disease, no microcephaly, hypotonia, cerebellar dysplasia, or developmental delay, which was inconsistent with the classic infantile PCH1A phenotype.

SMA: Excluded. The patient had adult-onset with purely distal involvement and normal proximal limb strength, without the typical proximal-predominant weakness pattern of SMA.

CMT2: Ruled out by neurophysiology. The patient had purely motor nerve involvement, with completely normal sensory nerve action potentials, and thus did not meet the diagnostic criteria for CMT2.

ALS: Excluded. There was no upper motor neuron involvement, no bulbar symptoms, and no fasciculations in cervical or bulbar-innervated muscles; the chronic progressive course with purely distal involvement was atypical for ALS.

HSP: Excluded. The patient had no spasticity, no hyperreflexia, and no pathological signs, with purely lower motor neuron involvement.

Kennedy disease (spinal and bulbar muscular atrophy, SBMA): Excluded. There was no bulbar involvement, no perioral fasciculations, and no endocrinological manifestations; the inheritance pattern was not consistent with X-linked transmission.

Diagnosis, Assessment, and Plan

Diagnosis

1. dHMN, confirmed by homozygous pathogenic variant c.1124G>A (p.Trp375Ter) in the VRK1 gene (autosomal recessive inheritance).

2. Pure motor axonal peripheral neuropathy, predominantly involving the distal lower limbs.

Assessment

The patient presented with slowly progressive distal motor weakness and muscle atrophy with preserved sensation, consistent with the classic adult-onset phenotype of VRK1-related dHMN.

Neurophysiological findings indicated diffuse motor axonal injury with normal sensory conduction, supporting a pure motor neuropathy.

Disease severity was mild to moderate at current presentation: the patient remained ambulatory and able to perform daily work independently, with no bulbar or respiratory involvement.

The pedigree showed autosomal recessive inheritance, with heterozygous carriers remaining asymptomatic.

Management and follow-up plan

1. Symptomatic management: Prescribe ankle-foot orthoses to improve gait stability and prevent fall injuries; refer to rehabilitation medicine for individualized strength training and stretching programs to maintain joint range of motion and prevent contractures.

2. Regular follow-up: Schedule annual neurological evaluations, including repeated nerve conduction studies, to monitor disease progression.

3. Genetic counseling: Provide detailed genetic counseling to the proband and family members regarding recurrence risk, reproductive planning options, and presymptomatic genetic testing for at-risk relatives.

4. Patient education: Advise avoidance of neurotoxic medications and excessive fatigue; provide psychological support to address disease-related anxiety.

Protocol

This study was approved by the Ethics Committee of Yunnan Provincial Hospital of Traditional Chinese Medicine (approval number: YYLW-2026-006). Written informed consent was obtained from the proband and all participating family members prior to enrollment.

1. Participant preparation and clinical assessment

1. The proband and available family members were enrolled after written informed consent was obtained.
2. Detailed demographic information, disease history, family history, and consanguinity information were collected using a standardized questionnaire.
3. A systematic neurological examination was performed, including assessments of muscle strength, muscle tone, tendon reflexes, sensory function, coordination, gait, and cranial nerve function.
4. The strength of all major muscle groups was documented using the Medical Research Council (MRC) scale.
5. Clinical photographs of the limbs were obtained after written consent was received from the participant.
   NOTE: All clinical evaluations were performed by a board-certified neurologist specializing in neuromuscular disorders to ensure diagnostic consistency.

2. Neurophysiological examination

1. Patient preparation
   1. The participant was instructed to avoid caffeine and strenuous exercise for at least 2 h before the examination.
   2. The examination room temperature was maintained at 24–26ºC, and the limb skin temperature was kept above 32ºC using a heating lamp when necessary.
   3. The participant was placed in a supine position in a quiet, dimly lit examination room.
2. Nerve conduction velocity study
   1. The skin at the electrode sites was cleaned with alcohol swabs to reduce impedance.
   2. Surface recording electrodes were placed over the motor point of the target muscle, and the reference electrode was positioned distally over the corresponding tendon.
   3. Stimulation electrodes were placed along the course of the nerve at standard anatomical landmarks.
   4. Motor nerve conduction studies were performed on the bilateral median, ulnar, tibial, and common peroneal nerves. Distal latency, compound muscle action potential (CMAP) amplitude, and motor conduction velocity were recorded.
   5. Sensory nerve conduction studies were performed on the bilateral median, ulnar, superficial peroneal, and sural nerves using orthodromic or antidromic techniques, as appropriate. Sensory nerve action potential (SNAP) amplitude and sensory conduction velocity were recorded.
   6. The filter bandpass was set to 20 Hz–10 kHz for motor studies and 20 Hz–2 kHz for sensory studies. Sweep speeds of 2 ms/division and 1 ms/division were used for motor and sensory studies, respectively.
3. Needle electromyography
   1. A concentric needle electrode was inserted into each target muscle under sterile conditions.
   2. Spontaneous activity at rest was assessed, including positive sharp waves, fibrillation potentials, and fasciculation potentials.
   3. Motor uni t potential (MUP) morphology was evaluated during mild voluntary contraction. Duration, amplitude, and polyphasia were measured.
   4. The recruitment pattern was assessed during maximum voluntary contraction.
   5. The tibialis anterior, medial gastrocnemius, abductor hallucis, first dorsal interosseous, deltoid, and rectus femoris muscles were examined bilaterally. The sternocleidomastoid and T10 paraspinal muscles were also examined.
      NOTE: All neurophysiological examinations were performed by an experienced electromyographer. Skin impedance was maintained below 5 kΩ to ensure recording quality. The reproducibility of all waveforms was confirmed across at least two consecutive stimulations.

3. Peripheral blood sample collection and genomic DNA extraction

1. Peripheral venous blood (3–5 mL) was collected from each participant in an ethylenediaminetetraacetic acid (EDTA) anticoagulant tube.
2. The blood samples were stored at 4ºC immediately after collection and processed within 24 h.
3. Genomic DNA was extracted from peripheral blood leukocytes using a genomic DNA extraction kit.
4. DNA concentration and purity were measured using a spectrophotometer.
5. DNA integrity was assessed using 1% agarose gel electrophoresis.
   NOTE: DNA samples with an A260/A280 ratio of 1.8–2.0 and a concentration of ≥50 ng/µL were accepted. Samples that failed to meet the quality-control criteria were re-extracted.

4. Whole-exome sequencing

1. Library preparation
   1. Genomic DNA (1 μg) was fragmented into 150–200 bp fragments by sonication.
   2. End repair, A-tailing, and adapter ligation were performed according to the library-preparation kit protocol.
   3. Exonic regions were enriched using a human whole-exome capture kit according to the manufacturer’s protocol.
   4. The captured library was amplified using polymerase chain reaction (PCR), and the resulting products were purified.
2. Sequencing
   1. Paired-end sequencing was performed on a high-throughput sequencing platform using a read length of 150 bp.
   2. A mean sequencing depth of ≥100 Χ across the target exome was targeted, with ≥98% of target bases covered at ≥10Χ depth and ≥95% covered at ≥20Χ depth.
   3. NOTE: A library concentration of ≥2 nM and an insert-size distribution of 250–350 bp were confirmed before sequencing. Q30 scores, GC content, and coverage depth were evaluated after sequencing to confirm data quality.

5. Bioinformatic analysis and variant filtering

1. Primary data analysis, including base calling, quality assessment, and adapter trimming, was performed.
2. Clean reads were aligned to the human reference genome GRCh37 (hg19) using sequence-alignment software.
3. Variant calling was performed to identify single-nucleotide variants (SNVs) and small insertions and deletions (indels).
4. Variants were annotated using public databases, including the Genome Aggregation Database (gnomAD), 1000 Genomes Project, ClinVar, and Online Mendelian Inheritance in Man (OMIM).
5. Variants were filtered and prioritized using the following criteria.
   1. Variants located in exonic or canonical splice-site regions were retained.
   2. Common variants with a minor allele frequency (MAF) of >0.01 in population databases were excluded.
   3. Nonsense, frameshift, splice-site, and missense variants predicted to be deleterious were prioritized.
   4. Candidate genes were compared with known neuromuscular disorder gene lists. Genes consistent with an autosomal recessive inheritance pattern were further prioritized.
   5. The VRK1 transcript NM_003384.3 was used as the reference sequence for variant nomenclature.

6. Sanger sequencing validation and familial segregation analysis

1. PCR primers flanking the candidate variant site were designed using primer-design software.
2. The target genomic region from each family member’s DNA was amplified using PCR.
3. The PCR products were purified, and bidirectional Sanger sequencing was performed.
4. Sequencing chromatograms were aligned with the reference sequence using sequence-alignment software.
5. The genotype of each family member was verified, and cosegregation of the variant with the disease phenotype in the pedigree was evaluated.
   NOTE: Clean, single-peak chromatograms without background noise were confirmed. PCR amplification and sequencing were repeated for samples that produced ambiguous results.

7. Variant pathogenicity classification and diagnostic confirmation

1. The verified variant was classified according to the 2015 standards and guidelines of the American College of Medical Genetics and Genomics (ACMG).
2. Clinical phenotypes, neurophysiological findings, genetic testing results, and familial segregation data were integrated to establish the final molecular diagnosis.
3. Genetic counseling regarding recurrence risk, reproductive options, and long-term prognosis was provided to the family.

Results

Genetic findings

WES was performed on the proband, with a mean target region depth of 112 ×; 98.7% of target bases were covered at ≥10 × depth, and 96.2% at ≥20 × depth. After variant filtering and prioritization, a homozygous nonsense variant c.1124G>A (p.Trp375Ter) in the VRK1 gene (transcript NM_003384.3, genome build GRCh37/hg19) was identified as the candidate pathogenic variant.

Sanger sequencing confirmed that the proband carried ...

Discussion

In this study, we applied an integrated diagnostic workflow combining clinical phenotyping, neurophysiological examination, WES, and familial segregation analysis to identify a homozygous VRK1 c.1124G>A pathogenic variant in a Chinese consanguineous family with adult-onset dHMN. This pedigree broadens the phenotypic spectrum of VRK1-related disorders and provides reference data for diagnosing dHMN in the Chinese population.

Methodological considerations for the diagnostic ...

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Reagents & Consumables
EDTA anticoagulant vacuum blood collection tube (5 mL)	BD Biosciences (Becton, Dickinson and Company)	367861
Genomic DNA Extraction Kit (spin-column format) for peripheral blood	QIAGEN; QIAamp DNA Blood Mini Kit	51106
Multiplex Ligation-dependent Probe Amplification (MLPA) Kit	MRC Holland	Custom probe set for target gene
PCR Amplification Kit (Taq polymerase, dNTPs, reaction buffer included)	Takara Bio; LA Taq PCR Kit	RR002A
Agarose powder (electrophoresis grade)	Biowest	111860
Sanger Sequencing Reaction Kit (BigDye Terminator v3.1)	Thermo Fisher Scientific; Applied Biosystems	4337455
Sequencing Product Purification Kit	Thermo Fisher Scientific; BigDye XTerminator Purification Kit	4376484
TAE electrophoresis buffer	Commercial grade / laboratory-prepared	—
PCR 8-strip tubes, centrifuge tubes and other routine consumables	Axygen	—
Instruments & Equipment
Capillary electrophoresis genetic analyzer (Sanger sequencing platform)	Applied Biosystems, Thermo Fisher Scientific	3500 Dx
Gradient thermal cycler (PCR instrument)	Bio-Rad Laboratories	C1000 Touch
Ultramicro spectrophotometer (for nucleic acid quantification)	NanoDrop One, Thermo Fisher Scientific	ND-ONE-W
Horizontal agarose gel electrophoresis system	Beijing Liuyi Biotechnology Co., Ltd.	DYY-6C
Gel documentation and analysis system	Bio-Rad Laboratories	GelDoc Go
Benchtop high-speed refrigerated centrifuge	Eppendorf	5424 R
Nerve conduction velocity / electromyography (NCV/EMG) system	Natus Medical	Keypoint 9033A07
Constant temperature metal bath / water bath	DLAB Scientific	—
Analysis Software
MLPA data analysis software (Coffalyser.Net)	MRC Holland	v140
Sanger sequencing peak analysis and sequence alignment software	Applied Biosystems, Thermo Fisher Scientific	Sequencing Analysis v6.0
Primer design software	Premier Biosoft	Primer Premier 6
Sequence alignment and variant interpretation tools	NCBI BLAST (online), ClinVar database	—