Correspondence

Phenotype in an Infant with SOD1 Homozygous Truncating Mutation

To the Editor:

Superoxide dismutase-1 (SOD1) is a ubiquitously expressed antioxidant enzyme. Mutations in SOD1 are associated with dominantly inherited amyotrophic lateral sclerosis, probably caused by the toxic effects of the protein rather than by loss of enzymatic activity.1,2 Here we report the clinical features of a girl with a homozygous truncating mutation and an absence of SOD1 activity.

The patient, who was 2 years 9 months of age at the time of this review, was born to consanguineous parents after an uneventful pregnancy. Axial hypotonia and loss of gross and fine motor function began at 6 months of age, after which severe, progressive spastic tetraparesis developed and Babinski’s sign was present in both feet. Atrophy, fasciculations, and other signs of lower motor neuron involvement were not noted. The results of motor- and sensory-nerve conduction studies performed when the patient was 9 months of age were normal. A needle electromyographic examination revealed a few fasciculations and myokymia but no other definite signs of lower motor neuron involvement. Cognitive development was age-appropriate. The sole abnormality noted on magnetic resonance imaging of the brain was mild frontoparietal atrophy. The serum lactate level was elevated on several occasions (maximum, 6.9 mmol per liter; normal range, 0.5 to 2.2) and normal on other occasions. The lactate level in the cerebrospinal fluid was normal, as were the results of all other laboratory analyses, including the serum activity of creatine kinase. A sibling with similar symptoms had died at the age of 6 years. No autopsy was performed.

Outline of the p.Cys112Trpfs*11 Protein, Expression of the Mutant Protein, and Oxygen Sensitivity of Patient Cells.

Panel A shows the alignment of a wild-type (WT) amino acid sequence and, from the patient in this report, the mutant superoxide dismutase-1 (SOD1) amino acid sequence. Mutant SOD1 was detected in extracts of fibroblasts obtained from the patient. Fibroblasts were grown in the presence (+) or absence (–) of the proteasome inhibitor bortezomib. Antibodies specific for peptides in the N-terminal part and the neopeptide sequence (in red) were used to detect the mutant SOD1 protein in Western blots. There was no reaction with an antibody against the C-terminal, indicating that the mutant protein (obtained from samples derived from two distinct biopsy specimens of the skin) was truncated. The mutant protein was detected in all four fibroblast colonies examined. HOM denotes the fibroblast extracts from the homozygous patient. The WT extract was obtained from an SOD1 fibroblast colony from a human control. Red arrowheads denote the position of the mutant protein, and black arrowheads the position of the larger, wild-type SOD1 protein. Panel B (scale bar, 10 mm) shows the results of culture of the biopsy specimens of the patient’s skin at 6 weeks, during which 2% oxygen and antioxidant protection were provided. The patient’s fibroblasts grew only as thin halos around the skin fragments. Panel C (scale bar, 50 μm), which is an enlargement of the periphery of the patient’s fibroblast colonies shown in Panel B, indicates that the patient’s fibroblasts were dysmorphic and that cell debris, indicative of cell death, was abundant. Panel D (scale bar, 50 μm) shows fibroblasts from the heterozygous father that were grown under conditions that were identical to those for the colonies illustrated in Panels B and C. The father’s fibroblast colonies show normal morphology and proliferation.

Trio-based whole-exome sequencing revealed homozygosity for a nucleotide duplication (c.335dupG) in SOD1, which was predicted to result in a defective protein, with truncation of 43 amino acids and the addition of a neopeptide (p.Cys112Trpfs*11) (Figure 1A) that consisted of 10 amino acids. The patient’s erythrocytes lacked SOD1 activity, and the level of activity in the healthy, heterozygous parents was half that of the normal level. Biopsy specimens of the skin were obtained. Fibroblasts from the girl did not grow when the specimens were cultured in ordinary conditions, in which the oxygen saturation of the culture medium is approximately 19%, but they did grow slowly when cultured with 2% oxygen, with a stabilized derivative of the alternative superoxide scavenger ascorbate in the medium (Figure 1B and 1C). The mutant 13-kD SOD1 protein was detected in the cultured fibroblasts obtained from the patient and both parents (Figure 1A; and Fig. S1 in the Supplementary Appendix, available with the full text of this letter at NEJM.org).

The complete absence of SOD1 activity conferred the patient’s cells in culture with extreme oxygen sensitivity. Her phenotype was remarkable for the predominant impairment of upper motor neurons, whereas other organ systems were unaffected. The mutant p.Cys112Trpfs*11 SOD1 protein was expressed and may have contributed to the motor disorder. However, the symptoms in this patient, with onset during infancy, were more severe than previously observed in patients homozygous for full-length SOD1 mutations who may have had some preserved SOD activity,2,3 which suggests that the total loss of SOD1 enzymatic activity was contributing to motor-neuron dysfunction. To the extent that events in humans can be compared with those in mice, the motor disturbances in this patient were more severe than those in SOD1 knockout mice,4,5 perhaps because in mice, in contrast with humans, the superoxide scavenger ascorbate is synthesized and can be replenished. Although we do not prove that there is a causal relationship between the loss of SOD activity from a truncating mutation and a severe, progressive phenotype characterized by loss of motor function, the present findings suggest that in clinical trials involving patients with amyotrophic lateral sclerosis, caution might be exercised regarding the use of gene therapies that may markedly depresses SOD1 activity.

Peter M. Andersen, M.D., Ph.D.
Ulrika Nordström, Ph.D.
Umeå University, Umeå, Sweden

Konstantinos Tsiakas, M.D.
Jessika Johannsen, M.D.
Alexander E. Volk, M.D.
Tatjana Bierhals, M.D.
University Medical Center Eppendorf, Hamburg, Germany

Per Zetterström, M.D., Ph.D.
Stefan L. Marklund, M.D., Ph.D.
Umeå University, Umeå, Sweden

Maja Hempel, M.D.
René Santer, M.D.
University Medical Center Eppendorf, Hamburg, Germany

Supported by the Swedish Brain Foundation, the Knut and Alice Wallenberg Foundation, and the Swedish Research Council.

Disclosure forms provided by the authors are available with the full text of this letter at NEJM.org.

This letter was published on July 17, 2019, at NEJM.org.

Drs. Andersen, Hempel, and Santer contributed equally to this letter.

  1. 1. Rosen DR, Siddique T, Patterson D, et al. Mutations in Cu/Zn superoxide dismutase gene are associated with familial amyotrophic lateral sclerosis. Nature 1993;362:59-62.

  2. 2. Andersen PM, Al-Chalabi A. Clinical genetics of amyotrophic lateral sclerosis: what do we really know? Nat Rev Neurol 2011;7:603-615.

  3. 3. Hayward C, Brock DJ, Minns RA, Swingler RJ. Homozygosity for Asn86Ser mutation in the CuZn-superoxide dismutase gene produces a severe clinical phenotype in a juvenile onset case of familial amyotrophic lateral sclerosis. J Med Genet 1998;35:174-174.

  4. 4. Reaume AG, Elliott JL, Hoffman EK, et al. Motor neurons in Cu/Zn superoxide dismutase-deficient mice develop normally but exhibit enhanced cell death after axonal injury. Nat Genet 1996;13:43-47.

  5. 5. Saccon RA, Bunton-Stasyshyn RKA, Fisher EMC, Fratta P. Is SOD1 loss of function involved in amyotrophic lateral sclerosis? Brain 2013;136:2342-2358.

Supplementary Material

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