Correspondence

Antibodies to L-Type Calcium Channels in Amyotrophic Lateral Sclerosis

N Engl J Med 1993; 328:1355-1357May 6, 1993

Article

To the Editor:

I was rather confused by the article by Smith et al. (Dec. 10 issue)1 and the related editorial by Rowland2. The authors suggest that high-titer antibodies to human motor-neuron voltage-gated calcium channels (VGCCs) present in patients with amyotrophic lateral sclerosis (ALS) cross-react with rabbit L-type VGCC epitopes. Skeletal-muscle L-type channels show only 15 to 45 percent homology with extracellular domains of neuronal L-type channels, and thus high titers are more likely to suggest a reaction to a primary muscle antigen3.

The assay used for L-type VGCCs is novel in that it relies on a sandwich enzyme-linked immunosorbent assay for gamma-subunit-containing L-type channels. There is evidence to suggest that gamma-subunit expression is low in central nervous system VGCCs and brain tissue4,5. Most assays for the Lambert-Eaton myasthenic syndrome are radioimmunoassays with neuronal cell-line or small-cell carcinoma targets and detection of binding of N-type channels with radiolabeled omega-conotoxin6. Would not a [³H]dihydropyridine assay using an L-type channel-expressing neuronal cell line as a target have been more specific for neuronal L-type channels? Assays for N-type channels do not detect elevated titers of antibodies to VGCCs in ALS,7 but N-type channels and possibly P-type, L-type, and T-type channels are found on human motor neurons6.

The pattern described by the authors would seem to fit a peripheral autoimmune process better, with extraneuronal tissues as the antigenic targets. If there was a continuing autoimmune process involving central nervous system motor neurons, would not one expect to find high antibody titers in cerebrospinal fluid associated with peri-motor-neuron lymphocytic infiltrates, oligoclonal bands, or positive immunoblots against purified L-channel VGCCs? None of these findings have been described in ALS.

The study implies that patients with familial ALS serve as adequate controls, yet different disorders can have similar clinical presentations. The study does not describe, for instance, the levels of antibodies found in patients with poliomyelitis or muscle atrophy due to denervation, which would help to define whether the process described was primary or secondary. There are marked differences between myasthenia gravis and motor neuron disease, so a primary process is unlikely. Could not a secondary reaction to exposed, degenerating, denervated muscle calcium channels give rise to similar findings? Such a process would expose large amounts of immunologically sequestered antigens to attack and would correlate with the rate of disease progression and agree with the findings of the study.

A.S. Wierzbicki, M.A., B.M.B.Ch., D.Phil.
Westminster Hospital, London SW1P 2AR, United Kingdom

7 References

1. 1

   Smith RG, Hamilton S, Hofmann F, et al. Serum antibodies to L-type calcium channels in patients with amyotrophic lateral sclerosis. N Engl J Med 1992;327:1721-1728
   Full Text | Web of Science | Medline

2. 2

   Rowland LP. Amyotrophic lateral sclerosis and autoimmunity. N Engl J Med 1992;327:1752-1753
   Full Text | Web of Science | Medline

3. 3

   Miller RJ. Voltage-sensitive Ca²⁺ channels. J Biol Chem 1992;267:1403-1406
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4. 4

   Jay SD, Ellis SB, McCue AF, et al. Primary structure of the gamma subunit of the DHP-sensitive calcium channel from skeletal muscle. Science 1990;248:490-492
   CrossRef | Web of Science | Medline

5. 5

   Biel M, Hullin R, Freundner S, et al. Tissue-specific expression of high-voltage-activated dihydropyridine-sensitive L-type calcium channels. Eur J Biochem 1991;200:81-88
   CrossRef | Medline

6. 6

   Sher E, Biancardi E, Passafaro M, Clementi F. Physiopathology of neuronal voltage-operated calcium channels. FASEB J 1991;5:2677-2683
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7. 7

   Lennon VA, Lambert EH. Autoantibodies bind solubilized calcium channel-omega-conotoxin complexes from small cell lung carcinoma: a diagnostic aid for Lambert-Eaton myasthenic syndrome. Mayo Clin Proc 1989;64:1498-1504
   Web of Science | Medline

To the Editor:

Colorful hypotheses about ALS are becoming as abundant and beautiful as cactus flowers after the spring rains, but are seemingly as evanescent (including some of our own). The intriguing finding by Smith et al. that serum antibodies diluted ≥ 1:1250 reacted with the L-type VGCCs derived from rabbit skeletal muscle raises several questions.

Patients with ALS usually have serum creatine kinase levels two to five times the upper limit of normal, often with the percentage of the MB fraction elevated, and the muscle biopsy specimen usually contains occasional alkaline phosphatase-positive “regen-degen” fibers1. In the authors' assay, are serum samples positive in various active myopathies involving elevated levels of creatine kinase and its MB fraction and regen-degen fibers? Only three (possibly treated?) patients with polymyositis were mentioned.

Do the positive samples from patients with ALS bind morphologically to sections of human (or rabbit) normal or regen-degen skeletal-muscle-fiber plasmalemma, or to human muscle plasmalemmal fractions?

Regarding the abscissa in Figure 4 of the article, what is the formula for calculating the interesting scale of values from 1 to 6?

Although I, too, have analogized dysimmune diabetes mellitus to explain why typical ALS could be basically dysimmune but not improved by immunosuppression,2 Smith et al.'s proposed mechanism and mine are different. They suggest an antibody attack directly on motor neurons, possibly from IgG within the cell, and “once the destructive processes have been initiated, the loss of remaining viable motor neurons may be independent of the initial antibody attack.” This would seem to require a conceptually new, non-antibody-destructive process continuously spreading from neuron to neuron, because lower motor neurons can repair themselves if a damaging mechanism is removed: we and others have seen in rare cases that immunosuppressive treatment -- e.g., with prednisone3 or intravenous immune globulin4 -- reverses ALS-like damage to lower motor neurons (associated with circulating abnormal polyclonal or monoclonal antibody), resulting in a remarkable improvement in strength. My hypothesis of dysimmune ALS2 posited, as the target of circulating antibody, a function of intermediate “cells X” (analogous to pancreatic beta cells), which then either fail to produce a “factor Y” (analogous to insulin) needed by motor neurons or fail to detoxify a substance detrimental to motor neurons. Irreversible immune destruction of the cells X before or very early in ALS would set a course of neuronal deterioration unalterable by immunosuppression. Although the hypothetical cells X are unknown, they have been proposed to be hepatocytes, because in them we have found steatosis and excessive mitochondrial paracrystalline inclusions in patients with ALS2.

W. King Engel, M.D.
University of Southern California School of Medicine, Los Angeles, CA 90017

4 References

1. 1

   Koller M, Engel WK. Increased serum creatine kinase MB isozyme (CK-MB) and alkaline phosphatase positive (AP+) regenerative muscle fibers in amyotrophic lateral sclerosis (ALS). Neurology 1984;34:Suppl:81-81 abstract.
   

2. 2

   Engel WK. RNA metabolism in relation to amyotrophic lateral sclerosis. Adv Neurol 1991;56:125-153
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3. 3

   Rai B, Engel WK. Rapidly progressing ALS-like fasciculating lower motor neuron disorder without dysimmune laboratory features, markedly benefitted following plasmapheresis plus prednisone. Muscle Nerve 1986;9:Suppl:106-106 abstract.
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4. 4

   Engel WK, Hanna CJ. Intravenous immunoglobulin: excellent benefit in otherwise refractory progressive muscular atrophy with IgM monoclonal gammopathy. Ann Neurol 1992;32:279-279 abstract.
   Web of Science

Author/Editor Response

The authors reply:

To the Editor: We appreciate the concern expressed by Drs. Wierzbicki and Engel regarding muscle denervation and atrophy as a possible cause of secondary production of antibodies to VGCCs in ALS. In reply to Dr. Engel: the titers of these antibodies do not correlate with the serum creatine kinase levels in the patients with ALS, nor are antibody titers elevated in the majority of subsequently tested patients with active polymyositis. Unfortunately, high background binding by human serum to skeletal-muscle plasmalemma has previously precluded meaningful immunohistochemical correlations. In reply to Dr. Wierzbicki: there are also low or absent anti-VGCC antibody titers in patients with longstanding denervation due to spinal-muscle atrophy, advanced ALS with slowly progressive disease, post-polio syndrome, or familial ALS, in which the extent of denervation is as pronounced as in sporadic ALS. These data, together with previous data on experiments involving passive transfer,1 still suggest a primary role for antibodies to VGCC in ALS.

Although we agree that caution is needed in extrapolating the effects of IgG in ALS from peripheral to central calcium channels, there is sufficient homology between portions of the α₁-subunits of skeletal-muscle L-type VGCCs and several neuronal VGCCs for immunologic cross-reactivity. Our previous demonstration of increased release of acetylcholine after passive transfer with IgGs of ALS1 would suggest that the relevant channel is more likely N-type or P-type. While the reported lack of ALS IgG reactivity with N-type channels from neuronal or small-cell tumor lines, as compared with motor-neuron channels, may reflect differences in antigenicity, more sensitive assays being evaluated in our laboratories do, in fact, suggest reactivity of ALS IgG with neuronal high-threshold calcium channels. Worries about the lack of evidence of central nervous system autoimmunity should be allayed by the recent demonstration of activated microglia and CD4 and CD8 T lymphocytes within the ventral horns of the spinal cord in ALS2,3.

No conceptually new mechanism of neuronal cell death need be implicated in ALS, since antibodies interacting with motor-neuron calcium channels could alter intracellular calcium homeostasis and lead to the death of the cell even after the antibody had departed. Furthermore, the same ALS IgGs that increase acetylcholine release could also activate microglia to release toxic agents or enhance glutamate release within the spinal cord. The common denominator may be altered intracellular calcium, explaining how autoimmune mechanisms could directly lead to destruction of motor neurons or could initiate excitotoxic cell death associated with decreased spinal cord glutamate levels4 and glutamate transport5.

R. Glenn Smith, M.D., Ph.D.
Enrico Stefani, M.D., Ph.D.
Stanley H. Appel, M.D.
Baylor College of Medicine, Houston, TX 77030

5 References

1. 1

   Appel SH, Engelhardt JI, Garcia J, Stefani E. Immunoglobulins from animal models of motor neuron disease and from human amyotrophic lateral sclerosis patients passively transfer physiological abnormalities to the neuromuscular junction. Proc Natl Acad Sci U S A 1991;88:647-651
   CrossRef | Web of Science | Medline

2. 2

   Kawamata T, Akiyama H, Yamada T, McGeer PL. Immunologic reactions in amyotrophic lateral sclerosis brain and spinal cord tissue. Am J Pathol 1992;140:691-707
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3. 3

   Engelhardt JI, Tajti J, Appel SH. Lymphocytic infiltrates in the spinal cord in amyotrophic lateral sclerosis. Arch Neurol 1993;50:30-36
   Web of Science | Medline

4. 4

   Plaitakis A, Constantakakis E, Smith J. The neuroexcitotoxic amino acids glutamate and aspartate are altered in the spinal cord and brain in amyotrophic lateral sclerosis. Ann Neurol 1988;24:446-449
   CrossRef | Web of Science | Medline

5. 5

   Rothstein JD, Martin LJ, Kuncl RW. Decreased glutamate transport by the brain and spinal cord in amyotrophic lateral sclerosis. N Engl J Med 1992;326:1464-1468
   Full Text | Web of Science | Medline

Citing Articles (2)

Citing Articles

1. 1

   Andrew Eisen. (1995) Amyotrophic lateral sclerosis is a multifactorial disease. Muscle & Nerve 18:7, 741-752
   CrossRef

2. 2

   Michael A. Meyer, Nicholas T. Potter. (1995) Issues & opinion. Sporadic ALS and chromosome 22: Evidence for a possible neurofilament gene defect. Muscle & Nerve 18:5, 536-539
   CrossRef