Fine specificity of anti-GQ1b IgG and clinical features

Journal of the Neurological Sciences 185 Ž2001. 5–9 www.elsevier.comrlocaterjns

Fine specificity of anti-GQ1b IgG and clinical features Keiichiro Susuki a,b, Nobuhiro Yuki a,) , Koichi Hirata a a

Department of Neurology, Dokkyo UniÕersity School of Medicine, Kitakobayashi 880, Mibu, Shimotsuga, Tochigi 321-0293, Japan b Department of Neurology, Yokohama City UniÕersity, Yokohama, Japan Received 26 September 2000; accepted 15 November 2000

Abstract Anti-GQ1b IgG frequently is present in sera of patients with Miller Fisher syndrome ŽMFS., Guillain-Barre´ syndrome ŽGBS. with ophthalmoplegia, Bickerstaff’s brainstem encephalitis ŽBBE., and acute ophthalmoparesis ŽAO. in the acute phase. Why various clinical signs develop under these conditions, however, has yet to be clarified. We investigated the fine specificity of anti-GQ1b IgG and its clinical correlation in sera from 82 patients: 56 with MFS, 11 with GBS, 13 with BBE, and 2 with AO. Anti-GQ1b IgG antibodies were absorbed by GT1a in 80 Ž98%. of the 82 sera, by GD1b in 11 Ž13%., and by the other b-series gangliosides GD3, GD2, or GT1b in 24 Ž29%.. The most frequent pattern of fine specificity was the cross-reaction with GT1a alone, seen in 56 Ž68%. samples. Of the 11 patients with anti-GQ1b IgG, cross-reacting with GD1b, 6 Ž55%. had impaired deep sense, and the association was significant Ž p s 0.02.. This is the first study to show that the fine specificity of anti-GQ1b IgG is heterogeneous and that the difference is correlated with the presence of a particular clinical sign. q 2001 Published by Elsevier Science B.V. Keywords: Anti-GQ1b IgG; Fine specificity; GD1b; Deep sensation

1. Introduction Anti-GQ1b IgG frequently is present in sera of patients with Miller Fisher syndrome ŽMFS., Guillain-Barre´ syndrome ŽGBS. with ophthalmoplegia w1x, Bickerstaff’s brainstem encephalitis ŽBBE. w2x, and acute ophthalmoparesis ŽAO. w3x in the acute phase. GQ1b ganglioside is so abundantly expressed in the oculomotor nerves w1x that the anti-GQ1b antibody may function in the development of ophthalmoparesis in these neurological disorders. Why various signs develop under these conditions, however, has yet to be clarified. Differences in the fine specificity of anti-ganglioside antibodies may account for the variety of clinical signs. Some patients with chronic sensory ataxic neuropathy have monoclonal IgM antibody against GD1b, as well as against GD3, GT1b, and GQ1b that have the wNeuAc a 2–8 NeuAc a 2–3 Gal b x structure in common Žreviewed in Ref. w4x.. In contrast, some patients with motor neuron disease-like disorder have IgM paraprotein to GD1b, GM1, and asialo-GM1, which reacts with the wGal b 1–3 GalNAcx

epitope common to these glycolipids w5x. Although the anti-GD1b IgG, which does not cross-react with GM1 is important in the pathogenesis of experimental sensory ataxic neuropathy, the antibody that cross-reacts with GM1 is not w6x. We hypothesized that the difference in the fine specificity of anti-GQ1b IgG produces the various signs of MFS, GBS, BBE, and AO. There are few reports on the fine specificity of anti-GQ1b IgG or on its clinical correlation. Anti-GQ1b IgG that cross-reacts with GT1a is associated with ophthalmoplegia in MFS and GBS w1x. In contrast, anti-GT1a IgG, which does not cross-react with GQ1b, has been detected in the pharyngeal-cervicalbrachial variant of GBS w7x. Although Carpo et al. w8x reported reactivity of anti-GQ1b-positive sera with other gangliosides, because of the lack of an absorption study they did not show its fine specificity. We therefore investigated the fine specificity of anti-GQ1b IgG and its correlation with particular clinical signs. 2. Materials and methods 2.1. Patients

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Corresponding author. Tel.: q81-282-86-1111 ext. 2578; fax: q81282-86-5884. E-mail address: [email protected] ŽN. Yuki..

We studied 82 patients who had anti-GQ1b IgG titers of 500 or more. Serum samples obtained during the acute

0022-510Xr01r$ - see front matter q 2001 Published by Elsevier Science B.V. PII: S 0 0 2 2 - 5 1 0 X Ž 0 1 . 0 0 4 6 4 - 6

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K. Susuki et al.r Journal of the Neurological Sciences 185 (2001) 5–9

Table 1 Diagnostic criteria for Miller Fisher syndrome, Bickerstaff’s brainstem encephalitis, and acute ophthalmoparesis I. Features required for diagnosis Miller Fisher syndrome Ž1. Progressive, relatively symmetric ophthalmoplegia and ataxia by 4 weeks Ž2. Hypo- or areflexia Ž3. Limb strength with 5 or 4 on the Medical Research Council Scale Bickerstaff’s brainstem encephalitis Ž1. Progressive, relatively symmetric ophthalmoplegia and ataxia by 4 weeks Ž2. Either consciousness disturbance Žcoma, semicoma, or stupor. or pyramidal signs Žhyperreflexia or pathological reflexes. Ž3. Limb strength with 5 or 4 on the Medical Research Council Scale Acute ophthalmoparesis Ž1. Progressive, relatively symmetric ophthalmoplegia by 4 weeks Ž2. Neither ataxia nor limb weakness II. Features strongly supportiÕe of the diagnosis for each condition Ž1. A history of infectious symptoms within 4 weeks before the onset of neurological symptoms Ž2. CSF albumino-cytological dissociation Ž3. Presence of anti-GQ1b IgG antibody III. Differential diagnosis Wernicke’s encephalopathy, vascular disease involving the brainstem, multiple sclerosis, neuro-Behc¸et’s disease, botulism, myasthenia gravis, brainstem tumor, pituitary apoplexy, vasculitis, and lymphoma IV. Appendix Ž1. Patients showing limb weakness Ž3 or less on the Medical Research Council Scale., in addition to ophthalmoplegia and ataxia, were diagnosed as having overlapping Miller Fisher syndrome and Guillain-Barre´ syndrome Ž2. Patients showing limb weakness, in addition to consciousness disturbance Žcoma, semicoma, or stupor. or pyramidal signs as well as ophthalmoplegia, were diagnosed as having overlapping Bickerstaff’s brainstem encephalitis and Guillain-Barre´ syndrome

phase of illness were assayed for anti-ganglioside antibodies at our neuroimmunological laboratory between August 1995 and June 1998. The clinical records of all the patients were reviewed in order to confirm the diagnoses. Diagnosis of GBS was based on the established clinical criteria w9x. Diagnoses of MFS, BBE, and AO were based on the criteria proposed by Odaka et al. w10x ŽTable 1.. 2.2. Absorption study The anti-GQ1b IgG absorption study was done as described elsewhere w11x using microtiter plates coated with 5-pmol portions of each ganglioside. b-Series gangliosides ŽGQ1b, GD3, GD2, GD1b, or GT1b. and a-series gangliosides ŽGM2, GM1, GD1a, or GT1a. were used. Purified GQ1b and GD3 were purchased from Dia-Iatron ŽTokyo, Japan.. GD2 and GM2 were purchased, respectively from Sigma ŽSt. Louis, MO, USA. and Funakoshi ŽTokyo, Japan.. GD1b, GT1b, GM1, and GD1a were gifts from Dr. Yasuo Suzuki ŽShizuoka University, Shizuoka, Japan.. GT1a was a gift from Dr. Toshio Ariga ŽTsukuba Research Laboratories, Eisai, Ibaraki, Japan.. The samples were added to the wells at the dilution, which gave an optical density between 0.5 and 2.5. After incubation at 48C overnight, the samples were used as the primary antibodies in the standard enzyme-linked immunosorbent assay w3x. Absorption rates were expressed as percentages of the optical densities obtained with and without absorption.

2.3. Correlation between clinical signs and the fine specificity of anti-GQ1b IgG antibodies We investigated the correlation between deep sense impairment and cross-reactivity with GD1b, which is recognized as a putative target molecule for serum antibodies in sensory ataxic neuropathy. We also investigated the correlation between limb weakness and cross-reactivity with GM1 or GD1a, which both are associated with acute motor axonal neuropathy. Their differences were examined with Fisher’s exact test.

3. Results 3.1. Clinical features The patient population was comprised of 56 with MFS, 11 with GBS, 13 with BBE, and 2 with AO. Acute paresis of the extraocular muscles was present in 81 Ž99%. of the 82 patients who had anti-GQ1b IgG ŽTable 2.. Ataxia was found in 72 Ž88%. patients. Deep tendon reflexes were absent or decreased in 74 Ž90%. patients. Thirty-three Ž40%. patients had limb weakness. Five patients with MFS and six with BBE, showing remarkable limb weakness later Ž3 or less on Medical Research Council Scale., were considered as overlap between MFS or BBE and GBS. A total of 9 Ž11%. patients had disturbance of superficial

K. Susuki et al.r Journal of the Neurological Sciences 185 (2001) 5–9 Table 2 Fine specificity of anti-GQ1b IgG antibodies and clinical signs Number

Absorbed by GT1a GD1b GD3rGD2rGT1b GM2rGM1rGD1a

Total

56

14

5

4

1

q

q

q q q

q q q q

q q

2 2

1

1

1

7

Table 3 Association between impaired deep sense and the anti-GQ1b IgG crossreacting with GD1b Deep sensation

82

Disturbed

q

Diagnosis MFS GBS BBE AO

43 3 9 1

7 3 3 1

3 2

Clinical signs Ophthalmoplegia Ataxia Areflexia or hyporeflexia Limb weakness Impaired superficial sense Impaired deep sense Disturbed consciousness Pathological reflex

56 53 53 20 8 12 6 5

14 10 11 5 1 2 3 2

5 4 4 4

4 4 4 2

1 1 1

4 1

1 1

1

80 Ž98%. 11 Ž13%. 24 Ž29%. 4 Ž5%.

q q

Anti-GQ1b IgG cross-reacting with GD1b Anti-GQ1b IgG not cross-reacting with GD1b

Intact

6

5

14

57

ps 0.02, odds ratios 4.9, confidence interval at 95% levels1.4–16.9. 1 1

1 1 1

1

1

56 11 13 2

81 Ž99%. 72 Ž88%. 74 Ž90%. 33 Ž40%. 9 Ž11%. 20 Ž24%. 12 Ž15%. 7 Ž9%.

sensation and 20 Ž24%. had disturbance of vibratory andror position sense. 3.2. Fine specificity of anti-GQ1b IgG antibodies Results of the absorption study of anti-GQ1b IgG antibodies by different gangliosides are shown in Table 2. Anti-GQ1b IgG antibody reacting with GQ1b alone was seen in only one patient with GBS. The antibodies from 80 Ž98%. of the 82 samples were absorbed by GT1a as well as by GQ1b. The most frequent pattern of fine specificity was the cross-reaction with GT1a alone, seen in 56 Ž68%.

samples. The antibodies were absorbed by the GD1b in samples from 11 Ž13%. patients; 6 with MFS, 4 with GBS, 1 with BBE, and in 24 Ž29%. by the other b-series gangliosides GD3, GD2, or GT1b. In addition, they were absorbed by GD1a in two patients and by GM2, GM1, or GD1a in two patients. Absorption patterns of anti-GQ1b IgG antibodies from representative patients are shown in Fig. 1. A few antibodies that do not cross-react with GQ1b were seen; anti-GD3 IgG in two patients, anti-GM1 IgG in one patient, and anti-GD1a IgG in one patient. 3.3. Correlation between clinical signs and the fine specificity of anti-GQ1b IgG antibodies The frequency of impaired deep sense in patients with anti-GQ1b IgG antibodies that cross-reacted with GD1b Ž6r11, 55%, 5 with MFS and 1 with GBS. was significantly higher Ž p s 0.02. than in the patients without these antibodies Ž14r71, 7%. ŽTable 3.. The frequency of limb weakness in patients with anti-GQ1b IgG antibodies that cross-reacted with GM1 andror GD1a Ž2r4, 50%. appeared to be higher than that in the other patients Ž20r78, 26%.. The difference did not, however, reach statistical significance.

Fig. 1. Absorption study results for three representative patients. Anti-GQ1b IgG was absorbed by GQ1b and GT1a in Patient 1 with Miller Fisher syndrome who had neither impaired deep sense nor severe limb weakness. Anti-GQ1b IgG was absorbed by GD1b as well as by GQ1b and GT1a in Patient 2 with Miller Fisher syndrome who had obvious disturbance of vibratory sense. Anti-GQ1b IgG was absorbed by GQ1b, GT1a, b-series gangliosides, except GT1b, and a-series gangliosides in Patient 3 with Guillain-Barre´ syndrome who had tetraplegia.

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K. Susuki et al.r Journal of the Neurological Sciences 185 (2001) 5–9

4. Discussion Except for one patient, all with anti-GQ1b IgG had paresis of the extraocular muscles in the acute phase of illness. This is further evidence that anti-GQ1b IgG is closely associated with external ophthalmoparesis w1x. Ataxia was found in 72 Ž88%. of the patients studied. The presence of anti-GQ1b IgG also is associated with ataxia w12,13x. The lesion responsible for ataxia in MFS has yet to be determined. IgG antibodies in sera from MFS patients with anti-GQ1b activity selectively bind to the molecular layer of the human cerebellum w14x. Some large neurons of human dorsal root ganglia, however, are immunostained by murine anti-GQ1b monoclonal antibody w13x. Postural body sway analysis findings suggest the selective involvement of the group 1a afferent system in 10 patients with MFS w15x. Ours is the first study to show that the fine specificity of anti-GQ1b IgG antibodies is heterogeneous. Similar to the findings of Chiba et al. w1x, 98% of the samples with anti-GQ1b IgG antibodies were absorbed by GT1a, which has a disialosyl residue linked to the external galactose common to GQ1b ŽFig. 2.. Anti-GQ1b IgG antibodies also cross-reacted with GD1b and other b-series gangliosides, which have a common disialosyl residue linked to the internal galactose ŽFig. 2.. These structures common to the gangliosides appear to be the binding sites of individual antibodies. Why anti-GQ1b IgG antibodies cross-react with the structurally different gangliosides GM2, GM1, or GD1a in two dimensions is not clear. These gangliosides may have a common unknown three-dimensional epitope, as suggested by the presence of molecular mimicry between GalNAc-GD1a and GM1b that differ structurally in two dimensions w16x. In some patients, anti-GQ1b IgG antibodies cross-reacted with b-series gangliosides as well as with GT1a, and those antibodies, which were absorbed by GM1 or GD1a, reacted with GT1a as well as b-series gangliosides ŽTable 2, Fig. 1.. These findings suggest the coexistence in individual patients of several anti-GQ1b IgG antibodies with different patterns of fine specificity. The presence of anti-GQ1b IgG, which cross-reacted with GD1b, was significantly associated with the disturbance of deep sensation. An immunohistochemical study has showed localization of GD1b in the neurons of human dorsal root ganglia w17x. These neurons were immunostained by serum IgM from a patient with sensory ataxic neuropathy who had IgM against GD1b, GD3, GT1b, or GQ1b w18x. Human IgM with anti-GD1b activity effectively killed rat dorsal root ganglion neurons in vitro w19x. Sensory ataxic neuropathy with loss of primary sensory neurons was induced in rabbits by sensitization with GD1b w20x. GD1b therefore is recognized as a putative target molecule for serum antibodies in sensory ataxic neuropathy. Our findings suggest that the cross-reactivity of antiGQ1b IgG with GD1b is involved in the development of impaired deep sense in MFS and GBS. Two of the four

Fig. 2. Structures of gangliosides GQ1b, GT1a, GD1b, and GD1a. GQ1b and GT1a have a common disialosyl residue linked to the external galactose Žsolid line ellipse.. GQ1b and GD1b have a disialosyl residue linked to the internal galactose Žbroken line ellipse. in common. GD1a differs structurally from GQ1b in two dimensions.

patients who had anti-GQ1b IgG antibodies which crossreact with GM1 or GD1a, GBS having been diagnosed for both, had remarkable limb weakness. Both anti-GM1 IgG w21x and anti-GD1a IgG w22x are associated with acute motor axonal neuropathy. The difference did not reach statistical significance in this study, possibly because of the small sample size. Our findings support the hypothesis that differences in the fine specificity of anti-GQ1b IgG antibodies have a role in the development of the variety of clinical signs seen in MFS, GBS, BBE, and AO.

Acknowledgements We are grateful to Drs. Y. Tagawa and M. Koga for the anti-GQ1b IgG routine assay, Dr. M. Odaka for summarizing the clinical records and making the diagnoses, and Ms. Y. Tsuchiya and Y. Ueno for their technical assistance. This research was supported in part by grants for Neuroimmunological Diseases from the Ministry of Health and Welfare of Japan and from the Ministry of Education, Science, Sports, and Culture of Japan Ž10780482, 10557063..

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