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. 2022 Oct;269(10):5420-5430.
doi: 10.1007/s00415-022-11178-9. Epub 2022 May 27.

Rho GTPase-activating protein 10 (ARHGAP10/GRAF2) is a novel autoantibody target in patients with autoimmune encephalitis

Sven Jarius  1 , Lars Komorowski  2 , Jens U Regula  3   4 , Jürgen Haas  5 , Stefanie Brakopp  2 , Brigitte Wildemann  5
Affiliations

Rho GTPase-activating protein 10 (ARHGAP10/GRAF2) is a novel autoantibody target in patients with autoimmune encephalitis

Sven Jarius et al. J Neurol. 2022 Oct.

Abstract

Background: In 2010, we described a novel immunoglobulin G (IgG) autoantibody (termed anti-Ca after the index case) targeting Rho GTPase-activating protein 26 (ARHGAP26, also termed GTPase regulator associated with focal adhesion kinase [GRAF], or oligophrenin-like protein 1 [OPHN1L]) in autoimmune cerebellar ataxia (ACA). Later, ARHGAP26-IgG/anti-Ca was reported in patients with limbic encephalitis/cognitive decline or peripheral neuropathy. In several of the reported cases, the syndrome was associated with cancer. ARHGAP10/GRAF2, which is expressed throughout the central nervous system, shares significant sequence homology with ARHGAP26/GRAF. Mutations in the ARHGAP10 gene have been linked to cognitive and psychiatric symptoms and schizophrenia.

Objective: To assess whether ARHGAP26-IgG/anti-Ca co-reacts with ARHGAP10.

Methods: Serological testing for ARHGAP10/GRAF2 autoantibodies by recombinant cell-based assays and isotype and IgG subclass analyses.

Results: 26/31 serum samples (84%) from 9/12 (75%) ARHGAP26-IgG/anti-Ca-positive patients and 4/6 ARHGAP26-IgG/anti-Ca-positive CSF samples from four patients were positive also for ARHGAP10-IgG. ARHGAP10-IgG (termed anti-Ca2) remained detectable in the long-term (up to 109 months) and belonged mainly to the complement-activating IgG1 subclass. Median ARHGAP26-IgG/anti-Ca and median ARHGAP10-IgG/anti-Ca2 serum titres were 1:3200 and 1:1000, respectively, with extraordinarily high titres in some samples (ARHGAP26-IgG/anti-Ca: up to 1:1000,000; ARHGAP10-IgG: up to 1:32,000). ARHGAP26/anti-Ca serum titres exceeded those of ARHGAP10-IgG in all samples but one. A subset of patients was positive also for ARHGAP10-IgM and ARHGAP10-IgA. CSF/serum ratios and antibody index calculation suggested intrathecal production of ARHGAP26-IgG/anti-Ca and anti-ARHGAP10. Of 101 control samples, 100 were completely negative for ARHGAP10-IgG; a single control sample bound weakly (1:10) to the ARHGAP10-transfected cells.

Conclusions: We demonstrate that a substantial proportion of patients with ARHGAP26-IgG/anti-Ca-positive autoimmune encephalitis co-react with ARHGAP10. Further studies on the clinical and diagnostic implications of ARHGAP10-IgG/anti-Ca2 seropositivity in patients with autoimmune encephalitis are warranted.

Keywords: Anti-Ca; Anti-Ca2; Antibodies; Antigen; Autoantibodies; Autoimmune encephalitis; Cerebellar ataxia; Cognitive decline; GRAF2; GTPase regulator associated with focal adhesion kinase (GRAF); Immunoglobulin G (IgG); Limbic encephalitis; Medusa head ataxia; Oligophrenin-like protein 1 (OPHN1L); Polyneuropathy; Rho GTPase-activating protein 10 (ARHGAP10); Rho GTPase-activating protein 26 (ARHGAP26).

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Conflict of interest statement

S.J., B.W., J.H. and J.U.R. report no conflicts of interest. L.K. and S.B. are employees of Euroimmun AG, Lübeck, Germany.

Figures

Fig. 1
Fig. 1
Binding of serum IgG1 (A-D), IgA (E) and IgM (F) from a patient with ACA to both ARHGAP26-transfected (A) and ARHGAP10-transfected HEK293 cells (B, E–F) and to primate cerebellum tissue sections (D) but not to mock-transfected HEK293 cells (C, E inset, F inset). Red (AF568) indicates bound IgG1 in (A-D); green (FITC) indicates bound IgA or IgM in (E–F); blue indicates nuclear staining by DAPI
Fig. 2
Fig. 2
Strong correlation of ARHGAP10-IgG serum and CSF titres with ARHGAP26-IgG (anti-Ca) serum and CSF titres (N = 13 serum [black squares] and 3 CSF [blue squares] samples with available end titres), as determined by means of cell-based assays. Samples yielding identical titres are shown slightly set-off (double-square). Note the use of a logarithmic (log2) scale
Fig. 3
Fig. 3
Demonstration of ARHGAP26 (GRAF1) and ARHGAP10 (GRAF2) and Arhgap10 mRNA expression in mouse brain and cerebellum. Panel A (reproduced from Safa Lucken-Ardjomande Häsler et al., GRAF1a is a brain-specific protein that promotes lipid droplet clustering and growth, and is enriched at lipid droplet junctions, J Cell Sci 2014, Fig. 1, under the terms of the Creative Commons Attribution License [http://creativecommons.org/licenses/by/3.0]) shows presence of the two antigens in mouse brain extracts from E16 and E18 embryos, P1 and P7 neonates, and from an adult, and demonstrates the developmentally regulated expression of both of the two proteins. Equal loading was verified on the same blot with an anti-Rab8 antibody. B Upper panel: ARHGAP10 and ARHGAP26 expression in mouse cerebellum extract as demonstrated by use of two rabbit antibodies (binding shown in red). Lower panel: While non-denatured ARHGAP10 expressed in HEK293 cells was clearly recognised by the patient’s serum IgG as shown in Fig. 1, no clear binding of the same patient’s IgG to ARHGAP10 (but to ARHGAP26) was seen in the Western blot, suggesting that the epitope detected by anti-ARHGAP10 might be sensitive to denaturing conditions. Red indicates binding of the two rabbit antibodies; green, binding of patient IgG; yellow, overlap of patient IgG and the respective rabbit antibodies. C Developmental changes in Arhgap10 mRNA levels in various brain regions in C57BL/6 J mice (n = 3 mice in each time point) (reproduced from Kazuhiro Hada et al. Mice carrying a schizophrenia-associated mutation of the Arhgap10 gene are vulnerable to the effects of methamphetamine treatment on cognitive function: association with morphological abnormalities in striatal neurons. Mol Brain 2021, Fig. 1, under the terms of the Creative Commons Attribution License [http://creativecommons.org/licenses/by/4.0])
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