Contributions
Abstract: P1773
Type: Poster Sessions
Abstract Category: N/A
In MS, an endogenous retroviral protein, pHERV-W-ENV (formerly MSRV-Env), has been shown to be abnormally expressed and to display effects relevant for MS pathogenesis (Küry et al. Trends Mol Med 2018). Immunohistochemistry studies independently detected pHERV-W-Env within demyelinating areas of all studied Multiple Sclerosis (MS) brains (van Horssen et al. Mult Scler Relat Disord 2016). A humanized antibody against pHERV-W Env is being evaluated in MS clinical trials (NCT01639300 ; NCT02782858).
We characterized the molecular organization and immunodetection profile of pHERV-W-Env from demyelinating MS lesions, compared to reference proteins from transfected cells.
Snap-frozen tissue blocks from MS active lesions, normal appearing white matter (NAWM) and white matter from controls were obtained from The Netherlands Brain Bank (VUMC ethics committee, Amsterdam The Netherlands). HEK-295T cells were transfected with clones encoding pHERV-W ENV, Syncytin and a GFP control. Soluble and insoluble fractions were separated after protein extraction. Analyses used automated capillary immunoelectrophoresis (WES, Proteinsimple, USA) and monoclonal antibodies (Geneuro, Geneva, Switzerland).
pHERV-W-Env expression extracted from transfected cells showed monomers and trimers in the insoluble fraction but a peak of hexamers around 400 KDa, in the soluble fraction. Oligomers were detected by two monoclonals targeting pHERV-W specific epitopes in N-ter (SU) and C-ter (TM) domains. Neither syncytin nor GFP cross-reacted. pHERV-W ENV deglycosylation disrupted hexamers and trimers with parallel increase in monomers. Similar oligomeric pattern was seen in proteins extracted from active MS lesions. The soluble fraction only contained the hexameric form detected by the anti-SU monoclonal but, conversely to transfected cells, the TM region was only detected after deglycosylation in a stable hexameric structure. This was not observed in corresponding NAWM and WM from non-MS controls.
This soluble high molecular weight HERV-W antigen from active MS lesions is immunologically different from syncytin and highly stable. Since HERV-W was first identified from virion-like particles produced by leptomeningeal and B-lymphocyte cultures (Perron et al. 1997. PNAS) and its envelope shown to block remyelination, this raises question about its possible identity with the MS-B-cell secreted factor (>300KDa) associated with sub-pial cortical demyelination (Lassmann, H. Exp Neurol 2014).
Disclosure: HP receives compensation for his work by Geneuro and is inventor on patents owned by bioMérieux, INSERM or Geneuro but has transferred all his rights to bioMérieux or to Geneuro under applicable laws for employed inventors.
BC, JP, SA and JB receive compensation for their work by Geneuro
Abstract: P1773
Type: Poster Sessions
Abstract Category: N/A
In MS, an endogenous retroviral protein, pHERV-W-ENV (formerly MSRV-Env), has been shown to be abnormally expressed and to display effects relevant for MS pathogenesis (Küry et al. Trends Mol Med 2018). Immunohistochemistry studies independently detected pHERV-W-Env within demyelinating areas of all studied Multiple Sclerosis (MS) brains (van Horssen et al. Mult Scler Relat Disord 2016). A humanized antibody against pHERV-W Env is being evaluated in MS clinical trials (NCT01639300 ; NCT02782858).
We characterized the molecular organization and immunodetection profile of pHERV-W-Env from demyelinating MS lesions, compared to reference proteins from transfected cells.
Snap-frozen tissue blocks from MS active lesions, normal appearing white matter (NAWM) and white matter from controls were obtained from The Netherlands Brain Bank (VUMC ethics committee, Amsterdam The Netherlands). HEK-295T cells were transfected with clones encoding pHERV-W ENV, Syncytin and a GFP control. Soluble and insoluble fractions were separated after protein extraction. Analyses used automated capillary immunoelectrophoresis (WES, Proteinsimple, USA) and monoclonal antibodies (Geneuro, Geneva, Switzerland).
pHERV-W-Env expression extracted from transfected cells showed monomers and trimers in the insoluble fraction but a peak of hexamers around 400 KDa, in the soluble fraction. Oligomers were detected by two monoclonals targeting pHERV-W specific epitopes in N-ter (SU) and C-ter (TM) domains. Neither syncytin nor GFP cross-reacted. pHERV-W ENV deglycosylation disrupted hexamers and trimers with parallel increase in monomers. Similar oligomeric pattern was seen in proteins extracted from active MS lesions. The soluble fraction only contained the hexameric form detected by the anti-SU monoclonal but, conversely to transfected cells, the TM region was only detected after deglycosylation in a stable hexameric structure. This was not observed in corresponding NAWM and WM from non-MS controls.
This soluble high molecular weight HERV-W antigen from active MS lesions is immunologically different from syncytin and highly stable. Since HERV-W was first identified from virion-like particles produced by leptomeningeal and B-lymphocyte cultures (Perron et al. 1997. PNAS) and its envelope shown to block remyelination, this raises question about its possible identity with the MS-B-cell secreted factor (>300KDa) associated with sub-pial cortical demyelination (Lassmann, H. Exp Neurol 2014).
Disclosure: HP receives compensation for his work by Geneuro and is inventor on patents owned by bioMérieux, INSERM or Geneuro but has transferred all his rights to bioMérieux or to Geneuro under applicable laws for employed inventors.
BC, JP, SA and JB receive compensation for their work by Geneuro