Contributions
Abstract: P1069
Type: Poster Sessions
Abstract Category: Pathology and pathogenesis of MS - Genetics/Epigenetics
Objective: To leverage a comprehensive, multidimensional RNA-sequencing pipeline to test the relative merits of three hypotheses regarding the role of B-cells in multiple sclerosis (MS) pathophysiology.
Background: Three prominent hypotheses for the role that B-cells play in MS are 1) clonally expanded B-cells in the central nervous system (CNS) target unidentified autoantigens resulting in pathogenic antibody secretion and/or antigen presentation to pathogenic T cells, 2) CNS B-cells display a pro-inflammatory phenotype or cause tissue injury through secreted factors other than pathogenic autoantibodies, and 3) virally-infected B-cells trigger CNS demyelination.
Methods: We isolated cerebrospinal fluid (CSF) and peripheral blood B-cells from treatment naïve, relapsing MS patients (n=7 [5 female], age=23-40 years, disease duration=37-252 days) and sorted them into the following B-cell categories: naïve, double negative, unswitched memory, switched memory, and plasmablast. We performed RNA extraction, reverse transcription, and amplification using random hexamer primers to generate a cDNA library for Illumina sequencing. We then used custom bioinformatics tools to assess for 1) shared clonal connections and/or similar immunoglobulin class restriction between B-cells across patients, 2) differential gene expression between peripheral and CNS B-cell subsets, and 3) presence of intracellular viral transcripts.
Results: We identified memory and plasmablast clonal connections (including identical clones) between patients across the CSF and periphery, dominated by IgA, IgG and IgM immunoglobulin subclasses. We identified pro-inflammatory profiles of CSF plasmablast B-cells that distinguish them from their peripheral counterparts and other CSF B-cell subsets. Finally, we detected no viral transcripts, including Epstein-Barr virus, in the B-cells (n=70 B-cell subsets).
Conclusion: We describe a multi-pronged bioinformatics protocol that mines a unified transcriptomic B-cell dataset to answer questions regarding clonal connections, B-cell phenotypes and infectious triggers of MS. Our findings support the hypothesis that B-cells originating in the periphery undergo clonal expansion and sustain a pro-inflammatory shift in the CSF in MS, without detectable viral infection.
Disclosure: Akshaya Ramesh has nothing to disclose.
Ryan D. Schubert has nothing to disclose.
Ravi Dandekar has nothing to disclose.
Ariele Greenfield has nothing to disclose.
Rita Loudermilk has nothing to disclose.
Edwina Tran has nothing to disclose.
Refujia Gomez has nothing to disclose.
Ari Green reports personal fees from Inception Sciences and Mylan Pharmaceuticals, and has reported serving on an end point adjudication committee for Biogen and Medimmune. He has served on trial steering committees for Novartis and serves on the Scientific Advisory Board for Bionure.
Riley Bove has received personal compensation for medical legal consulting and for consulting or serving on the advisory boards of F. Hoffmann-La Roche Ltd, Sanofi-Genzyme and Novartis.
Xiaoming Jia has nothing to disclose.
Joseph L. DeRisi has nothing to disclose.
Jeffrey M. Gelfand receives research support from Genentech and MedDay, currently consults for Biogen has received personal compensation for medical legal consulting.
Bruce A C. Cree receivers personal compensation for consulting from Abbvie, Biogen, EMD Serono, GeNeuro, Novartis and Sanofi Genzyme.
Stephen L. Hauser currently serves on the SAB of Symbiotix, Annexon, Bionure, and Molecular Stethoscope and on the BOT of Neurona. Dr. Hauser also has received travel reimbursement and writing assistance from F. Hoffman-La Roche Ltd. for CD20-related meetings and presentations.
Michael R. Wilson receives research grant support from Roche and received a research speaking honorarium from Merck.
Abstract: P1069
Type: Poster Sessions
Abstract Category: Pathology and pathogenesis of MS - Genetics/Epigenetics
Objective: To leverage a comprehensive, multidimensional RNA-sequencing pipeline to test the relative merits of three hypotheses regarding the role of B-cells in multiple sclerosis (MS) pathophysiology.
Background: Three prominent hypotheses for the role that B-cells play in MS are 1) clonally expanded B-cells in the central nervous system (CNS) target unidentified autoantigens resulting in pathogenic antibody secretion and/or antigen presentation to pathogenic T cells, 2) CNS B-cells display a pro-inflammatory phenotype or cause tissue injury through secreted factors other than pathogenic autoantibodies, and 3) virally-infected B-cells trigger CNS demyelination.
Methods: We isolated cerebrospinal fluid (CSF) and peripheral blood B-cells from treatment naïve, relapsing MS patients (n=7 [5 female], age=23-40 years, disease duration=37-252 days) and sorted them into the following B-cell categories: naïve, double negative, unswitched memory, switched memory, and plasmablast. We performed RNA extraction, reverse transcription, and amplification using random hexamer primers to generate a cDNA library for Illumina sequencing. We then used custom bioinformatics tools to assess for 1) shared clonal connections and/or similar immunoglobulin class restriction between B-cells across patients, 2) differential gene expression between peripheral and CNS B-cell subsets, and 3) presence of intracellular viral transcripts.
Results: We identified memory and plasmablast clonal connections (including identical clones) between patients across the CSF and periphery, dominated by IgA, IgG and IgM immunoglobulin subclasses. We identified pro-inflammatory profiles of CSF plasmablast B-cells that distinguish them from their peripheral counterparts and other CSF B-cell subsets. Finally, we detected no viral transcripts, including Epstein-Barr virus, in the B-cells (n=70 B-cell subsets).
Conclusion: We describe a multi-pronged bioinformatics protocol that mines a unified transcriptomic B-cell dataset to answer questions regarding clonal connections, B-cell phenotypes and infectious triggers of MS. Our findings support the hypothesis that B-cells originating in the periphery undergo clonal expansion and sustain a pro-inflammatory shift in the CSF in MS, without detectable viral infection.
Disclosure: Akshaya Ramesh has nothing to disclose.
Ryan D. Schubert has nothing to disclose.
Ravi Dandekar has nothing to disclose.
Ariele Greenfield has nothing to disclose.
Rita Loudermilk has nothing to disclose.
Edwina Tran has nothing to disclose.
Refujia Gomez has nothing to disclose.
Ari Green reports personal fees from Inception Sciences and Mylan Pharmaceuticals, and has reported serving on an end point adjudication committee for Biogen and Medimmune. He has served on trial steering committees for Novartis and serves on the Scientific Advisory Board for Bionure.
Riley Bove has received personal compensation for medical legal consulting and for consulting or serving on the advisory boards of F. Hoffmann-La Roche Ltd, Sanofi-Genzyme and Novartis.
Xiaoming Jia has nothing to disclose.
Joseph L. DeRisi has nothing to disclose.
Jeffrey M. Gelfand receives research support from Genentech and MedDay, currently consults for Biogen has received personal compensation for medical legal consulting.
Bruce A C. Cree receivers personal compensation for consulting from Abbvie, Biogen, EMD Serono, GeNeuro, Novartis and Sanofi Genzyme.
Stephen L. Hauser currently serves on the SAB of Symbiotix, Annexon, Bionure, and Molecular Stethoscope and on the BOT of Neurona. Dr. Hauser also has received travel reimbursement and writing assistance from F. Hoffman-La Roche Ltd. for CD20-related meetings and presentations.
Michael R. Wilson receives research grant support from Roche and received a research speaking honorarium from Merck.