Using Next Generation Sequencing to Unravel the Pathogenesis of Sporadic Inclusion Body Myositis - The International IBM Consortium Genetic Study

Lead Research Organisation: University College London
Department Name: Institute of Neurology

Abstract

The commonest muscle disease that occurs in patients over the age of 45 years is a muscle wasting disease called inclusion body myositis (IBM). Patients typically develop progressive muscle wasting and weakness that progresses and causes marked disability and ultimately death from immobility over the course of around 10 years. There are no effective treatment for patients with IBM. The precise cause of this muscle disease is not known. However, on muscle biopsies from patients there seems to be a combination of some mild inflammation in the muscle and also an accumulation of abnormal proteins, similar to the accumulated proteins that are seen in the brains of patients with neurodegenerative diseases such as Alzheimer's, fronto-temporal dementia and motor neurone disease.

Previous research has indicated that there may be genetic factors that predispose people to getting IBM but the previous studies have been quite small and not conclusive. In this research we have brought together experts in IBM from all over the world including Europe, USA and Australia to generate increased awareness of IBM, define diagnostic criteria, collect clinical information and DNA. Over the last three years we have been able to collect the largest group ever of IBM patients and DNA samples - approximately 950 cases and this number will be over 1000 once this study begins. The patient DNA and muscle tissue has been carefully stored for this work. This very large collection of DNA has put us in a very good position to undertake much more detailed genetic studies than have ever been done before to try and work out what the genetic risks factors and genes are that predispose people to this devastating disease.

We plan to use the latest next generation sequencing techniques to unravel all the coding variants (those that alter proteins) that are present in 200 IBM patients DNA samples in comparison with 200 patients that are controls with normal muscles. We will analyze the DNA that we have already extracted from patients muscle tissue as this is the best diagnostic group. We will replicate the variants found in a further 700 IBM cases and over 2200 other controls. We are highly experienced in next generation sequencing technology and this has been strengthened by the recent award of a Wellcome Trust equipment grant to purchase the latest next generation sequencer. Recently we have used these techniques to identify the genetic causes of other neuromuscular disorders.

In comparison with other disorders like Alzheimer's disease, where proteins are aggregated in the brain as opposed to the muscle as in IBM, the greatest advancement have been made with the identification of disease genes and genetic risk factors. If we can work out what the key genes are and how these disease causing pathways function, we will pave the way for new therapies and treatments to help patients.

Technical Summary

1. Library preparation for exome sequencing
In each case high quality DNA (A260/280 ratio >1.8) has been extracted. We have switched over to the Illumina TruSeq Enrichment Kit as it allows six samples to be pooled for exome sequencing and is therefore far more cost effective. The enrichment method is a standard protocol from Illumina starting with the TruSeq sample preparation, pooling of six samples, capture of targeted regions with the TruSeq exome kit and then sequencing. The HiSeq has a very large capacity for sequencing, with the TruSeq enrichment our current most cost effective run is 100bp paired-end reads which gives 60 million reads per sample, at around 68 fold coverage (2 samples per lane). Depending on cost we can modify the runs in several ways, usually by extending read length and changing samples per lane.

2. Exome sequence data processing and variant calling
After the genome analyser generates the raw sequencing images (image analysis) there are several steps involved in the read conversion to variant calling. These include initial base calling, sequence alignment, building, indexing, sorting SAM files, recalibrating quality scores, removing duplicates and merging lanes. Variants are called and filtered based on quality control (read depth, quality scores etc). A genotype file is produced and then annotation and prediction of consequence (amino-acid change, predicted damage of variant) is carried out using the package Sorting Intolerant From Tolerant (SIFT).

3. Statistical analysis
This will primarily focus on gene level comparisons testing a reduced set of genes identified as harbouring non-synonymous coding variants or splice variants that may contribute to risk of IBM. Exome sequence data will be filtered first to remove common variants found in publicly available data from dbSNP and the 1000 Genomes Project. A second level of filtering will be used to extract non-synonymous coding variants and splice variants using existing exome annota

Planned Impact

Who will benefit
Importance and unmet health need for patients and the economy: Inclusion body myositis is the most common acquired muscle disease in patients over the age of forty years. It causes progressive major disability and often premature death after 5-10 years from the consequences of immobility. It represents a significant unmet health need in the UK-worldwide. Since onset is often in prime working life and patients often become significantly disabled quickly and cannot contribute to the economy. A systematic genetic study using the latest genetic technology has not been undertaken. Limited studies have suggested a HLA association, however the disease is unresponsive to immunosuppression and discovery of new mechanisms pathways is required. Key beneficiaries from this research include 1. The immediate and wider science communities, 2. Key established and new industry partners who can develop new paradigms for drug testing based on new genetic discovery 3. Patient organisations and patients. Until now IBM patients have been a relatively neglected group experiencing delayed and misdiagnosis. Professor Hanna and the MRC Centre for translational research have existing partnerships with the major patient organisations linked to IBM namely the myositis support group where Professor Hanna is an advisor helped plan annual patient meetings and the Muscular Dystrophy Campaign that also support IBM patient groups.

How will they benefit
Potential areas for exploitation from this research: Discovery of new genetic associations for IBM will have significant potential to build on existing collaborations with industry and develop new industry collaborations. Potential new areas arising from this research include- identification of new target pathways that might be druggable, the possibility of developing new or taking advantage of existing animal models and of developing new cellular models. All these possibilities will be of interest to industry especially for testing new therapies based on the genetic discovery of new targets. We have established collaborations with the Senexis (Cambridge biotech) to screen anti protein aggregate small molecules and we are currently using a nematode and a myoblast line for screening. However, Senexis- (see MRC Centre Senexis collaboration at www.cnmd.ac.uk) have access to very large number of potential agents- new genetic discovery could direct selection. We have an existing collaboration with Cytrx USA and completing a proof of principle heat shock protein upregulation experimental medicine study in IBM- this relationship will be developed by new genetic discovery. New IBM genetic associations has potential for new genetic testing paradigms for risk stratification and potential for commercialisation of genetic testing.
With Patients and Patient Organisations- the MRC Centre for Neuromuscular Diseases has established effective and regular communication channels with the key patient groups representing IBM. Results of this research along with implications and impact for patients and families will be discussed at the annual Myositis Support Group meeting, the annual MRC Centre patient group meeting and the Muscular Dystrophy Campaign patient group meetings and local branch meetings. All our latest IBM research is posted on the MRC Centre website, the MDC website and the Myositis support group newsletter. With policy makers and NHS commissioners-Professor Hanna is actively engaged with the cross party parliamentary group for neuromuscular diseases including IBM and has given evidence in parliament about the impact of neuromuscular diseases including IBM and has worked with this group and the commissioners to develop standards of care for patients- this work is ongoing and can include implications of this genetic research discovery for patient care eg genetic risk assessment.

Publications

10 25 50
 
Description Improved and larger range of neurogenetic tests
Geographic Reach National 
Policy Influence Type Influenced training of practitioners or researchers
Impact Improved and larger range of neurogenetic tests. This leads to greater research and a better service for patients
 
Description BMA project grant
Amount £21,000 (GBP)
Organisation British Medical Association (BMA) 
Sector Learned Society
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Start 01/2007 
End 01/2010
 
Description CRBC project grant/CRBC
Amount £98,000 (GBP)
Organisation National Institute for Health Research 
Department UCLH/UCL Biomedical Research Centre
Sector Public
Country United Kingdom
Start 06/2009 
End 05/2011
 
Description Equipment award Wellcome Trust
Amount £661,363 (GBP)
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 08/2011 
End 07/2016
 
Description MRC Project Grant
Amount £522,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 01/2011 
End 01/2015
 
Description MRC Research Grant (The Pathophysiology of Spinocerebellar degeneration)
Amount £1,600,000 (GBP)
Funding ID G0802760 
Organisation Medical Research Council (MRC) 
Sector Academic/University
Country United Kingdom
Start 02/2010 
End 01/2015
 
Description MYOPROSP Consortium
Amount £66,301 (GBP)
Organisation Medical Research Council (MRC) 
Sector Academic/University
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Start  
 
Description PhD studentship to work on IBM and neuromuscular disorders from the Saudi Government
Amount £150,000 (GBP)
Organisation Government of Saudi Arabia 
Sector Public
Country Saudi Arabia
Start 09/2016 
End 09/2020
 
Description Project grant: A randomised controlled trial of efficacy of heat shock protein upregulation in IBM
Amount $1,543,444 (USD)
Organisation Food and Drug Administration (FDA) 
Sector Public
Country United States
Start  
 
Description Proof of concept trial
Amount $100,000 (USD)
Organisation Higher Education Funding Council for England 
Sector Public
Country United Kingdom
Start 05/2016 
End 07/2016
 
Description UCL CBRC equipment grant
Amount £339,000 (GBP)
Organisation National Institute for Health Research 
Department UCLH/UCL Biomedical Research Centre
Sector Public
Country United Kingdom
Start 08/2011 
End 08/2016
 
Description Wellcome Trust Equipment Grant
Amount £661,363 (GBP)
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 08/2011 
End 08/2016
 
Description Wellcome Trust Strategic Award
Amount £980,000 (GBP)
Funding ID The Wellcome Trust (equipment and the Synaptopathies strategic award (104033/z/14/z) 
Organisation Wellcome Trust 
Sector Charity/Non Profit
Country United Kingdom
Start 05/2015 
End 04/2020
 
Title Genome sequencing and GeCIP 
Description Genome sequencing and GeCIP 
Type Of Material Technology assay or reagent 
Year Produced 2015 
Provided To Others? Yes  
Impact Genome sequencing and GeCIP 
URL http://www.genomicsengland.co.uk
 
Title Muscle international registry and biobank 
Description Muscle international registry and biobank 
Type Of Material Database/Collection of Data/Biological Samples 
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Provided To Others? Yes  
Impact Muscle international registry and biobank 
 
Title Synaptopathies collaboration 
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Provided To Others? Yes  
Impact Synaptopathies collaboration 
 
Title SOLVE-RD, Coriell, Neurobiobank database of samples, tissue 
Description SOLVE-RD, Coriell, Neurobiobank database of samples, tissue: all anonymous Important for genetics and analysis 
Type Of Material Database/Collection of data 
Provided To Others? No  
Impact SOLVE-RD, Coriell, Neurobiobank database of samples, tissue: all anonymous Important for genetics and analysis 
 
Title Synaptopathies collaboration 
Description Synaptopathies collaboration 
Type Of Material Database/Collection of data 
Year Produced 2015 
Provided To Others? Yes  
Impact Synaptopathies collaboration 
 
Description Ataxia UK 
Organisation Ataxia UK
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution Information and genetic analysis of patients
Collaborator Contribution Made members aware of our work, identified patients and families
Impact Publications, made UK patients with Ataxia more aware
Start Year 2006
 
Description Athens collaboration on ataxia and neuropathy 
Organisation National and Kapodistrian University of Athens
Department Neurology Athens
Country Greece 
Sector Academic/University 
PI Contribution Large Greek study on ataxia and neuropathy in Greece
Impact Large Greek study on ataxia and neuropathy in Greece
Start Year 2010
 
Description BMA 
Organisation British Medical Association (BMA)
Country United Kingdom 
Sector Learned Society 
PI Contribution Funding and publication
Collaborator Contribution Funding and good press for our research
Impact Funding and publication
Start Year 2007
 
Description Cambridge collaboration on Dog ataxia and neuropathy. 
Organisation University of Cambridge
Department Department of Veterinary Medicine
Country United Kingdom 
Sector Academic/University 
PI Contribution Cambridge collaboration on Dog ataxia and neuropathy. We are working on the genetics of neuropathy and ataxia in dog models of human disease
Collaborator Contribution Dog tissue
Impact Ongoing work
Start Year 2010
 
Description Childhood Motor Neuron Disease consortium 
Organisation Great Ormond Street Hospital (GOSH)
Department Department of Neurology
Country United Kingdom 
Sector Hospitals 
PI Contribution Collaborative group that have collected clinical details, cells, DNA and treatment responses on Childhood Motor Neuron Disease
Collaborator Contribution Collaborators have provided clinical details, cells, DNA for exome sequencing and gene identification
Impact A number of DNA samples and fibroblasts collected
Start Year 2011
 
Description Childhood Motor Neuron Disease consortium 
Organisation University of Pennsylvania
Department Department of Neurology
Country United States 
Sector Academic/University 
PI Contribution Collaborative group that have collected clinical details, cells, DNA and treatment responses on Childhood Motor Neuron Disease
Collaborator Contribution Collaborators have provided clinical details, cells, DNA for exome sequencing and gene identification
Impact A number of DNA samples and fibroblasts collected
Start Year 2011
 
Description Childhood Motor Neuron Disease consortium 
Organisation University of Sydney
Department Brain and Mind Research Institute
Country Australia 
Sector Academic/University 
PI Contribution Collaborative group that have collected clinical details, cells, DNA and treatment responses on Childhood Motor Neuron Disease
Collaborator Contribution Collaborators have provided clinical details, cells, DNA for exome sequencing and gene identification
Impact A number of DNA samples and fibroblasts collected
Start Year 2011
 
Description EUROSCA 
Organisation European Commission
Department EC FP6 Collaborative Projects
Country European Union (EU) 
Sector Academic/University 
PI Contribution Identification, screening and functional characterization of ataxia genes
Collaborator Contribution EUROSCA is a collaboration between researchers and clinicians working on ataxia. These has brought cases and techniques that have benefited my research.
Impact Publications as already give. Patients and clinical details of cases with ataxia
Start Year 2006
 
Description European Brain Bank Network 
Organisation Medical Research Council (MRC)
Department MRC UK Brain Banks Network
Country United Kingdom 
Sector Public 
PI Contribution Brain tissue for our research
Collaborator Contribution Brain tissue for research
Impact Brain tissue for our research
Start Year 2006
 
Description European and American Brain Bank Network 
Organisation Medical Research Council (MRC)
Department MRC UK Brain Banks Network
Country United Kingdom 
Sector Public 
PI Contribution This collaboration has given valuable patient brain tissue to our research
Collaborator Contribution Collaborated with tissus
Impact Publications and the addition of important tissue resources
Start Year 2006
 
Description Genomics England Genome Sequencing (100,000 genomes) and Neurology GeCIP (HH is co-lead) 
Organisation Genomics England
Country United Kingdom 
Sector Public 
PI Contribution Genomics England Genome Sequencing (100,000 genomes) and Neurology GeCIP (HH is co-lead)
Collaborator Contribution Genomics England Genome Sequencing (100,000 genomes) and Neurology GeCIP (HH is co-lead)
Impact Genomics England Genome Sequencing (100,000 genomes) and Neurology GeCIP (HH is co-lead)
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Description Greek Collaboration on neurogenetics - Athens, Thessaloniki and Larisa 
Organisation National and Kapodistrian University of Athens
Department Neurology Athens
Country Greece 
Sector Academic/University 
PI Contribution Greek Collaboration on neurogenetics - Athens, Thessaloniki and Larisa Sharing of DNA and clinical details on series and families
Collaborator Contribution Greek Collaboration on neurogenetics - Athens, Thessaloniki and Larisa Sharing of DNA and clinical details on series and families
Impact Sharing of families and data
Start Year 2011
 
Description Laboratory of Neurogenetics, NIA 
Organisation National Institute on Aging
Country United States 
Sector Public 
PI Contribution Collaborating on techniques such as developing DNA arrays and exome sequencing in our lab
Collaborator Contribution Collaboration on techniques and patients
Impact Joint publications and techniques
Start Year 2006
 
Description Laboratory of Neurogenetics, NIA and NIH, USA 
Organisation National Institute on Aging
Country United States 
Sector Public 
PI Contribution Data, cases, publications
Collaborator Contribution Sharing of data, cases and techniquesData and publications
Impact Publications, assistance with grants
 
Description Laboratory of Neurogenetics, NIA and NIH, USA 
Organisation National Institute on Aging
Country United States 
Sector Public 
PI Contribution Data, cases, publications
Collaborator Contribution Sharing of data, cases and techniquesData and publications
Impact Publications, assistance with grants
 
Description Larisa Greek parkinsonism study 
Organisation University of Thessaly
Department Neurology Thessaly
Country Greece 
Sector Academic/University 
PI Contribution Larisa Greek parkinsonism study. Over 1200 Greek parkinsonian patients and controls collected and DNA extracted
Impact Over 1200 Greek parkinsonian patients and controls collected and DNA extracted. Work ongoing, GWAS underway
Start Year 2010
 
Description MRC Centre for Neuromuscular Diseases 
Organisation University College London
Department MRC Centre for Neuromuscular Diseases
Country United Kingdom 
Sector Public 
PI Contribution The MRC Centre for Neuromuscular Diseases is an MRC funded centre set up to investigate the causes and identify treatments for neuromuscular diseases.
Collaborator Contribution The MRC Centre for Neuromuscular Diseases is an MRC funded centre set up to investigate the causes and identify treatments for neuromuscular diseases. I am a member and collaborator in the The MRC Centre for Neuromuscular Diseases.
Impact The MRC Centre for Neuromuscular Diseases is an MRC funded centre set up to investigate the causes and identify treatments for neuromuscular diseases. I am a member and collaborator in the The MRC Centre for Neuromuscular Diseases.
Start Year 2006
 
Description MRC NMC 
Organisation University College London
Department MRC Centre for Neuromuscular Diseases
Country United Kingdom 
Sector Public 
PI Contribution I am a member of the MRC Centre for Neuromuscular Diseases which has brought in very important collaborations between myself and Mike Hanna and Mary Reilly. We have generated considerable data, patient information and publications.
Collaborator Contribution I am a member of the MRC Centre for Neuromuscular Diseases which has brought in very important collaborations between myself and Mike Hanna and Mary Reilly. We have generated considerable data, patient information and publications.
Impact I am a member of the MRC Centre for Neuromuscular Diseases which has brought in very important collaborations between myself and Mike Hanna and Mary Reilly. We have generated considerable data, patient information and publications.
Start Year 2008
 
Description MSA Trust 
Organisation Sarah Matheson Trust for MSA
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution Data, tours of the lab, newsletter information
Collaborator Contribution Patients, we write a research update in the newletter, many patients have given blood and donated their brain as a result of the research
Impact Publications, data, tours of the lab, newsletter information
Start Year 2006
 
Description NORD 
Organisation National Organization for Rare Disorders (NORD)
Country United States 
Sector Charity/Non Profit 
PI Contribution Patient's referred and a grant
Collaborator Contribution Patient's referred and a grant
Impact Patient's referred and a grant
Start Year 2008
 
Description Neuromics FP7 collaboration 
Organisation European Commission
Department Seventh Framework Programme (FP7)
Country European Union (EU) 
Sector Public 
PI Contribution Neuromics FP7 collaboration where we received funding for a post-doc (to prof hanna) and also funding for sequencing. There was significant collaboration and added value from this collaboration with shared results and materials
Collaborator Contribution Shared results and materials - genome/exome sequencing, functional data and patient materials
Impact Publications, collaboration and preparation for other rare disease grants
Start Year 2012
 
Description Neuromics neurogenetics collaboration 
Organisation Eberhard Karls University of Tubingen
Department Neuromics
Country Germany 
Sector Academic/University 
PI Contribution We are the channelopathy and ataxia part of the collaboration. Our role is exome sequencing, genetics and expression
Collaborator Contribution Groups working on several areas of genetics
Impact Recently started
Start Year 2012
 
Description PENN collaboration on Dog ataxia and neuropathy 
Organisation University of Pennsylvania
Department School of Veterinary Medicine (UPenn)
Country United States 
Sector Academic/University 
PI Contribution PENN collaboration on Dog ataxia and neuropathy. We are working on the genetics of neuropathy and ataxia in dog models of human disease
Collaborator Contribution Provided tissue and clinical details
Impact Joint grant submitted to NIH
Start Year 2010
 
Description Sakhalin Universitycollaboration 
Organisation Sakhalin State University
Country Russian Federation 
Sector Academic/University 
PI Contribution Collaboration on patients with Neuropathy in Russia
Impact ongoing
Start Year 2010
 
Description Sarah Matheson Trust for MSA 
Organisation Sarah Matheson Trust for MSA
Country United Kingdom 
Sector Charity/Non Profit 
PI Contribution Brought patients and encouraged brain donation of MSA patients for our research
Collaborator Contribution Brought patients and encouraged brain donation of MSA patients for our research
Impact Publications, Brought patients and encouraged brain donation of MSA patients for our research
Start Year 2006
 
Description Synaptopathies consortium 
Organisation Partnership for Advanced Computing in Europe (PRACE)
Country Belgium 
Sector Charity/Non Profit 
PI Contribution Synaptopathies consortium: Wellcome Trust strategic award Sequencing in paroxysmal disorders with collaboration with Rothman, Kullmann, Hanna, Sisodiya, Goadsy and others
Collaborator Contribution Families and samples
Impact Built up a significant cohort
Start Year 2015
 
Description University of Tehran 
Organisation University of Tehran
Country Iran, Islamic Republic of 
Sector Academic/University 
PI Contribution Prof Elahe has provided many inportant families for our research and we currently share a PhD student
Collaborator Contribution Prof Elahe has provided many inportant families for our research and we currently share a PhD student
Impact Prof Elahe has provided many inportant families for our research and we currently share a PhD student
Start Year 2009
 
Description Wayne State University 
Organisation Wayne State University
Country United States 
Sector Academic/University 
PI Contribution Sharing of CMT1A patient data and DNA
Collaborator Contribution Sharing of CMT1A patient data and DNA
Impact Sharing of CMT1A patient data and DNA
Start Year 2008
 
Description dystonia genetics and functional gene collaboration 
Organisation Sanford Heart Hospital
PI Contribution Exome sequencing, fibroblast and expression studies
Collaborator Contribution dystonia genetics and functional gene collaboration
Impact DMRF joint grant
Start Year 2010
 
Description Genomics England Genome Sequencing (100,000 genomes) and Neurology GeCIP (HH is co-lead) 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Genomics England Genome Sequencing (100,000 genomes) and Neurology GeCIP (HH is co-lead)
Year(s) Of Engagement Activity 2015
URL http://www.genomicsengland.co.uk
 
Description HSP society 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation Keynote/Invited Speaker
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact HSP yearly meet and section in HSP booklet
Year(s) Of Engagement Activity 2012,2013
 
Description Muscle Study group UK and international 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation Keynote/Invited Speaker
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Genetics and exome sequencing in Muscle Disease

Collaboration
Year(s) Of Engagement Activity 2013
 
Description Neuromuscular research day 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Schools
Results and Impact Around 100 patients

More patients involved in our research
Year(s) Of Engagement Activity 2007,2008,2009,2010