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School of Clinical Sciences
EXPLORING THE NEUROPROTECTIVE AND
ALERTING EFFECTS OF MODAFINIL IN
MULTIPLE SCLEROSIS AND EXPERIMENTAL
AUTOIMMUNE ENCEPHALOMYELITIS
Rashid Hamid Bibani
M.B.CH.B., M.I.M.
Thesis submitted to the University of Nottingham for the degree of Doctor of
Philosophy in Clinical Neurology
MAY 2013
II
"In the name of ALLAH, the Entirely Merciful, the
Especially Merciful"
DEDICATION
I dedicate this thesis to my wonderful family. Particularly to my understanding and
patient wife, Qhadamkhir, who has put up with these many years of research. This
was all possible thanks to her continuous encouragement and her moral support. To
my precious Children: Rasti, Rawsht, and Asuda who are the joy of our lives. Finally,
I dedicate this work to those who believe in diligence, science, art, and the pursuit of
academic excellence.
III
DECLARATION OF ORIGINALITY
I hereby declare that this thesis is my own work based on research that was
undertaken during my study in the Clinical Neurology Division, School of Clinical
Sciences, the University of Nottingham and Queens Medical Centre. to the best of
my knowledge it contains no material previously published or written by another
person, or no material which to a substantial extent has been accepted for the
award of any other degree except where due acknowledgement is made in the
thesis. Any contribution made to the research by others, with whom I have worked at
the university of Nottingham or elsewhere, is explicitly acknowledged in the thesis.
IV
TABLE OF CONTENTS
Page
DEDICATION………………………………………….…………………………….……..II
DECLARATION OF ORIGINALITY…………………………………………………….III
TABLE OF CONTENTS……………...………………………………………………..…IV
LIST OF PUBLICATIONS AND CONFERENCE PRESENTATIONS DERIVED
FROM THE WORK.............................................................................................…..XI
ABSTRACT……………………………………………………………………………….XII
ACKNOWLEDGMENTS…………………………………………………………….…..XV
LIST OF ABBREVIATION………………………………………..…...……....….…..XVII
LIST OF FIGURES………………………………………………................................XXI
LIST OF TABLES………………………………………………………………….….XXIII
CHAPTER 1 INTRODUCTION…………………………………….................................1
Overview of the chapter……………………………………………….............................2
1.1 MULTIPLE SCLEROSIS……............................................................................3
1.1.1 History and background of Multiple Sclerosis………………….……….…..….3
1.1.2 Epidemiology of Multiple Sclerosis……………………………….………..……6
1.1.3 Pathogenesis of Multiple Sclerosis……………………………….………..……7
1.1.3.1 Plaque formation…………………………………………………………..……7
1.1.3.2 Neurodegeneration in Multiple Sclerosis.............................................…...9
1.1.3.2.1 Axonal Degeneration……………………………………………………..…11
1.1.3.2.1.1 Mechanism of axonaldegeneration…………………………………..….11
1.1.3.2.1.1.1 Axonal degeneration in acute inflammatory process……………..…11
1.1.3.2.1.1.2 Progressive axonal damage in chronic plaques……………………..12
1.1.3.3 Gray matter lesions in Multiple Sclerosis……………………………………13
1.1.4 Clinical Courses of Multiple Sclerosis…………………………………………13
1.1.5 Clinical Features of Multiple Sclerosis…………………………………………14
1.1.6 Disabilities in Multiple Sclerosis………………………………………………..16
1.1.6.1 Physical Disabilities……………………………………………………………16
1.1.6.2 Cognitive Disabilities…………………………………………………………..17
1.1.7 Fatigue in Multiple Sclerosis……………………………………………………17
1.1.7.1 Definition and overview……………………………………………………….17
1.1.7.2 Types of Fatigue……………………………………………………………….19
1.1.7.2.1 Motor Fatigue………………………………………………………………..19
1.1.7.2.2 Cognitive Fatigue ……………………………………………………………20
1.1.7.3 Measurement of Fatigue……………………………………………………...20
1.1.7.4 Pathogenesis of Fatigue……………………………………………………...20
V
1.1.7.5 Management of Fatigue……………………………………………………....22
1.1.7.5.1 Non-Pharmacological therapies……………………………………………23
1.1.7.5.2 Pharmacological drug therapies…………………………………………...23
1.1.8 Autonomic dysfunction in Multiple Sclerosis………………………………….24
1.1.9 Diagnosis of Multiple Sclerosis…………………………………………………25
1.1.9.1 Diagnostic criteria for Multiple Sclerosis…………………………………….25
1.1.9.1.1 Poser Criteria………………………………………………………………...25
1.1.9.1.2 The MacDonald Criteria…………………………………………………….26
1.1.9.1.2.1 Revised 2005………………………………………………………………26
1.1.9.1.2.2 Revised 2010………………………………………………………………27
1.1.10 Treatment of Multiple Sclerosis……………………………………………….27
1.1.10.1 Acute Treatments…………………………………………………………….27
1.1.10.2 Disease Modifying Therapies…………………………………………….…28
1.1.10.3 Combination Therapies……………………………………………………...32
1.1.10.4 Investigational Therapies……………………………………………………33
1.1.10.5 Symptomatic Therapies……………………………………………………..37
1.1.10.6 Neuroprotective Agents……………………………………………………..39
1.1.11 Prognosis and Complications of Multiple Sclerosis………………………...43
1.2 EXPERMINAT AUTOIMMUNE ENCEPHALOMYELITIS (EAE)……………..45
1.2.1 EAE Induction……………………………………………………………………45
1.2.2 Pathogenesis of EAE ……………………………………………………………45
1.2.3 Clinical scores of EAE…………………………………………………………..46
1.2.4 EAE and Multiple Sclerosis…………………………………………………….46
1.2.4.1 EAE and multiple sclerosis treatments………………………………….…..47
1.2.4.2 Major differences between EAE and multiple sclerosis…………………...47
1.3 MODAFINIL (PROVIGIL)…………………………………………………….……48
1.3.1 Introduction……………………………………………………………………….48
1.3.2 Pharmacodynamic properties of Modafinil……………………………………49
1.3.3 Clinical efficacy and Tolerability of Modafinil……………………………….…50
1.3.4 Mechanism of Action of Modafinil……………………………………………...52
1.3.4.1 Effects of Modafinil on the Dopaminergic Pathways………………………52
1.3.4.2 Effects of Modafinil on Noradrenergic Pathways…………………………..53
1.3.4.3 Effects of Modafinil on Glutamate……………………………………………54
1.3.4.4 Effect of Modafinil on gama amino butyric acid (GABA)…………………..54
1.3.4.5 Effect of Modafinil on serotonin………………………………………………54
1.3.4.6 Effects of Modafinil on Histaminergic Pathways……………………………55
1.3.4.7 Effects of Modafinil on Orexinergic Pathways…...…………………………55
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1.3.5 Approved Indications of Modafinil……………………………………………...55
1.3.5.1 Narcolepsy……………………………………………………………………..55
1.3.5.2 Obstructive Sleep Apnoea (OSA)……………………………………………56
1.3.5.3 Shift-Work Sleep Disorder (SWSD)………………………………………….56
1.3.6 Investigational Uses of Modafinil……………………………………………….57
1.3.6.1 Neurological Disorders………………………………………………………..57
1.3.6.1.1 Parkinson's disease…………………………………………………………57
1.3.6.1.2 Myotonic Dystrophy…………………………………………………………57
1.3.6.2 Psychiatric Disorders………………………………………………………….58
1.3.6.2.1 Attention Deficit Hyperactivity Disorder (ADHD) …………………………58
1.3.6.2.2 Depression…………………………………………………………………...58
1.3.6.2.3 Schizophrenia………………………………………………………………..58
1.3.6.2.4 Alzheimer‘s Disease………………………………………………………...59
1.3.6.2.5 Effect of modafinil on addiction and substances dependency………….59
1.3.6.2.5.1 Cocaine …………………………………………………………………….59
1.3.6.2.5.2 Amphetamines…………………………………………………………….60
1.3.6.2.5.3 Nicotine…………………………………………………………………….60
1.3.6.3 Effect of modafinil on Disorders Associated with Fatigue…………………60
1.3.6.3.1 Chronic Fatigue Syndrome…………………………………………………60
1.3.6.3.2 Fatigue in Post-Polio Syndrome…………………………………………...60
1.3.6.3.3 Fatigue in Multiple Sclerosis……………………………………………….61
1.3.6.3.4 Fatigue in Parkinson's disease…………………………………………….61
1.3.6.3.5 Cancer-related Fatigue……………………………………………………..62
1.3.6.4 Recovery from General Anaesthesia………………………………………..62
1.3.6.5 Sleep-Deprived Emergency Room Physicians……………………………..62
1.3.6.6 Effects of modafinil on quality of life (QoL)…………………………………62
1.3.6.7 Effects of modafinil on Cognitive Performance…………………………….63
1.3.6.8 Effects of Modafinil in Healthy Volunteers………………………………….63
1.3.6.8.1 Non-sleep deprived volunteers……………………………………………63
1.3.6.8.2 Sleep-deprived volunteers…………………………………………………63
1.3.7 Neuroprotective aspects of modafinil………………………………………….64
1.4 SUMMARY AND CONCLUSIONS……………………………………………….67
CHAPTER 2 EXPLORING THE POTENTIAL NEUROPROTECTIVE EFFECTS
OF MODAFINIL IN MULTIPLE SCLEROSIS (RETROSPECTIVE STUDY)………68
2.1 Introduction…………………………………………………………………………69
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2.2 Methods…………………………………………………………………………….69
2.2.1 Patients……………………………………………………………………….…..69
2.2.2 Data Analysis and Statistics……………………………………………………70
2.3 Results………………………………………………………………………………71
2.3.1 Demographic and Clinical Characteristics…………………………………….71
2.3.1.1 Patient demographics…………………………………………………………71
2.3.2 Effect of modafinil on EDSS progression……………………………………..71
2.3.3 Evaluation the role of DMTs concomitantly received with modafinil on
EDSS changes…………………………………………………………………………….74
2.4 Discussion…………………………………………………………………………..76
2.4.1 Limitations of the study..…..…………………………………………………….77
2.4.2 Conclusions……………………………………………………………………....79
2.4.3 Future work……………………………………………………………………….79
CHAPTER 3 MODULATION OF EXPERIMENTAL AUTOIMMUNE
ENCEPHALOMYELITIS (EAE) BY MODAFINIL……………………………………..80
3.1 Introduction………………………….……………………………..……………….81
3.2 Methods and Animals………….…………………………………………………..83
3.2.1 Animals…………………………….……………………………………………..83
3.2.2 Peptide……………………………………………………………………………83
3.2.3 Induction of EAE…………………………………………………………………83
3.2.4 Treatment of mice…………………………..……………………………………84
3.2.5 Clinical evaluation…………………………………..……………………………85
3.2.6 Immunohistopathological evaluation of neuroprotection potential of
modafinil in EAE …………………………………………………………….…………….85
3.2.6.1 Histopathological Examination of EAE……………………….……………..86
3.2.6.2 Determination of Various Cytokines/Chemokines in Serum……………..87
3.2.6.3 Detection of modafinil-Induced Apoptosis in Primary T Cells………...…..87
3.2.6.4 Statistical analysis of immunohistopathological data………………….…..88
3.2.7 Statistical Analysis.......................................................................................88
3.3 Results…………………………………………………………….….…….….……88
3.3.1 Modafinil ameliorated clinical severity of EAE mice………….….…..………88
3.4 Discussion…………………………………………………………….……….……91
CHAPTER 4 ASSOCIATION OF A DEFICIT OF AROUSAL WITH FATIGUE
IN MULTIPLE SCLEROSIS: EFFECT OF MODAFINL…………………….………...94
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4.1 Introduction……………………………………………………………………..….95
4.2 Material and Methods………………………………………………………..……99
4.2.1 Subjects……………………………………………………………………….….99
4.2.1.1 Patients…………………………………………………………………….….100
4.2.1.2 Healthy controls…………………………………………………………..….101
4.2.2 Drugs…………………………………………………………………………….101
4.2.3 Design……………………………………………………………………….…..101
4.2.4 Procedure………………………………………………………………….……102
4.2.5 Tests and apparatus…………………………………………………………...102
4.2.5.1 Tests of alertness……………………………………………………….……102
4.2.5.1.1 Self-rating of alertness…………………………………………………….102
4.2.5.1.2 Instrumental measurements of alertness………………………………..103
4.2.5.1.2.1 Critical flicker fusion frequency (CFFF)……………………….………103
4.2.5.1.2.2 Pupillographic sleepiness test (PST)…………………………….........103
4.2.5.1.3 Psychomotor tests……………………………………………………...….104
4.2.5.1.3.1 Choice reaction time (CRT)…………………………………………….104
4.2.5.2 Tests of autonomic function………………………………………..……….104
4.2.5.2.1 Cardiovascular measures…………………………………………….…..104
4.2.5.2.2 Pupil diameter………………………………………………………………105
4.2.6 Data analysis and statistics……………………………………………………105
4.3 Results……………………………………………………………………………..105
4.3.1 Measures of alertness…………………………………………………………105
4.3.1.1 Comparison of groups prior to treatment………………………………….105
4.3.1.2 Effect of modafinil…………………………………………………………….107
4.3.2 Autonomic measures…………………………………………………………..108
4.3.2.1 Comparison of groups prior to treatment………………………………….108
4.3.2.2 Effect of modafinil…………………………………………………………....108
4.3.2.2.1 Cardiovascular measures…………………………………………………108
4.3.2.2.2 Pupil diameter………………………………………………………..…….110
4.3.3 Subjects‘ verbal reports………………………………………………..………111
4.4 Discussion…………………………………………………………………………111
CHAPTER 5 CEREBROSPINAL FLUID OLIGOCLONAL BAND AND
MULTIPLE SCLEROSIS PROGRESSION: A RETROSPECTIVE STUDY………118
5.1 Introduction…………………………………………………………………….….119
5.2 Methods…………………………………………………………………………...122
5.2.1 Subjects and Setting…………………………………………………..……….122
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5.2.2 Data used and main outcome measures……………………………….…..122
5.2.3 Study design……………………………………………………………………123
5.2.4 Statistical analyses……………………………………………………….……123
5.3 Results…………………………………………………………………………….123
5.3.1 Association between OCB status and disease progression………………124
5.3.2 OCB results over time………………………………………………………....125
5.3.3 Effects of OCB status on time to diagnosis and disease initial type……..126
5.4 Discussion…………………………………………………………………………126
5.5 Conclusion………………………………………………………………………...128
CHAPTER 6 A META-ANALYSIS OF fMRI STUDIES ON FATIGUE IN
MULTIPLE SCLEROSIS………………………………………………………………..130
6.1 Introduction……………………………………………...………………………..131
6.1.1 Structural MRI and Fatigue in Multiple Sclerosis………………………..….131
6.1.1 fMRI and fatigue in Multiple Sclerosis………………………….…………….132
6.2 Methods……………………………………………………………………………134
6.2.1 Local activation likelihood estimate (LocalALE)……………………….……134
6.2.2 Study inclusion…………………………………………………………..……..135
6.2.3 Data extraction…………………………………………………………..……..136
6.3 Results………………………………………………………………………….….137
6.3.1 Multiple sclerosis patients with fatigue…………………………………….…137
6.3.2 Multiple sclerosis patients without reported subjective fatigue……………138
6.3.3 Multiple sclerosis patients with fatigue vs. MS patients without fatigue….139
6.3.4 Healthy controls under same tasks as MS patients with fatigue……….…140
6.3.5 Brain activations in multiple sclerosis patients (with or without fatigue)
vs. healthy controls……………………………………………………………………...141
6.3.6 Comparing ALE in Multiple sclerosis patients and healthy controls……..142
6.3.7 Activation clusters related to modafinil exposure in people without
conventional MRI-detectable brain morphological lesions (healthy; drug
addicts; narcoleptic patients)……………………………………………………………142
6.3.8 Activation or deactivation clusters in people with chronic fatigue
syndrome vs. healthy controls………………………………………………….………144
6.4 Discussion…………………………………………………………………………145
6.4.1 Brain activation in MS with fatigue, without fatigue or undergoing
fatiguing tasks vs. healthy controls………………………………………………….…147
6.4.2 MS patients with fatigue: role of thalamo-striate loop…………..………….148
6.4.3 Modafinil-related activations and deactivations: possible implications
for fatigue in Multiple Sclerosis…………………………………………………………150
6.4.4 Conclusions……………………………………………………..……………….152
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CHAPTER 7 GENERAL SUMMARY AND CONCLUSIONS…………..…………..153
7.1 General summary…………………………………………………….…..………154
7.2 Conclusions……………………………………………………………...………..158
7.3 Limitations and Strengths of the studies…………………………………….…160
7.3.1 Limitations of the studies………………………………………………………160
7.3.2 Strengths of the studies………………………………………………….…….161
7.4 Clinical implementation and importance of the findings………………...……161
7.5 Difficulties in clinical implication OF modafinil…………………………………162
7.6 Recommendations for future research…………………………………………163
BIBILOGRAPHY………………………………………………………………..……….164
APPENDICES………………………………………………………………………...….206
Appendix 1 Ethics approval………………………………………………………….207
Appendix 2 Expanded disability status scale (EDSS)…………………….………207
Appendix 3 McDonald criteria for diagnosis of multiple sclerosis (2001)………208
Appendix 4 The 2005 revisions to the McDonald diagnostic criteria for
multiple sclerosis…………………………………………………………………..…….208
Appendix 5 Magnetic resonance imaging criteria to demonstrate
dissemination of lesions in time……………………………………………………...…210
Appendix 6 Magnetic resonance imaging criteria to demonstrate brain
abnormality and demonstration of dissemination in space…………...……………..210
Appendix 7 Diagnosis of multiple sclerosis in disease with progression
from onset…………………………………………………………………………...……211
Appendix 8 Revised McDonald diagnostic criteria (2010)………………………..212
Appendix 9 Brodmann areas in the brain of human beings………………………213
Appendix 10 Protocol of a randomised, assessor-blind, non-treatment
controlled, parallel group design exploratory trial to explore the
neuroprotective potential of modafinil in multiple sclerosis……………………….…215
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LISTS OF PUBLICATIONS CONFERENCE PRESENTATIONS DERIVED FROM
THE WORK
Publications
Bibani, R. H., Tench, C. R., George, J., Manouchehrinia, A., Palace, J.,
Constantinescu, C. S., 2012. Reduced EDSS progression in multiple sclerosis
patients treated with modafinil for three years or more compared to matched
untreated subjects. Multiple Sclerosis and Related Disorders 1, 131-135.
Niepel, G., Bibani, R. H., Vilisaar, J., Langley, R. W., Bradshaw, C. M., Szabadi, E.,
Constantinescu, C. S., 2012. Association of a deficit of arousal with fatigue in
multiple sclerosis: Effect of modafinil. Neuropharmacology 64, 380-388.
Conference Presentations:
R.H. Bibani. C.R. Tench, C.S. Constantinescu, 2011. Positive effect of modafinil on
EDSS progression in multiple sclerosis. 21th Meeting of the European Neurological
Society, Lisbon, Portugal / 28 - 31 May 2011.
Rashid H Bibani, Christopher R Tench and Cris S Constantinescu, A drug used to
treat fatigue in Multiple Sclerosis may help cure the disease completely. Research
Showcase, the University of Nottingham 7th June 2011.
Rashid H Bibani, Christopher R Tench and Cris S Constantinescu, Modafinil and
EDSS progression in multiple sclerosis. The Former Institute of Neuroscience, the
University of Nottingham, Medical School Foyer 28th September 2011.
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ABSTRACT
Multiple sclerosis (MS) is the most common demyelinating disease. It is
characterised by a great variety of neurological deficits, which most commonly
present initially in a relapsing remitting fashion and then take on a gradually
progressive course. MS is incurable, since present medications do not counteract
progression of the disease. Therefore, an additional strategy aims to focus on
prevention of the neuronal loss in an attempt to stop or slow down the progression
of the disease.
In this thesis the neuroprotective potential of modafinil is tested in MS in a
retrospective study. The ability of modafinil to reduce neurological dysfunction in the
MS animal model is also investigated.
In retrospective study the expanded disability status scale (EDSS) progression of
thirty patients with MS who received modafinil for the treatment of MS-related
fatigue for an uninterrupted period of 3 years or more was compared with ninety
matched patients not treated with modafinil, followed up for a matching period of
time. We found that the EDSS increase in patients not treated with modafinil was
greater than in those treated with modafinil in both relapsing/remitting and
progressive MS.
In another experiment, we evaluated the effect of two treatment doses (low dose
and high dose) of modafinil on the level of disability in experimental autoimmune
encephalomyelitis (EAE) in a placebo controlled study. Modafinil decreased the
severity of EAE at both treatment doses and the effect was greater in high dose.
The study in chapter 4 was aimed to explore the anti-fatigue and alerting effects of
modafinil in MS in an attempt to link these with the potential neuroprotective effects
of modafinil. This was a detailed reanalysis of a prospective placebo controlled
study (based on prospectively collected data), in which we examined whether there
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is any difference between MS patients with fatigue, MS patients without fatigue, and
healthy controls on measures of alertness and autonomic function. We found that
MS patients with fatigue, compared with healthy controls, had reduced level of
alertness on all the tests used, MS patients with fatigue had a reduced level of
autonomic function compared to the other two groups. Furthermore, we found that
Modafinil displayed alerting and sympathomimetic effects in all three groups of
subjects.
In Chapter 5, we assessed a problem relevant to the progression of MS. We take
advantage of the methods and data used in the chapter 2 to apply the same
retrospective study methodology and statistical retrospective modeling of EDSS
progression using the linear regression model to look at the role of oligoclonal band
(OCB) positivity or negativity in EDSS progression. Unlike previous studies in
smaller cohorts, we did not find that OCB negative patients have a more benign
course of disease.
The meta-analysis study in chapter 6 was designed to generate some knowledge
regarding the central mechanism of fatigue in general and fatigue related to MS,
using a novel functional magnetic resonance imaging (fMRI) meta-analysis method
developed by CR Tench in our group. The study has also aimed to explore the brain
areas which could be activated by modafinil treatment. The conclusion of this study
was that the thalamus and striate are central and relevant nodes for the
pathogenesis of fatigue in MS. The study has not detected the specific brain area to
be activated by modafinil and showed multiple brain activations.
With regard to the promising findings in our previous experiments, the protocol of a
prospective phase II clinical trial was designed and detailed in appendix 10 using
radiological primary and clinical secondary outcome measures.
In conclusion, modafinil may slow down the progression of disability in patients with
MS and decrease disease severity in EAE. Modafinil can display alerting and
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sympathomimetic effects in MS patients as well as in healthy subjects. The
thalamus and striate are central and relevant nodes for the pathogenesis of fatigue
in MS. These are also areas affected by the MS gray matter pathology and may be
targets for neuroprotection by modafinil in MS. Finally, we have not reported a
significant difference in disease progression measured by EDSS and MSSS
between OCB negative and OCB positive in our patients with MS.
This seemingly heterogeneous group of experiments, primarily centred on
modafinil‘s potential as mechanistic therapy in MS, bring, I hope, new knowledge of
aspects of disease progression and pharmacological neuroprotection in a stage of
the disease where therapeutic options are currently limited and the need for new
treatments is great.
XV
ACKNOWLEDGMENTS
First of all, praise is due to almighty ALLAH with His compassion and mercifulness
to allow me finalizing this Ph.D. project.
I am in deep gratitude to the Iraqi Ministry of Higher Education and Scientific
Research, and to my employers, Hawler Medical University in Kurdistan, for
such a wonderful opportunity to travel to distant lands, in comfort and security. It is
my duty to repay with my service for many years to come.
I am extremely grateful to the University of Nottingham for all of the education and
experiences I have gained over the past four years. This has been an intense time
of personal development and learning to deal with some of life's trials and
tribulations.
My sincerest and everlasting thanks to my supervisor, Professor Cris
Constantinescu, for his ever cheerful guidance through these years and for doing
the hard pioneering work, paving the way for my little research. His insightful
comments have helped sharpen my scientific writing, and I am grateful for his advice
in conducting and publishing research.
Thanks to my co-supervisor Dr Christopher Tench for his guidance through the
intricacies of research is much appreciated, especially during the statistical analysis
of the data he provided me with statistical advice and helped me with statistical
calculations and interpretation of results.
There have been some other people who have helped me over the last four years
and some of them contributed one way or another to this work I would particularly,
like to thank;
Dr Graham Niepel who produced the initial database for the body of work contained
within the chapter four in this thesis, as well as Professor Szabadi who had an
excellent support in statistical analysis and interpretation of the results as well as the
XVI
invaluable advice in the planning and execution of this study. I would like to stress
that this study would not have been possible to be published without his contribute.
Dr Bruno Gran and James Crooks for their immense technical support regarding
induction of EAE and monitoring the clinical scores of the mice and invaluable
advice through most part of this experiment.
Dr Radu Tananescu for his enormous contribution who shared me a lot of
knowledge and opinions during conducting the meta-analysis study.
Ali Manuocheherinia for letting me to have access into his pooled database which I
used in part, in the study described in chapter five as well as his help in the
statistical analysis of the data.
My deepest gratitude to my family without whom none of this would be possible: To
my wife Qhadamkhir, for her essential support, especially at times when I could do
nothing else other than this work, my three children: Rasti, Rawsht and Asuda for
their love, and my family back home for their understanding and patience, especially
my mum, Rahma, for her love, care and support.
Finally, thanks to colleagues and staffs in the Division of Clinical Neurology for the
camaraderie without which office life would probably be unbearable.
XVII
LIST OF ABBERVIATION
AD autonomic dysfunction
ABM autologous bone marrow
ACTH adrenocorticotrophic hormone
ADHD attention deficit-hyperactivity disorder
ALE activation likelihood method
AMPA 2-amino-3-(3-hydroxy-5-methyl-soxazol-4-yl) propanoic acid
ANS autonomic nervous system
APC antigen-presenting cells
ATP adenosine triphosphate
BA brodmann area
BBB blood brain barrier
BDI Beck Depression Inventory
BOLD blood oxygen level-dependent
BP blood pressure
CBT cognitive behaviour therapy
CC corpus callosum
CD cluster of differentiation
CDMS clinically definite multiple sclerosis
CFA complete Freund‘s adjuvant
CFFF critical flicker fusion frequency
CFS chronic fatigue syndrome
CHMP Committee for Medicinal Products for Human Use
CIS clinically isolated syndrome
CNS central nervous system
CNTF cilliary neurotrophic factor
CPAP continuous positive airway pressure
CRT choice reaction time
CSF cerebro spinal fluid
DA dopamine
DAT dopamine transporters
DMSO dimethylsulfoxide
DMTs disease modifying therapies
DPCC dorsal posterior cingulate cortex
DR dopamine receptor
EAE experimental autoimmune encephalomyelitis
XVIII
EDS excessive daytime sleepiness
EDSS expanded disability status scale
ESS Epworth sleepiness scale
FAI fatigue assessment instrument
FCDR false cluster discovery rate
FDA food and drug administration
FDS fatigue descriptive scale
FWER family wise error rate
FIS fatigue impact scale
FMRI functional magnetic resonance imaging
FODOS fields of dead oligodendrocytes
FOSQ functional outcomes of sleep questionnaire
FSS fatigue severity scale
GA glatiramer acetate
GABA gamma-amino butyric acid
GM gray matter
GNDS Guy's Neurological Disability Scale
HLA human leukocyte antigen
IEF isoelectric focusing
IGF insulin like growth factor
IgG immunoglobulin G
IL interleukin
IM intramuscular
INF interferon
IV intravenous
LH/PF lateral hypothalamic/prefronatal
LIF leukaemia-inhibitory factor
LP lumbar puncture
MBP myelin basic protein
MFG medial frontal gyrus
MFIS modified fatigue impact scale
MHC major histocompatibility complex
MHRA Medicines and Healthcare Products Regulatory Agency
MNI Montreal Neurological Institute
MOG myelin oligodendrocyte glycoprotein
MPTP 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine
MRI magnetic resonance imaging
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MRS magnetic resonance spectroscopy
MRT motor reaction time
MS multiple sclerosis
MSC mesenchymal stem cell
MSFC multiple sclerosis functional composite
MSLT multiple sleep latency test
MSSS multiple sclerosis severity scale
MWT maintenance of wakefulness test
NAA N-acetylaspartate
NAA/Cr N-acetylaspartate/creatinine
NAT noradrenalin transporters
NE norepinephrine
NET norepinephrine transporters
NFI-MS neurological fatigue index
NMO neuromyelitis opica
NO nitric oxide
OCB oligo clonal band
OSA obstructive sleep apnoea
PASAT paced auditory serial addition test
PBS phosphate-buffered saline
PD Parkinson‘s disease
PET positron emission tomography
PLP proteolipid protein
PPAR peroxisome proliferators-activated receptor
PPMS primary progressive multiple sclerosis
PRMS relapsing remitting multiple sclerosis
PST pupillographic sleepiness test
PUI papillary unrest index
QoL quality of life
ROI region of interest
RRMS relapsing remitting multiple sclerosis
RRT recognition reaction time
SDT symbol digit substitution test
SPMS secondary progressive multiple sclerosis
SRIs serotonin receptor inhibitors
SSR sympathetic skin response
SSS Stanford Sleepiness Scale
XX
SWSD shift works sleep disorder
TCR T-cell receptor
TGF tumor growth facto
TH T helper
TMN tuberomamillary nucleus
TNF tumour necrosis factor
TRT Total Reaction Time
VAS-F visual analogue scales for fatigue
VFQ25 visual function questionnaire
VLA-4 very late antigen
VLPO ventrolateral preoptic nucleus
WM white matter
XXI
LIST OF FIGURES
Figure 1.1 Dr Martin Charcot (1825-1893)………………………………………………4
Figure 1.2 Geography of multiple sclerosis and migration………………….…………7
Figure 1.3 Characteristics brain pathology in multiple sclerosis………………...…….8
Figure1.4 Immune-mediated demyelination and axonal transaction……….……….10
Figure 1.5 Clinical courses of multiple sclerosis………………………………..……..14
Figure 1.6 Chemical structure of modafinil…………………………………………….50
Figure 2.1 Mean ±SEM EDSS changes at baseline EDSS and EDSS after 3
or more years treatment or follow-up in Modafinil-treated group and
non-modafinil group with progressive and RRMS……………………………...…..….73
Figure 2.2 Mean ±SEM EDSS changes at baseline EDSS and EDSS after 3
or more years treatment in modafinil-treated group concomitantly received or
not received DMTs……………………………………….………………………….……75
Figure 3.1 Effect of modafinil treatment on clinical score of EAE associated
disease activity……………………………………………………….………………..…..89
Figure 3.2 The maximum Severity of clinical course in three groups of EAE………90
Figure 4.1 Measures of alertness: comparison of groups prior to
treatment………………………………………………………………………………….106
Figure 4.2 Measures of alertness: effect of modafinil……………………….………107
Figure 4.3 Autonomic measures: comparison of group prior to treatment…..……109
Figure 4.4 Relationship between light intensity and Pupil diameter…………….…110
Figure 4.5 Schematic diagram of the dopaminergic arousal system showing
XXII
the possible sites of action of modafinil…………………………………………..……116
Figure 6.1 ALE maps for the independent activation likelihood analysis in
MS patients with fatigue undergoing motor tasks-significant clusters.ALE
maps for the independent activation likelihood analysis in MS patients with
fatigue undergoing motor tasks - significant clusters………………………...………137
Figure 6.2 ALE maps for the independent activation likelihood analysis in
MS patients without fatigue undergoing motor tasks-significant clusters
(FCDR level 0.06)…………………………………………………………………..……139
Figure 6.3 ALE maps for the independent activation likelihood analysis in
MS patients with fatigue vs. without fatigue undergoing motor
tasks-significant clusters (FCDR level 0.07)………………………………..…………140
Figure 6.4 ALE maps for independent activation likelihood analysis in
MS patients with fatigue vs. without fatigue undergoing motor tasks-significant
clusters………………………………………………………………………………...….142
Figure 6.5 ALE maps for the independent activation likelihood analysis in
subjects under modafinil (healthy; drug addicts; narcoleptic patients)
all clusters…………………………………………………………………………...……144
Figure 6.6 ALE for the independent activation likelihood in subjects under
modafinil (healthy; drug addicts; narcoleptic patients) -significant clusters of
activations……………………………………………………………………….………..144
XXIII
LIST OF TABLES
Table 1.1 Clinical features of multiple sclerosis…………………………………………5
Table1.3 Symptomatic treatments for multiple sclerosis……………………..……….37
Table 2.1 Demographic characteristics and clinical epidemiology of the
patients……………………………………………………………………………………..71
Table 2.2 Patient's Characteristics according to the multiple sclerosis clinical
types…………………………………………………………………………………….…..72
Table 2.3 Fisher's exact test analysis in the modafinil-treated and untreated
groups have baseline EDSS 0-5 with EDSS increase by ≥1.0 point and the
patients have baseline EDSS ≥5.5 score with EDSS increase by ≥0.5 point…….74
Table 2.4 Mean±SD of the EDSS1 (baseline) and EDSS2 (post treatment) in
modafinil-treated patients received or not received DMTs…………………..………75
Table 3.1 Mean ± SEM of the subject groups………………………………………….89
Table 4.1 Characteristics of the subjects……………………………………………..100
Table 5.1 Demographic and clinical characteristics of the patients………………..124
Table 5.2 Results of linear regression models analysis to calculate the
effects of OCB on EDSS and MSSS……………………………..……………………125
Table 5.3 Prevalence of the OCB positive and OCB negative patients at the
time of the diagnosis………………………….…………………………………………125
Table 6.1 Studies providing data on activations in MS patients with fatigue….….137
Table 6.2 Studies providing data on activations in MS patients without fatigue.....138
Table 6.3 Studies providing data on activations in MS patients with fatigue vs.
patients without fatigue……………………………………………………………...…..139
XXIV
Table 6.4 Studies providing data on activation in healthy control………………….140
Table 6.5 Studies providing data on activations in MS patients
(with fatigue/without fatigue) vs. healthy controls…………………………………….141
Figure 6.6 Studies providing data on activations in subjects under modafinil
(healthy; drug addicts; narcoleptic patients)……………………………………….….143
Table 6.7 Studies providing data on activations or deactivations clusters in
people with CFS vs. healthy controls…………………………………………….……145
1
CHAPTER 1 INTRODUCTION
2
Overview of the chapter
This chapter begins with a general review of multiple sclerosis (MS), regarding its
history and background, epidemiology, immunopathology, clinical courses, clinical
features, diagnosis, current therapies for MS and the future treatment strategies. As
experimental autoimmune encephalomyelitis (EAE) is a useful model for predicting
success with clinical trials in MS, and it is considered a valuable model for aiding the
development of new treatments for MS, a section of this chapter is an overview of
EAE, focusing on: history, EAE induction, pathophysiology and its contribution to the
development, validation, and testing of MS drugs. This is followed by a review of
modafinil, the wakefulness-promoting drug, which focuses on general description of
the drug, mode of the action, the effect of modafinil in MS and other neurodegerative
diseases, and the possible neuroprotective properties of modafinil.
3
1.1 MULTIPLE SCLEROSIS
MS is the most common demyelinating disease. It is characterised clinically by a
great variety of neurological deficits, which most commonly present initially in a
relapsing remitting fashion and then take on a gradually progressive course.
Pathologically, MS is characterised by inflammation, demyelination, axonal loss, and
gliosis.
1.1.1 History and background of Multiple Sclerosis
In the United Kingdom the case of Elizabeth Foster, dating to 1757, likely,
represents the first reasonably convincing case of MS in the medical literature. She
was presented with paralytic disorders in the left side of the body. She was treated
by electrical stimulations. This case was reported by Dr Patrick Brydone, and the
report was published in the leading scientific journal of the day (Philosophical
Transactions) (Lincoln and Ebers, 2012).
Two cases have been reported from the late 13th century, a woman in Iceland
(Poser, 1994), and a Dutch woman (Medaer, 1979) both with chronic, multifocal,
and partially remitting neurologic illnesses that might have been MS.
In 1868 MS was pathologically described by Jean-Martin Charcot (Figure 1.1) a
French neurologist at the University of Paris, who examined a young woman with a
tremor and some other neurological features including slurred speech and abnormal
eye movements, which were different from neurological features in other reported
neurological conditions. Post-mortem, he examined her brain and found the
characteristic plaques of MS (Murray, 2009). In the USA MS was recognized by Dr.
Edward Seguin in 1878. In 1916 Dr James Dawson at the University of Edinburgh
performed microscopic examinations of the MS patient‘s brain post-mortem.
4
Figure 1.1 Dr Martin Charcot (1825-1893). Source: (Paciaroni et al., 2008).
MS is a chronic progressive inflammatory and degenerative disease of the central
nervous system (CNS). It is characterised by the presence of areas of multifocal
demyelination (plaques) that result from damage the protective coat (myelin) of
nerve fibres. Also there is destruction of oligodendroglia, perivascular inflammation,
and chemical changes in lipid and protein constituents of myelin in and around the
plaques.
In the mid-1990s the understanding of MS changed. The results of clinical trials and
findings from neuropathology of MS demonstrated a neurodegenerative process
with axonal injuries that follows demyelination, which are responsible for progressive
neurological impairment.
Spinal cord lesions in MS are common, particularly in the cervical spine, and usually
occur early in the disease. The first description of cervical spinal cord MS by MRI
was performed in 1988 (Honig and Sheremata, 1989). Spinal MS is often associated
with concomitant brain lesions; however, as many as 20% of patients with spinal
lesions do not have intracranial plaques (Noseworthy et al., 2000).
An increasing amount of evidence suggests that MS is heterogeneous (Compston,
2007). Genetic, immunological and unknown environmental factors are known to
5
contribute to the development of MS, but a specific cause for this disease is not
identified (Compston and Coles, 2002). Potentially, it is the most common cause of
non-traumatic neurological disability in young adults and is a tremendous burden for
years to come (Compston and Coles, 2002). Any age group can be affected but its
peak is in the most economically productive years of life.
MS is more common in temperate climates in people of Northern European descent
and it is infrequent in equatorial areas.
Currently, the four major clinical types of MS include relapsing-remitting (RRMS),
primary progressive (PPMS), secondary progressive (SPMS) and progressive
relapsing (PRMS) (Lublin and Reingold, 1996).
Benign MS is a variant of RRMS where patients remain fully functional in all
neurologic systems 10-15 years after disease onset. Clinically isolated syndrome
(CIS) is described as the first neurological episode and may or may not progress to
clinically definite MS (CDMS).
Neuromyelitis optica (NMO) (Devic‘s disease) is an MS-like inflammatory
demyelinating disease, extensively affecting the spinal cord and optic nerves
(Compston and Coles, 2008; Weinshenker et al., 2006; Wingerchuk et al., 2006).
Despite many similarities, current data strongly suggest NMO is an entity distinct
from MS.
Although progressive neurological disability might be present from the onset of MS,
the initial attack of MS is generally mild and self-limiting, but relapsing is common
after a variable duration (Crayton et al., 2004).
Diagnosis of MS is based on evidence of the dissemination in space, dissemination
in time. History and neurological assessment are the cornerstone for the diagnosis
of MS. MRI is the most sensitive method for showing white matter (WM) lesions in
6
patients suspected of having MS. Lumbar puncture (LP) and clinical
neurophysiological tests may be necessary to establish the diagnosis of MS.
So far, there is no curable treatment for MS. Currently approved MS therapeutics
have a mainly anti-inflammatory mode of action. The aim of treatment in MS is to
reduce the frequency, and limit the lasting effects, of relapses, relieve symptoms,
prevent disability arising from disease progression, and promote tissue repair.
The expected future course of the disease mainly depends on subtype. Individuals
with progressive subtype, particularly the primary progressive subtype, have a more
rapid decline in neurological and cognitive functions. The prognosis in females
generally is better than in males. Initial MS symptoms of visual loss or sensory
problems are thought to be markers for a relatively better prognosis. In general, one
third of patients will still be able to work after 15–20 years of the onset of the disease
(Ebers, 2005).
MS is not lethal by itself but death is the result of remarkable disability and disease
complications such as repeated respiratory and urinary tract infections.
1.1.2 Epidemiology of Multiple Sclerosis
MS is recognised throughout the world with high prevalence in the Northern
Europeans, the North of America and Southern Australia. It is seen less frequently in
Asians and is very rare among indigenous people of Africa and Australia (Figure
1.2) Although genetic susceptibility and ethnic group pattern are likely involved, no
concrete data have been shown as to why certain regions have a higher incidence
of MS (Wallin et al., 2000).
The disease has an incidence of about seven per 100000 every year, prevalence of
around 120 per 100000, and lifetime risk of one in 400 (Compston and Coles, 2008).
It has been estimated that within 15 years more than 50% of non-treated MS
patients need assistance with their daily household and employment responsibilities
7
Figure 1.2 Geography of multiple sclerosis and migration.
The five continents are depicted to show medium prevalence of multiple sclerosis
(orange), areas of exceptionally high frequency (red), and those with low rates
(grey-blue). Some regions are fairly uncharted and these colours are only intended
to provide an impression of the geographical trends. Major routes of migration from
the high-risk zone of northern Europe, especially including small but informative
studies, are shown as dotted arrows. Studies involving migrants from low-risk to
high-risk zones are shown as solid arrows. Source: (Compston and Coles, 2008).
(Pugliatti et al., 2006). Most of the people with MS usually die of complications such
as pneumonia and repeated urinary tract infection rather than of MS itself (especially
in bedridden patients) (Ebers, 2005). MS is more common in females, according to
Pugliatti's review article the women-to-men ratio for MS in Europe varies from 1.1 to
3.4 (Pugliatti et al., 2006). However recent reports have stated an increase in
incidence of MS in women. The basis for this difference is unknown, but hormonal
components may be responsible (Debouverie et al., 2007).
1.1.3 Pathogenesis of Multiple Sclerosis
1.1.3.1 Plaque formation
The mechanisms of the initial pathogenic events leading to plaque formation are
controversial (Lucchinetti et al., 2000). The most commonly held view is that the
peripherally activated T-cells migrate into the CNS and attack myelin and
8
oligodendrocytes resulting in production of focal inflammatory lesions (Lassmann et
al., 2007). Barnett and Prineas (2004) had an alternative view: they suggested that
in some cases the earliest lesions comprise large areas of apoptotic
oligodendrocytes, termed fields of dead oligodendrocytes (FoDOs). The
pathogenesis of FoDOs is tentative, but could involve humoral factors or
oligodendrocyte degeneration in response to viral infection. In any case, the
evolution of active lesions involves widespread, focal loss of myelin, the presence of
large numbers of activated macrophages digesting myelin degradation products,
and a T-cell infiltrate, with CD8+ T-cells predominating (Figure 1.3).
Figure 1.3 Characteristic brain pathology in multiple sclerosis.
A / Low-power image of active demyelinating white matter lesion, showing
macrophages with myelin degradation products (arrows) and reactive gliosis
(arrowheads). B/ Higher-magnification image of the active lesions shown in (A)
reveals demyelinated axons (arrows), macrophages with myelin debris (arrowheads)
and dystrophic axons (asterisk) within the myelin sheath. Source: (Lassmann et al.,
2012).
Although the pathogenesis of MS is not fully understood involvement of cell-
mediated immune and humoral immune response to undetermined antigen(s) is
doubtless. Pathologically MS is characterized by perivenular and parenchymal
infiltration of lymphocytes and macrophages in the parenchyma of the brain, brain
stem, optic nerves, and spinal cord. In general the accepted view of MS
9
pathogenesis has linked the disease course to sensitisation a myelin-specific, CD4+
T lymphocyte in the peripheral in response to macrophage presentation of a foreign
antigen in association with major histocompatibility complex (MHC) class I and class
II (Höftberger et al., 2004; Wucherpfennig and Strominger, 1995). This results in
peripherally activated T cells expressing, and recognising, vascular adhesion
molecules facilitating their entry through blood brain barrier (BBB). Inside the CNS
activated T cells release pro-inflammatory cytokines resulting in up regulation of
local antigen-presenting cells (APC) with the capacity to present self-myelin proteins
(Lassmann et al., 2007).
In addition to T cells the autoimmune B cells and humoral immune mechanisms are
now believed also to play key roles in the pathogenesis of MS and plaque initiation
(Gay and Esiri, 1991; Owens et al., 2006), and demyelination in patients with
established MS (Wucherpfennig and Strominger, 1995). This component has been
recognised previously in MS diagnosis through the presence of oligoclonal bands
(OCB) in the cerebrospinal fluid (CSF) and increased intrathecal Immunoglobulin G
(IgG) synthesis (Link and Huang, 2006). Variable degrees of clonally expanded
populations of memory B cells and plasma cells are found in lesions and CSF from
patients with MS (Bartoš et al., 2007; Magliozzi et al., 2007; Owens et al., 2003). It
has been shown that depletion of B-cells in MS lesions results in a reduction in
gadolinium enhanced lesions on MRI and reduced relapse frequency (Bar-Or et al.,
2008).
1.1.3.2 Neurodegeneration in Multiple Sclerosis
Besides the inflammatory activity in the CNS the degenerative process in MS
appears to start early in the disease (Figure 1.4). Significant brain atrophy has been
found in early diagnosed MS patients with little disability (Chard et al., 2002).
Atrophy of CNS is most pronounced in the progressive phase of MS, and correlates
with the rate of decline in neurological function (Losseff et al., 1996). A study
10
showed low level of N-acetylaspartic acid (NAA), a marker for axonal damage
shown by magnetic resonance spectroscopy (MRS) in MS patients (De Stefano et
al., 2002).Pathologically, in these stages, the lesions are characterised by
Figure 1.4 Immune-mediated demyelination and axonal transaction.
Axonal ovoids are hallmark of transacted axons. Abundant axonal ovoids were
detected in MS tissue (a) when stained for myelin protein (red) and axons (green).
There are areas of demyelination (arrowheads), mediated by microglia and
haematogenous monocytes. One of the axons ends in a large swelling (arrow) or
axonal retraction bulb (arrow). (b and c) Schematic of axonal response during and
following transaction. Demyelination is an immune-mediated or immune cell assisted
process leading to axonal transaction. When transacted, the distal end of the axon
rapidly degenerates while the proximal end connected to the neuronal cell body
survives and transported organelles accumulate at the transaction site and form an
ovoid (arrows). Source: (Trapp and Nave, 2008).
demyelination, activated microglia, apoptotic death of neurons, interlaced with
macrophages and myelin debris, making up the glial scar tissue. The lesions have
less leukocyte infiltrations and there is marked depletion of oligodendrocytes
(Lucchinetti et al., 2003).
11
1.1.3.2.1 Axonal Degeneration in Multiple Sclerosis
Although the MS lesion includes both inflammatory and demyelinating components
their relative influence on axonal loss is unclear. Axonal pathology was mentioned in
early reports that included description of axonal swelling, axonal transaction and
Wallerian degeneration (Kornek and Lassmann, 1999). Some studies have
demonstrated a high incidence of acute axonal injury within both early and chronic
MS lesions (Ferguson et al., 1997; Kornek and Lassmann, 1999; Trapp et al., 1998).
Axonal degeneration is occurs in the setting of acute inflammatory demyelination
(Trapp et al., 1998) and/or as a consequence of chronic demyelination (Bjartmar et
al., 2000; Dutta et al., 2006) (Figure 1.4).
1.1.3.2.1.1 Mechanism of Axonal Degeneration
1.1.3.3.1.1.1 Axonal degeneration in acute inflammatory process
The most accepted contributing causal factors for axonal damage in the acute
lesions are:
Immune-cell mediated injury: There is a close link between axonal injury and
cytotoxic effect of T-cell in human which initiated through direct T cell mediated
cytotoxicity with the target axon (Neumann et al., 2002). Axonal transection has
been reported in vitro in an antigen dependent immunological reaction with Class I
MHC restricted T lymphocytes (Medana et al., 2001). Also the interaction of
activated macrophages or microglia cells with axons in the course of axonal injury
has been suggested in EAE, such cells are consistently found in close contact with
degenerating axons in EAE (Brunn et al., 2008).
Glutamate in acute axonal injury: Increased levels of glutamate after inflammatory
injury leads to excess excitatory activation of inotropic subtypes of glutamate
receptors such as α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)
receptors. This results in toxic accumulation of intracellular sodium and calcium
12
during normal electrical activation (Ouardouz et al., 2009). Evidence of increased
oligodendrocyte and axonal survival, after treatment with a glutamate antagonist in
animal and cellular models, further supports a role for glutamate in acute axonal
injury (Pitt et al., 2000).
Nitric oxide and acute axonal injury: Evidence suggests that nitric oxide (NO), which
is released from inflammatory cells, at higher concentrations may lead to irreversible
destruction of axons (Smith et al., 2001). Also it has been reported that NO may
even directly damage nerve cell bodies and dendrites and can play a role in
demyelination and oligodendroglia damage (Bolanos et al., 1997).
1.1.3.3.1.1.2 Progressive axonal damage in chronic plaques
The most accepted contributing causal factors for axonal damage in the chronic
plaques are:
Remyelination failure: In vitro, trophic factors such as insulin-like growth factor-type
1 (IGF-1) which is a polypeptide growth factor similar in structure to insulin and
neuregulin provided by oligodendrocyte promote normal axon function and survival
to axons. Lack of these factors in chronic lesion result in neurodegeneration and
death of the axons (Compston, 1996; Wilkins and Compston, 2005).
Conduction defects: Axonal conduction is a continuous energy dependent process
that is essential for maintaining cell function. Demyelination disrupts axonal
conduction. Studies have shown that conduction defects along chronically
demyelinated axons contribute to the progression of neurological disability (Kornek
et al., 2001; Waxman, 2001).
Toxic level of intracellular calcium: Studies have shown that stimulation of glutamate
receptors results in Ca2+ influx from both the extracellular space, and from
ryanodine-dependent intracellular stores. The processes result in abnormally
increased intracellular levels of Ca2+ that culminates in the activation of degradation
13
enzymes, inhibition of mitochondrial function and cellular death (Ouardouz et al.,
2009; Trapp and Stys, 2009).
1.1.3.3 Gray Matter lesions in Multiple Sclerosis
MS is generally believed to be a WM disease but conclusions from advanced MRI
techniques and histopathological findings have indicated prominent gray matter
(GM) changes suggestive of both demyelination and axonal damage. These have
been detected in MS cortical lesions (Chard et al., 2002). Generally, GM lesions are
a more prominent feature of PPMS and SPMS, where they can be extensive,
suggesting it is a predominantly late phenomenon in MS pathology (Kutzelnigg et
al., 2005). However, it is also documented that cortical lesions are present from the
earliest stages of MS, accumulate over time, and exceed WM lesions in progressive
MS (Brownell and Hughes, 1962; Lassmann and Lucchinetti, 2008). Pathologically
the lesions are characterised by demyelination, activated microglia, apoptotic death
of neurons, and have less leukocyte infiltrations. Furthermore it is believed that
cortical plaques have important role in contributing to the disease burden in patients
with MS (Peterson et al., 2001).
1.1.4 Clinical courses of Multiple Sclerosis
MS is divided into four clinical subtypes (Lublin and Reingold, 1996) (Figure 1.5).
Relapsing-remitting MS: It is defined as more than one clinical attack of
demyelination, that is an initial episode followed by at least one (relapse), separated
by period(s) of complete or partial recovery (remission). Approximately 80-85% of
patient with MS have RRMS at the onset.
Secondary progressive MS: It is the stage that follows RRMS; symptoms are
continuous and gradually worsen, without remission. Relapses may occur, but less
frequent than in the RR phase. After several years about 50% of patients with
RRMS progress into SPMS (Rovaris et al., 2006).
14
Primary progressive MS: less than 20% people with MS experience continuous
worsening from disease onset with no preceding relapses, although relapses may
subsequently occur, but at low frequency.
Progressive relapsing MS: It is an uncommon form of MS characterised by acute
relapses superimposed on progressive course.
Figure 1.5 Clinical courses of multiple sclerosis.
A/ Relapsing/remitting multiple sclerosis: Clearly-defined disease relapses with full
recovery or with squeal and residual deficit upon recovery; periods between disease
relapses characterised by a lack of disease progression. B/ Secondary progressive
multiple sclerosis: Initial relapsing/remitting disease course followed by progression
with or without occasional relapses, minor remissions or plateaux. C/ Primary
progressive multiple sclerosis: Disease progression from onset, with occasional
plateaux and temporary minor improvements. D/ Progressive-relapsing multiple
sclerosis: Progressive disease from onset, with clear acute relapses, with or without
full recovery.
1.1.5 Clinical Features of Multiple Sclerosis
Clinical features of MS are varied and capricious, depending on location and degree
of the lesions affecting the CNS (Table 1.1). Symptoms start with beginning of
interruption of myelinated tracts in the CNS. Insidious or abrupt weakness in one or
more limbs, a sensory disturbance, monocular visual loss (optic neuritis), double
vision (diplopia), gait instability, and ataxia are the possible initial symptoms of MS.
15
Early symptoms may be severe or trivial. With progression of the disease bladder
dysfunction, heat intolerance and fatigue occur in most patients. Additional
Table 1.1 Clinical features of multiple sclerosis. Source: (Compston and Coles,
2008)
Cerebrum
Cognitive impairment
Deficits in attention,
reasoning, and executive
function (early); dementia
(late)
Hemisensory and motor
Upper motor neuron signs
Affective (mainly
depression)
Epilepsy (rare)
Focal cortical deficits (rare)
Optic nerve
Unilateral painful loss of
vision
Scotoma, reduced visual
acuity, colour vision, and
relative afferent pupillary
defect
Cerebellum and cerebellar
pathways
Tremor
Postural and action tremor,
dysarthria
Clumsiness and poor
balance
Limb incoordination and
gait ataxia
Brainstem
Diplopia, oscillopsia
Nystagmus, internuclear
and other complex
ophthalmoplegias
Vertigo
Impaired swallowing
Dysarthria
Impaired speech and
emotional lability
Pseudobulbar palsy
Paroxysmal symptoms
Spinal cord
Weakness
Upper motor neuron signs
Stiffness and painful
spasms
Spasticity
Bladder dysfunction
Erectile impotence
Constipation
Other
Pain
Fatigue
Temperature sensitivity
and exercise intolerance
16
symptoms include Lhermitte's symptom, hemifacial weakness, vertigo, and tonic
spasms and other paroxysmal symptoms. Cognitive deficits commonly occur in late
onset. Depression and suicide ideation are more common than in age-matched
controls (Compston and Coles, 2008).
1.1.6 Disabilities in Multiple Sclerosis
MS is associated with physical and cognitive disabilities. They have clear impact on
quality of life (QoL) (Janardhan and Bakshi, 2000). Several scales are used to
measure disability in MS such as expanded disability status scale (EDSS), The
Guy's Neurological Disability Scale (GNDS), Multiple sclerosis severity score
(MSSS), paced auditory serial addition test (PASAT), symbol digit substitution test
(SDT), multiple sclerosis functional composite (MSFC), etc.
1.1.6.1 Physical Disabilities
MS Patients vary in the severity of their illness from no obvious physical disability to
being severely disabled. It has become increasingly important both in the clinical
setting and in therapeutic trials to measure disability levels repeatedly in order to
assess progression of disability. The EDSS is a gold-standard measure for
assessing level of disability (Kurtzke, 1983). It is an ordinal scale with 19 disease
steps between 0 and 10 (Appendix 3) The scale measures activity limitation based
on the examination of eight functional systems (pyramidal, cerebellar, brainstem,
sensory, bowel and bladder, visual, cerebral, other) plus ambulation. It does have
some well documented limitations, the most important of which are: it is biased
towards locomotor function, not a sensitive measure to define irreversible
progression of disease and has only moderate inter- and intra-rater reliability (Ebers
et al., 2005; Sharrack et al., 1999). To address this limitations Sharrack and Hughes
established a new disability scale, Guy‘s Neurological Disability Scale (GNDS),
which is a simple clinical disability scale capable of embracing the whole range of
disabilities which could be encountered in the course of MS (Sharrack and Hughes,
17
1999). Identification of sustained disability progression is an important outcome
measure in therapeutic trials in MS. An increase of 1 point on the EDSS above
baseline (or 1.5 EDSS points if the baseline EDSS is 0), subsequently confirmed at
repeat assessment either 3 or 6 months later are the most commonly used
measures (Kappos et al., 2006b). Clinically important change in the EDSS was
deemed to be 1 point change in the range 0–5.0 and 0.5 point change in the range
5.5–8.5. There are some difficulties with these definitions; relapses may produce
neurological changes persisting for many months still followed by full recovery and
people with RRMS exhibit day-to-day fluctuation in neurological signs and
symptoms unrelated to relapses (Leary et al., 2005). EDSS does not take into
account the important aspect of disease duration, which is a major factor in
accumulation of CNS damage over time and the accumulation of functional
disability. To address this deficiency, Roxburgh with his colleagues based on
databases from 11 countries have introduced the MSSS as a method for comparing
disease progression in MS using single assessment at a single point in time
(Roxburgh et al., 2005).
1.1.6.2 Cognitive Disabilities
Cognitive dysfunction is common in MS. It does not strongly correlate with the
physical disability and EDSS score (Miller et al., 1998). Cognitive impairments are
evident in tests measuring attention, vigilance, processing speed, working memory
and executive function. Tests such as PASAT, SDT and MSFC are useful tools to
detect cognitive disability progression in MS and they are sensitive to change over
time.
18
1.1.7 Fatigue in Multiple Sclerosis
1.1.7.1 Definition and overview
As fatigue is a subjective feeling, there is no unique definition for fatigue. Initially,
fatigue in MS has been defined as "an abnormal sense of tiredness or lack of
energy, out of proportion to the degree of effort or level of disability, that significantly
interferes with routine physical or intellectual functioning" (Weinshenker et al.,
1992). The UK Multiple Sclerosis Society defines MS fatigue as ―an overwhelming
sense of tiredness for no apparent reason.‖ Krupp has described fatigue as an
overwhelming sense of tiredness that is out of proportion to ―normal‖ tiredness
(Krupp, 2006). Medically, fatigue in MS has been defined as a ―reversible, motor and
cognitive impairment with reduced motivation and desire to rest, either appearing
spontaneously or brought on by mental or physical activity, humidity, acute infection
and food ingestion. It can occur at any time but is usually worse in the afternoon. In
MS, fatigue can be daily, has usually been present for years and has greater
severity than any premorbid fatigue‖ (Mills and Young, 2008). Fatigue is one of the
most common symptoms in patients with MS. It is reported that 50% to 92% of
patients with MS experience significant fatigue (Kaynak et al., 2006; Lerdal et al.,
2007; Zajicek et al., 2010). It has been described as chronic and the most
debilitating feature of the disease by 15% to 40% of MS patients (Fisk et al., 1994b;
Giovannoni, 2006; Krupp, 2003).
Fatigue has a significant negative impact on daily work, family life, and social
activities of persons with MS and is associated with the perception of an impaired
general health, mental state (Janardhan and Bakshi, 2002; Ritvo et al., 1996). The
majority of patients with MS experience worse fatigue when temperature is higher,
especially those with severe fatigue (Leavitt et al., 2012; Lerdal et al., 2007), and
clearly carries a major physical and psychological burden, especially when
completing everyday tasks (Leocani et al., 2008; Mills and Young, 2008). MS
19
patients often report specific triggers for fatigue, such as heat (Krupp and
Christodoulou, 2001).
The common fatigue symptoms are: reduced energy, malaise, motor weakness
during sustained activity, and difficulty maintaining concentration. Fatigue is
diagnosed when the presence of fatigue symptoms lasts for at least 50% of days for
more than 6 weeks (Multiple Sclerosis Clinical Practice Guideline, 1999). Self-report
questionnaires such as the fatigue severity scale (FSS) may be useful in the
diagnosis of MS fatigue and as a surrogate outcome measure.
Fatigue in MS may manifest itself in a variety of forms, including acute fatigue
localized to specific muscle groups and persistent, global fatigue. Fatigue affects
both motor and cognitive ability.
1.1.7.2 Types of Fatigue
1.1.7.2.1 Motor Fatigue
Motor fatigue is defined as a decline in motor performance during sustained muscle
activity (Bigland-Ritchie et al., 1998). Motor fatigue worsens during MS
exacerbations involving the motor system and improves during remission, but does
not change during exacerbations in which the motor system is unaffected (Djaldetti
et al., 1996). Studies have found that motor fatigue during intermittent voluntary
submaximal contractions of the tibialis anterior muscle was associated neither with
self-reported fatigue in MS patients nor with overall neurologic impairment/disability,
but it was associated with pyramidal signs on examination (Djaldetti et al., 1996;
Sharma et al., 1995). The pathophysiologic basis for motor fatigue in MS patients
remains unclear. Both peripheral and central mechanisms have been suggested.
Studies using transcranial magnetic stimulation have suggested that there may be
decreased central activation as fatigue occurs in MS patients (Brasil-Neto et al.,
1994; Sheean et al., 1997). On the other hand studies focusing on exercise-induced
20
biochemical changes in muscle have suggested that peripheral mechanisms are
involved, producing alterations in muscle metabolism (Hainut and Duchateau, 1989;
Kent-Braun et al., 1994; Miller et al., 1990).
1.1.7.2.2 Cognitive fatigue
Cognitive fatigue can be defined as a decrease in, or inability to sustain, task
performance throughout the duration of a continuous information processing speed
task (Schwid et al., 2002). Cognitive fatigue can occur in all stages of the disease
and usually does not correlate with demographic or disease characteristics such as
age, gender, depression, disability or disease severity, or disease duration
(Parmenter et al., 2003).
Comparing with healthy control, during continuous information processing speed
task patients with MS become cognitively fatigued sooner, reflected by a breakdown
in task performance (Bryant et al., 2004).
1.1.7.3 Measurement of Fatigue
Available measurements for fatigue so far are; FSS, Fatigue Descriptive Scale
(FDS), Modified Fatigue Impact Scale (MFIS), Neurological Fatigue Index (NFI-MS)
and Visual Analogue Scale for Fatigue (VAS-F) (Johnson, 2008). These measures
are mainly self-report questionnaires, and they are not specific to MS. One of the
most commonly used self-report scales is FSS, which is a self-report questionnaire
designed to assess fatigue in general (Krupp et al., 1989). It has shown that FSS
has ability to highlight the approach towards appropriate and individualised
treatments (Valko et al., 2008).
1.1.7.4 Pathogenesis of MS fatigue
The exact aetiology and pathophysiology of fatigue in MS patients are not well
understood, it appears to be complex and multifactorial. Both peripheral and central
mechanisms have been suggested but no satisfactory conclusion has been
21
achieved so far (Kos et al., 2008). Fatigue may be directly related to the underlying
MS disease process and the disease mechanisms such as proinflammatory
cytokines, CNS lesion load, cerebral quantitative imaging abnormalities and patterns
of cerebral activation, endocrine influences and axonal injury (primary fatigue),
[reviewed in (Induruwa et al., 2012)] or may be secondary to non-disease-specific
factors such as secondary effects of inflammation on neuromodulation, disruption of
neural pathways necessary for brain activity, and daytime somnolence due to
nocturnal sleep disturbances such as sleep problems, urinary problems, spasms,
pain, anxiety or depression (secondary fatigue) (Bakshi, 2003; Krupp, 2003; Schwid
et al., 2002). MS fatigue has not been shown to be correlated with disease duration,
gender, psychosomatic mechanisms, physical disability, or sleep dysfunction. A
study has showed that obvious fatigue has been observed in patients with benign
MS with no disability and it was also showed that MS fatigue is not related to some
markers of systemic inflammation (Giovannoni et al., 2001).
A study by Bakshi et al. showed a significant relationship between fatigue and
depression in MS independent of physical disability (Bakshi et al., 2000). Kaminska
et al (2011) have found that sleep disturbances in MS may also result in or
exacerbate fatigue in MS.
Using conventional MRI, only a weak correlation between MRI lesion load and
fatigue has been reported (Bakshi et al., 1999; Colombo et al., 2000). In contrast, by
using more advanced MRI techniques other studies have found that GM pathology
(Cantor, 2010) and the basal ganglia (Téllez et al., 2008) may be a contributing
factor to the development of MS related fatigue. The results of the Niepel et al study
had supported the role of the GM in the pathogenesis of fatigue in MS (Niepel et al.,
2006). The relationship between MS fatigue and brain atrophy has been suggested
by several studies. Yaldizli et al (2011) have found that corpus callosum (CC)
atrophy was present in subjects with MS and may play a role in the evolution of MS-
22
related fatigue. Other studies have suggested that patients with higher levels of
fatigue have higher WM and GM atrophy (Marrie et al., 2005; Pellicano et al., 2010;
Tedeschi et al., 2007). In a comparative study with healthy control a strongest
correlation between cortical atrophy and fatigue in the MS patient has reported
(Pellicano et al., 2010).
It has been found that the fatigued MS patients have significantly increased
adrenocorticotropic hormone (ACTH) levels in the combined dexamethasone-
corticotrophin releasing hormone (Dex-CRH) test, compared to those without fatigue
(Gottschalk et al., 2005). In a similar study with 73 progressive MS patients, Téllez
et al. proposed that fatigue could be related to low serum levels of
dehydroepiandrosterone (Tellez et al., 2006). There is also evidence suggest that
increased activation of central neural circuits is associated with MS fatigue. Several
studies have suggested that performing motor function increases loss of strength
and increased cortex excitability in a wider cerebral area than in control subjects and
led to early fatigue (Benwell et al., 2007; Leocani et al., 2001; Thickbroom et al.,
2008).
Axonal damage is also suggested as being a factor for fatigue in MS. A study by
Tartaglia et al used MRS, found that the N-acetyl aspartate (NAA): Creatinine
(NAA/Cr) ratio used as marker of CNS axonal damage was significantly lower in a
high-fatigue than in a low-fatigue group of MS patients. There was also a significant
inverse linear correlation between the FSS scores and the NAA/ Cr ratio (Tartaglia
et al., 2004).
1.1.7.5 Management of fatigue
Fatigue in MS is different from fatigue in healthy subjects and it is one of the most
challenging symptoms to treat (Krupp et al., 2010). Despite various non-
pharmacological and pharmacological treatments or combinations trials definitive
evidence of their relative efficacy and tolerability is unavailable.
23
1.1.7.5.1 Non-pharmacologic therapies
Non-pharmacological approaches include aerobic exercise programmes, energy
conservation strategies and cognitive behavioural therapy (CBT). The benefit of
cooling therapies has been tested which been reported to be effective in reduction of
fatigue and improvements in physical, cognitive, and psychosocial function
(Flensner and Lindencrona, 2002). Improvement of sleep has been evaluated to
treat fatigue in MS (Heesen et al., 2006) . Aerobic exercise program found to be
effective in reduction of MS fatigue and improvement of health (Mostert and
Kesselring, 2002).
The use of CBT to treat fatigue in MS is still under investigation. A randomised
control trial of patients with MS related fatigue receiving either CBT or relaxation
therapy showed that at 6 months after treatment, both groups described clinically
significant decreases in fatigue levels equivalent to those of the healthy comparison
group (van Kessel et al., 2008).
1.1.7.5.2 Pharmacologic Drug therapy
Several medications have been tried for treatment of fatigue in MS. Amantadine is a
dopaminergic agent that has been evaluated for fatigue. A significant efficacy on
some of the studies, but not all, has been found (Cohen and Fisher, 1989; Krupp et
al., 1995; Murray, 1985; Rosenberg and Appenzeller, 1988). Modafinil is a wake-
promoting agent has been studied in patients with MS, and was effective on several
measures of fatigue (Rammohan et al., 2002; Zifko et al., 2002). Aminopyridines are
potassium channel-blocking agent exert their effect through enhancing conduction in
demyelinated nerve fibres. Its effects on MS fatigue have been suggested but not
definitively demonstrated (Rossini et al., 2001; Schwid et al., 1997).
Improvement of fatigue score in MS patients by Prokarin, which is a proprietary
blend of histamine and caffeine, has been found in a placebo controlled study
24
(Gillson et al., 2002). Metz et al. have provided evidence that MS fatigue may be
improved with immune modulating treatment with either glatiramer acetate (GA) or
interferon beta (IFN-β), shown by improved total fatigue impact scale (FIS) scores
(Metz et al., 2004).
1.1.8 Autonomic dysfunction in multiple sclerosis
Autonomic dysfunction (AD) in people with MS is well documented, but, the
significance of these abnormalities and the relationship to clinical characteristics is
not yet established (Flachenecker et al., 2003; Merkelbach et al., 2001).
AD particularly affects the bladder, bowel, cardiovascular function, sleep, sexual and
sweat glands. This may be clinically evident such as bladder disturbances or may
be subclinical, when abnormal sympathetic skin response (SSR) or decreased heart
rate variation is estimated (Linden et al., 1995; Linden et al., 1997).
AD has an important impact on the disability in MS patients and is considered as
one of the crucial components that have an impact on the QoL outcomes in these
patients.
The pathophysiology behind AD remains unclear but plaques located adjacent to the
pathways significant for autonomic function in the hypothalamus involving fornix,
anterior commissure, internal capsule, optic system and spinal cord might be the
basis for autonomic disturbances in MS patients (Huitinga et al., 2001). It has been
suggested that demyelination may disrupt the central autonomic network in the
insular, anterior cingulate and ventromedial prefrontal cortices, central nucleus of
the amygdala, paraventricular hypothalamus and the medulla or interfere with the
descending autonomic nervous system (ANS) pathways during their course in the
brainstem or spinal cord (Vita et al., 1993).
The autonomic nerve activity is not assessed directly, but the response of the
effector organs can be measured. Electrophysiological evaluations for assessing AD
25
in MS patients has been established as a diagnostic tool for AD, some studies have
suggested the use of some self-completed questionnaires on the symptoms of
patients with AD (Flachenecker et al., 2001; Nasseri et al., 1999).
AD may not only be a consequence of the disease but may also in itself play a
pathogenetic role; evidence from animal and clinical studies suggest interactions
between the immune system and the ANS (Chelmicka-Schorr and Arnason, 1994;
Zoukos et al., 1994).
1.1.9 Diagnosis of Multiple Sclerosis
Diagnosing MS is complex and sometimes lengthy process. Clinical findings and
supporting evidence from supplementary tests, such as MRI of the brain, CSF
examination, and clinical neurophysiology are the bases for diagnosis of MS.
Clinical ground is a cornerstone for diagnosis of MS. MRI has become a valuable
test for confirming the probable cases of MS. The diagnosis depends on detection of
lesions which are disseminated in time and space. CSF examination is used for
detection of OCBs and IgG level in the CSF. Finally the clinical neurophysiology has
a role in supporting the diagnosis especially the visual and somatosensory evoked
potentials, which are helpful in identifying additional, silent lesions (Polman et al.,
2005b).
1.1.9.1 Diagnostic Criteria for Multiple Sclerosis
1.1.9.1.1 The Poser criteria
In 1983 Poser with his colleagues established a new diagnostic criteria for MS
(Poser et al., 1983):
Clinically definite MS
2 attacks and clinical evidence of 2 separate lesions
2 attacks, clinical evidence of one and paraclinical evidence of another separate
lesion
26
Laboratory supported Definite MS
2 attacks, either clinical or paraclinical evidence of 1 lesion, and CSF immunologic
abnormalities
1 attack, clinical evidence of 2 separate lesions & CSF abnormalities
1 attack, clinical evidence of 1 and paraclinical evidence of another separate lesion,
and CSF abnormalities
Clinically probable MS
2 attacks and clinical evidence of 1 lesion
1 attack and clinical evidence of 2 separate lesions
1 attack, clinical evidence of 1 lesion, and paraclinical evidence of another separate
lesion
Laboratory supported probable MS
2 attacks and CSF abnormalities
1.1.9.1.2 The McDonald criteria
In 2001 an international panel in association with the National Multiple Sclerosis
Society of America recommended revised Diagnostic criteria of MS. They make use
of advances in MRI imaging techniques in order to facilitate in diagnosis of MS and
using MS, possible MS or not MS (McDonald et al., 2001). Currently, McDonald
criteria are regarded as the gold standard for MS diagnosis (Appendix 2).
1.1.9.1.2.1 Revised 2005
The McDonald criteria were revised in 2005 to clarify some terms such as exactly
what is meant by an "attack," "dissemination," a "positive MRI," etc. (Polman et al.,
2005b) (Appendix 4).
27
1.1.9.1.2.2 Revised 2010
In 2010, the International Panel on Diagnosis of MS revised the McDonald
diagnostic criteria. This revision had simplified the demonstration of CNS lesions in
space and time by MRI techniques and made the criteria for all people including
the non-Western Caucasian populations (Polman et al., 2011) (Appendix 5).
1.1.10 Treatment of Multiple Sclerosis
MS is a progressive disease that has no cure. Treatment categories are:
Acute treatment
Disease-modifying therapies
Combination therapies
Investigational Therapies
Symptomatic therapy,
Neuroprotective agents
1.1.10.1 Acute Treatment
Treatment of acute attacks will shorten the duration and possibly decrease the
severity of the attack.
Corticosteroids: Corticosteroids are a mainstay of treatment for acute exacerbations
associated with MS. The most commonly used corticosteroids are
methylprednisolone and prednisone. There are several potential modes of action,
which include reducing oedema, stabilising the BBB, decreasing pro-inflammatory
cytokines, and T cell apoptosis (Gold et al., 2001).
Plasmapheresis: Patients have been treated with plasmapheresis for acute, severe
attacks, were reported to exhibit moderate or marked functional improvement after
the initial treatment. In cases steroids are contraindicated or not effective, plasma
28
exchange can be an alternative for short term use in severe attacks (Meca-Lallana
et al., 2003; Weinshenker, 2001).
The 2011 American Academy of Neurology (AAN) guideline confirms that
plasmapheresis is probably effective in relapsing forms of MS as second-line
treatment for exacerbations that resist steroid treatment.
1.1.10.2 Approved disease modifying therapies
The disease modifying therapies (DMT) for MS currently approved for use in
relapsing forms of MS include the following:
Interferons (INF): INFs are natural proteins that are produced by the body in
response to infectious stimuli. They were first described in 1957. Based on the type
of receptor through which they signal, human INFs have been classified into three
major types: INF type I; bind to a specific cell surface receptor complex known as
the INF- α receptor. The type I INFs present in humans are IFN-α, INF-β and IFN-ω.
INF type II: Binds to interferon-gamma receptor. In humans this is IFN-γ. INF type
III: Signal through interleukin 28 receptor, alpha subunit (Papatriantafyllou, 2013).
The INF currently approved for treatment of MS are INF β-1b (Betaseron, Extavia)
and INF β-1a (Rebif1, Avonex). INF β has been shown to inhibit T-cell activation and
reduce BBB permeability to inflammatory cells, Pivotal phase III studies of INF β
have all demonstrated a significant reduction in relapse rate and improvement in
MRI measures of disease activity in RRMS (Ebers, 1998; Jacobs et al., 1996; The
Ifnb Multiple Sclerosis Study Group, 1993).
Glatiramer Acetate (GA) : In experimental models, the immunomodulatory
mechanism of action for GA involves binding to MHC molecules and consequent
competition with various myelin antigens for their presentation to T cells (Arnon and
Aharoni, 2004). In addition, GA is a potent inducer of specific T helper 2 type
29
suppressor cells that migrate to the brain and lead to bystander suppression; these
cells also express anti-inflammatory cytokines.
The benefit of GA was first established in patients with RRMS. Two placebo-
controlled studies have shown that GA significantly lowered relapse rate (Johnson et
al., 1995), and significantly reduced disability progression (Johnson et al., 1998) as
compared to the placebo. Another trial found that GA treatment led to a significant
reduction in the number of new T2 lesions on brain MRI (Comi et al., 2001a). A
recent head-to-head comparison trial (Betaferon ⁄ Betaseron vs. GA) (O'Connor et
al., 2009) has shown largely similar efficacy between the INF β treatments and GA.
Mitoxantrone (Novantrone): Mitoxantrone is an antineoplastic drug. USA food and
drug administration (FDA) has approved this drug for patients suffering from
secondary-progressive, progressive-relapsing, or worsening RRMS. Mitoxantrone
decreases proinflammatory cytokines, augments suppressor cell function and
decreases the migration of T cells into the CNS by suppressing the activity of T
cells, B cells, and macrophages. In Progressive MS mitoxantrone can alter the
disease course and also suggested benefit in the treatment in RRMS (Mahdavian et
al., 2010).
Natalizumab (Tysabri): Natalizumab is a monoclonal antibody against the cell
adhesion molecule α4-integrin. It inhibits the migration of T and B cells into the CNS,
resulting in a reduction of inflammatory demyelinating lesions. Natalizumab can
slow the disease progression and decreases the number of relapses (Mahdavian et
al., 2010). An uncommon, but potentially deadly, side effect of treatment of MS
patients with natalizumab is the development of progressive multifocal
leukoencephalopathy (PML). Clinically, PML manifests with subacute progressive
cognitive decline and focal neurological deficits, and it is usually fatal (Sahraian et
al., 2012).
30
Fingolimod: In 2010, fingolimod became the first oral DMT approved for treatment of
MS and categorized in a new class called sphingosine 1-phosphate receptor (S1P-
R) modulators. This S1P-R modulator deprives T cells of the signal they need to
leave lymph nodes, thus inhibiting them from circulating and entering the brain.
Studies have found that fingolimod reduces the number of lesions detected on MRI
and clinical disease activity in patients with MS (Kappos et al., 2006a; Mahdavian et
al., 2010).
Teriflunomid: Teriflunomide is an oral reversible inhibitor of dihydroorotate
dehydrogenase (DHODH), a mitochondrial membrane protein essential for
pyrimidine synthesis (Palmer, 2010). DHODH blocks de-novo pyrimidine synthesis
leading to an inhibition of the proliferation of autoreactive B and T cells. In the
presence of teriflunomide, replication of hematopoietic and memory cells is
preserved through metabolism of the existing pyrimidine pool. Teriflunomide has
been shown to have modulation of immunoglobulin class switching, IL-2 production,
and IL-2 receptor expression (Siemasko et al., 1996)
Teriflunomide was compared with placebo in a phase II trial in patients with RRMS
and SPMS still experiencing relapses. Patients who received teriflunomide had
significantly fewer T1-enhancing lesions or new or enlarging T2 lesions than those
treated with placebo. Patients receiving teriflunomide had significantly reduced T2
disease burden. The proportion of patients with increased disability by EDSS at 36
weeks was significantly lower with teriflunomide compared with placebo (O'Connor
et al., 2006)
Two phase II studies evaluated teriflunomide as adjunctive therapy in persons with
MS (Freedman et al., 2010; Freedman et al., 2009). In these studies, patients
receiving glatiramer acetate or a β-IFN were randomised to add placebo or
teriflunomide their current therapy. In both studies, teriflunomide had good safety
and tolerability and was associated with improved disease control manifested as
31
reduced number and volume of T1 gadolinium-enhancing lesions, compared with
placebo.
Results from the phase III, TEMSO study demonstrated significant reduction in
annualized relapse rate (ARR) and disability progression with teriflunomide
compared with placebo (Miller et al., 2012). The numbers of gadolinium-enhancing
T1 lesions and unique active lesions per scan were also reduced with teriflunomide
vs. placebo (Nelson et al., 2011).
Teriflunomide is also being evaluated as an adjunctive therapy in combination with
IFN-β in the phase III, TERACLES study, with estimated completion in 2014. Two
additional studies are underway; TOWER and TENERE are monotherapy studies
comparing teriflunomide with placebo and IFN-β-1a subcutaneous, respectively
(ClinicalTrials.gov., 2011a). TOPIC is an ongoing phase 3 trial evaluating the
efficacy and safety of once daily teriflunomide vs. placebo in patients with clinically
isolated syndrome (ClinicalTrials.gov., 2011b).
BG-12 (Dimethyl Fumarate): BG-12 is a fumaric acid ester with immunomodulatory
properties. BG-12 has demonstrated benefits in animal models of EAE. Fumaric
acid esters may decrease leukocyte passage through the BBB and exert
neuroprotective properties by the activation of antioxidative pathways(Lee et al.,
2008).
DEFINE was a phase III, placebo-controlled, comparative study of BG-12 in RRMS
patients (Gold et al., 2012). Patients with were randomised to BG-12 at two different
doses dose or to placebo. Both BG-12 doses were associated with a significant
decrease in the proportion of patients who relapsed at 2 years compared with
placebo. Both BG-12 doses were significantly superior to placebo in reducing ARR,
the number of new or newly enlarging T2 hyperintense lesions, and the number of
new gadolinium-enhancing lesions. BG-12 was also superior to placebo in slowing
32
the rate of disability progression as measured by EDSS scores at 2 years (Gold et
al., 2012).
CONFIRM was a phase III, study, investigated the efficacy and safety of oral BG-12,
at two different doses, as compared with placebo in patients with RRS. An active
agent, glatiramer acetate, was also included as a reference comparator. In patients
with RRMS, BG-12 (at both doses) and glatiramer acetate significantly reduced
relapse rates and improved neuroradiologic outcomes relative to placebo (Fox et al.,
2012).
1.1.10.3 Combination therapies
The combinations of intravenous (IV) methylprednisolone and methotrexate with
intramuscular (IM) INF β-1a have been tested in clinical trials (Cohen et al., 2008;
Cohen et al., 2009). The results have revealed a non-significant trends favouring IV
methylprednisolone for new or enlarging T2-hyperintense lesions, gadolinium-
enhancing lesions, relapse rate, EDSS change, MSFC score change, and a
combined measure of clinical and MRI disease activity. The benefit of methotrexate
has not been suggested.
The results of another clinical trial of oral methylprednisolone as add-on therapy to
INF β-1a for the treatment of RRMS (Sorensen et al., 2009) have suggested that the
mean yearly relapse rate was less in the methylprednisolone group compared to the
placebo group while EDSS progression was not significantly different between the
two groups.
Calabresi et al (2002) in a pilot study have investigated the effect of adding
methotrexate to INF β-1a in MS patients. They found a significant reduction in
gadolinium-enhancing lesion number and mean relapse number. A similar open-
label trial had investigated the combination of azathioprine with INF β-1a (Pulicken
et al., 2005) found a reduction in gadolinium-enhancing MRI lesions.
33
Combination of IM INF β-1a with IV natalizumab has been studied in a clinical trial
where it was found that the risks of relapse, number of new or enlarging T2-
hyperintense lesions, and mean number of gadolinium-enhancing lesions were
significantly lower in the natalizumab group than in the placebo group. This was a
largest combination trial so far and data from this trial indicated a clear advantage of
natalizumab plus intramuscular INF β-1a over intramuscular INF β-1a alone on
clinical and imaging endpoints (Rudick et al., 2006).
A monthly infusion of IV natalizumab along with GA has been evaluated in a clinical
trial. The results have indicated that combination group had superiority compared to
placebo group for the primary and most secondary imaging outcomes (Goodman et
al., 2009).
In some pilot studies a short course of mitoxantrone is used to induce
immunosuppression, followed by immunomodulation with first-line drugs such as
INF-β or GA. These trials have suggested a promising findings with both drugs
[reviewed in (Boggild, 2009)].
Despite evidence from many preliminary studies that lends support to the safety,
tolerability, and efficacy of several combination regimens, many of larger trials of
these combinations have yielded negative or conflicting results. Combination
therapy remains an attractive option in MS treatment, however, the neuroprotection
strategy in MS was rarely studied. The future efforts should focus on combining anti-
inflammatory and neuroprotective or reparative strategies.
Investigation therapies in multiple sclerosis
Research into additional treatment options continues to advance. Multiple
approaches are being investigated based on the increasing knowledge about
immune system abnormalities and CNS lesion formation in MS. These include
34
approaches to counteract or reduce immune system activation, BBB disruption,
neuronal loss and myelin loss.
The development of new pharmacologic agents for the treatment of MS has led to
changes in the treatment of MS. To date, six drugs have entered or completed
phases II and III clinical trials. These include laquinimod, alemtuzumab, daclizumab,
rituximab, ocrelizumab and ofatumumab.
MS requires lifelong DMTs, and all of the currently available first-line DMTs are
parenteral formulations only. As the advent of new oral drugs will lead to increased
patient compliance and contribute to longer sustain symptom-free periods and less
marked disability.
Recent approval of fingolimod, teriflunomide and dimethyl fumarate, as the oral
drugs to treat MS has marked a new frontier in the treatment of MS. Their entry onto
the market will provide additional treatment options.
Laquinimod: Laquinimod is an immunomodulator has been shown to promote anti-
inflammatory cytokine profiles in human peripheral blood mononuclear cells. In EAE
models, laquinimod had effectively reduced inflammation, demyelination, and axonal
damage (Bruck and Wegner, 2011).
Laquinimod has been evaluated in phase III trials, in patients with RRMS who were
randomised to receive laquinimod or placebo. Laquinimod treatment resulted in
reduction in ARR vs. placebo and decrease in the risk for disability progression, as
measured by EDSS. Treatment with laquinimod was also associated with reduction
in progression of brain atrophy vs. placebo (Comi, 2013).
In the second phase III study, laquinimod was compared with placebo in patients
with RRMS. Laquinimod was associated with a statistically significant reduction of
ARR , risk of disability progression and of brain volume loss compared with placebo
(Consortium of Multiple Sclerosis Centers, 2011)..
35
Alemtuzumab: Alemtuzumab is a humanised monoclonal antibody against CD52, a
glycoprotein antigen found on the surface of mature lymphocytes and monocytes.
The exact biological function of CD52 remains unclear but some evidence suggests
that it may be involved in T-cell migration (Watanabe et al., 2006). Alemtuzumab
has also been shown to induce production of neurotrophic factors in reconstituted
autoreactive T cells (Jones et al., 2010).
Efficacy of alemtuzumab for the treatment of MS has been assessed through a
number of clinical trials. In a Phase II study (CAMMS223), Treatment with
alemtuzumab was associated with a significant reduction of annualized relapse rate
compared to IFN-β-1a as well as significantly decreased T2-weighted lesion burden
than IFN-β-1a Patients who were treated with alemtuzumab experienced a
significantly lower rate of sustained disability accumulation versus IFN-β-1a as
evidenced by improvements of the EDSS score (Coles, 2008).
Two Phase III studies [CARE-MS I and CARE-MS II (Cohen et al., 2012; Coles et
al., 2012) evaluated the safety and efficacy of alemtuzumab compared with INF-β in
patients with RRMS. In both studies, a significant reduction in relapse rate
compared with interferon-beta 1a was observed. In one of the trials, a significant
reduction in disease progression compared with interferon-beta 1a was also seen.
Daclizumab: Daclizumab is a humanised monoclonal antibody directed against the
high-affinity IL-2 receptor. This receptor is present on activated, but not resting, T
cells. Binding of IL-2 to this receptor is necessary for clonal expansion and
continued viability of activated T cells (Vincenti et al., 1998) .
Daclizumab was evaluated for the treatment of RRMS in the phase II CHOICE trial.
It was a placebo-controlled study in patients with active disease despite IFN-β
treatment. Patients were randomised to receive two different subcutaneous doses of
daclizumab or placebo as an adjunct to their current IFN-β therapy. The mean
number of new or enlarged gadolinium-enhancing lesions was 4.75 in the IFN-β–
36
placebo group vs. 1.32 for patients who received IFN-β with high-dose daclizumab
and 3.58 for those treated with IFN-β with low-dose daclizumab) (Wynn et al., 2010).
SELECT is a phase II clinical trial that evaluated two doses of daclizumab in patients
with RRMS (Business Wire., 2011). At 1 year, daclizumab was associated with
significant reductions in ARR for both dose groups,
Daclizumab is also being compared with i.m. IFN-β-1a in a phase III study in
patients with RRMS (ClinicalTrials. gov., 2011).
Rituximab: Rituximab is a chimeric monoclonal antibody that depletes CD20-positive
B cells through cell-mediated and complement-dependent cytotoxic effects and
promotion of apoptosis. A phase II clinical trial assessed efficacy of rituximab in
patient with RRMS, rituximab treatment resulted in significantly decreased numbers
of gadolinium-enhancing lesions vs. placebo as well as a significantly decreased risk
for relapse (Hauser et al., 2008). The results of a phase II/III placebo–controlled trial
in PPMS revealed no significant difference in the time to confirmed disease
progression between the placebo and rituximab groups (Hawker et al., 2009).
Ocrelizumab: Ocrelizumab is a humanised anti-CD20 monoclonal antibody that
results in B cell depletion. It has been evaluated in patients with RRMS who were
randomised to treatment with i.v. ocrelizumab and i.m. IFN-β-1a or placebo. The
mean number of gadolinium-enhancing lesions was reduced in the treated group
compared to placebo (Kappos et al., 2011).
Ocrelizumab is also being evaluated in phase III, placebo-controlled trial in patients
with PPMS (Montalban et al., 2011). The primary outcome measure of this trial is
time to onset of sustained disability progression (Hauser et al., 2008). Two large
global studies will compare ocrelizumab with IFN-β-1a subcutaneous (OPERA I and
II) in patients with RRMS (Clinical Trials. gov., 2012).
37
Ofatumumab: Ofatumumab is a third anti-CD20 antibody being developed for the
treatment of MS. A phase II safety and pharmacokinetics study in with RRMS
indicated no dose-limiting toxicities and no unexpected safety findings. Active
treatment also resulted in significant reductions in the number of gadolinium-
enhancing T1 lesions and new/enlarging T2 lesions in patients treated with
ofatumumab vs. placebo (Genmab., 2011).
1.1.10.4 Symptomatic Treatment
Symptomatic treatment is an essential component of the management of MS. The
aims of symptomatic treatment are ; elimination or reduction of symptoms impairing
the functional abilities and QoL of the affected patients and avoiding development
of a secondary physical impairment due to an existing disease effects. Many
therapeutic techniques as well as different pharmacological agents have tried for the
treatment of MS symptoms (Table1.2).
Table1.2 Symptomatic treatments for multiple sclerosis. Source: (Compston
and Coles, 2008)
Symptoms
Signs
Treatment
Established
efficacy
Equivocal
efficacy
Speculative
Cognitive
impairment
Deficits in
attention,
reasoning, and
executive function
(early); dementia
(late)
Cognitive
training
Hemisensory
and motor
Upper motor
neuron signs
Affective
(mainly
depression)
Antidepressant
drugs
Epilepsy (rare)
Anticonvulsants
Focal cortical
deficits (rare)
Unilateral
painful loss of
Scotoma, reduced
visual acuity,
Low vision aids
38
vision
colour vision, and
relative afferent
pupillary defect
Tremor
Postural and
action tremor,
dysarthria
Carbamazepine
, B.blockers,
clonazepam,
thalamomectom
y,and thalamic
stimulation
Clumsiness
and poor
balance
Limb
incoordination
and gait ataxia
Diplopia,
oscillopsia
Nystagmus,
internuclear
ophthalmoplegias
Baclofen,
gabapentin
Vertigo
Prochloperazine
, cinnarazine
Impaired
swallowing
Dysarthria
Anticholinergic
drugs
Speech therapy
Impaired
speech
Pseudobulbar
palsy
Tricyclic
antidepressants
Speech therapy
Paroxysmal
symptoms
Carbamazepine,
gabapentin
Weakness
Upper motor
neuron signs
Stiffness and
painful
spasms
Spasticity
Tizanidine,
baclofen,dantrol
-ene,
benzodiazepine,
intrathecal
baclofen
Botulinum toxin,
corticosteroids
Cannabinoids
Bladder
dysfunction
Anticholinergic
drugs and/or
intermittent self-
catheterisation,
Decompressing,
intrvesical
botulinum toxin
Abdominal
vibration,
cranberry juice
Erectile
impotence
Sildenafil
Constipation
Bulk laxatives,
enema
Pain
Carbamazepine,
gabapentin
Tricyclic
antidepressant
drugs,
transcutaneous
electrical nerve
stimulation
Fatigue
Amantadine
Modafinil
Pemoline,
fluoxetine
Temperature
sensitivity and
exercise
intolerance
Cooling suit, 4-
aminopyridine
39
1.1.10.5 Neuroprotective agents
There is increasing evidence that degenerative mechanisms are present in all the
progressive forms of MS. Therefore restorative therapies that improve function of
damaged neural pathways, as well as neuroprotective and repair strategies, will be
necessary.
There are several agents which may show promise as potential neuroprotective
therapies that could prevent axonal and neuronal damage either directly or indirectly
after CNS insults. These include:
Disease modifying therapies: Results from several studies have suggested
improvement disability outcomes in DMTs treated patients but the actual benefit of
long term treatment in the later stages of the disease is unclear (Van der Walt et al.,
2010). Importantly, the available DMTs are only partially effective in preventing the
onset of permanent disability in MS patient (Trojano et al., 2007). The current
existing DMTs predominantly target the recruitment of systemic immune responses
and, as such, they would not be expected to modulate significantly the pathogenesis
of axonal degeneration once it is established.
Growth Factors:
Insulin-like growth factor-1 (IGF-1): IGF-1 promotes oligodendrocytes
growth and maturation (McMorris and McKinnon, 1996) and also enhances
neuronal development (Ozdinler and Macklis, 2006). The results of studies of
IGF-1 in EAE models are conflicting, initial studies showed an improvement
in disability in acute and chronic demyelinating EAE (Li et al., 1998; Yao et
al., 1996). Subsequent studies showed a transient effect only (Cannella et
al., 2000), or failed to show a sustained benefit of IGF-1 in EAE (Genoud et
al., 2005). In a pilot study IGF-1 had administered to few patients with
40
SPMS showed no improvement in the primary MRI endpoints, including new
enhancing lesions, WM lesion load (Frank et al., 2002).
Erythropoietin: Erythropoietin is a haematopoietic growth factor. Its anti-
inflammatory and neuroprotection effects has been established in
experiments in different models of EAE (Agnello et al., 2002). Both Li et al
and Diem et al have found that Erythropoietin decrease in axonal loss in
EAE compared to controls (Diem et al., 2005; Li et al., 1998). In an open-
label pilot study Erythropoietin has been tested in humans suffering from
chronic progressive MS. Clinical and neurophysiological improvement of
motor function and cognitive performance was reported (Ehrenreich et al.,
2007)
The neuropoietic cytokines (leukaemia-inhibitory factor (LIF) and ciliary
neurotrophic factor (CNTF): There is a large amount of evidence to suggest
that LIF and CNTF enhance neuronal survival in the context of axonal injury
(Hagg et al., 1993). It has been found that survival after axotomy (transaction
of the axon from the nerve cell body) can be improved in new born rats by
the administration of either LIF (Hughes et al., 1993) or CNTF (Sendtner et
al., 1992). Recent work highlights the fact that CNTF exerts more robust
effects on neuronal survival and growth when applied in combination with its
soluble form of CNTF-receptor- α (Ozog et al., 2008).
Sodium Channel Blockers: Evidence from EAE studies has shown a beneficial effect
of sodium-blocking agents, such as flecainide and lamotrigine to improve axonal
survival and decrease disability (Bechtold et al., 2004; Bechtold et al., 2006). In
contrast, in a phase II placebo-controlled clinical trial Kapoor et al have not found
neuroprotective effects of treatment with lamotrigine in SPMS (Kapoor et al., 2008).
Calcium Channel Blockers: A study suggested that calcium channel blockers
prevent axonal loss and disability in treated EAE animals (Brand-Schieber and
41
Werner, 2004). Another study has suggested a possible neuroprotective effect of
some of the calcium channel blockers such as nimodipine, nifedipine and ryanodine
(Ouardouz et al., 2009).
Mesenchymal Stem Cells: Autologous bone marrow (ABM) derived mesenchymal
stem cell (MSC) can promote neuroprotection by inhibiting gliosis, apoptosis, and
stimulate local progenitor cells (Yang et al., 2009). Evidence shows a specific
immunomodulatory effect of MSCs through inhibition of T and B cells and
maturation of antigen presenting cells (Uccelli et al., 2006). On the other hand
several EAE experiments have shown that treatment with ABM derived MSC
significantly improved clinical outcomes (Bai et al., 2009; Gordon et al., 2008; Kassis
et al., 2008; Zappia et al., 2005; Zhang et al., 2005).
Glutamate Antagonists: Treatment with glutamate antagonist in EAE result in
substantial amelioration of disease, increased oligodendrocyte survival and reduced
neurofilament H, an indicator of axonal damage (Pitt et al., 2000). Memantine, a N-
methyl-D-aspartate antagonist has shown amelioration of disability in EAE
(Wallstrom et al., 1996). Kalkers et al. has assessed the effect of riluzole in a small
cohort of PPMS patients in an open-label study. The study revealed a nonsignificant
reduction in the rate of cervical cord atrophy and decrease in the development of T1
hypointense lesions (Kalkers et al., 2002).
HMG-CoA Reductase Inhibitors (Statins): Evidence of neuroprotection due to statin
therapy in preclinical studies has been demonstrated in several studies through a
possible reduction in axonal loss (Neuhaus and Hartung, 2007; Paintlia et al., 2009;
Sena et al., 2003; Youssef et al., 2002). Available clinical data regarding statins in
the treatment of MS are not entirely consistent. Most of the studies have showed no
benefit (Rudick et al., 2009; Sorensen et al., 2011; Wang et al., 2011). In contrast, a
study enrolled 30 patients with active RRMS (Vollmer et al., 2004) has showed a
42
significant decrease in the number of gadolinium-enhancing lesions in brain MRI
scans compared with pre-treatment brain MRI scans.
Cannabinoids: Cannabis is used by MS patients for relief from a variety of
symptoms (Clark et al., 2004). In vitro studies have suggested effect of
cannabinoids on several potential mechanisms of axonal injury, including glutamate
release (Fujiwara and Egashira, 2004), oxidative free radicals as well as damaging
calcium flux (Kreitzer and Regehr, 2001). Which, in excess, can cause neuronal
death in neuroinflammatory disease (Kapoor et al., 2003; Pitt et al., 2000).
Furthermore, exogenous agonists of the cannabinoid CB1-receptor have possible
neuroprotective effects in EAE animal models (Pryce et al., 2003). Subsequent
clinical studies on cannabinoids for symptomatic treatment of MS (Rog et al., 2005;
Zajicek et al., 2003; Zajicek et al., 2012) , and understanding of the biology of
cannabis shows that cannabis signals to an endogenous cannabinoid system via
cannabinoid receptors which can regulate neurotransmission and cell death
pathways (Howlett et al., 2002). Despite these promising results, neuroprotective
effects in MS by cannabinoids and the modulation of the endocannabinoid system
must still be established.
Modafinil: Modafinil is a wakefulness-promoting agent. Besides the already FDA
approved uses, modafinil also has potential non-approved clinical uses which some
of them increasingly believed to be neuroprotection. Modafinil prevents glutamate
toxicity in cultured cortical cells (Antonelli et al., 1998). Another study conducted on
rat had suggested that modafinil can decrease toxic aspartate and glutamate levels
after striatal ischemic injury caused by endothelin-1 (Ueki et al., 1993a).
Furthermore, it was found that modafinil can prevents development of lesions in the
hippocampus induced by the neurotoxic nerve gas soman (Lallement et al., 1997).
Clinically, in a recent retrospective study, we suggest that modafinil significantly
reduces the disease severity in MS measured by EDSS score (Bibani et al., 2012).
43
The neuroprotective potential of modafinil has been tested in other
neurodegenerative diseases, in particular Parkinson‘s disease (PD). The result of
some studies found that modafinil could prevent degeneration of the nigrostriatal
pathway in1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induced PD models
and mechanical injury of the nigrostriatal pathway (Fuxe et al., 1992; Jenner et al.,
2000; van Vliet et al., 2006; Xiao et al., 2004). Modafinil will be discussed further in
this thesis.
1.1.11 Prognosis and complications of Multiple Sclerosis
The clinical subtype of the disease; the individual's sex, race, age, and initial
symptoms; and the degree of disability the person experiences were shown to have
contribution with the expected future course of MS. Individuals with progressive
subtypes of MS, particularly the primary progressive subtype, have a more rapid
decline in physical and mental functions. Older individuals when diagnosed are
more likely to experience a chronic progressive course, with more rapid progression
of disability. Females generally have a better prognosis than males. Initial MS
symptoms of sensory problems or visual symptoms, are predictors for a relatively
good prognosis, whereas motor problems are markers for a relatively poor
prognosis. Better outcomes are also associated with the presence of only a single
symptom at onset.
The degree of disability varies among individuals with MS. In general, one of three
individuals will still be able to work after 15–20 years. 15% of people diagnosed with
MS never have a second relapse, and these people have minimal or no disability
after ten years (Pittock et al., 2004).
The life expectancy of people with MS is 5 to 10 years lower than that of unaffected
people and two-thirds of the deaths in people with MS are directly related to the
consequences of the disease (Compston and Coles, 2008). Despite improvement in
management of MS, along with some successful treatment infection such as
44
pneumonia and urinary tract infection are common complications of MS. The risk of
suicide is common in MS patients. Young patients are the most likely victim
(Sadovnick et al., 1992).
Interestingly, it has been found that deaths from malignancy are less common than
in age-matched controls (Sadovnick et al., 1991).
Higher EDSS scores are associated with increased mortality. Median time from
disease onset to reaching a disability level when one needs a walking-aid is about
20 years (Confavreux et al., 2003; Myhr et al., 2001; Phadke, 1987). MS has heavy
economic and personal burden. The costs are highly correlated with disease
severity (Kobelt et al., 2006; Parkin et al., 2000).
45
1.2 EXPERIMENTAL AUTOIMMUNE ENCEPHALOMYELITIS AND MULTIPLE
SCLEROSIS
In the second decade of 20th century, Experimental autoimmune encephalomyelitis
(EAE) was first described by Koritschoner and Schweinburg (Koritschoner and
Schweinburg, 1925),They induced spinal cord inflammation in rabbits by inoculation
with human spinal cord. Subsequently, researchers have attempted to reproduce
the encephalitic complications associated with rabies vaccination by repetitive
immunisation of rhesus monkeys with CNS tissue (Rivers et al., 1933). Moreover,
studies have showed that EAE can be elicited in many different species, including
rodents and primates, and from these studies it became clear that EAE can
reproduce many of the clinical, neuropathological and immunological aspects of the
neurodegenerative disease including MS (Hohlfeld and Wekerle, 2001). At the
present time, EAE studies have provided an insight into general neuroscience and
immunology concepts, and developing general therapeutic strategies.
1.2.1 EAE induction
EAE can be induced in susceptible strains of different species by sensitisation with
CNS myelin antigens (Active EAE) (Williams et al., 1994), or by the adoptive transfer
of CNS myelin antigen-specific CD4+ T cells into naive syngeneic recipients
(Passive EAE) (Pender, 1995; Pettinelli and McFarlin, 1981).
1.2.2 Pathogenesis of EAE
There is substantial evidence that the initiating effector lymphocyte in EAE
pathogenesis is the autoreactive CNS specific CD4+ T cell; however, it has been
found that myelin specific CD8+ T cells also play a role in EAE pathogenesis
[reviewed in (Goverman et al., 2005)].
46
Besides T cells the role of B cells in EAE pathogenesis has been demonstrated
(Wolf et al., 1996). However, the role of B cell and its contributions to EAE
pathogenesis appears to be contradictory and may reflect the involvement of
multiple roles for B cells or different B cell subsets during disease pathogenesis
(Bouaziz et al., 2008).
1.2.3 Clinical scores of EAE
In the classic EAE model, animals develop an ascending flaccid paralysis, which –
depending on disease severity – The clinical scoring scale is as follows; 0—healthy,
1—flaccid tail, 2—impaired righting reflex and/or impaired gait, 3—partial hind-leg
paralysis, 4—total hind-leg paralysis, 5—any sign of front-leg paralysis, and 6—
moribund/ dead (O'Brien et al., 2010).
1.2.4 EAE and multiple sclerosis
EAE is primarily used as an animal model of autoimmune inflammatory diseases of
the CNS. It has become a well characterised model for organ-specific autoimmune
disease in general. EAE contributed enormously to our understanding of
autoimmunity, neuroinflammation, cytokine biology and immunogenetics, and the
development of MS therapeutics.
Mice and rats have been used commonly for EAE. In addition certain monkey
species like marmosets are used for specific questions that cannot be easily
assessed in rodents (t Hart et al., 2011). Most studies are presently done using
C57BL/6 mice, where disease is induced by immunisation with myelin
oligodendrocyte glycoprotein (MOG) peptide, representing residues 35–55,
emulsified in Freund's adjuvant that is supplemented with Mycobacterium
tuberculosis extract. This protocol is used because it works reproducibly and
because it allows one to take advantage of the wealth of genetic resources on the
C57BL/6 background. There are some limitations of this protocol when translated to
47
the MS: In most cases, the C57BL/6 model of EAE is monophasic, without
relapses; the T cell component is predominantly CD4+; and spinal cord is affected
out of proportion to brain.
1.2.5 EAE and multiple sclerosis treatments
EAE has contributed to the development, validation, and testing of MS drugs. One
major MS treatment (Natalizumb) came directly, in a mechanism-based fashion,
from EAE research (Polman et al., 2006). EAE has also played a successful role in
assurance of the currently licensed and used DMT: IFN-beta (Abreu, 1982) GA
(Johnson et al., 1995; Teitelbaum et al., 1971) and the anti-VLA-4 antibody (Polman
et al., 2006). A substantial number of other studies have shown treatment success
with concordant results in EAE and MS, using a variety of compounds. Some of
these agents, like azathioprine, mitoxantrone and fingolimod are licenced or well-
established therapies for specific groups of patients with MS. Others, like laquinimod
have reached late phase clinical trials [reviewed in (Constantinescu et al., 2011a)].
However, numerous other therapeutics that showed promise in EAE were found to
be ineffective or detrimental in MS (Denic et al., 2011).
1.2.6 Major differences between EAE and multiple sclerosis
Beside all promising achievement from EAE studies in relation to MS, there are
differences in aetiopathogenesis, immunohistopathology, and genetic components
between EAE and MS. To reduce the gap between EAE and MS creating new and
refined EAE models in humanized mice or perhaps by switching to species more
closely related to humans, such as the common marmoset (Callithrix jacchus). The
MS-like disease phenotype of marmoset EAE is particularly useful to investigate
treatment approaches in relapsing-remitting and chronic forms of MS (t Hart et al.,
2011).
48
1.3 MODAFINIL (PROVIGIL)
1.3.1 Introduction
In the late 1970s scientists working with the French pharmaceutical company Lafon
have generated a novel wake-promoting agent known as Adrafinil. In the early
1990s the primary metabolite of Adranifil, Modafinil, was derived which had similar
activity. Modafinil has been prescribed in France since 1994 under the name
Modiodal, and in the US since 1998 as Provigil. Its approval for use in the UK was in
December 2002. In 1998 modafinil was approved by the United State food and drug
administration (USFDA) for excessive daytime sleepiness (EDS) associated with
narcolepsy. Almost a decade later evidence emerged showing its effectiveness in
treating several sleep disorders (Ballon and Feife, 2006). Modafinil was approved by
USFDA in 2004 for the treatment of obstructive sleep apnoea/hypopnoea (OSA)
syndrome, and shift work sleep disorder (SWSD) (Minzenberg and Carter, 2007).
Modafinil that is chemically and pharmacologically different from other central
nervous system (CNS) stimulants (Saper and Scammell, 2004) has a large potential
for many uses in psychiatry, neurology and general medicine. Because of its ability
to improve several clinical symptoms in different diseases modafinil seems to be
one of the important drugs. The pharmacologic effects of modafinil are complex and
it is thought to alter various neurotransmitters in the brain (Minzenberg and Carter,
2007). A clear mode of action of modafinil has not been established so far but
interaction of modafinil with dopaminergic, noradrenergic, glutamatergic, gamma-
aminobutyric acid (GABA)ergic, serotoninergic, orexinergic, and histaminergic
pathways have been suggested in several animal studies (Ballon and Feife, 2006;
Ferraro et al., 1999; Ferraro et al., 1998; Ferraro et al., 2002; Madras et al., 2006;
Minzenberg and Carter, 2007).
Modafinil has been investigated in healthy volunteers, and in individuals with clinical
disorders. Its beneficial effect has been shown in many clinical disorders associated
49
with excessive sleepiness, fatigue, impaired cognition and other symptoms such as
myotonic dystrophy (MacDonald et al., 2002), attention deficit hyperactivity disorder
(ADHD) in children and adolescents (Lindsay et al., 2006), depression (DeBattista et
al., 2004), Parkinson‘s disease (PD), multiple sclerosis (MS) (Bibani et al., 2012;
Littleton et al., 2010; Rammohan et al., 2002), and hastening recovery from general
anaesthesia (Larijani et al., 2004). In sleep-deprived healthy volunteers, modafinil
improves mood, fatigue, sleepiness and cognition (Wesensten et al., 2005).
Modafinil improves the ability of the on call physicians to attend lectures after a full
night shift in emergency departments (Gill et al., 2006).
The potential interactions of modafinil with a variety of drugs through induction and
inhibition several cytochrome P450 isoenzymes has been reported (Robertson et
al., 2000). Reduction of the modafinil dose in old age groups and in patients with
hepatic and renal impairment is mandatory. Insomnia, headache, nausea,
nervousness and hypertension are commonly reported adverse effects of modafinil
(Robertson and Hellriegel, 2003). Other less common side effects of modafinil are
decrease of appetite, weight loss and dermatological problems. Children and
adolescents have greater risk to get these side effects. Modafinil may theoretically
have some abuse/addictive potential. The results seen in two trials on cocaine
addiction treated with modafinil were inconclusive (Dackis et al., 2005; Umanoff,
2005).
1.3.2 Pharmacodynamic Properties of Modafinil
Modafinil is (2-[(diphenylmethyl) sulfinyl] acetamide. The molecular formula is
C15H15NO2S and the molecular weight is 273.35. The chemical structure is shown in
(Figure 1.6).
Modafinil has two enantiomers (R-modafinil and S-modafinil). The R-modafinil also
known as armodafinil has longer half-life than S-modafinil while the S-modafinil has
50
Figure 1.6 Chemical structure of modafinil.
a faster rate of clearance. The wake-promoting activity is likely due primarily to the
R-modafinil (Robertson and Hellriegel, 2003). Modafinil is used in a once-daily
dosing. It is readily absorbed from gastrointestinal tract and maximum plasma
concentrations occur at 2–4 hours (Robertson and Hellriegel, 2003). Metabolism
occurs primarily through the liver, with renal elimination of metabolites (Robertson
and Hellriegel, 2003). Less than 10% of the administered dose is excreted in urine
as unchanged drug (Robertson and Hellriegel, 2003; Wong et al., 1999). Modafinil
induces the cytochrome P450 enzymes CYP1A2, and CYP3A4 and has potential to
inhibit CYP2C19 (Robertson et al., 2000), thus it may prolong elimination and
increase circulating levels of drugs that are primarily metabolized via this enzyme
(e.g., diazepam, phenytoin, and propranolol). Modafinil suppressed CYP2C9 activity
in cultures of human hepatocytes, suggesting a potential for drug interactions
between modafinil and enzyme substrates such as warfarin and phenytoin
(Robertson Jr et al., 2002). Modafinil also enhances the effects of antidepressants
(Menza et al. 2000, Ninan et al. 2004).
1.3.3 Clinical Efficacy and Tolerability of Modafinil
The efficacy and tolerability of modafinil were recognised in several studies with
different designs. These studies have been conducted in patients with OSA (Black
and Hirshkowitz, 2005; Pack et al., 2001), SWSD (Czeisler et al., 2005), and
51
narcolepsy (Mitler et al., 2000). However, a significant greater frequency of adverse
effects has been reported with modafinil in placebo-controlled clinical trials, and
these were more frequent with fixed dose studies than the in flexible dose study
(Swanson et al., 2006). Increase in both systolic and diastolic blood pressure (BP)
with modafinil has been reported (Muller et al., 2004; Turner et al., 2003). In
contrast, several studies have reported no significant changes in BP (systolic and
diastolic), pulse rate and/or electrocardiography (ECG) (Biederman et al., 2005;
Black and Hirshkowitz, 2005; Broughton et al., 1997; Greenhill et al., 2006; Saletu et
al., 1989) . A significantly higher rating for somatic anxiety and several physical
symptoms such as tremor, palpitations, dizziness, muscular tension, physical
tiredness and irritability have been reported with modafinil compared with placebo
(Randall et al., 2003). Decreased appetite and weight loss with modafinil was
reported in studies of ADHD (Biederman et al., 2005; Greenhill et al., 2006).
Different types of skin lesions related to modafinil taking have been reported inform
of maculopapular/morbiliform rash and a case of possible erythema
multiforme/Stevens-Johnson Syndrome (Biederman et al., 2005). Two patients with
major depressive disorder developed suicidal ideation in the second week of a trial
of combined modafinil and serotonin reuptake inhibitors (SRI) therapy (Dunlop et al.,
2007). Psychosis has been reported with modafinil in schizophrenia, post-polio
fatigue patients and in SWSD patient without any history of psychiatric disorder
(Mariani and Hart, 2005; Spence et al., 2005; Vasconcelos et al., 2007). Withdrawal
symptoms of modafinil have not been observed in subjects with ADHD (Greenhill et
al., 2006). There is no conclusive evidence for abuse potential of modafinil so far,
as the psychomotor effects of modafinil do not appear to be mediated via a
catecholamine mechanism (Ferraro et al., 1996), which might account for modafinil's
reduced side-effect profile and low abuse potential (Deroche-Gamonet et al., 2002).
A single fatal case of multi-organ hypersensitivity reaction has been described
(Sabatine et al., 2007).
52
1.3.4 Mechanism of Action of Modafinil
Modafinil: has been named ―a drug in search of a mechanism‖ (Saper and
Scammell, 2004). Despite several years of studies a well-defined biochemical
mechanism of action of modafinil has not yet been elucidated. The pharmacologic
effects of modafinil are complex and it is thought to alter various neurotransmitters in
the brain (Minzenberg and Carter, 2007). Briefly, in animal studies, modafinil has
been shown to interact with dopaminergic, noradrenergic, glutamatergic,
GABAergic, serotoninergic, orexinergic, and histaminergic pathways (Ballon and
Feife, 2006; Madras et al., 2006; Minzenberg and Carter, 2007). It has been
demonstrated that modafinil activates the hippocampus, which receives afferent
innervation from the sleep-wake centre of the hypothalamus (Becker et al., 2004;
Kim et al., 2007).
1.3.4.1 Effects of Modafinil on the Dopaminergic Pathways
The evidence regarding the interaction of dopaminergic pathway in modafinil's mode
of action has changed over time. The initial animal studies showed modafinil had
only a weak affinity for dopamine receptors (Mignot et al., 1994), and had not
stimulated release of dopamine in the mouse caudate nucleus (De Sereville et al.,
1994). Furthermore, it has no effect on the firing rate of the dopaminergic neurons
in the rat midbrain (Akaoka et al., 1991). In other studies it was also found that
various dopamine D1 and D2 receptor antagonists and inhibition of dopamine
synthesis does not affect on the modafinil-induced hyperactivity in mice (Duteil et al.,
1990; Simon et al., 1995), importantly, a slight reduction of the arousal with
modafinil has been reported in cats (Lin et al., 1992). In contrast subsequent animal
studies showed that modafinil administration in different doses and routes leads to
increased extracellular levels of dopamine in the prefrontal cortex (Hilaire et al.,
2001), caudate nucleus (Wisor et al., 2001), nucleus accumbens (Murillo-Rodriguez
et al., 2007), and striatal slices preloaded with [3H]dopamine (Dopheide et al., 2007).
53
Also it has been found that modafinil inhibits the dopaminergic neurons in the ventral
tegmental area and the substantia nigra (Korotkova et al., 2006). Evidence from
preclinical studies suggests that Modafinil increases dopamine in brain by targeting
the dopamine transporters (DAT) (Greenhill, 2006). On the other hand the role of
dopamine receptors (D1 and D2) in the mode of action of modafinil have been
evaluated and it was found that D1 and D2 receptors are involved in alerting effects
of modafinil (Qu et al., 2008).
By using recent MRI techniques (positron emission tomography (PET)) the idea
about interaction of dopamine in the mode of action of modafinil was further
expanded. Madras et al (2006) used this technique and documented the significant
occupancy of striatal DAT by modafinil in monkeys and in vitro modafinil inhibits
DATs . Furthermore, in a supporting study it was found that mice lacking DAT do not
respond to the wake-promoting effects of modafinil (Wisor et al., 2001). Findings
from a human study documented the crucial role of dopamine in the wake-promoting
effects of modafinil, and the blockage of DATs and increased dopamine in the
human brain (including the nucleus accumbens) (Volkow et al., 2009).
1.3.4.2 Effects of Modafinil on Noradrenergic Pathways
There is considerable pharmacological evidence that modafinil, acts through
adrenergic mechanisms to promote waking. Animal studies found that modafinil
increases levels of noradrenaline in the rat prefrontal cortex and medial
hypothalamus (de Saint Hilaire et al., 2001). In rat brain slices, modafinil potentiates
noradrenergic inhibition of the sleep active neurons of the ventrolateral preoptic area
of the hypothalamus (Gallopin et al., 2004). Various α-adrenoceptor antagonists
attenuate the modafinil-induced arousal in cats (Lin et al., 1992), and locomotor
activity in mice (Stone et al., 2002) and monkeys (Duteil et al., 1990). Evidence has
strongly suggested that modafinil promotes waking by activating α1-adrenergic
receptors. Response to modafinil was significantly reduced in genetically α1-
54
adrenoceptor-deficient mice (Stone et al., 2002). Furthermore, modafinil occupies
noradrenaline transporter (NAT) sites in the thalamus of rhesus monkeys in vivo and
blocks noradrenaline transport via NAT in vitro (Madras et al., 2006). On the other
hand, it has been found that Modafinil does not bind to adrenergic receptors at
physiological doses (Mignot et al., 1994), and it does not affect the firing rate of the
rat pontine noradrenergic neurons (Akaoka et al., 1991) and it does little to reduce
cataplexy that normally responds to α1-receptor agonists or to agents that block the
reuptake of noradrenaline by NAT (Mignot et al., 1993; Nishino et al., 1993).
1.3.4.3 Effects of Modafinil on Glutamate
Ferraro et al in series of studies found that modafinil increases levels of the
glutamate in the thalamus and hippocampus (Ferraro et al., 1997), striatum (Ferraro
et al., 1998) and medial pre-optic area and the posterior hypothalamus (Ferraro et
al., 1999) of the rat brain.
1.3.4.4 Effect of Modafinil on gama amino butyric acid (GABA)
Animal studies have reported the effect of modafinil in reducing GABA levels in the
cortex (Tanganelli et al., 1994), medial pre-optic area and posterior hypothalamus
(Ferraro et al., 1999), hippocampus (Ferraro et al., 1997), nucleus accumbens,
striatum, globus pallidus and substantia nigra (Ferraro et al., 1998). This might lead
to the conclusion that via GABA reductions, modafinil is able to improve motor
activity and cortical functions (Della Marca et al., 2004).
1.3.4.5 Effect of Modafinil on serotonin
There is an inverse effect of modafinil on the level of serotonin and GABA in
different brain areas. Studies have found that modafinil decreases levels of GABA,
but increases levels of serotonin (Ferraro et al., 2000; Ferraro et al., 2002). Also, it
has found that SRIs and serotonin selective neurotoxins abolish the effect of
modafinil on GABA release (Tanganelli et al., 1992; Tanganelli et al., 1995). SRIs
55
enhance the effect of modafinil on serotonin levels (Ferraro et al., 2005; Ferraro et
al., 2002).
1.3.4.6 Effects of Modafinil on Histaminergic Pathways
Ishizuka et al (2008) found that modafinil increases histamine levels in the anterior
hypothalamus in rats . They also found that enhancement of the locomotor activity in
treated rats with modafinil is reversible with depletion of neuronal histamine.
1.3.4.7 Effects of Modafinil on Orexinergic Pathways
The interaction of modafinil with orexin neurons in the brain is complicated and not
clear yet. Although modafinil activates the orexin neurons (Scammell et al., 2000), it
is also useful for narcolepsy deficient in orexin neurons (Nishino, 2003). It has also
been found that modafinil is more effective in producing wakefulness in orexin
knockout mice than in wild-type litter mates (Willie et al., 2005).
1.3.5 Approved Indications of Modafinil
The use of modafinil has been approved for ameliorating the excessive sleepiness
associated with narcolepsy, SWSD and residual sleepiness in OSA.
1.3.5.1 Narcolepsy
The main symptoms of narcolepsy are EDS, cataplexy (an abrupt loss of muscle
tone triggered by emotion), hypnagogic hallucinations and sleep paralysis. Four
randomised, double-blind, placebo-controlled trials have assessed the usefulness of
modafinil in treatment of EDS in narcolepsy (Billiard et al., 1994; Broughton et al.,
1997; Fry, 1998; US Modafinil in Narcolepsy Multicenter Study, 1998). Improvement
of EDS by the objective measures in all four studies have demonstrated and
improvement by the subjective Epworth Sleepiness Scale (ESS) was also seen
except the Billiard et al study.
56
1.3.5.2 Obstructive Sleep Apnoea (OSA)
EDS is one of the main symptoms of OSA and continuous positive airway pressure
(CPAP) is the gold-standard treatment for OSA. Modafinil is approved by the FDA
for treating residual sleepiness despite optimal treatment of OSA (in November 2010
The Agency‘s Committee for Medicinal Products for Human Use (CHMP)
recommended that this indication should remove from the product information).
Several studies have carried out for evaluation the role of Modafinil in OSA. The
largest of these studies was a double-blind, randomised, placebo-controlled study
(Pack et al., 2001) The primary efficacy measures were ESS, multiple sleep latency
test (MSLT) results, and CPAP use. The positive effects of modafinil have
demonstrated in both the ESS and MSLT results, while there was no difference
noted in CPAP use between groups. In another relatively similar size study Black
and Hirshkowitz, have confirmed the effectiveness of modafinil in the clinical
situation which showed that the efficacy of modafinil, as measured subjectively by
the ESS and objectively by the maintenance of wakefulness test (MWT), does not
change over an extended period of the study which was 12 weeks (Black and
Hirshkowitz, 2005). Although reduction in the CPAP usage was not decreased in
both mentioned studies this was noted in the smaller randomised study of modafinil
(Kingshott et al., 2001) and in an open-label extension trial (Schwartz et al., 2003).
Moreover, modafinil improved symptoms of depression, anxiety, and irritability in
patients with OSA (Kumar, 2008).
1.3.5.3 Shift-Work Sleep Disorder (SWSD)
Three randomised, double-blind, placebo-controlled, parallel group, multicentre trials
have evaluated the usefulness of modafinil in SWSD with different measures of
efficacy. In the first trial (Czeisler et al., 2005), The modafinil group had a statistically
significant increase in mean sleep latency compared with the placebo group,
Patients taking modafinil also had a significant improvement in performance on a
57
vigilance test. In the second trial (Erman and Rosenberg, 2007), a statistically
significant greater increase in the mean Functional Outcomes of Sleep
Questionnaire score in patients received modafinil compared with placebo was
seen, also a significant improvements in the activity and in the vigilance and
productivity domain scores with modafinil were reported. Furthermore Modafinil
significantly improved the mental and emotional scores compared with placebo. In
the third study (Walsh et al., 2004) a significant improvement on vigilance testing
and the Maintenance of Wakefulness Test (MWT) have been reported.
1.3.6 Investigational Uses of Modafinil
1.3.6.1 Neurological Disorders
1.3.6.1.1 Parkinson's disease
EDS is one of the main symptom of PD (Ondo et al., 2005). Significant improvement
in ESS scores were seen in two small, randomised, double-blind, placebo-controlled
studies (Adler et al., 2003; Happe et al., 2001). In contrast this achievement was not
found in another study with similar design (Ondo et al., 2005). Also striatal activation
in PD by modafinil has been shown (Scammell et al., 2000).
Two studies were conducted by Ferraro et al indicating that modafinil could have
anti-parkinsonian effects on the motor symptoms of PD (Ferraro et al., 1997; Ferraro
et al., 1998).
1.3.6.1.2 Myotonic Dystrophy
EDS is common symptom in myotonic dystrophy (Laberge et al., 2004). Three
randomised, double-blind, placebo-controlled, crossover trials suggested positive
effects of modafinil in improving EDS in patients with myotonic dystrophy
(MacDonald et al., 2002; Talbot et al., 2003; Wintzen et al., 2007).
58
1.3.6.2 Psychiatric Disorders
1.3.6.2.1 Attention Deficit Hyperactivity Disorder (ADHD)
Three placebo-controlled, open-label trials have assessed modafinil for treatment of
ADHD in children. A beneficial effect of modafinil in ADHD symptoms in children in
terms of increasing attention and decreasing hyperactive and impulsive behaviour
was observed (Amiri et al., 2008; Boellner et al., 2006; Rugino and Copley, 2001).
Two other studies also found that modafinil progressively decreases the ADHD
symptoms (Biederman et al., 2006; Swanson et al., 2006). A small comparative
study had conducted to explore the efficacy of modafinil with dexamphetamine and
placebo in adults with ADHD (Taylor and Russo, 2000). They found that both
dexamphetamine and modafinil significantly reduced the ADHD symptoms. Another
study suggested a beneficial effects of a single dose of modafinil on working
memory, visual memory, planning, response inhibition and sustained attention in
adults with ADHD (Turner et al., 2004a).
1.3.6.2.2 Depression
Three studies have assessed the efficacy of modafinil in major depression by using
different combinations of instruments (DeBattista et al., 2004; Dunlop et al., 2007;
Fava et al., 2005). Two of these studies have shown considerable placebo effect,
with reductions in ESS and FSS scores with both modafinil and placebo (DeBattista
et al., 2004; Fava et al., 2005). However in a small study the efficacy of modafinil as
adjunctive treatment for bipolar depression has been assessed (Frye et al., 2007).
The result has suggested that modafinil may be helpful in bipolar depression.
1.3.6.2.3 Schizophrenia
Several studies have assessed the effect of modafinil on clinical measures in
schizophrenia. Modafinil showed no greater effect on fatigue than placebo (Pierre et
al., 2007; Sevy et al., 2005) and no effect (Sevy et al., 2005) or only limited effect
59
(Hunter et al., 2006; Spence et al., 2005; Turner et al., 2004b) on cognition
performance.
1.3.6.2.4 Alzheimer's disease
A link between systemic inflammation and Alzheimer‘s disease has been suggested
(Rogers et al., 1988). A recent study has found that the modafinil derivatives exhibit
anti-inflammatory activity as evidenced by lowering of lipopolysaccharide-induced
nitric oxide (NO) generation and of inflammation-related enzymes in BV2 microglial
cells. They have also reported that the anti-inflammatory activity of modafinil
derivatives is better than that of aspirin in the cultured cells used. These results
suggest that modafinil derivatives can be developed as potential anti-inflammatory
agents and a treatment strategy for Alzheimer‘s disease related dementia (Jung et
al., 2012). In contrast, Frakey et, al., (2012) have reported that the addition of
modafinil to the standard of care treatment (cholinesterase inhibitor medication) in
individuals with Alzheimer's disease has not resulted in significant additional
reductions in apathy or improvements in performance of activities of daily living.
1.3.6.2.5 Effect of modafinil on addiction and substances dependency
1.3.6.2.5.1 Cocaine
Modafinil has been assessed for potential treatment of cocaine addiction. Modafinil
could induce a decline in cocaine use (Dackis et al., 2005; Hart et al., 2008) and this
more specifically in a subset of cocaine without alcohol dependence (Anderson et
al., 2009). Another supporting study found that the decrease of use also was
associated with longer periods of abstinence (Dackis et al., 2005).
In contrast to the previous positive results Dakis et al in a recent study had
concluded that modafinil has no significant differences compared to placebo on the
cocaine abstinence, cocaine craving, cocaine withdrawal, retention, and tolerability
(Dackis et al., 2010, 2012).
60
1.3.6.2.5.2 Methamphetamine
Two studies have suggested that modafinil can increases in the number drug-free
days in amphetamine dependence and it can decrease the withdrawal syndrome
(McGregor et al., 2008; Shearer et al., 2009). However Shearer et al found no effect
of modafinil on craving for methamphetamine (Shearer et al., 2009).
1.3.6.2.5.3 Nicotine
A positive role of modafinil in abstinent smokers has not been confirmed in two
clinical trials; in contrast, they have reported an increase of negative effects and
depressive symptoms after ingestion of modafinil (Schnoll et al., 2008; Sofuoglu et
al., 2008). Furthermore, one of the trials reported that nicotine-abstinent participants
smoked more with modafinil and had more withdrawal symptoms reported than non-
abstinent participants with placebo, which resulted in the trial being discontinued
(Schnoll et al., 2008).
1.3.6.3 Effect of modafinil on Disorders Associated with Fatigue
1.3.6.3.1 Chronic Fatigue Syndrome
In a single randomised, double-blind, placebo-controlled, crossover study in patients
with chronic fatigue syndrome (CFS) modafinil had inconsistent effects on the
primary efficacy measure of cognition (Randall et al., 2005a), and no improvement
was seen in the secondary efficacy measures of fatigue, quality of life (QoL) or
mood.
1.3.6.3.2 Fatigue in Post-Polio Syndrome
A placebo-controlled study, conducted by Chan et al revealed no significant
difference between the two treatments in the terms of the ESS scores and other
secondary efficacy measures (Chan et al., 2006). Another study with relatively
similar design revealed improvements in primary efficacy measures of ESS,visual
analogue scale for fatigue (VAS-F) and fatigue impact scale (FIS) with both placebo
61
and modafinil without significant differences between the two treatments
(Vasconcelos et al., 2007).
1.3.6.3.3 Fatigue in Multiple Sclerosis
Fatigue is the most troublesome symptom in MS (Fisk et al., 1994a). Modafinil has
been reported to improve fatigue in patients with MS (Lange et al., 2009; Littleton et
al., 2010; Rammohan et al., 2002; Zifko et al., 2002). The benefits of modafinil on
the FSS were more pronounced than those previously reported with other commonly
used medications, including amantadine. In patients with RR or progressive forms of
MS, modafinil was associated with significant improvements on several measures of
fatigue, including the fatigue severity scale (FSS), the modified fatigue impact scale
(MFIS), and the VAS-F (Rammohan et al., 2002). In a supporting study Zifko et al
found that a low-dose regimen of modafinil is significantly improves fatigue and
sleepiness and is well tolerated by patients with MS (Zifko et al., 2002). Littleton et al
(2010) have suggested that modafinil may be useful for treatment of fatigue in MS,
particularly when the fatigue is associated with sleepiness. In contrast Stankoff et al
found no improvement of fatigue in patients with multiple sclerosis treated with
modafinil vs. placebo according to the MFIS (Stankoff et al., 2005).
1.3.6.3.4 Fatigue in Parkinson’s disease
Lou (2009) had conducted a study to evaluate the subjective mental and physical
fatigue in PD patients by using self-report questionnaires. The findings revealed that
Levodopa and modafinil could improve physical fatigability in PD subjects. In
another study Lou et al demonstrated that although modafinil may be effective in
reducing physical fatigability in PD, it did not improve fatigue symptoms (Lou et al.,
2009).
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1.3.6.3.5 Cancer-related Fatigue
Morrow et al (2005) found that using modafinil was associated with significant
improvement in fatigue severity and other measures of QoL in women who reported
persistent fatigue after completion of breast cancer treatment . Kaleita et al (2006)
found that modafinil significantly improves fatigue scores in people with malignant
and benign brain tumours who were treated with surgery, radiotherapy, and/or
chemotherapy. Spathis et al (2009) found a statistically and clinically significant
reduction in fatigue and improvement of daytime sleepiness and depression/anxiety
in cancer patients whom treated with Modafinil. This finding has further supported by
other studies (Cooper et al., 2009; Rabkin et al., 2009).
1.3.6.4 Recovery from General Anaesthesia
A study found that patients who receive modafinil have significantly less exhaustion
and they will be more alert and energetic during the stage of recovery from general
anaesthesia compared with the control (Larijani et al., 2004). On the other hand
studies found that the sedative effects of anti-psychotics and opiates after general
anaesthesia can be reduced by modafinil (Larijani et al., 2004; Makela et al., 2003;
Webster et al., 2003).
1.3.6.5 Sleep-Deprived Emergency Room Physicians
It has found that modafinil improves mood, fatigue, sleepiness and cognition in
sleep-deprived healthy volunteers, and in full night shift duty physicians in
emergency department (Gill et al., 2006).
1.3.6.6 Effects of modafinil on quality of life (QoL)
A study found that modafinil significantly improves the QoL (Black and Hirshkowitz,
2005), but this effect was not found in the another study (Kingshott et al., 2001).
63
1.3.6.7 Effects of modafinil on Cognitive Performance
A positive effect of modafinil on cognition has been found in healthy young and
elderly volunteers (Makris et al., 2007; Turner et al., 2003). A significant
improvement in the level of alertness has been found with modafinil (Niepel et al.,
2012), while a significant effectiveness on cognitive performance was not shown in
the small crossover trial by using the Steer Clear (a computerised driving simulator
with road obstacles which can be avoided by pressing a key) (Dinges and Weaver,
2003).
1.3.6.8 Effects of Modafinil in Healthy Volunteers
1.3.6.8.1 Non-sleep deprived volunteers
Effects of modafinil have been studied in healthy volunteers under differing
conditions. Studies have found improvement of cognition with modafinil in non-
sleep-deprived, healthy young and elderly volunteers (Makris et al., 2007; Turner et
al., 2003). In contrast, results from three studies conducted by Randall et al suggest
that the benefits of modafinil are insufficient to be considered as a cognitive
enhancer in non-sleep-deprived individuals (Randall et al., 2004; Randall et al.,
2005b; Randall et al., 2003).
1.3.6.8.2 Sleep-deprived volunteers
The effect of modafinil in healthy sleep-deprived subjects has been evaluated in
several studies. Modafinil led to improved subjective measures such as mood,
fatigue, sleepiness, vigilance and improved objective measures such as reaction
times, logical reasoning and short-term memory, and the Maintenance of
Wakefulness Test (MWT) (Pigeau et al., 1995; Wesensten et al., 2002; Wesensten
et al., 2005).
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1.3.7 Neuroprotective aspects of modafinil
Evidence from preclinical studies suggests neuroprotective effects of modafinil. The
neuroprotective mechanisms of modafinil are unknown. Generally two groups of
theories exist: the protective effect could be via modulation of neurotransmitters or
could be via interference with cell death processes. The ability of modafinil to protect
against degeneration of nigrostriatal dopamine neurons induced by 1-methyl-4-
phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) has been suggested to be related to
actions on GABAergic mechanisms of modafinil (Fuxe et al., 1992; Ueki et al.,
1993a). It has also been suggested that modafinil has the ability to restore the
locomotor activity in neurons already injured by MPTP, but not during the initial
phase (Jenner et al., 2000). Findings from another study suggest that administration
of a high dose modafinil with MPTP can selectively alter GABA binding density in
the internal globus pallidus and prevents the MPTP toxicity (Zeng et al., 2004).
Another supporting study suggested that modafinil prevents the decline in motor
activity induced by MPTP treatment (Xiao et al., 2004). This potential
neuroprotective role for modafinil in rodents was found to be mediated by anti-
oxidant effects and modulation of nigrostriatal GABA and striatal nor-adrenaline and
oxitriptan release, decreasing the GABA release and increasing the glutathione
through an antioxidative process which may be independent of its wake-promoting
effects (Xiao et al., 2004). These findings were supported by another study, which
showed that modafinil significantly prevented the MPTP-induced change in
locomotor activity, hand-eye coordination and small fast movements (van Vliet et al.,
2006).
In two studies Ferraro et al have found that modafinil could normalise the disturbed
balance of neurotransmitters by affecting glutamate and GABA release in specific
areas of the basal ganglia through a maximal increase in glutamate release in these
brain regions, associated with a lack of effect on GABA release. Furthermore, they
65
found that modafinil inhibits dose-dependently the activity of GABA neurons in the
cerebral cortex and in the nucleus accumbens, sleep-related brain areas such as
the medial preoptic area and the posterior hypothalamus (Ferraro et al., 1997;
Ferraro et al., 1998).
The second group of theories addresses the interference of modafinil with cellular
processes. The explanation for the mechanism of action of modafinil in the neuronal
cellular processes has been argued in several studies. Taking together, the
modulation of neurotransmitters by modafinil may be related to improvement of
energy metabolism, synthesis and release of neurotrophic factors, recovery of
calcium homeostasis, improvement in metabolic activity, free radical scavenging or
stimulation of repair processes such as axonal regeneration from the surviving cell
bodies (Antonelli et al., 1998; Fuxe et al., 1992; Jenner et al., 2000; Lallement et al.,
1997; Ueki et al., 1993b). Modafinil stimulates the enzymatic breakdown of
glutamate resulting in an increase in glutamine and a reduction in the cytotoxic
effects of glutamate (Touret et al., 1994). Modafinil inhibits the cytochrome P450
enzymes particularly CYP2C9 (Robertson et al., 2000). Inhibition of cytochrome
P450 enzymes reduces damage in arterial ischemia and reperfusion (Fleming et al.,
2001; Granville et al., 2004). Modafinil‘s suppression of brain cytochrome P450
could occur through a direct intracellular site of action to suppress CYP2C9 or
through enhancement of serotonin release (Ferraro et al., 2005; Tanganelli et al.,
1995). These effects of modafinil could explain its ability to reduce the production of
reactive oxygen species and to promote better mitochondrial function.
Piérard et al (1995) suggested that the neuroprotective effect of modafinil is due to
its ability to increase the cortical pool of creatine-phosphocreatine.
Furthermore, modafinil also protects noradrenergic and serotonergic neurons
against mechanical trauma induced by partial hemitransection as well as neostriatal
neurons against ischemic lesions associated with local endothelin-1 microinjection
66
and prevent increases in toxic aspartate and glutamate levels after striatal ischemic
injury (Ueki et al., 1993a).
Modafinil prevents development of lesions in the hippocampus induced by the
neurotoxic nerve gas soman (Lallement et al., 1997). A MRS study has shown the
neuroprotective ability of modafinil to prevent neuronal death and prevent glutamate
toxicity in cultured cortical cells (Antonelli et al., 1998). Modafinil inhibits GABA
release in areas involved in the direct and indirect pathways of the basal ganglia-
thalamus-cortex loop (Ferraro et al., 1997). The direct or indirect protective effects
or the sustained administration of modafinil could have increased the activity of the
remaining serotonergic neurons in the striatum, as modafinil does affect 5-HT levels
in the brain (Ferraro et al., 2002).
Modafinil activates the histaminergic system and increases hypothalamic histamine
release and c-Fos expression provided there are intact orexinergic neurons
(Ishizuka et al., 2010). It has been shown that histamine, via its H3 receptors
has wakefulness-promoting effects, improves cognition and is neuroprotective
against brain ischemia and neurodegenerative disorders (Fan et al., 2011; Stocking
and Letavic, 2008). The presence of the central H3 receptor is CNS-protective
against experimental autoimmune encephalomyelitis (EAE), an experimental model
of MS. H3 receptor activation reduce the susceptibility to autoimmune inflammatory
disease of the CNS (Parmentier et al., 2007; Teuscher et al., 2007).
Evidence suggests that modafinil may also act via a mechanism similar to the
neuropeptides orexin-A and -B to promote histamine release (Chemelli et al., 1999;
Ishizuka et al., 2003; Scammell et al., 2000).
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1.1.4 Summary and conclusions
The primary aim of this thesis is to gain greater insight into the potential
neuroprotective effects of modafinil in MS. In order to investigate this, we have
reviewed MS in general and the literature related to the principal aim of this thesis.
MS is a debilitating CNS disease in which neurodegeneration is the major
determinant of the accumulation of irreversible (progressive) disability. MS has
many physical and mental health consequences that limit the independence and
QoL of those living with the disease. MS is a challenging disease to treat.
Successful early on, DMTs eventually become partially ineffective in reducing
relapses and slowing disease progression, resulting in long term disability
accumulation. However, DMT may not or may only partially confer neuroprotection.
Neuroprotective agents that impact directly on neuronal survival would be desirable,
particularly since axonal loss and neuronal injury have been shown to be the
histological correlates of neurological disability.
Evidence from preclinical studies suggests a potential neuroprotective effect of
modafinil. The symptomatic benefits of modafinil have been studied in neurological,
psychiatric, general medicine and even in healthy volunteers. However, its potential
neuroprotection has not been extensively evaluated in persons with MS.
Part of this chapter was a review of EAE. From the pathogenesis point of view, EAE
is a good model for studying MS mechanisms (Farooqi et al., 2010). The possible
role of EAE in exploring the neuroprotection strategy for MS was also reviewed in
this chapter and in chapter three.
Taken together, understanding the defects in the current MS treatment strategies,
and the potential neuroprotective effect of modafinil encouraged us to look for
developing and evaluating a new therapeutic strategy for MS. The following
chapters in this thesis will attempt to shed light on this topic.
68
CHAPTER 2 EXPLORING THE POTENTIAL
NEUROPROTECTIVE EFFECTS OF MODAFINIL IN MULTIPLE
SCLEROSIS (RETROSPECTIVE STUDY).
69
2.1 Introduction
Modafinil is a wakefulness-promoting agent. It has been used for treatment of
narcolepsy, obstructive sleep apnoea, and shift-work sleep disorder (Robertson and
Hellriegel, 2003). Modafinil has been used with varying success for symptomatic
treatment in MS. The majority of these studies focused on the effects of modafinil on
fatigue (Brioschi et al., 2009; Lange et al., 2009). A recent study supports a
beneficial effect of modafinil in MS fatigue, particularly in the considerable proportion
MS patients whose fatigue was associated with excessive daytime sleepiness
(Littleton et al., 2010).
Evidence from preclinical studies suggests neuroprotective effects of modafinil. The
neuroprotective potential of modafinil has been studied in animal models of
neurodegenerative diseases (Antonelli et al., 1998; Jenner et al., 2000; Piérard et
al., 1995; van Vliet et al., 2006; Xiao et al., 2004) (see chapter1).
These findings led us to explore, in a retrospective study, the potential
neuroprotective potential of modafinil, as inferred through the impairment/disability
progression, in MS, as it has been used widely in the treatment of MS related
fatigue. The neuroprotective potential of modafinil, if confirmed clinically, may lead to
new modalities for treating neurodegenerative diseases.
2.2 Methods
2.2.1 Patients
The MS clinic database at the Nottingham University Hospital was interrogated for
selection of patients who had received or were receiving modafinil.
Of these, we selected patients who had been on modafinil for 3 years or more, on
the assumption that a subtle neuroprotective effect may require this length of time to
be recognised clinically in terms of expanded disability status scale (EDSS) change.
The demographic and clinical characteristics (age, sex, type of MS, disease
70
duration, follow up period and concomitant disease modifying therapies (DMT)) of
these patients were obtained. For each modafinil subject three best matched MS
control subjects, who had no exposure to modafinil at all, were selected based on
the clinical characteristics of the patients mentioned above.
EDSS scores were recorded before the start of the modafinil treatment and at a
follow-up point, at least 3 years later, in the modafinil group; and at matching time
points in the non-modafinil group.
The primary parameter investigated was change in EDSS after ≥ 3 years of
modafinil treatment or ≥ 3 years of follow up in the non-modafinil group.
2.2.2 Data Analysis and Statistics
Descriptive statistics were used to describe the sample characteristics. R (a
language and environment for statistical computing) (http://CRAN.R-project.org/)
was used to assess differences between treated and untreated groups when other
relevant covariates were considered. These covariates were age, gender, disease
duration, MS type (relapsing or progressive), duration of follow up, baseline EDSS,
and any history of treatment with DMTs. Covariates that did not contribute
significantly to the model were removed one at a time and least significant first.
The number of patients who had an increase in EDSS of 1 step or more for EDSS
≤5 and of 0.5 or more for EDSS ≥5.5 was compared between treatment groups. For
this I used Fisher‘s exact test using SPSS version 18 (www.ibm.com/uk/SPSS).
2.3 Results
2.3.1 Demographic and Clinical Characteristics
2.3.1.1 Patient demographics
Of 65 patients with clinically definite MS, according to Poser and/or MacDonald
criteria, (Poser et al. 1983; McDonald et al. 2001) who had exposure to modafinil,
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thirty had received modafinil for the treatment of MS-related fatigue for an
uninterrupted period of 3 years or more. Modafinil dose ranged between 100mg and
400mg. Ninety matched non-modafinil treated patients were also included in this
study. Patient demographics are provided in (Table 2.1) and (Table 2.2).
Table 2.1 Demographic characteristics and clinical epidemiology of the
patients.
Modafinil-treated patients
(n=30)
Non-modafinil
patients (n=90)
P value
Median(range)
Age in years
DD in years
Follow-up
EDSS (baseline)
EDSS (follow up)
44.5 (27-61)
45(28-61)
0.995*
9 (4-30)
9 (3-32)
0.870**
4 (3-7)
4 (3-8)
0.607*
3 (1-6.5)
3 (0-7.5)
0.751*
3 (0-7.0)
4 (0-8.5)
Gender n(%)
Female
19 (63.3%)
60 (66.6%)
MS
clinical
types
n(%)