© 2014 Direct One Communications, Inc. All rights reserved. 1 Understanding the Immunology of Multiple Sclerosis Kathleen Costello, MS, ANP-BC, MSCN, and

© 2014 Direct One Communications, Inc. All rights reserved. 1

Understanding the Immunology of Multiple Sclerosis

Kathleen Costello, MS, ANP-BC, MSCN, and Anne Gocke, PhDJohns Hopkins Multiple Sclerosis Center and Johns Hopkins University School of Medicine, Baltimore, Maryland

A REPORT FROM THE 28TH ANNUAL MEETING OF THE CONSORTIUM OF MULTIPLE SCLEROSIS CENTERS AND THE 19TH ANNUAL MEETING OF THE AMERICAS COMMITTEE FOR TREATMENT AND RESEARCH IN MULTIPLE SCLEROSIS


How the Normal Immune System Works


How the Normal Immune System Works

The normal immune system:» Protects against infectious threats

» Responds to active infections

» Prevents development of autoimmune conditions

The body’s natural defense barriers (skin, body fluids, mucus, and cilia in the lungs and gut) work together to keep infectious threats out of the system.

A breech in this barrier system provokes the cellular activity of the innate immune system.

Activation of neutrophils, monocytes, macrophages, and natural killer cells leads to eradication of most pathogens from the body.


Adaptive Immunity

When an infection requires additional immunologic intervention, the adaptive immune system is called to action.

Various messengers signal T and B cells to begin to defend the body.

Cell-surface molecules are critical to activation of the adaptive immune system, as are co-stimulatory molecules that allow for full activation of adaptive immune cells.

Adhesion molecules on blood vessels and cell walls are upregulated, allowing cells to move from the bloodstream to the site of infection in tissues.


Immune-System Cells:

Macrophages

Macrophages under the skin, in the lungs, and in the tissues surrounding the gut collect waste products and minimize cellular debris from dead cells.

Receptors on the surface of macrophages recognize signals from the pathogen that stimulate a “seek eat destroy” mechanism.

Macrophages send out signals called cytokines to recruit neutrophils and monocytes from the blood to the site of infection.

Macrophages also can engulf an invader and display a piece of it on the cell surface so that it can be recognized by the adaptive immune system.



Neutrophils and Monocytes

Along with macrophages, the chemokines IL-1 and TNF-a signal neutrophils circulating in the blood to slow down, sense the location of the infection, and migrate from the blood vessel to the infected tissue.

Monocytes mature into activated macrophages, which stimulate repair mechanisms and produce IL-1, TNF, and reactive oxygen species.

Monocytes also act as antigen-presenting cells (APCs) for lymphocyte functioning in adaptive immunity, and they become further stimulated by the mechanisms of this protective mechanism.



Natural Killer (NK) Cells

Circulating NK cells use the same mechanism as neutrophils to exit the circulation and migrate into tissues at the infection site.

Upregulation of adhesion molecules allows NK cells to enter the tissue and become killers.

NK cells deliver powerful enzymes to target cells infected by viruses, which causes the cells to die, thereby reducing the number of infected cells.

NK cells also secrete cytokines that help activate macrophages and prime more macrophages to participate in the killing.


Complement

Complement is a series of proteins that is activated very rapidly in a coordinated and orderly way by the innate and adaptive immune systems.

In innate immunity, proteins in the complement system can become active when they recognize common chemical groups on the infected cell surface.

Complement can ”tag” an invader for destruction or bore a hole in invading cells to destroy them.


Antigens and Antibodies

Activation of the adaptive immune system induces:» Cell-mediated activity involving specialized T cells

known as T helper (Th) cells

» Humoral activity, involving B and T cells, antibodies, and complement

Antigen recognition and binding allows antibodies to perform four important effector functions that are important in eliminating invading pathogens: » Opsonization

» Complement activation

» Toxin neutralization

» Blocking attachment


T-Cell Activation

T cells are activated only if a recognizable antigen is presented to them by an antigen-presenting cell, such as a dendritic cell or a macrophage.

Naïve T cells are stimulated by antigen presentation and differentiate into various T-cell subsets:» Th1 differentiation results from stimulation by IL-

23 and interferon γ.

» Th2 differentiation results from stimulation by IL-4.

» Th17 differentiation results from stimulation by IL-6, IL-1, and IL-23.

T cells become activated and multiply based upon the antigen presented and the cellular environment.


T-Cell Activation continued


B-Cell Activation

B cells recognize antigen in the circulation and bind to the antigen and internalize it.

After internal processing, a small piece or peptide of the antigen is presented on the cell surface in a groove of the major histocompatibility complex (MHC) molecule, where an activated or memory T cell can recognize and interact with it.

This contact sends signals to the B and T cells to allow the T cells to stimulate production of receptors and cytokines and provide a second co-stimulatory signal for B cells.


B-Cell Activation continued


Summary

A highly complex immune system protects the body from pathogens and purges invasive proteins from the circulation.

The normal immune system is characterized by specificity, diversity, and memory, and it can differentiate the body’s own cells from foreign cells.

If the immune system malfunctions, however, autoimmunity and disease may develop.

© 2014 Direct One Communications, Inc. All rights reserved. 15© 2014 Direct One Communications, Inc. All rights reserved. 15

What Causes Inflammation of the Central Nervous System in MS?


Pathogenesis of MS

Multiple sclerosis (MS) is an immune-mediated inflammatory disease characterized by myelin destruction, damage to CNS-resident cells, and loss of mobility and cognition.

In acute and chronic active lesions, axons commonly are preserved; macrophages that have taken up myelin debris are evident.

In contrast, inactive lesions feature a loss of axons and oligodendrocytes and few macrophages.

Cortical plaques, which involve the gray matter, also may be found.


Risk Factors for Development of MS

The antigens that are targeted in MS may include neuronal proteins and astrocytic proteins.

Genetic predisposition has been linked with mutations in cytokine receptor genes (IL2RA, IL7R) as well as HLA-DR2B.

Environmental factors, including vitamin D, Epstein-Barr virus, and gut and lung immunity, also play an important role in MS susceptibility.

Gender is another risk factor for development of the disease; approximately two thirds of MS patients are female.


Actions of T and B Cells in MS

CD4+ T cells and cytotoxic CD8+ T cells play a pathogenic role in MS, likely due to their ability to:» Secrete pro-inflammatory cytokines

» Recruit peripheral monocytes and B cells to MS lesions

B cells may secrete antibodies that can mediate direct damage to axons.

The presence of lymphoid follicles in the meninges of some patients points toward a pathogenic role for B cells in MS.

B cells also help in neuronal repair and remyelination by promoting clearance of myelin debris via opsonization.


Other Pathologic Changes

Microglia sense changes in the CNS and release cytokines and chemokines that pave the way for entry of other immune cells into the lesion site.

Peripheral monocytes infiltrate the CNS and secrete proinflammatory cytokines and toxic molecules, such as nitric oxide, IL-1, IL-6, and matrix metalloproteinases, that can directly damage oligodendrocytes and neurons.

The most important determinants of permanent neurologic disability in MS patients are axonal damage and loss; axonal damage may occur even early in the course of the disease.


Hypothetical Pathologic Mechanisms

Activation of CD8+ T cells to target neurons directly

Vigorous CD4+ T-cell responses that recruit macrophages, resulting in the release of inflammatory mediators and toxic molecules

Binding of antibodies to neuronal surface antigens, followed by complement fixation or antibody-mediated phagocytosis of axons

Invasion by immune cells resulting in secondary, inflammation-independent neurodegeneration

Chronic inflammation leading to mitochondrial dysfunction, dysregulation of ion channels, and release of glutamate or nitric oxide


Summary

The etiology of MS is unknown; however, the immune system is important to the development of the disease, and lesions affect both the gray and white matter.

MS may begin with an invasion of the CNS by T and B cells; these events may be secondary to activation of microglia and macrophages and the local release of self or foreign antigens.

A small number of antigens present in the CNS may drive the highly focused, persistent acquired immune response in MS.


How Current Disease-Modifying Therapies Affect the Altered Immune Response in MS


Disease-Modifying Therapies (DMTs)


Interferon b-1a and b-1b

Reduce T-cell activation and proliferation Reduce secretion of matrix

metalloproteinases that disrupt the blood-brain barrier (and thus allow fewer immune cells entry into the CNS)

Inhibit interferon g release (reduces antigen presentation to T cells)

Limit T-cell migration across the blood-brain barrier

Reduce antigen processing and antigen presentation to T cells


Interferon b-1a and b-1b continued

At low doses, minor side effects include flu-like symptoms, headache, transaminitis, and depression; major side effects include suicidal ideation, anaphylaxis, hepatic injury, blood dyscrasias, seizures, and autoimmune hepatitis.

At high doses, all of these effects may be seen, in addition to injection-site reactions and skin necrosis.

Patients should be followed with complete blood counts with differential, liver and thyroid function tests, and interferon-neutralizing antibodies, if clinically warranted.


Glatiramer Acetate

Causes migration of Th2 cells into the CNS, modification of antibody production by plasma cells, and regulation of B-cell properties.

Recent evidence suggests that glatiramer acetate may produce cytokine modulation, inhibition of antigen presentation to T cells, and effects on oligodendrocyte precursor cells (myelin repair).

Minor side effects include injection-site reactions and post-injection vasodilatory reactions.

Major side effects include lipoatrophy, skin necrosis, and anaphylaxis.


Fingolimod

Antagonizes sphingosine 1-phosphate receptors, blocking lymphocyte egress from secondary lymphoid organs to the peripheral blood circulation

Minor side effects include mild lymphopenia and transaminitis.

Major side effects include bradycardia, heart block, hypertension, increased risk of herpetic infections, lymphopenia, macular edema, skin cancer, reactive airway, and posterior reversible encephalopathy syndrome.


Teriflunomide

Mimics pyrimidine as a DNA building block, interfering with DNA synthesis and inhibiting dihydro-orotate dehydrogenase.

Reduces T-cell proliferation and activation and production of cytokines, and it interferes with the interaction between cells and APCs.

Minor side effects include diarrhea, nausea, and thinning of the hair.

More severe effects include transaminitis, lymphopenia, teratogenicity, latent tuberculosis, neuropathy, and hypertension.


Dimethyl Fumarate

Changes the balance of Th1 to Th2 cells Activates the transcription factor Nrf2

transcriptional pathway, reducing oxidative stress.

Minor side effects include flushing and GI distress

More severe effects include transaminitis and leukopenia.


Mitoxantrone

Inhibits DNA topoisomerase II, suppressing the proliferation of T and B cells and macrophages.

Side effects range from nausea, vomiting, hair thinning, infections, liver dysfunction, and menstrual irregularities to cardiotoxicity, acute myelogenous leukemia, serious infections, and infertility.

Owing to its high-risk profile and the availability of other effective DMTs with fewer and less severe side effects, mitoxantrone currently is used infrequently for the treatment of MS.


Natalizumab

Block the expression of a4-integrin on the surface of lymphocytes, limiting the number of lymphocytes that enter the CNS.

Side effects include headaches, joint pain, fatigue, and a wearing-off phenomenon, wherein patients feel a temporary recrudescence of their MS symptoms just prior to each infusion.

The major risk of taking natalizumab is a severe, potentially fatal infection known as progressive multifocal leukoencephalopathy (PML)

Infusion reactions, hypersensitivity reactions, and other serious infections also may occur.


Peginterferon b-1a

Peginterferon b-1a is a chemically modified version of interferon b-1a where polyethylene glycol has been attached to the interferon molecule, thus allowing it to remain active in the circulation for 2–4 weeks after a single subcutaneous injection.

Adverse effects include injection-site reactions and flu-like symptoms.

Laboratory monitoring of MS patients receiving this peginterferon b-1a should be similar to that of other beta interferons.


How Does Progressive MS Differ from Relapsing MS?


Disease Progression

Most patients with MS will experience progression at some stage of their disease.

A definitive disease mechanism for progressive and relapsing MS has not been identified, although the role of environmental triggers, genetics, and other factors has been scrutinized.

A number of interlinked pathways may contribute to pathogenesis of the disease.


Primary-Progressive MS (PPMS)

Patients with PPMS exhibit a steady increase in disability without attacks.

Men are more likely to be affected by PPMS than by any other form of the disease.

This form of disease progression typically beings about 10 years after the onset of relapsing-remitting MS (RRMS).

Genetic susceptibility and the pathology of this disorder are similar to those of other forms of MS, and an underlying neurodegenerative problem could be involved.


Primary-Progressive MS (PPMS) continued

Meningeal infiltrates of B and T cells are particularly prominent in patients with PPMS.

In addition, lymphoid follicles associated with underlying microglia activation and cortical plaques may be evident.

White-matter plaques often are neuroinflammatory at their center, but microglia, macrophages, and ongoing simmering and possibly concentrically expanding axonopathy may be found.

Diffuse, low-grade parenchymal inflammation also has been reported.



In PPMS, brain damage is driven by inflammation similar to that of RRMS but with an intact blood-brain barrier.

This condition starts as an inflammatory disease; however, chronic inflammation then leads to neurodegeneration or disease progression independent of inflammation.

This neurodegeneration might cause intact neurons to lose control over microglial activation.

In early stages of the disease, inflammation amplifies progression of primarily neurodegenerative disease.



Therapeutic failures could be explained by perivascular inflammation, which often occurs without associated disruption of the blood-brain barrier.

Axonal and neuronal death may result from glutamate-mediated excitotoxicity, oxidative injury, iron accumulation, and/or mitochondrial failure.

Currently, a lack of full understanding about the biology of MS impedes development of effective treatments for PPMS.


Differences Between RRMS and PPMS

RRMS is characterized by acute focal inflammatory lesions in the white matter and is associated with inflammatory demyelinating lesions in the brain and spinal cord, which herald axonal damage and loss related to the foci of inflammation.

The gray matter is involved less prominently than it is in progressive MS.

In contrast, PPMS is characterized by diffuse (rather than focal) inflammation and involves more prominent cortical demyelination, diffuse axonal injury, and, frequently, the presence of microglial nodules in the brain.


Pathogenesis:

Inflammation

Inflammation occurs in all stages of MS and is characterized by perivascular and parenchymal infiltrates of lymphocytes and macrophages.

The initial inflammatory response in patients with RRMS mainly involves the proliferation of CD8+ T cells in active lesions, as abundant macrophages are recruited and activated.

In the secondary response, T and B cells and macrophages are recruited as a result of myelin destruction.


Pathogenesis:

Inflammation continued

Profound damage to the blood-brain barrier, as evidenced by the presence of gadolinium-enhancing lesions on MRI, results from infiltration of inflammatory cells into the CNS.

The relationship between inflammation and damage to the blood-brain barrier is less obvious in patients with progressive MS, because impairment can occur with or without inflammatory infiltrates.

Features of lymphoid follicle are found in large aggregates of inflammatory cells in the meninges; as the disease progresses, inflammation becomes compartmentalized behind the intact blood-brain barrier.


Pathogenesis:

Demyelination

The pathology of both acute and relapsing MS is dominated by focal inflammatory demyelinated plaques in the white matter.

In progressive MS, slow expansion of preexisting lesions results in pronounced cortical demyelination associated with extensive diffuse injury in white and gray matter that appears to be normal.

Cortical lesions occur most abundantly during disease progression; these lesions are most prominent in subpial cortical layers and can be linked to local meningeal inflammation.

Activated microglia are associated with active lesions.


Pathogenesis:

Tissue Injury

The brains of patients with MS exhibit widespread inflammation, microglial activation, astrogliosis, and mild demyelination and axonal loss in normal-appearing white matter.

The extent and severity of these changes increase with disease duration and most closely are asociated with progressive MS; small-caliber axons are most affected.

However, the extent of diffuse injury does not correlate with the number, size, or destructiveness of focal lesions; instead, it correlates moderately with the extent of cortical demyelination.


Pathogenesis:

Tissue Injury continued

RRMS is related to distinct patterns of demyelination and tissue injury, probably because the inflammatory lesions present in RRMS possess distinct immune processes.

In contrast, patterns of tissue injury largely are homogeneous in secondary-progressive MS (SPMS) and PPMS, probably because of the slow expansion of existing lesions with sparse remyelination.


Disease Processes:

Microglial Activation

In both RRMS and progressive MS, active tissue injury is associated with microglial activation.

In addition, microglial nodules are seen in progressive MS.

Oxidative bursts by activated microglia my help to induce demyelination and progressive axonal injury.

Microglia may be neuroprotective and could promote remyelination after debris is removed and neurotrophic molecules are secreted.


Disease Processes:

Mitochondrial Injury

The pathogenesis of MS may be related to energy deficiency or “virtual hypoxia.”

Impaired NADH dehydrogenase activity and increases in complex IV activity have been noted in the mitochondria of MS lesions.

Mitochondrial injury may reflex oxidative damage in areas of initial tissue injury.

The increased susceptibility to neurodegeneration seen in patients with SPMS and PPMS may be explained partially by an accumulation of mitochondrial DNA deletions.


Disease Processes:

Mitochondrial Injury continued

Oxidative stress encourages mitochondrial dysfunction in a number of ways: » Free radicals disrupt mitochondrial enzyme

function.

» Oxidative stress modifies mitochondrial proteins and accelerates their degeneration.

» Further, oxidative stress interferes with de novo synthesis of respiratory chain components and induces mitochondrial DNA damage.

Oxidative injury is pronounced in progressive MS lesions, even though inflammation is low; therefore, it may be driven by factors other than inflammatory processes.


Disease Processes:

Iron Accumulation

In the aging brain, iron accumulates and is stored in oligodendrocytes; subsequently, it is detoxified when it binds to ferritin.

Intracytoplasmic iron accumulation may explain why these cells are so susceptible to degeneration during oxidative stress.

In MS lesions, activated macrophages and microglia take up iron, leading to dystrophy, fragmentation, and cellular degeneration.

The process depends upon patient age and may be more pronounced among patients with progressive MS than among those with RRMS.


Summary

A number of interlinked pathways contribute to development of MS, and inflammation mediated by T and B cells and macrophages orchestrate the demyelination and degeneration seen in all patients.

Tissue injury may be affected by microglia activation, oxidative injury, and damage to mitochondria.

In progressive MS, liberation of iron may enhance oxidative damage and result in increased neurodegeneration.

Accumulated tissue damage exhausts functional reserve brain capacity and may accelerate clinical deterioration, with slow, progressive tissue injury.

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© 2014 Direct One Communications, Inc. All rights reserved. 1 Understanding the Immunology of Multiple Sclerosis Kathleen Costello, MS, ANP-BC, MSCN, and