Mechanisms of action of interferons and
`glatiramer acetate in multiple sclerosis
`
`Suhayl Dhib-Jalbut, MD
`
`Articles
`
`Article abstract—MS is an immunologically mediated disease, as determined by observation of the response to immuno-
`therapy and the existence of an animal model, experimental autoimmune encephalitis. Interferon (IFN) ␤-1b, IFN ␤-1a,
`and glatiramer acetate, the therapies used for relapsing or remitting MS, have mechanisms of action that address the
`immunologic pathophysiology of MS. The IFNs bind to cell surface-specific receptors, initiating a cascade of signaling
`pathways that end with the secretion of antiviral, antiproliferative, and immunomodulatory gene products. Glatiramer
`acetate, a synthetic molecule, inhibits the activation of myelin basic protein-reactive T cells and induces a T-cell repertoire
`characterized by anti-inflammatory effects. Although the two classes of drugs have some overlapping mechanisms of
`action, the IFNs rapidly block blood–brain barrier leakage and gadolinium (Gd) enhancement within 2 weeks, whereas
`glatiramer acetate produces less rapid resolution of Gd-enhanced MRI activity. IFN ␤ has no direct effects in the CNS, but
`glatiramer acetate-specific T cells are believed to have access to the CNS, where they can exert anti-inflammatory and possibly
`neuroprotective effects.
`NEUROLOGY 2002;58(Suppl 4):S3–S9
`
`Multiple sclerosis is a challenging disease in terms of
`our understanding of the etiology and underlying
`pathophysiology and in the design of effective thera-
`pies. Several factors, such as exposure to certain vi-
`ruses, are proposed to be involved in the etiology of MS.
`In addition, genes that encode HLA or T-cell receptor
`phenotypes may be important predisposing factors.1
`Regardless of etiology, MS appears to be immuno-
`logically mediated, as demonstrated by the observa-
`tion that MS responds to immunotherapy and by the
`existence of an animal model, experimental autoim-
`mune encephalomyelitis (EAE). The prevailing hy-
`pothesis is that autoreactive T cells of the CD4⫹ T
`helper (Th)1 population orchestrate the pathogenetic
`process in MS.2 These cells recognize antigen(s) pre-
`sented by macrophages or dendritic cells and are
`consequently activated to secrete proinflammatory
`cytokines: interleukin (IL)-1, interferon (IFN)␥, and
`tumor necrosis factor (TNF). These allow the upregu-
`lation of adhesion molecules and their ligands on the
`blood–brain barrier (BBB) endothelial cells and lym-
`phocytes, respectively. Autoreactive T cells can then
`adhere to the BBB endothelium and secrete metallo-
`proteinases. This leads to the digestion of the BBB
`matrix membrane, allowing activated T cells to in-
`vade the CNS. This phase of the process is believed
`to correlate with the appearance of gadolinium (Gd)-
`enhancing lesions on MRI.
`Amplification of immunoreactivity takes place in
`
`the CNS, where T cells are further activated by anti-
`gen(s) presented on microglia, resulting in the secre-
`tion of proinflammatory cytokines and chemokines
`that attract and retain inflammatory cells in the
`CNS. Effector mechanisms that mediate demyelina-
`tion include the ability of activated macrophages to
`strip myelin and to secrete myelinotoxic substances
`such as TNF␣, nitric oxide (NO), and free radicals.
`Additional mechanisms may include complement-
`dependent antibody-mediated damage and a direct at-
`tack on oligodendrocytes by CD8⫹ cytotoxic T cells.
`The extensive inflammation and the chronicity of
`the process may result in damage to axons, a marker
`of irreversible disability. Recovery is believed to be
`mediated by Th2 helper cells, which secrete anti-
`inflammatory cytokines, such as IL4, IL10, and
`transforming growth factor (TGF)␤, that can deacti-
`vate macrophages.
`Recent evidence suggests that MS may be a heter-
`ogeneous disease with various pathologic subtypes.3
`Therefore, it follows that increased understanding of
`the pathophysiologic processes of MS should enhance
`the design of more effective therapies.
`Over the past decade MS patients have benefited
`enormously from therapeutic research efforts. In
`1990 there were no drugs to treat MS, but today
`there are four FDA-approved treatments: IFN ␤-1a
`(Avonex), IFN ␤-1b (Betaseron), glatiramer acetate
`(Copaxone), and mitoxantrone (Novantrone). An-
`
`From the Department of Neurology, University of Maryland School of Medicine, and the Baltimore VA Medical Center, Baltimore, MD.
`Publication of this supplement was supported by an unrestricted educational grant from Teva Neuroscience. The sponsor has provided S.D-J. with honoraria
`and grant support, both in excess of $10,000, during his professional career.
`Address correspondence and reprint requests to: Dr. Suhayl Dhib-Jalbut, Department of Neurology, University of Maryland Hospital, 22 S. Greene St., Rm.
`N4W46, Baltimore, MD 21201.
`
`Copyright © 2002 by AAN Enterprises, Inc. S3
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`Figure 1. Type I and type II interferons
`(IFN␣, -␤, and -␥) bind to species-
`specific cell surface receptors. Binding
`induces a cascade of signaling path-
`ways that eventually lead to the secre-
`tion of IFN-stimulated gene products
`such as MxA protein and PKR. These
`gene products have immunomodulatory,
`antiviral, and antiproliferative actions
`that account for the usefulness of IFNs
`in the treatment of cancer, viral infec-
`tions, and MS.
`
`other formulation of IFN ␤-1a, Rebif, is available in
`many countries and is expected to become available
`in the United States. IFN ␤-1a and IFN ␤-1b are
`type I IFNs indicated for relapsing–remitting (RR)
`MS. Glatiramer acetate is also approved for RRMS,
`and mitoxantrone has been approved for worsening
`RRMS and for progressive relapsing and secondary
`progressive MS. This article reviews the current
`state of knowledge about the mechanisms of action of
`the drugs approved for RRMS, the IFNs and glati-
`ramer acetate.
`
`Interferons.
`IFNs are proteins secreted by cells in
`response to invading organisms. In general, they
`have antiviral and anti-inflammatory effects, and
`they modulate the immune system. The type I IFNs
`include IFN ␣, and IFN ␤. These are primarily pro-
`duced by fibroblasts and have
`strong anti-
`inflammatory properties. Type II IFN includes IFN␥,
`which is produced primarily by cells of the immune
`system. This review focuses on the type I IFNs,
`which are used clinically in the treatment of MS.
`The commercially available IFNs include IFN
`␤-1b and IFN ␤-1a. The difference between the two
`IFNs is that IFN ␤-1a is glycosylated, whereas IFN
`␤-1b is not. In addition, IFN ␤-1b contains one amino
`acid substitution from the natural molecule. Their
`biologic effects are probably quite similar, although
`there are purported differences in terms of antigenic-
`ity and other properties.4,5 Moreover, differences in
`S4
`NEUROLOGY 58(Suppl 4) April 2002
`
`dosage regimens and route of administration may
`produce different responses.6-11
`All IFNs bind to cell surface species-specific recep-
`tors. Binding induces a cascade of signaling path-
`ways, the end result of which is secretion or
`production of a number of proteins called IFN-
`stimulated gene products. These gene products are
`antiviral, antiproliferative, and immunomodulatory
`(figure 1). Experimental evidence in animal models
`and in humans indicates that IFN ␤ has several
`potential mechanisms of action in MS (table 1).12,13
`T-cell activation. T-cell activation occurs as a re-
`sult of T-cell receptor recognition of processed anti-
`gen in the context of HLA class II molecules
`expressed on antigen-presenting cells. The formation
`of this trimolecular complex alone is insufficient for
`T-cell activation. A second signal delivered by co-
`stimulatory molecular interaction, such as B7/CD28
`or CD40/CD40L, is required for T-cell activation. In
`
`Table 1 IFN␤ mechanisms of action in MS
`
`Reduction in T-cell activation
`Inhibition of IFN␥ effects
`Induction of immune deviation
`Inhibition of blood–brain barrier leakage
`CNS effects (?)
`Antiviral effect (?)
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`the absence of a second signal, T cells become aner-
`gic. Activated T cells can proliferate and differentiate
`into effector T cells, including Th and cytotoxic T
`cells. Th cells can be divided into two phenotypes.
`Th1 cells secrete inflammatory cytokines, which lead
`to macrophage activation and, in the case of MS,
`mediate destruction of myelin. Th2 cells secrete anti-
`inflammatory cytokines, inhibit the inflammatory ef-
`fects of Th1 cells, and activate B cells to produce
`antibodies.14
`Evidence from our work and that of others indi-
`cates that IFN ␤ can interfere with T-cell activation
`in several ways.12 First, IFN ␤ counteracts many of
`the proinflammatory effects of IFN ␥. This occurs
`primarily because of competition for shared signaling
`and transcription factors induced by these cytokines
`(figure 1). For example, IFN ␥ enhances HLA class II
`molecules, and this may be the mechanism by which
`IFN ␥ worsens MS.15 IFN ␤ inhibits the upregulation
`of HLA class II, which is believed to interfere with
`antigen processing and presentation, and conse-
`quently with T-cell activation.16 Second, IFN ␤ may
`have an effect on co-stimulatory molecule interac-
`tion, including B7/CD2817 and CD40:CD40L.18 By in-
`terfering with these two groups of molecules, IFN ␤
`could inhibit T-cell activation, including the activa-
`tion of myelin-reactive T cells.16
`Immune deviation. Several studies12,13 have
`shown that IFN␤ can tilt the balance in favor of an
`anti-inflammatory response either by inhibiting Th1
`or by promoting Th2 cytokine production. For exam-
`ple, IFN ␤ enhances peripheral blood mononuclear
`cell secretion of IL1019 and inhibits IL12,20 a key
`proinflammatory cytokine. It is unclear whether
`changes in these cytokines correlate with response to
`therapy. Immune deviation as a therapeutic mecha-
`nism for IFN ␤ in MS is controversial, in view of
`recent findings indicating that IFN ␤ can upregulate
`a number of proinflammatory gene products in hu-
`man peripheral blood mononuclear cells.21 This ap-
`pears to indicate that a proinflammatory response to
`IFN ␤ may in some way be beneficial, or that the net
`balance is in favor of an anti-inflammatory response,
`or that its mechanism of action may be entirely un-
`related to cytokine changes.
`Blood–brain barrier effects. MRI indicates that
`IFN ␤ has a prominent effect on the BBB. In NIH
`studies,22 almost 90% of MS patients treated with
`IFN showed a rapid and robust decrease in the num-
`ber of Gd-enhancing lesions on MRI. Although this
`may be the dominant mechanism of action of IFN␤
`in MS, the decrease in enhancing lesions does not
`necessarily correlate with the clinical response to
`IFN ␤ in the long run.
`IFN ␤ probably affects the BBB by two mecha-
`nisms: by interfering with T-cell adhesion to the en-
`dothelium, although the evidence for this is not
`strong,23 and by inhibiting the ability of T cells to get
`into the brain. Some MS patients treated with IFN ␤
`demonstrated a rise in serum soluble vascular cell
`adhesion molecule (sVCAM), which correlated with a
`
`reduction in the number of MRI Gd-enhancing le-
`sions.24 sVCAM may act as a decoy by binding VLA-4
`on T cells, thus inhibiting their attachment to the
`endothelium of the BBB.
`IFN ␤ may also interfere with T-cell/endothelial-
`cell adhesion by inhibiting HLA class II expression
`on endothelial cells, which can also function as li-
`gands for T cells.25 T cells secrete proteases and gela-
`tinases, one of which is matrix metalloproteinase 9
`(MMP9). MMP9 digests the matrix membrane and
`allows T cells to enter the brain. That MMP9 may be
`involved in MS pathogenesis is suggested by evi-
`dence of elevated levels of MMP9 in the spinal fluid
`of MS patients and its correlation with the number
`of enhancing lesions on MRI.26,27
`IFN ␤ inhibits MMP9 production by activated T
`cells,28,29 which may explain the dramatic effect of
`IFN ␤ in inhibiting the opening of the BBB. How-
`ever, the possibility that MMP9 may be a marker of
`injury rather than a therapeutic target for IFN ␤
`should be kept in mind.30
`Despite speculation that IFN ␤ may have an effect
`in the CNS because of in vitro evidence of an effect
`on glial cells in humans16,31 and the demonstration of
`accessibility to the CNS in healthy mice,32 there is no
`clinical evidence that IFN ␤ enters the brain in
`humans.
`Antiviral effects. MS may be caused by a virus in
`a subset of patients, on the basis of circumstantial
`evidence.33,34 The clinical courses of human herpesvi-
`rus (HHV) infections and MS have some similarities,
`such as chronicity, dormancy, reactivation in the
`case of herpes and relapses in the case of MS, and
`vulnerability to stress and hormonal imbalance.
`HHV6 in particular may be involved in the patho-
`genesis of MS in a subset of patients.35 It was re-
`cently reported that treatment with valaciclovir can
`decrease the number of MRI-enhancing lesions in a
`subset of MS patients with active disease.36 There-
`fore, if a viral infection is a cause of MS in some
`patients, IFN ␤ may have an additional therapeutic
`effect in this group through its antiviral properties.
`
`Glatiramer acetate. Glatiramer acetate (copoly-
`mer-I) is a synthetic molecule composed of four
`amino acids: glutamine, lysine, alanine, and ty-
`rosine. These four amino acids are represented in
`myelin basic protein (MBP), which is a suspect anti-
`gen involved in the induction of autoimmunity in
`MS.37 The polypeptide was originally produced in an
`attempt to mimic MBP and to induce EAE.38 Instead
`of inducing disease, the copolymer prevented the in-
`duction of EAE in animals.39 This finding triggered
`clinical studies of the use of glatiramer acetate in
`MS.40,41 Because of the randomness of the amino acid
`composition and the relatively short length of the
`peptide, glatiramer acetate has the ability to bind to
`HLA class II (DR) molecules, including HLA DR2.
`This binding property suggests several mechanisms
`of action, based on experimental evidence in EAE
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`Table 2 Potential mechanisms of action of glatiramer acetate in
`MS
`
`Inhibition of myelin-reactive T cells
`Induction of anergy in myelin-reactive T cells
`Induction of anti-inflammatory Th2 cells
`Bystander suppression in the CNS
`Neuroprotection
`
`and more recently in MS patients treated with the
`drug (table 2; figure 2).
`Inhibition of myelin-reactive T cells. Studies
`have demonstrated the ability of glatiramer acetate
`to inhibit the activation of MBP-reactive T cells.42
`Myelin-reactive Th1 clones exposed to increasing
`doses of glatiramer acetate manifest dose-dependent
`inhibition of proliferation and IFN ␥ production with
`relative antigen specificity.43 In addition, glatiramer
`acetate induces anergy in MBP-reactive T cells in
`vitro43 and modulates T-cell receptor recognition of
`the MBP-immunodominant peptide82–100.44 Collec-
`tively, these findings suggest that glatiramer acetate
`can interfere with T-cell activation.
`Induction of anti-inflammatory Th2 cells. Stud-
`ies both in EAE and in humans indicate that a likely
`in vivo mechanism of action of glatiramer acetate
`involves the induction of immunomodulatory Th2
`cells.42 Such glatiramer acetate-specific T cells may ex-
`ert their protective action by entering the CNS com-
`partment45 and by the production of anti-inflammatory
`cytokines in response to cross-recognition of myelin
`antigens (bystander suppression).
`Induction of glatiramer acetate-reactive Th2 cells
`presumably occurs because glatiramer acetate can
`act as an altered peptide ligand that delivers a weak
`signal to T cells, resulting in preferential Th2 cell
`activation.46 Our recent studies (unpublished data)
`indicate that glatiramer acetate-reactive Th2 cells
`are generated as early as 2 months after treatment
`is initiated47 and are sustained for up to 9 years
`(figure 3), despite a drop in the precursor frequency
`
`of these cells. A possible explanation for the sus-
`tained Th2 phenotype despite reduced proliferation
`is that glatiramer acetate results in a progressive
`deletion of high-affinity T cells or, alternatively, may
`induce a subset of nonproliferating immunoregula-
`tory T cells with anti-inflammatory properties.
`Bystander suppression in the CNS. Bystander
`suppression implies that glatiramer acetate-reactive
`T cells are capable of entering the CNS and recognizing
`cross-reactive antigen(s), probably myelin antigen(s).
`These T cells can then secrete anti-inflammatory cyto-
`kines and suppress inflammation. Although it is
`technically difficult
`to demonstrate glatiramer
`acetate-reactive T cells in the human CNS, recent
`EAE evidence supports this mechanism.45 Further-
`more, glatiramer acetate-reactive T cells may have a
`neuroprotective effect on neurons and axons.48 In MS
`patients, glatiramer acetate treatment reduces the
`proportion of new MS lesions that evolve into “black
`holes,”49 suggesting a potential neuroprotective ef-
`fect. Therefore, glatiramer acetate may have a bene-
`ficial clinical effect in the long term because axonal
`degeneration is believed to cause irreversible dam-
`age in MS. Whether glatiramer acetate acts system-
`ically, centrally, or both in humans is unclear. The
`fact that the drug inhibits the appearance of new
`MRI Gd-enhancing lesions50 could suggest a signifi-
`cant systemic effect.
`
`IFN␤
`IFN␤ and glatiramer acetate compared.
`and glatiramer acetate have different but overlap-
`ping mechanisms of action, and both ultimately re-
`sult in a decreased proinflammatory response in the
`periphery and the CNS (table 3). However, IFN␤
`rapidly blocks BBB leakage and Gd enhancement
`within 2 weeks, whereas glatiramer acetate activity
`on the BBB produces less rapid and dramatic resolu-
`tion of Gd-enhanced MRI activity.
`The clinically evident effects of glatiramer acetate
`may appear to be delayed compared with the onset of
`IFN effects. The time-dependent effect of treatment
`with glatiramer acetate was observed in the United
`
`Figure 2. Mechanism of action of glati-
`ramer acetate (GA) summarized. After
`SC injection, GA binds HLA class II
`(DR) on antigen-presenting cells in
`lymph nodes. As a result, GA can block
`the activation of myelin-reactive T cells
`or render these cells anergic. In addi-
`tion, GA induces GA-specific Th2 cells
`that cross the blood–brain barrier
`(BBB) and produce bystander suppres-
`sion as a result of cross-recognition of
`myelin antigens. These cells may also
`have a neuroprotective function.
`
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`Table 3 Comparison of activities of glatiramer acetate and INF␤
`
`Glatiramer
`acetate
`
`IFN␤
`
`Yes
`Yes
`
`Interference with T-cell activation
`Decrease in Th1 and enhancement
`of Th2 cytokines
`Yes
`Induces Th2 cells*
`Inhibits T-cell/BBB transmigration* No
`CNS effects
`Yes
`Neuroprotection
`Yes?
`Antibodies
`Inert
`
`* Critical differences.
`
`Yes
`Yes
`
`No
`Yes
`No
`No?
`Neutralizing
`
`with the delayed MRI effects. Therefore, although
`IFN ␤ has the desired effect of rapidly blocking in-
`flammation, it is less likely to have an effect in the
`CNS compartment once inflammation sets in be-
`cause there is no evidence that IFN ␤ can access the
`CNS in pharmacologically relevant concentrations.
`Therefore, glatiramer acetate-specific T cells may
`have a distinct ability to enter the CNS, downregu-
`late inflammation at the lesion site, and perhaps
`contribute to neuroprotection.
`
`Combination therapy. Few studies have ad-
`dressed the possibility of combining IFN ␤ and glati-
`ramer acetate.53-55 In vitro evidence53 suggests a
`possible additive effect on the inhibition of myelin-
`reactive T cells. There is also evidence that the com-
`bination of the two drugs is safe, as can be measured
`clinically and by MRI.55 On the other hand, IFN ␤
`has significant antiproliferative effects and therefore
`has the potential to inhibit the generation of glati-
`ramer acetate-reactive T cells (unpublished data). In
`addition, work in EAE mice has suggested that the
`combination of the two drugs is counterproductive.54
`Furthermore, because IFN ␤ blocks BBB leakage,22
`this could interfere with the migration of glatiramer
`acetate-specific T cells into the brain. If this is the
`case, the combination would be counterproductive.
`We recently addressed these concerns in a group
`of five MS patients receiving a combination of IFN
`␤-1a and glatiramer acetate treatment as part of a
`multicenter safety study.55 Specifically, we addressed
`the question of whether IFN ␤ interferes with the
`generation of glatiramer acetate-reactive Th2 cells,
`which are believed to underlie the mechanism of ac-
`tion of this drug. The data were compared with data
`obtained from a group of 12 MS patients receiving
`glatiramer
`acetate monotherapy. Glatiramer
`acetate-reactive T-cell lines from patients receiving
`monotherapy or combination therapy both showed
`Th2 bias, as reflected by increased levels of IL-5 and
`decreased levels of IFN ␥ (figure 4).56
`These findings suggest that the combination of the
`two drugs is unlikely to compromise the ability of
`glatiramer acetate to induce a Th2 response. What is
`unclear is whether IFN ␤ blocks the entry of the
`April 2002 NEUROLOGY 58(Suppl 4) S7
`
`Figure 3. Percentages of glatiramer acetate-reactive T-cell
`lines (TCL) classified as Th1, Th0, or Th2 were compared
`in patients who had had short-term (1–10 months; 73
`TCL) or long-term (6–9 years; 32 TCL) glatiramer acetate
`therapy, or who had not taken glatiramer acetate (52 TCL).
`Classification of Th phenotype was based on the ratio of
`IFN␥ (a Th1 marker) to IL5 (a Th2 marker). A ratio ⬎2 was
`classified as Th1, 0.5 to 2 as Th0, and ⬍0.5 as Th2.
`
`States pivotal trial51 and in the European/Canadian
`trial,50 and is consistent with the immunologic activ-
`ity of glatiramer acetate. That is, in vivo studies
`have suggested that, over time, glatiramer acetate
`treatment induces glatiramer acetate-specific T cells
`to proliferate and secrete anti-inflammatory cyto-
`kines that are typical of Th2 regulatory or suppres-
`sor T cells. A proportion of the glatiramer acetate-
`specific T cells can be cross-stimulated by MBP and
`its immunodominant fragments to secrete the same
`regulatory cytokines. In the EAE model, these cells
`confer protection from clinical disease.50 Patients
`treated with glatiramer acetate demonstrated a re-
`duction in proinflammatory cytokines and an in-
`crease in anti-inflammatory cytokines.52 Anti-
`inflammatory cytokines peaked during the first 6
`months of treatment and then gradually decreased,
`whereas proinflammatory cytokine levels continued
`to decrease.
`These immunologic observations are consistent
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`have a macrophage-dominant disease, and some
`might have a primary oligodendrocyte pathology.
`This pathologic diversity has implications for treat-
`ment. Some patients with antibody-mediated disease
`may respond well to plasmapheresis, for example,
`whereas patients with a macrophage-mediated dis-
`ease may respond to TNF inhibitors. The likelihood
`that MS is a syndrome of different diseases with
`different etiologies would explain the partial re-
`sponse to monotherapy and suggests a potential
`value for combination therapy. It also suggests that
`defining the pathologic process in individual patients
`may allow specific targeting of the process.
`
`Acknowledgment
`Dr. Dhib-Jalbut’s research is supported by grants from TEVA
`Pharmaceuticals, Berlex Laboratories, Serono, the National Insti-
`tute of Neurological Disorders and Stroke (K-24-NS02082), and
`the Department of Veterans Affairs.
`
`References
`1. Martin R, Dhib-Jalbut S. Immunology and etiologic concepts.
`In: Burks J, Johnson K, eds. Multiple sclerosis: diagnosis,
`medical management and rehabilitation. New York: Demos
`Medical Publishing, 2000:141–165.
`2. Martin R, Sturzebecher CS, McFarland HF. Immunotherapy
`of multiple sclerosis: where are we? Where should we go?
`Nature Immunol 2001;2:785–788.
`3. Lucchinetti C, Bruck W, Parisi J, Scheithauer B, Rodriguez
`M, Lassmann H. Heterogeneity of multiple sclerosis lesions:
`implications for the pathogenesis of demyelination. Ann Neu-
`rol 2000;47:707–717.
`4. Alam J, Goelz S, Rioux P, et al. Comparative pharmacokinetics and
`pharmacodynamics of two recombinant human interferon beta-1a
`(IFN beta-1a) products administered intramuscularly in healthy
`male and female volunteers. Pharmacol Res 1997;14:546–549.
`5. Allain H, Schuck S. Observations on differences between in-
`terferons used to treat multiple sclerosis. J Clin Res 1998;1:
`381–392.
`6. Khan OA, Xia Q, Bever CT Jr, Johnson KP, Panitch HS,
`Dhib-Jalbut SS. Interferon beta-1b serum levels in multiple
`sclerosis patients following subcutaneous administration.
`Neurology 1996;46:1639–1643.
`7. Khan OA, Dhib-Jalbut SS. Serum interferon beta-1a (Avonex)
`levels following intramuscular injection in relapsing-remitting
`MS patients. Neurology 1998;51:738–742.
`8. Buraglio M, Trinhcard-Lugan, Munafo A, Macnamee M. Re-
`combinant human interferon beta-1a (Rebif) vs recombinant
`interferon beta-1b (Betaseron) in healthy volunteers. Clin
`Drug Invest 1999;18:27–34.
`9. Durelli et al., and the Independent Comparison of Interferon
`(INCOMIN) Trial Study Group. A multicenter trial comparing
`clinical and MRI efficacy of Betaseron and Avonex in multiple
`sclerosis. Neurology 2001;56:A148. Abstract.
`10. OWIMS (Once Weekly Interferon for MS) Study Group. Evi-
`dence for interferon beta-1a dose response in relapsing remit-
`ting MS. Neurology 1999;53:679–686.
`11. Deisenhammer F, Mayringer I, Harvey J, et al. A comparative
`study of the relative bioavailability of different interferon beta
`preparations. Neurology 2000;54:2055–2060.
`12. Dhib-Jalbut S. Mechanisms of interferon beta action in multi-
`ple sclerosis. Mult Scler 1997;3:397–401.
`13. Yong VW, Chabot S, Stuve O, Williams G. Interferon beta in
`the treatment of multiple sclerosis: mechanisms of action.
`Neurology 1998;51:682–689.
`14. Delves PJ, Roitt IM. The immune system. Second of two parts.
`N Engl J Med 2000;343:108–117.
`15. Panitch HS, Hirsch RL, Schindler J, Johnson KP. Treatment
`of multiple sclerosis with gamma interferon: exacerbations
`associated with activation of the immune system. Neurology
`1987;37:1097–1102.
`16. Jiang H, Milo R, Swoveland P, Johnson KP, Panitch H, Dhib-
`Jalbut S. Interferon beta-1b reduces interferon gamma-
`
`Figure 4. Interferon␤-1a does not interfere with the gener-
`ation of glatiramer acetate-specific T cells when the two
`drugs are used in combination. Patients on monotherapy
`and combination therapy both showed increased levels of
`IL-5 and decreased levels of IFN␥.
`
`glatiramer acetate Th2 cells into the CNS. This pos-
`sibility is speculative, because activated T cells can
`cross the BBB and focal permeability is not required.57
`Recent evidence also indicates that Th2 cells preferen-
`tially migrate to the CNS.58 Furthermore, there is no
`evidence in human patients that the therapeutic ef-
`fect of glatiramer acetate necessarily involves a di-
`rect CNS effect, and no studies in EAE have
`indicated that an IFN ␤ countereffect takes place.
`
`Comment. As our understanding of the patho-
`physiology of MS continues to develop, we hope that
`we will be able to apply new knowledge to the ratio-
`nale of the drug development process. Therapies for
`MS are only partially effective, and there is no cure
`as yet. In addition, the available therapies have the
`drawback of administration by injection, a factor that
`can affect long-term compliance. Therefore, new drugs
`are needed that effectively target the immunologic pro-
`cesses of MS, including agents that can be adminis-
`tered orally. Despite variable results in early trials,56,59
`studies of oral formulations of IFN, glatiramer ace-
`tate (i.e., the CORAL study), and myelin continue.
`MS is probably a heterogeneous disease, suggest-
`ing that the pathologic process may differ signifi-
`cantly from patient to patient. Some patients may
`have primarily T-cell–mediated disease, others may
`S8
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`MYLAN PHARM. v YEDA
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`induced antigen-presenting capacity of human glial and B
`cells. J Neuroimmunol 1995;61:17–25.
`17. Genc K, Dona DL, Reder AT. Increased CD80(⫹) B cells in
`active multiple sclerosis and reversal by interferon beta-1b
`therapy. J Clin Invest 1997;99:2664–2671.
`18. Teleshova N, Bao W, Kivisakk P, Ozenci V, Mustafa M, Link
`H. Elevated CD40 ligand expressing blood T-cell levels in
`multiple sclerosis are reversed by interferon-beta treatment.
`Scand J Immunol 2000;51:312–320.
`19. Rep MH, Schrijver HM, van Lopik T, et al. Interferon-beta
`treatment enhances CD95 and interleukin 10 expression but
`reduces interferon-gamma producing T cells in MS patients.
`J Neuroimmunol 1999;96:92–100.
`20. Wang X, Chen M, Wandinger KP, Williams G, Dhib-Jalbut S.
`IFN beta-1b inhibits IL-12 production in peripheral blood
`mononuclear cells in an IL-10-dependent mechanism: rele-
`vance to IFN beta-1b therapeutic effects in multiple sclerosis.
`J Immunol 2000;165:548–557.
`21. Wandinger KP, Sturzebecher CS, Bielekova B, et al. Complex
`immunomodulatory effects of interferon beta in multiple scle-
`rosis include the upregulation of T helper 1-associated marker
`genes. Ann Neurol 2001;50:349–357.
`22. Calabresi PA, Stone LA, Bash CN, et al. Interferon beta re-
`sults in immediate reduction of contrast-enhanced MRI le-
`sions in multiple sclerosis patients followed by weekly MRI.
`Neurology 1997;48:1446–1448.
`23. Dhib-Jalbut S, Jiang H, Williams GJ. The effect of interferon
`beta-1b on lymphocyte-endothelial cell adhesion. J Neuroim-
`munol 1996;71:215–222.
`24. Calabresi PA, Tranquill LR, Dambrosia JM, et al. Increases in
`soluble VCAM-1 correlate with a decrease in MRI lesions in
`multiple sclerosis treated with interferon beta-1b. Ann Neurol
`1997;41:669–674.
`25. Huynh HK, Oger J, Dorovini-Zis K. Interferon beta downregulates
`interferon gamma-induced class II MHC molecule expression and
`morphological changes in primary cultures of human brain mi-
`crovessel endothelial cells. J Neuroimmunol 1995;60:63–73.
`26. Lee MA, Smith S, Palace J, et al. Spatial mapping of T2 and
`gadolinium-enhancing T1 lesion volumes in multiple sclerosis:
`evidence for distinct mechanisms of lesion genesis? Brain
`1999;122(pt 7):1261–1270.
`27. Waubant E, Goodkin DE, Gee L, et al. Serum MMP-9 and
`TIMP-1 levels are related to MRI activity in relapsing multi-
`ple sclerosis. Neurology 1999;53:1397–1401.
`28. Stuve O, Dooley NP, Uhm JH, et al. Interferon beta-1b de-
`creases the migration of T lymphocytes in vitro: effects on
`matrix metalloproteinase-9. Ann Neurol 1996;40:853–863.
`29. Leppert D, Waubant E, Burk MR, Oksenberg JR, Hauser SL.
`Interferon beta-1b inhibits gelatinase secretion and in vitro
`migration of human T cells: a possible mechanism for treatment
`efficacy in multiple sclerosis. Ann Neurol 1996;40:846–852.
`30. Bever CT Jr, Rosenberg GA. Matrix metalloproteinases in
`multiple sclerosis: targets of therapy or markers of injury?
`Neurology 1999;53:1380–1381.
`31. Ransohoff RM, Devajyothi C, Estes ML, et al. Interferon beta
`specifically inhibits interferon gamma-induced class II major
`histocompatibility complex gene transcription in a human as-
`trocytoma cell line. J Neuroimmunol 1991;33:103–112.
`32. Pan W, Banks WA, Kastin AJ. Permeability of the blood-brain
`and blood-spinal cord barriers to interferons. J Neuroimmunol
`1997;76:105–111.
`33. Gilden DH, Devlin ME, Burgoon MP, Owens GP. The search for
`virus in multiple sclerosis brain. Mult Scler 1996;2:179–183.
`34. Zhao ZS, Granucci F, Yeh L, Schaffer PA, Cantor H. Molecu-
`lar mimicry by Herpes simplex virus-type 1: autoimmune dis-
`ease after viral infection. Science 1998;279:1344–1347.
`35. Challoner PB, Smith KT, Parker JD, et al. Plaque-associated
`expression of human herpesvirus 6 in multiple sclerosis. Proc
`Natl Acad Sci USA 1995;92:7440–7444.
`36. Jakobsen J, Bech E, Gadelberg P, et al. A placebo-controlled,
`randomized, double-blind, MRI trial of antiherpes therapy in
`patients with relapsing-remitting multiple sclerosis (RRMS).
`Neurology 2001;56(suppl 3):A74. Abstract.
`37. Paterson PY. Experimental allergic encephalomyelitis and au-
`toimmune disease. Adv Immunol 1966;5:131–208.
`38. Arnon R. The development of Cop 1 (Copaxone威), an innova-
`tive drug for the treatment of multiple sclerosis: personal
`reflections. Immunol Lett 1996;50:1–15.
`
`39. Teitelbaum D, Meshorer A, Hirshfeld T, et al. Suppression of
`experimental allergic encephalomyelitis by a synthetic
`polypeptide. Eur J Immunol 1971;1:242–248.
`40. Bornstein MB, Miller A, Slagle S, et al. A pilot trial of Cop 1
`in exacerbating-remitting multiple sclerosis. N Engl J