Table of Contents For Ex. 1011A
`PCT Publication no. 2008/097596 A2
`
`Front page ------------------------------------------------------------- page 2
`
`Specification ---------------------------------------------------------- pages 3 - 36
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`Claims ------------------------------------------------------------------ pages 37 - 39
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`Drawings --------------------------------------------------------------- pages 40 - 43
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`
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`Page 1 of 43
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`(12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT)
`
`(19) World Intellectual Property Organization
`International Bureau
`
`(43) International Publication Date
`14 August 2008 (14.08.2008)
`
`(51) International Patent Classification:
`GOIN 33/50 (2006.01)
`
`peT
`
`(21) International Application Number:
`PCTIUS2008/001602
`
`(22) International Filing Date: 7 February 2008 (07.02.2008)
`
`(25) Filing Language:
`
`(26) Publication Language:
`
`English
`
`English
`
`(30) Priority Data:
`60/888,921
`
`8 February 2007 (08.02.2007) US
`
`(71) Applicant (jor all designated States except US): BIOGEN
`IDEC MA INC. [USIUS]; 14 Cambridge Center, Cam(cid:173)
`bridge, MA 02142 (US).
`
`(72) Inventor; and
`(75) Inventor/Applicant (for us only):
`Matvey, E. [USIUS]; 3 Louis Road,
`01876 (US).
`
`LUKASHEV,
`Tewksbury, MA
`
`11111111111111111111111111111111111111111111111111111111111111111111111111111111111111111111111
`
`(10) International Publication Number
`WO 2008/097596 A2
`(81) Designated States (unless otherwise indicated, for every
`kind of national protection available): AE, AG, AL, AM,
`AO, AT, AU, AZ, BA, BB, BG, BH, BR, BW, BY, BZ, CA,
`CH, CN, CO, CR, CU, CZ, DE, DK, DM, DO, DZ, EC, EE,
`EG, ES, Fl, GB, GD, GE, GH, GM, GT, HN, HR, HU, ID,
`IL, IN, IS, JP, KE, KG, KM, KN, KP, KR, KZ, LA, LC,
`LK, LR, LS, LT, LU, LY, MA, MD, ME, MG, MK, MN,
`MW, MX, MY, MZ, NA, NG, NI, NO, NZ, OM, PG, PH,
`PL, PT, RO, RS, RU, SC, SD, SE, SG, SK, SL, SM, SV,
`SY, TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN,
`ZA, ZM,ZW.
`
`(84) Designated States (unless otherwise indicated, for every
`kind of regional protection available): ARIPO (BW, GH,
`GM, KE, LS, MW, MZ, NA, SD, SL, SZ, TZ, UG, ZM,
`ZW), Eurasian (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM),
`European (AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, Fl,
`FR, GB, GR, HR, HU, IE, IS, IT, LT, LU, LV, MC, MT, NL,
`NO, PL, PT, RO, SE, SI, SK, TR), OAPI (BF, BJ, CF, CG,
`CI, CM, GA, GN, GQ, GW, ML, MR, NE, SN, TD, TG).
`
`(74) Agent: GARRETT, Arthur, S.; Finnegan, Henderson,
`Farabow, Garrett & Dunner L.L.P., 901 New York Avenue,
`NW, Washington, DC 0001-4413 (US).
`
`Published:
`without international search report and to be republished
`upon receipt of that report
`
`-iiiiiiii
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`iiiiiiii
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`--!!!!!!!!
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`--
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`!!!!!!!!
`iiiiiiii
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`iiiiiiii ----
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`\0
`0\
`In
`t---
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`Q
`' - ------------------------------------------------------------------------------------------
`~ (54) Title: NRF2 SCREENING ASSAYS AND RELATED METHODS AND COMPOSITIONS
`Q
`o ment of neurological diseases, such as, e.g., multiple sclerosis (MS). The compounds described upregulate the cellular cytoprotective
`M
`(57) Abstract: Provided are certain methods of screening, identifying, and evaluating neuroprotective compounds useful for treat(cid:173)
`> pathway regulated by Nrf2. Also provided are certain methods of utilizing such compounds in therapy for neurological disease, par(cid:173)
`
`~ ticularly, for slowing or reducing demyelination, axonal loss, or neuronal and oligodendrocyte death.
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`Nrf2 SCREENING ASSAYS
`AND RELATED METHODS AND COMPOSITIONS
`
`[0001]
`
`Provided are certain compounds for treating neurological diseases,
`
`including demyelinating neurological diseases, such as, e.g., multiple sclerosis.
`
`[0002]
`
`Multiple sclerosis (MS) is an autoimmune disease with the autoimmune
`
`activity directed against central nervous system (eNS) antigens. The disease is
`
`characterized by inflammation in parts of the eNS, leading to the loss of the myelin
`
`sheathing around neuronal axons (demyelination), loss ofaxons, and the eventual
`
`death of neurons, oligodenrocytes and glial cells.
`
`[0003] An estimated 2,500,000 people in the world suffer from MS. It is one of
`
`the most common diseases of the eNS in young adults. MS is a chronic, progressing,
`
`disabling disease, which generally strikes its victims some time after adolescence, with
`diagnosis generally made between 20 and 40 years of age, although onset may occur
`
`earlier. The disease is not directly hereditary, although genetic susceptibility plays a
`part in its development. Relapsing-remitting MS presents in the form of recurrent
`
`attacks of focal or multifocal neurologic dysfunction. Attacks may occur, remit, and
`
`recur, seemingly randomly over many years. Remission is often incomplete and as
`
`one attack follows another, a stepwise downward progression ensues with increasing
`permanent neurological deficit.
`
`[0004] Although various immunotherapeutic drugs can provide relief in
`
`patients with MS, none is capable of reversing disease progression, and some can
`
`cause serious adverse effects. Most current therapies for MS are aimed at the
`
`reduction of inflammation and suppression or modulation of the immune system. As of
`
`2006, the available treatments for MS reduce inflammation and the number of new
`
`episodes but not a/l have an effect on disease progression. A number of clinical trials
`
`have shown that the suppression of inflammation in chronic MS rarely significantly
`limits the accumulation of disability through sustained disease progression, suggesting
`
`that neuronal damage and inflammation are independent pathologies. Promoting eNS
`
`remyelination as a repair mechanism and otherwise preventing axonal loss and
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`neuronal death are some of the important goals for the treatment of MS. For a
`
`comprehensive review of MS and its current therapies, see, e.g., McAlpine's Multiple
`
`Sclerosis, by Alastair Compston et aI., 4th edition, Churchill Livingstone Elsevier,
`
`2006.
`
`[0005]
`
`"Phase 2 enzymes" serve as a protection mechanism in mammalian
`
`cells against oxygen/nitrogen species (ROS/RNS), electrophiles and xenobiotics.
`
`These enzymes are not normally expressed at their maximal levels and, their
`
`expression can be induced by a variety of natural and synthetic agents. Nuclear factor
`
`E2-related factor 2 (Nrf2) is a transcription factor responsible for the induction of a
`
`variety of important antioxidant and detoxification enzymes that coordinate a protective
`
`cellular response to metabolic and toxic stress.
`
`[0006] ROS/RNS are most damaging in the brain and neuronal tissue, where
`
`they attack post-mitotic (Le., non-dividing) cells such as glial cells, oligodendocytes,
`
`and neurons, which are particularly sensitive to free radicals. This process leads to
`
`neuronal damage. Oxidative stress has been implicated in the pathogenesis of a
`
`variety of neurodegenerative diseases, including ALS, Alzheimer's disease (AD), and
`
`Parkinson's disease (PO). For review, see, e.g., van Muiswinkel et aI., Curr. Drug
`
`Targets CNS--Neurol. Disord., 2005,4:267-281. An anti-oxidative enzyme under
`
`control of Nrf2, NQ01 (NAD(P)H dehydrogenase, quinone (1), was recently reported
`
`to be substantially upregulated in the brain tissues of AD and PO subjects (Muiswinkel
`
`et aI., Neurobiol. Aging, 2004, 25: 1253). Similarly, increased expression of NQ01
`
`was reported in the ALS subjects' spinal cord (Muiswinkel et aI., Curr. Drug
`
`Targets--CNS. Neural. Disord., 2005,4:267-281) and in active and chronic lesions in
`
`the brains of patients suffering from MS (van Horssen et aI., Free Radical BioI. & Med.,
`
`2006,41 311-311). These observations indicate that the Nrf2 pathway may be
`
`activated in neurodegenerative and neuroinflammatory diseases as an endogenous
`
`protective mechanism. Indeed, most recently, it has been reported that induced
`
`activation of Nrf2-dependent genes by certain cyclopenanone-based compounds
`
`(NEPP) counters the toxic effects of metabolic inhibition and ROS/RNS production in
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`the brain and protects neurons from death in vitro and in vivo (see Satoh et aI., PNAS,
`
`2006, 103(3):768-773).
`
`[0007] Additionally, many publications have reported neuroprotective effects of
`
`compounds in natural plant-derived compounds ("phytochemicals"), including
`
`a-tocopherol (vitamin E), Iycopene (tomatoes), resveratrol (red grapes), sulforaphane
`
`(broccoli), EGCG (green tea), etc. For review, see Mattson and Cheng, Trends in
`
`Neurosci., 2006, 29(11 ):632-639. Originally, the action of these compounds was
`
`attributed to their anti-oxidant properties. However, while most anti-oxidants are
`
`effective only at high concentrations, at least some of these compounds appear to
`
`exert neuroprotective effects at much lower doses. Emerging evidence suggests that
`
`these compounds may exert their neuroprotective effects by activating cellular
`
`stress-response pathways, including the Nrf2 pathway, resulting in the upregulation of
`
`neuroprotective genes. However, the exact mechanism of action of these compounds
`remains poorly understood.
`
`[0008] To date, more than 10 different chemical classes of inducers of Nrf2
`
`pathway have been identified including isothiocyanates and their thiol addition
`
`products, dithiocarbamates, as well as 1 ,2-dithiole-3-thiones, trivalent arsenic
`
`derivatives (e.g., phenyl arsenoxide), heavy metals, certain conjugated cyclic and
`
`acyclic polyenes (including porphyrins, chlorophyllins, and chlorophyll), and vicinal
`
`dimercaptans. These inducers have few structural similarities. They are mostly
`
`electrophiles, and all can react chemically with thiol groups by alkylation, oxidation, or
`
`reduction, suggesting that the intracellular sensor for inducers is likely to contain very
`
`highly reactive (cysteine) thiols. The inducers can modify thiol groups by a variety of
`
`mechanisms including: alkylation (Michael addition acceptors, isothiocyanates,
`
`quinones); oxidation (e.g., peroxides and hydroperoxides); and direct reaction with
`
`thiolldisulfide linkages (e.g., vicinal dithiols such as 1,2-dimercaptopropanol, lipoic
`
`acid). These diverse response mechanisms provide plasticity for cellular responses to
`a variety of electrophilic and oxidant stressors.
`
`[0009] Provided are methods that comprise at least one of the following
`methods:
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`1) methods of screening for at least one new candidate compound for
`
`treating a neurological disease;
`
`2) methods of evaluating neuroprotective properties of at least one drug
`candidate for treating a neurological disease;
`
`3) methods of comparing (e.g., for bioequivalence) at least two
`
`pharmaceutical compositions which comprise fumaric acid derivatives;
`
`4) methods of treating a neurological disease by administering to the subject
`
`in need thereof at least one compound that is partially structurally similar to
`
`DMF or MMF; and
`
`5) methods of treating a neurological disease by a combination therapy that
`
`comprises administration of at least one first compound that upregulates
`
`the Nrf2 pathway and at least one second compound that does not
`
`upregulate the Nrf2 pathway.
`
`[0010]
`
`In some embodiments, the neurological disease is a neurodegenerative
`
`disease such as, for example, ALS, Parkinson's disease, Alzheimer's disease, and
`
`Huntington's disease. In some embodiments the neurological disease is MS or
`
`another demyelinating neurological disease.
`
`[0011]
`
`In some embodiments, the methods 1-3 further comprise:
`
`a) contacting a cell with the test compound, and
`b) determining whether the Nrf2 pathway is upregulated in the cell.
`
`In some embodiments, the methods may further comprise:
`
`c) determining whether the test compound slows or prevents demyelination,
`
`axonal loss, and/or neuronal death, and/or
`
`d) selecting the test compound as a candidate for treating neurodegeneration
`
`in a neurological disease if 1) the Nrf2 pathway is upregulated and 2)
`
`demyelination, axonal loss, and/or neuronal death are/is prevented or
`
`slowed.
`[0012]
`In some embodiments, the methods 1-3 comprise contacting a cell with
`
`at least one test compound and determining whether the Nrf2 pathway is upregulated
`
`in the cell. In such methods, an upregulation of the Nrf2 pathway above a threshold
`
`-4-
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`(e.g., by at least 30% over a control) indicates that the at least one compound has at
`
`least one biological property beneficial in treating a neurological disease (e.g.,
`
`neuroprotective properties). In some embodiments, the upregulation of the Nrf2
`
`pathway is assessed (in vivo and/or in vitro) by at least one of the following:
`
`i) expression levels of endogenously produced and/or exogenously
`
`introduced Nrf2;
`
`ii) subcellular localization and/or nuclear translocation of Nrf2;
`
`iii) expression levels and/or activity of one or more genes under control of
`
`Nrf2 (e.g., endogenous NQ01) or an Nrf2-regulated reporter gene in an
`
`artificial reporter construct;
`
`iv) levels of Nrf2 binding to the Nrf2-binding DNA element ARE;
`
`v) stability of Nrf2/Keap1 complexes; and
`
`vi) modification (e.g., alkylation) levels of Keap1 and/or at least one other
`
`Nrf2/Keap1-associated proteins.
`
`[0013]
`
`In some embodiments of methods 1-3, the compounds that are being
`
`screened, evaluated, or compared comprise at least one member of at least one of the
`
`following classes of compounds: mild alkylating agents, Michael addition acceptors,
`
`and compounds that are metabolized upon administration to Michael addition
`
`acceptors. In some embodiments, the Michael addition acceptor has the structure of
`
`Formula I, II, III, or IV set forth below.
`
`[0014]
`
`In some embodiments method 1 comprises:
`
`a) contacting a cell with a plurality of test compounds,
`
`b) determining whether the Nrf2 pathway is upregulated in the cell, and
`
`c) selecting from the plurality of compounds at least one compound that
`
`upregulates the Nrf2 pathway,
`
`wherein an upregulation of the Nrf2 pathway by the selected at least one compound
`
`indicates that the selected at least one compound may be useful for treating a
`
`neurological disease. The plurality of compounds may be represented, e.g., by a
`
`combinatorial chemical library, and the method may be performed, e.g., by
`
`high-throughput screening.
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`[0015]
`
`In some embodiments method 2 comprises:
`
`a) contacting a cell with the at least one drug or drug candidate, and
`
`b) determining whether the Nrf2 pathway is upregulated in the cell,
`
`wherein an upregulation of the Nrf2 pathway by the at least one drug or drug
`
`candidate indicates that the at least one drug or drug candidate is useful for
`
`neuroprotection in treating a human having a neurological disease.
`
`[0016]
`
`In some embodiments method 3 comprises:
`
`a) contacting a cell with a first composition comprising at least one test
`
`compound, and
`b) comparing the level of Nrf2 pathway upregulation in the cell by the at least
`
`one test compound to the corresponding level of the Nrf2 pathway
`upregulation in a control cell treated with a second composition comprising
`
`at least one of DMF and MMF.
`
`[0017]
`
`In some embodiments of method 3, the test compound is fumaric acid,
`
`a salt thereof, or a fumaric acid derivative. In some embodiments, the first
`
`composition comprises DMF, MMF, or both. In some embodiments, the dose and/or
`
`the formulation of the first composition differs from the dose and/or the formulation of
`
`the second composition.
`
`[0018]
`
`In some embodiments, method 3 further comprises:
`
`c) comparing at least one pharmacokinetic parameter (e.g., serum-half-life) of
`
`the first and the second compositions.
`
`[0019]
`
`In some embodiments method 4 comprises administering to the
`
`mammal a therapeutically effective amount of at least one neuroprotective compound
`
`having Formula I, II, III, or IV, e.g., a fumaric acid derivative (e.g., DMF or MMF).
`
`[0020]
`
`In some embodiments method 4 provides a method of slowing or
`
`preventing neurodegeneration in a patient in need thereof, by administering the
`
`compound in an amount and for a period of time sufficient to slow or prevent
`
`demyelination, axonal loss, and/or neuronal death, e.g., by at least 30% relative to a
`
`control.
`
`[0021]
`
`In some embodiments method 5 comprises:
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`a) administering to the mammal a therapeutically effective amount of at least
`
`one first compound that upregulates the Nrf2 pathway, and
`
`b) administering a therapeutically effective amount of at least one second
`
`compound that does not upregulate the Nrf2 pathway.
`
`[0022]
`
`In some embodiments of method 5, the at least one first compound,
`
`used in step (a), is a compound of Formula I, II, III, or IV, e.g., a fumaric acid derivative
`
`(e.g., DMF or MMF); and the at least one second compound, which is used in step (b),
`
`is an immunosuppressive or an immunomodulatory compound that does not
`
`upregulate the Nrf2 pathway (e.g., by more than 30% over a control).
`
`[0023]
`
`In some embodiments method 5 comprises administering to the
`
`mammal a therapeutically effective amount of a compound of Formula I, II, III, or IV.
`
`[0024]
`
`In some embodiments of methods 1-5, the at least onecompound being
`
`screened, identified, evaluated, or used for treating a neurological disorder is not
`
`fumaric acid or its salt, or a fumaric acid derivative (e.g., DMF or MMF).
`
`[0025] Other features and embodiments of the invention will be apparent from
`
`the following description and the claims.
`
`BRIEF DESCRIPTION OF THE FIGURES
`
`[0026] Figure 1 demonstrates that DMF and MMF are activators of Nrf2 at
`
`concentrations within clinical exposure range (cells in culture).
`
`[0027] Figure 2 shows results of RNAi experiments.
`[0028] Figure 3 shows evidence of Nrf2 activation by DMF and MMF In vivo.
`
`[0029] Figure 4 shows evidence of Nrf2 activation by DMF and MMF In vivo.
`
`[0030] Fumaric acid esters, such as DMF, have been proposed for treatment
`of MS (see, e.g., Schimrigk et a/., Eur. J. Neuro/., 2006,13(6):604-10; Drugs R&D,
`2005,6(4):229-30).
`
`[0031] Provided are, among other things, means for identifying compounds
`
`with a new therapeutic modality useful in at least one of multiple neurological
`
`indications and, optionally, complementary to other drugs for the treatment of a
`
`neurological disease, including a number of currently used immunomodulators.
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`[0032] DMF is a member of a large group of anti-oxidant molecules known for
`
`their cytoprotective and anti-inflammatory properties. These molecules also share the
`
`property of the Nrf2 pathway activation. Thus, the finding that DMF activates the Nrf2
`
`pathway in conjunction with the neuroprotective effects of DMF further offers a
`
`rationale for identification of structurally and/or mechanistically related molecules that
`would be expected to be therapeutically effective for the treatment of neurological
`
`disorders, such as, e.g., MS.
`
`[0033] Certain terms are defined in this section; additional definitions are
`
`provided throughout the description.
`
`[0034] The terms "activation" and "upregulation," when used in reference to
`
`the Nrf2 pathway, are used interchangeably herein.
`
`[0035] The terms "disease" and "disorder" are used interchangeably herein.
`
`[0036] The term "a drug for treating a neurological disease" refers to a
`
`compound that has a therapeutic benefit in a specified neurological disease as shown
`
`in at least one animal model of a neurological disease or in human clinical trials for the
`
`treatment of a neurological disease.
`
`[0037] The term "neuroprotection" and its cognates refer to prevention or a
`
`slowing in neuronal degeneration, including, for example, demyelination and/or axonal
`
`loss, and/or, neuronal and/or oligodendrocyte death. Neuroprotection may occur
`
`through several mechanisms, e.g., through reducing inflammation, providing
`
`neurotrophic factors, scavenging free radicals, etc. As used herein, a compound is
`
`considered neuroprotective if it (1) upregulates the Nrf2 pathway above a certain
`
`threshold and (2) provides neuroprotection, regardless of possible other mechanisms
`of action.
`
`[0038] The terms "treatment," "therapeutic method," "therapeutic benefits," and
`
`the like refer to therapeutic as well as prophylactic/preventative measures. Thus,
`
`those in need of treatment may include individuals already having a specified disease
`
`and those who are at risk for acquiring that disease.
`
`[0039] The terms "therapeutically effective dose" and "therapeutically effective
`
`amount" refer to that amount of a compound which results in at least one of prevention
`
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`or delay of onset or amelioration of symptoms of a neurological disorder in a subject or
`
`an attainment of a desired biological outcome, such as reduced neurodegeneration
`
`(e.g., demyelination, axonal loss, and neuronal death) or reduced inflammation of the
`
`cells of the eNS.
`[0040]
`In one aspect, provided are methods of evaluating neuroprotective
`properties of test compounds, including the following methods:
`
`1) methods of screening for new candidate compounds that may be
`
`useful for treating a neurological disease;
`
`2) methods of evaluating neuroprotective properties of drugs and
`
`candidates that are used or proposed for treating a neurological
`
`disease;
`
`3) methods of comparing (e.g., for bioequiva/ence) two or more
`
`pharmaceutical compositions which contain fumaric acid
`
`derivatives;
`
`[0041]
`
`In some embodiments, methods 1-3 may comprise:
`
`a) contacting a cell with the test compound,
`
`b) determining whether the Nrf2 pathway is upregulated in the cell,
`
`and, in some embodiments, additionally performing the following step(s):
`
`c) determining whether the test compound slows or prevents demyelination,
`
`axonal loss, and/or neuronal death, and/or
`
`d) selecting the test compound as a candidate for treating neurodegeneration
`
`in a neurological disease if 1) the Nrf2 pathway is upregulated and 2)
`
`demyelination, axonal loss, and/or neuronal death arelis prevented or
`
`slowed.
`
`Method 1
`[0042]
`
`In some embodiments the methods of screening for a candidate
`
`compound for treating a neurological disease comprise:
`
`a) contacting a cell with a plurality of test compounds,
`
`b) determining whether the Nrf2 pathway is upregulated in the cell, and
`
`c) selecting from the plurality of compounds at least one compound that
`
`upregulates the Nrf2 pathway,
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`wherein an upregulation of the Nrf2 pathway by the selected at least one compound
`
`indicates that the selected at least one compound may be useful for treating a
`
`neurological disease. For example, the plurality of compounds may be represented by
`
`a combinatorial chemical library, and the screening method may be performed by a
`
`high-throughput screening as described in, e.g., High-Throughput Screening in Drug
`
`Discovery (Methods and Principles in Medicinal Chemistry), by Jorg HOser (ed.), John
`
`Wiley & Sons (2006).
`
`[0043] Combinatorial libraries of compounds are also described in, e.g.,
`
`Solid-Supported Combinatorial and Parallel Synthesis of Small-Molecular-Weight
`
`Compound Libraries (Tetrahedron Organic Chemistry) Ian Salusbury (ed.), Elsevier
`
`(1998); Combinatorial Libraries: Synthesis, Screening and Application Potential
`
`(Library Binding), by Riccardo Cortese (ed.), Walter de Gruyter (1995). The libraries of
`
`compounds may be, for example, quinone libraries and other libraries as described in
`
`Mittoo, Comb. Chem. & High Throughput Screen, 2006, 9:421-423.
`
`[0044]
`
`In some embodiments, the at least one compound or plurality of
`
`compounds being screened and/or selected comprises at least one compound
`
`selected from at least one of the following groups of compounds: mild alkylating
`
`agents, Michael addition acceptors or compounds that are metabolized to Michael
`
`addition acceptors, including compounds of Formulas I, II, III, or IV.
`
`[0045]
`
`In some of the embodiments, the at least one compound is selected
`
`from fumaric acid, its salts, and fumaric acid derivatives.
`
`Method 2
`
`[0046] Also provided are methods of evaluating neuroprotective properties of
`
`at least one drug or drug candidate for treating at least one neurological disease.
`
`Such methods comprise:
`
`a) contacting a cell with the at least one drug or drug candidate, and
`
`b) determining whether the Nrf2 pathway is upregulated in the cell,
`
`wherein the upregulation of the Nrf2 pathway by the at least one drug or drug
`
`candidate indicates that the at least one drug or drug candidate is neuroprotective in
`
`treating a human having a neurological disease.
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`[0047]
`
`In some embodiments, the upregulation of the Nrf2 pathway by the at
`
`least one drug or drug candidate indicates that the at least one drug or drug candidate
`
`has at least one activity selected from slowing demyelination, slowing the loss of
`
`axons, and slowing the rate of neuronal death.
`
`[0048]
`
`In some embodiments, the method of evaluating at least one drug or
`
`drug candidate comprises an additional step:
`
`c) evaluating demyelination, loss ofaxons, and/or neuronal death.
`
`[0049]
`
`In some embodiments, steps a) and c) are performed in vivo in at least
`
`one model of a neurological disease, e.g., as described below.
`
`[0050]
`
`In other embodiments, particularly those in which the neurological
`
`disease is multiple sclerosis or another demyelinating disease, the evaluated at least
`
`one drug or drug candidate for a neurological disease is chosen from the following:
`
`FTY720 (2-(4-octylphenethyl)-2-aminopropane-1 ,3-diol; Novartis); anti-IL 12 antibody
`
`(e.g., ABT-874; Abbott Laboratories); GSK683699 (GSK/Tanabe); NeuroVax (Immune
`
`Response Corp.; Darlington, Curr. Opin. Mol. Ther., 2005, 7(6):598-603); anti-CCR2
`
`antibody (e.g., MLN 1202; Millennium); interferon 13-1 a (e.g., Avonex®; Biogen Idec);
`
`anti-a4-integrin antibody (e.g., Tysabri®; Biogen Idec/Elan); anti-CD20 antibody (e.g.,
`
`Rituxan® (Biogen Idec/Genentech); TV 5010 (Teva); NBI-788 (Neurocrine); MBP8298
`(BioMS (see Warren et aI., Eur. J. NeuroL, 2006, 13(8):887-95); Mylinax (Oral
`Cladribine; 2-chlorodeoxyadenosine; Serono/lV AX); Teriflunomide
`
`«Z)-2 -cyano-N-( 4-(trifluoromethyl )phenyl )-3-hyd roxybut -2 -enam ide; Sanofi-Aventis);
`
`Temsirolimus (Wyeth); Laquinimod
`
`(5-chloro-N-ethyl-1,2 -dihyd ro-4-hyd roxy-1-methyl-2 -oxo-N-phenylqui noline-3-carboxa
`
`mide; Active BiotechlTeva); and interferon tau (Tauferon; Pepgen).
`
`[0051]
`
`In some embodiments, the at least one drug or drug candidate being
`
`evaluated is at least one compound selected from at least one class selected from a
`
`mild alkylating agent, a Michael addition acceptor, and a compound that is
`
`metabolized to a Michael addition acceptor, including compounds of Formulas I, II, III,
`
`or IV.
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`[0052]
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`In some of the embodiments, the compound is fumaric acid, its salt, or
`
`a fumaric acid derivative.
`
`Method 3
`
`[0053] Also provided are methods of comparing (e.g., for bioequivalence) at
`
`least two pharmaceutical compositions. Such methods comprise:
`a) contacting a cell with at least one first composition comprising a test
`
`compound, and
`
`b) comparing the level of the Nrf2 pathway upregulation in the cell by the test
`
`compound to the corresponding level of the Nrf2 pathway upregulation in a
`cell treated with at least one second composition ("comparator
`
`composition") comprising DMF, MMF, or both.
`
`[0054]
`
`In some embodiments, substantially dissimilar levels of upregulation by
`
`the at least one first and at least one second compositions indicate that the
`compositions are not bioequivalent.
`
`[0055]
`
`In some embodiments, the test compound is fumaric acid, its salt
`
`thereof, a fumaric acid derivative, or mixtures thereof. In some embodiments, the first
`
`composition comprises at least one of DMF, MMF, and both DMF and MMF. In some
`
`embodiments, the dose and/or the formulation of the at least one first composition
`
`differs from the dose and/or the formulation of the at least one second composition.
`The at least one first composition may be a controlled release composition such as,
`
`e.g., compositions described in WO 2006/037342.
`
`[0056]
`
`In some embodiments, the method further comprises and additional
`
`step:
`
`c) comparing at least one pharmacokinetic parameter of the at least one first
`
`and the at least one second compositions.
`
`[0057] Pharmacokinetic parameters and methods for evaluating the same are
`
`well known and are described in, e.g., Pharmacokinetics, Second Edition (Drugs and
`
`the Pharmaceutical Sciences) by Milo Gibaldi et al. (eds.), Informa Healthcare (1982).
`
`Examples of such pharmacokinetic parameters that can be evaluated include serum
`half-life, clearance, and volume distribution.
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`[0058]
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`In some embodiments, substantially dissimilar pharmacokinetic
`
`parameter(s) of the a least one first and at least one second compositions indicate that
`
`the compositions are not bioequivalent.
`
`[0059]
`
`In some embodiments, the test compound being evaluated is a mild
`
`alkylating agent, and more specifically, a Michael addition acceptor, or a compound
`
`that is metabolized to a Michael addition acceptor.
`
`[0060]
`
`In some of the embodiments, the test compound is fumaric acid or its
`
`salt, or a fumaric acid derivative.
`
`[0061J Also provided are methods of treating a mammal who has or is at risk
`
`for developing a neurological disease, including the following methods:
`
`4) methods of treating a neurological disease by administering to the subject
`
`in need thereof at least one compound that is partially structurally similar
`
`to DMF or MMF (including compounds selected using methods 1-3
`
`described above) ; and
`
`5) methods of treating a neurological disorder by a combination therapy that
`
`includes administration of a first compound that does not upregulate the
`
`Nrf2 pathway and a second compound that upregulates the Nrf2 pathway.
`
`Method 4
`
`[0062] Also provided are methods of treating a neurological disease by
`
`administering to the subject in need thereof at least one compound that is at least
`
`partially structurally similar to DMF and/or MMF.
`
`[0063]
`
`In some embodiments of method 4, a method of treating a mammal
`
`who has or is at risk for a neurological disease is provided. The methods comprises
`
`administering to the mammal a therapeutically effective amount of at least one
`
`neuroprotective compound which has Formula I, II, III, or IV, e.g., a fumaric acid
`derivative (e.g., DMF or MMF).
`
`In some embodiments of method 4, a method of slowing or preventing
`[0064J
`neurodegeneration (more specifically, e.g., demyelination, axonal loss, and/or
`
`neuronal death) in a subject in need thereof, by administering the at least one
`
`compound in an amount and for a period of time sufficient to do at least one of slow or
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`prevent demyelination, slow or prevent axonal loss, and alow or prevent neuronal
`
`death, e.g., by at least 30%,50%, 100% or higher over a control over a period of at
`
`least 5, 10, 12, 20, 40, 52, 100, or 200 weeks, or more.
`
`Method 5
`
`[0065] Also provided are methods of treating a mammal having a neurological
`
`disease by combination therapy. In some embodiments such methods comprise:
`
`a) administering to the mammal a therapeutically effective amount of at least
`
`one first compound that upregulates the Nrf2 pathway, and
`
`b) administering a therapeutically effective amount of at least one second
`
`compound that does not upregulate the Nrf2 pathway.
`
`[0066]
`
`In some of embodiments of method 5, the at least one first compound,
`
`used in step (a), is a compound of Formula I, II, III, or IV, e.g., DMF or MMF; and the
`
`at least one second compound, which is used in step (b), is an immunosuppressive or
`
`an immunomodulatory compound that does not upregulate the Nrf2 pathway (e.g., by
`
`more than 30%, 50%, 100% over a control).
`
`[0067]
`
`In some embodiments of method 5, the method comprises
`
`administering to