WO2006105795A1 - Novel indolinone derivatives - Google Patents

Abstract

The invention relates to novel indolinone derivatives of formula I wherein X, Ri, R2, R3, R4, Rs, Re, Ri', FV/ FV/ IV and R5' are as shown herein as well as their use in the treatment of inflammatory autoimmune diseases such as multiple sclerosis, in particular for administration by the oral route.

Description

NOVEL INDOLINONE DERIVATIVES

FIELD OF INVENTION

The present invention relates to derivatives of indolinone compounds and their inclusion in pharmaceutical compositions intended for oral administration as well as their use in the treatment of inflammatory autoimmune diseases such as multiple sclerosis.

BACKGROUND OF THE INVENTION

Multiple sclerosis is an autoimmune inflammatory disease of the central nervous system characterised by T-cell infiltration, demyelination of white matter and axonal injury. The disease mostly affects young adults with an onset at 20-40 years of age and affects twice as many women as men (A. Compton and A. Coles, The Lancet 359, 6 April 2002, pp. 1221-1231). Multiple sclerosis is more common in temperate climate zones and thus has a prevalence of 50-130 out of 100,000 in northern Europe and North America (N. Hellings et al., Immunologic Research 25(1), 2002, pp. 27-51). While the higher incidence and prevalence of multiple sclerosis in certain European populations has not been adequately explained, it is believed that increased genetic susceptibility in these populations is partly responsible. The presence of a genetic element in the etiology of the disease is supported by family studies showing that first-degree relatives of multiple sclerosis patients have a 20-40 times increased risk of developing the disease relative to the general population (J. H. Noseworthy et al., New England Journal of Medicine 343(13), 2000, pp. 938-952). Furthermore, it has been recognised that populations with a high frequency of for instance the HLA-DR2 allele have a significantly higher risk of developing multiple sclerosis (Hellings et al., supra; Noseworthy et al., supra). However, no single major susceptibility gene for multiple sclerosis has been identified so far, and the results of genome screens conducted to identify susceptibility genes rather point to multiple genes exerting a moderate effect (Hellings et al., supra).

Based on these studies, it would appear that genetic susceptibility is not enough in itself to provoke multiple sclerosis. This theory is given credence by the fact that the rate of prevalence of multiple sclerosis among people of European descent living outside Europe is half of that persisting in parts of northern Europe and that the low frequency of multiple sclerosis in Africans increases significantly among first-generation decendants living in Europe (Compton and Coles, supra). Environmental factors have therefore also been proposed as contributing to the development of multiple sclerosis. In particular, it is believed that certain antigens present on pathogenic organisms such as viral or bacterial epitopes which structurally resemble autoantigenic epitopes of, for instance, myelin basic protein, proteolipid protein, myelin-associated glycoprotein or oligodendrocyte glycoprotein, which are all components of the myelin sheath, may lead to activation of T-cells that are reactive with such antigenic epitopes and initiating the inflammatory process eventually resulting in clinical manifestations of multiple sclerosis. This phenomenon is generally referred to as molecular mimicry (Hellings et al., supra; A. Bar-Or et al., J. Neuroimmunol. 100, 1999, pp. 252-259; A. Kami and H. L. Weiner, "Organ-Specific Inflammatory Diseases" Chapter 77 in Clinical Immunology; Principles and Practice, 2^nd Ed. (R.R. Rich et al., Eds.), Mosby, London, 2001).

Multiple sclerosis is usually defined as either a relapsing-remitting or a progressive disease. The relapsing-remitting form with which 80% of the patients are initially afflicted (Compton and Coles, supra) is characterised by discrete attacks with full or partial recovery between relapses. In 40-50% of the patients, the disease eventually becomes progressive (secondary progressive stage). The disease may also be progressive from the outset (primary progressive form) characterised by a gradual decline in neurological function with no periods of remission. The clinical symptoms of the relapsing-remitting form of multiple sclerosis may vary widely from one patient to the other, but commonly affected individuals initially experience some degree of visual and sensory impairment, limb paresthesias, limb weakness, clumsiness, fatigue and gait ataxia, while in the later stages cognitive impairment, progressive quadriparesis, sensory loss, ataxic tremors, pain and spasticity are more common (Noseworthy et al., supra). The primary progressive form may initially manifest as one or more of these symptoms, gradually declining into quadriparesis, cognitive decline, visual loss, brainstem syndromes and cerebellar, bowel and bladder dysfunction (Noseworthy et al., supra).

Pathologically, multiple sclerosis is characterised by the presence of demyelinated plaques or sclerotic lesions where the myelin sheath surrounding the axons is destroyed. The inflammatory infiltrate in the lesions is composed of T-cells, B-cells, microglia and macrophages which interact with the myelin sheath and participate in the demyelinating process by local production of immune-related molecules such as adhesion molecules, cytokines and chemokines as well as demyelinating antibodies, oxygen free radicals and nitric oxide (Kami and Weiner, supra). While axonal destruction is not pronounced in the early stages of the disease (although more pronounced in patients suffering from the primary progressing form), demyelination of the axons results in slowing and blocking conductivity (Noseworthy et al., supra). Regression of the symptoms may be associated with partial remyelination after the initial inflammation has subsided showing that oligodendrocytes (myelin-producing cells) are present in the lesions (Kami and Weiner, supra). In later stages, irreversible axonal injury, gliotic scarring and gradual loss of oligodendrocyte progenitor cells may result from repeated episodes of inflammatory attack and lead to permanent loss of neurological function (Noseworthy et al., supra).

While the immunopathogenesis of multiple sclerosis is still largely unknown, it has been shown that autoreactive T-cells specific for myelin basic protein and other antigens of the central nervous system exist in the periphery of healthy individuals as wells as individuals who later develop multiple sclerosis (Bar-Or et al., supra; O'Connor et al., J. Clin. Immunol. 21(2), 2001, pp. 81-93). Thus, the presence of myelin-reactive T-cells in the periphery is not enough in itself to explain the development of multiple sclerosis. In multiple sclerosis patients, these T-cells become activated, possibly by cross-reactivity with bacterial or viral antigens that structurally resemble myelin antigens (i.e. the phenomenon known as molecular mimicry) and/or by bacterial superantigens, and persist in an enhanced state of activation (Hellings et al., supra). It has been found that the autoreactive T-cells are predominantly CD4+ T helper cells type 1 (ThI) producing interleukin-2 (IL-2), interferon^ (IFN-γ) and tumour necrosis factor (TNF-α) (B. Gran and A. Rostami, Current Neurology and Neuroscience Reports 1, 2001, pp. 263-270). In order for such proinflammatory T-cells to migrate to the central neurvous system, they express chemokine receptors, adhesion molecules and matrix metalloproteinases that enable them to cross the blood-brain barrier. Thus, it has been found that expression levels of the chemokines which are chemotactic for ThI cells, IP-10 and RANTES, and their corresponding receptors, CXCR3 and CCR5, are elevated in sclerotic lesions and cerebrospinal fluid of multiple sclerosis patients (Bar-Or et al., supra). Altered levels of the adhesion molecules ICAM-I and VCAM-I have been identified on endothelial cells of multiple sclerosis lesions (O'Connor et al., supra). ICAM-I and VCAM-I are important for endothelial-leukocyte interactions and leukocyte extravasation. Matrix metalloproteinases expressed by activated T-cells, monocytes and astrocytes may disrupt the basement membrane of the blood-brain barrier and facilitate transmigration of T-cells and breakdown of the extracellular matrix (O'Connor et al., supra).

Once the T-cells have entered the central nervous system they become reactivated on encountering the autoantigen, e.g. myelin basic protein, presented by MHC class II expressing antigen presenting cells (microglia and dendritic cells), and the ThI cells respond by producing proinflammatory cytokines such as TNF-α, IFN-γ and IL-2, while the Th2 cells produce anti-inflammatory cytokines such as IL-4, IL-5 and IL-10 (Bar-Or et al., supra). In turn, the inflammatory process leads to up-regulation of MHC class II expression and adhesion molecules on the blood-brain barrier endothelium, facilitating a further influx of T-cells, B-cells and macrophages and hence an amplification of the inflammatory response (Hellings et al., supra). This theory is supported by the finding that myelin basic protein reactive T-cell clones from multiple sclerosis patients were found to secrete increased amounts of different cytokines such as TNF-α, IL-2 and IL-IO (Hellings et al., supra). Demyelination (myelin destruction) is believed to be brought about by the combined effects of cytotoxic cells (macrophages and T-cells), oxygen free radicals, demyelinating autoantibodies and cytokine-induced toxicity (Hellings et al., supra).

Traditionally, corticosteroids such as prednisolone have been administered intravenously to multiple sclerosis patients during acute relapses in order to attenuate the inflammatory response. It has been found that treatment with corticosteroids during relapses reduces the duration of relapses and their short-term morbidity, but not the permanent disabilities resulting from repeated relapses (Compton and Coles, supra). Furthermore, treatment with potent corticosteroids at high doses has serious side effects, notably osteoporosis, aseptic bone necrosis, skin atrophy, striae cutis, insomnia, myopathy, posterior and capsular cataract and glaucoma as well as reactivation of the disease upon cessation of treatment. More recently, interferon-β (IFN-β) has been introduced as a treatment of relapsing-remitting multiple sclerosis and has been found to decrease the rate of relapse, increase the proportion of patients who were relapse free and reduce the number of patients who had moderate to severe relapses. On the other hand, INF-β treatment is extremely costly and its long-term efficacy has not been established. There is concern that the treatment may induce the formation of neutralising antibodies that may reduce the activity of IFN-β (Noseworthy et al., supra). Most of the patients initially experience flu-like symptoms when treated with IFN-β. Glatiramer acetate is another recent treatment based on a mixture of random synthetic peptides intended to mimic myelin basic protein. In a double-blind trial of relapsing- remitting multiple sclerosis, glatiramer acetate was found to decrease the rate of relapse. Glatiramer acetate is believed to be most effective for mildly disabled patients with a recent diagnosis of multiple sclerosis. Fewer treatment options exist for patients in the progressive phase of the disease. Immunosuppressive therapy, e.g. with cyclophosphamide or methotrexate, is frequently attempted, but it is generally recognised that once the disease enters the progressive stage treatment is very difficult. IFN-β has been in clinical trials for secondary progressive multiple sclerosis but the results did not show that the treatment slowed progression of disability and the benefits of this treatment in secondary progressive disease are controversial.

Moreover, IFN-β and glatiramer acetate are peptidic in nature and as such must be administered parenterally, e.g. by injection. This is clearly less convenient to patients than an oral treatment. It would therefore constitute a significant improvement compared to existing multiple sclerosis therapies if a medicament suitable for oral administration were to be developed.

SUMMARY OF THE INVENTION

In the course of research leading to the present invention, certain compounds within the class of indolinones were found to exhibit a surprisingly high level of activity in experimentally induced autoimmune encephalomyelitis (EAE), making them potentially useful as immunomodulatory agents in the treatment of autoimmune inflammatory diseases such as multiple sclerosis. However, these compounds are generally highly lipophilic and are therefore less suitable for administration by the oral route. Further experimental work has led to the development of derivatives of indolinone compounds showing a high activity in vivo when administered orally.

Accordingly, the present invention relates to a compound of formula I

I wherein X is O or S

Ri is -OC(O)R₉, -NR₁₀Ru, -C(O)NR₁₀Ru, -OC(O)NRi₀Rn, -NHC(O)Ri₀, -NHC(O)ORi₀, - NHC(O)NR₁₀Ru, -S(O)R₉, -S(O)₂R₉, -S(O)OR₉, -S(O)₂NR₁₀R₁₁, -OR₁₂, -C(O)R₁₂, - C(O)OR₁₂, -OC(O)OR₁₂, -P(O)(OR₁₀)(OR₁₁), -OP(O)(OR₁₀)(ORu), polyoxyethylene, C_1-10 alkyl, C_2-I0 alkenyl, C₂-io alkynyl, aryl, carbocyclyl, heteroaryl or heterocyclyl, said Ci_-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, carbocyclyl, heteroaryl or heterocyclyl being substituted with one or more substituents selected from the group consisting of -OR₁₀, - C(O)R₁₀, -C(O)OR₁₀, -OC(O)R₁₀, -OC(O)OR₁₀, -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -OC(O)NR₁₀R₁₁, - OP(O)(OR₁₀)(OR₁₁), -P(O)(OR₁₀)(OR₁₁), -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, - SR₁₀, -S(O)R₁₀, -S(O)₂R₁₀, -S(O)₂NR₁₀Rn, -S(O)₂OR₁₀, polyoxyethylene, aryl, heteroaryl, carbocyclyl and heterocyclyl, said aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with -OR₁₀, -C(O)R₁₀, -C(O)OR₁₀, -OC(O)R₁₀, -OC(O)OR₁₀, - P(O)(ORi₀)(OR_n), -OP(O)(OR₁₀)(OR₁₁), -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -NHC(O)R₁₀, - NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, -S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁, -S(O)₂OR₁₀ or polyoxyethylene, and C_1-6 alkyl substituted with polyoxyethylene, -OR₁₀, -C(O)R₁₀, - C(O)OR₁₀, -OC(O)R₁₀, -OC(O)OR₁₀, -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -P(O)(OR₁₀)(OR₁₁), - OP(O)(OR₁₀)(OR₁₁), -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, - S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁ or -S(O)₂OR₁₀, wherein

Rg is C_1-10 alkyl, C_2-10 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl or heterocyclyl, said C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl or heterocyclyl being optionally substituted with one or more substituents selected from the group consisting of halogen, trihalomethyl, C_1-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, polyoxyethylene, -ORi₃, -C(O)Ri₃, -C(O)ORi₃, -OC(O)R₁₃, - OC(O)OR₁₃, -NR₁₃R₁₄, -C(O)NR₁₃R₁₄, -OC(O)NR₁₃R₁₄, -NHC(O)R₁₃, -NHC(O)OR₁₃, - NHC(O)NR₁₃R₁₄, -P(O)(OR₁₃)(OR₁₄), -OP(O)(OR₁₃)(OR₁₄), -SR₁₃, -S(O)R₁₃, -S(O)₂R₁₃, - S(O)₂NR₁₃R₁₄ and -S(O)₂OR₁₃; R₁₀ and R₁₁ are the same or different and independently selected from the group consisting of hydrogen, polyoxyethylene, C_1-I0 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₀ and R₁₁, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, said Ci- ₁₀ alkyl being substituted with one or more, same or different substituents selected from the group consisting of halogen, trihalomethyl, C_1-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, polyoxyethylene, -OR₁₃, -C(O)R₁₃, -C(O)OR₁₃, -OC(O)R₁₃, - OC(O)OR₁₃, -N₃, -NR₁₃Ri₄, -C(O)NR₁₃R₁₄, -OC(O)NR₁₃R₁₄, -NHC(O)R₁₃, -NHC(O)OR₁₃, - NHC(O)NR₁₃R₁₄, -P(O)(OR₁₃)(OR₁₄), -OP(O)(OR₁₃)(OR₁₄), -SR₁₃, -S(O)R₁₃, -S(O)₂R₁₃, - S(O)₂NR₁₃R₁₄ and -S(O)₂OR₁₄, said aryl, heteroaryl, carbocyclyl or heterocyclyl being optionally substituted with one or more substituents selected from the group consisting Of -OR₁₃, -C(O)R₁₃, -C(O)OR₁₃, -C(O)NRi₃Ri₄ and polyoxyethylene, and said C_2-I0 alkenyl, C₂-_I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, Ci_-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, polyoxyethylene, -OR₁₃, -C(O)Ri₃, -C(O)ORi₃, -OC(O)Ri₃, -OC(O)ORi₃, - NRi₃R₁₄, -C(O)NRi₃Ri₄, -OC(O)NRi₃R₁₄, -NHC(O)Ri₃, -NHC(O)OR₁₃, -NHC(O)NRi₃Ri₄, - P(O)(OR₁₃)(OR₁₄), -OP(O)(OR₁₃)(OR₁₄), -SRi₃, -S(O)Ri₃, -S(O)₂Ri₃, -S(O)₂NRi₃Ri₄ and - S(O)₂ORi₄, said aryl, heteroaryl, carbocyclyl or heterocyclyl being optionally substituted with one or more substituents selected from the group consisting of -ORi₃, -C(O)Ri₃, - C(O)ORi₃, -C(O)NRi₃Ri₄ and polyoxyethylene, provided that R_i0 and Ru are not both hydrogen;

Ri₂ is Ci-io alkyl, C_2- io alkenyl, C_2-I0 alkynyl, aryl or carbocyclyl substituted with hydroxy, amino, cyano, halogen, carboxy, nitro, amido, polyoxyethylene, -ORi₃, -C(O)Ri₃, - C(O)ORi₃, -OC(O)Ri₃, -OC(O)ORi₃, -NR₁₃Ri₄, -C(O)NRi₃Ri₄, -OC(O)NRi₃Ri₄, -NHC(O)Ri₃, -NHC(O)ORi₃, -NHC(O)NRi₃Ri₄, -P(O)(ORi₃)(OR₁₄), -OP(O)(ORi₃)(ORi₄), -SR_i3, -S(O)R_i3, -S(O)₂R₁₃, -S(O)₂NRi₃Ri₄ and -S(O)₂OR₁₄, or heteroaryl or heterocyclyl optionally substituted with hydroxy, amino, cyano, halogen, carboxy, nitro, amido, polyoxyethylene, -OR₁₃, -C(O)R₁₃, -C(O)OR₁₃, -OC(O)R₁₃, -OC(O)OR₁₃, -NR₁₃Ri₄, - C(O)NRi₃Ri₄, -OC(O)NRi₃R₁₄, -P(O)(ORi₃)(ORi₄), -OP(O)(ORi₃)(ORi₄), -NHC(O)R₁₃, - NHC(O)OR₁₃, -NHC(O)NRi₃Ri₄, -SRi₃, -S(O)Ri₃, -S(O)₂Ri₃, -S(O)₂NRi₃Ri₄ and - S(O)₂OR₁₄;

Ri₃ and R_i4 are the same or different and independently selected from the group consisting of hydrogen, Ci_-6-alkyl, C_2-6-alkenyl, C_4-6-alkadienyl, C_2-6-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R_i3 and Ri₄, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, each Ci- ₆ alkyl, C_2-6 alkenyl, C_2-6 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl substituent being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, hydroxy, Ci_-4 alkyl, Ci_-4 alkoxy, nitro, cyano, amino, oxo, halogen, trihalomethyl, Ci_-4 alkylthio, Ci_-4 alkylamino, Ci_-4 alkoxycarbonyl, carboxy, -CONH₂ or -S(O)₂NH₂;

R_2/ R₃, R₄ and R₅ are the same or different and independently selected from the group consisting of hydrogen, halogen, trihalomethyl, Ci_-10 alkyl, C_2-I0 alkenyl, C_2-io alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, -ORi₅, -C(O)Ri₅, -C(O)ORi₅, -OC(O)Ri₅, -OC(O)OR₁₅, - NR₁₅R₁₆, -C(O)NR₁₅R₁₆, -OC(O)NR₁₅R₁₆, -NHC(O)R₁₅, -NHC(O)ORi₅, -NHC(O)NRi₅Ri₆, - SRi₅, -S(O)Ri₅, -S(O)₂Ri₅, -S(O)₂NRi₅Ri₆ and -S(O)₂ORi₅, wherein R₁₅ and Ri₆ are the same or different and independently selected from the group consisting of hydrogen, Ci- ₁₀ alkyl, C_2-I0 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₅ and Ri₆, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, said Ci_-I0 alkyl, C_2-I0 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, Ci_-6-alkyl, C₂-₆-alkenyl, C_2-6-alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, - ORi₇, -C(O)R₁₇, -C(O)ORi₇, -OC(O)ORi₇, -OC(O)Ri₈, -NRi₇Ri₈, -C(O)NRi₇Ri₈, - OC(O)NRi₇Ri₈, -NHC(O)Ri₇, -NHC(O)OR₁₇, -NHC(O)NRi₇Ri₈, -SR₁₇, -S(O)R₁₇, -S(O)₂R₁₇, - S(O)₂NR₁₇R₁₈ and -S(O)₂OR₁₇, wherein R₁₇ and R₁₈ are the same or different and independently selected from the group consisting of hydrogen, C_1-6-alkyl, C_2-6-alkenyl, C_2-6-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₇ and R₁₈, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, each d_-6-alkyl, C_2-6-alkenyl, C_2-6-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl substituent being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, hydroxy, C_x-4 alkyl, C_1-4 alkoxy, nitro, cyano, amino, oxo, halogen, trihalomethyl, C_1-4 alkylthio, C_1-4 alkylamino, C_1-4 alkoxycarbonyl, carboxy, -CONH₂ or -S(O)₂NH₂;

Re is hydrogen, C_1-6 alkyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, halogen, -OR₇, - C(O)R₇, -C(O)OR₇, -NR₇R₈, S(O)₂NR₇R₈, wherein R₇ and R₈ are independently hydrogen, C_1-6 alkyl, aryl or heterocyclyl, said C_1-6 alkyl or heterocyclyl being optionally substituted by heterocyclyl, -OR₇, -C(O)R₇ or -C(O)OR₇;

Ri', R₂' R₃', R₄' and R₅' are the same or different and independently selected from the group consisting of hydrogen, halogen, trihalomethyl, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, -OR₁₅, -C(O)R₁₅, -C(O)OR₁₅, -OC(O)R₁₅, - OC(O)OR₁₅, -NR₁₅R₁₆, -C(O)NR₁₅R₁₆, -OC(O)NR₁₅R₁₆, -NHC(O)R₁₅, -NHC(O)OR₁₅, -

NHC(O)NR₁₅R₁₆, -SR₁₅, -S(O)R₁₅, -S(O)₂R₁₅, -S(O)₂NR₁₅R₁₆ and -S(O)₂OR₁₆, wherein R₁₅ and R₁₆ are the same or different and independently selected from the group consisting of hydrogen, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₅ and R₁₆, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, said C_1-10 alkyl, C_2-I0 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, C_1-6-alkyl, C_2-6-alkenyl, C_2-6-alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, -OR₁₇, -C(O)R₁₇, -C(O)OR₁₇, -OC(O)R₁₈, -OC(O)OR₁₇, -NR₁₇R₁₈, -C(O)NR₁₇R₁₈, -OC(O)NR₁₇R₁₈, -NHC(O)R₁₇, -NHC(O)OR₁₇, -NHC(O)NR₁₇R₁₈, -SR₁₇, -S(O)R₁₇, -S(O)₂R₁₇, -S(O)₂NR₁₇R₁₈ and -S(O)₂OR₁₇, wherein R₁₇ and R₁₈ are the same or different and independently selected from the group consisting of hydrogen, C_1-6-alkyl, C_2-6-alkenyl, C_2-6-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₇ and R₁₈, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, each C_1-6-alkyl, C_2-6-alkenyl, C_2-6-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl substituent being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, hydroxy, C_1-4 alkyl, C_1-4 alkoxy, nitro, cyano, amino, oxo, halogen, trihalomethyl, C_1-4 alkylthio, C_1-4 alkylamino, C_1-4 alkoxycarbonyl, carboxy, -CONH₂ or -S(O)NH₂; or pharmaceutically acceptable salts thereof.

In another aspect, the invention relates to a pharmaceutical composition comprising one or more compounds of formula I together with a pharmaceutically acceptable excipient or vehicle.

In a further aspect, the invention relates to the use of a compound of general formula I for the preparation of a medicament for preventing, treating or ameliorating multiple sclerosis, or delaying the onset of or reducing the relapse rate in multiple sclerosis.

In a still further aspect, the invention relates to a method of preventing, treating or ameliorating multiple sclerosis, or delaying the onset of or reducing the relapse rate in multiple sclerosis, the method comprising administering to a patient in need thereof an effective amount of a compound of formula I.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

In the present context, the term "Ci-u-alkyl" is intended to mean a linear or branched hydrocarbon group having 1 to 12 carbon atoms, such as methyl, ethyl, propyl, iso- propyl, butyl, te/t-butyl, /so-butyl, pentyl, hexyl, etc. Analogously, the terms "C_1-10 alkyl" and "C_1-6-alkyl" are intended to denote a linear or branched hydrocarbon group having 1 to 10 or 1 to 6 carbon atoms, respectively, such as methyl, ethyl, propyl, /so-propyl, pentyl or hexyl, and the term "C_1-4-alkyl" is intended to cover linear or branched hydrocarbon groups having 1 to 4 carbon atoms, e.g. methyl, ethyl, propyl, /so-propyl, butyl, /so-butyl, te/t-butyl.

Similarly, the terms "C_2-i₂-alkenyl" are intended to cover linear, cyclic or branched hydrocarbon groups having 2 to 12, 4 to 12 or 6 to 12 carbon atoms and comprising one, two or three unsaturated bonds. Examples of alkenyl groups are vinyl, allyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl, heptadecaenyl, butadienyl, pentadienyl, hexadienyl, heptadienyl, heptadecadienyl, hexatrienyl, heptatrienyl, octatrienyl and heptadecatrienyl. Preferred examples of alkenyl are vinyl, allyl, butenyl, especially allyl.

Similarly, the term "C_2-i₂-alkynyl" is intended to mean a linear or branched hydrocarbon group having 2 to 12 carbon atoms and comprising a triple bond. Examples hereof are ethynyl, propynyl, butynyl, octynyl, and dodecaynyl.

The term "halogen" or "halo" includes fluoro, chloro, bromo, and iodo.

In the present context the term "aryl" is intended to mean a fully or partially aromatic carbocyclic ring or ring system, such as phenyl, naphthyl, 1,2,3,4-tetrahydronaphthyl, anthracyl, phenanthracyl, pyrenyl, benzopyrenyl, fluorenyl and xanthenyl, among which phenyl is a preferred example.

The term "heteroaryl" is intended to mean a fully or partially aromatic carbocyclic ring or ring system, usually a mono- or bicyclic ring system comprising 5-12 ring atoms, where one or more of the carbon atoms have been replaced with heteroatoms, e.g. nitrogen (=N- or -NH-), sulphur, and/or oxygen atoms. Examples of such heteroaryl groups are oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrrolyl, imidazolyl, pyrazolyl, pyridinyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazinyl, coumaryl, furyl, thienyl, quinolyl, benzothiazolyl, benzotriazolyl, benzodiazolyl, benzooxazolyl, phthalazinyl, phthalanyl, triazolyl, tetrazolyl, isoquinolyl, acridinyl, carbazolyl, dibenzazepinyl, indolyl, benzopyrazolyl, phenoxazonyl. Particularly interesting heteroaryl groups are oxazolyl, isoxazolyl, oxadiazolyl, oxatriazolyl, thiazolyl, isothiazolyl, thiadiazolyl, thiatriazolyl, pyrrolyl, imidazolyl, pyrazolyl, pyridinyl, pyrimidinyl, pyrazinyl, pyridazinyl, furyl, thienyl, quinolyl, triazolyl, tetrazolyl, isoquinolyl, indolyl in particular pyrrolyl, imidazolyl, pyridinyl, pyrimidinyl, thienyl, quinolyl, tetrazolyl, and isoquinolyl. The term "carbocyclyl" is intended to indicate a cyclic hydrocarbon radical, which may be a saturated or unsaturated, non-aromatic, mono- or bicyclic ring comprising 5-12 ring atoms, such as C_3-8 cycloall<yl, e.g. cyclopropyl, cyclopentyl, cyclohexyl or cyclooctyl, or a C_3-8 cycloalkylene radical, e.g. cycloprop-2-enyl, cyclobut-2-enyl, cyclopent-2-enyl, cyclohex-3-enyl, cycloocta-4-enyl or cyclohex-3,5-dienyl.

The term "heterocyclyl" is intended to mean a non-aromatic ring or ring system, usually a mono- or bicyclic ring system comprising 5-12 ring atoms, where one or more of the carbon atoms have been replaced with heteroatoms, e.g. nitrogen (=N- or -NH-), sulphur, and/or oxygen atoms. Examples of such heterocyclyl groups are imidazolidine, piperazine, hexahydropyridazine, hexahydropyrimidine, diazepane, diazocane, pyrrolidine, piperidine, azepane, azocane, aziridine, azirine, azetidine, pyroline, tropane, oxazinane (morpholine), azepine, dihydroazepine, tetrahydroazepine, hexahydroazepine, oxazolane, oxazepane, oxazocane, thiazolane, thiazinane, thiazepane, thiazocane, oxazetane, diazetane, thiazetane, tetrahydrofuran, tetrahydropyran, oxepane, tetrahydrothiophene, tetrahydrothiopyrane, thiepane, dithiane, dithiepane, dioxane, dioxepane, oxathiane, oxathiepane and dioxolane. The most interesting examples are imidazolidine, piperazine, hexahydropyridazine, hexahydropyrimidine, diazepane, diazocane, pyrrolidine, piperidine, azepane, azocane, azetidine, tropane, oxazinane (morpholine), oxazolane, oxazepane, thiazolane, thiazinane, and thiazepane, in particular imidazolidine, piperazine, hexahydropyridazine, hexahydropyrimidine, diazepane, pyrrolidine, piperidine, azepane, dioxolane and thiazinane.

The term "alkoxy" is intended to indicate a radical of formula OR*, wherein R* is alkyl as defined above, e.g. methoxy, ethoxy, propoxy, butoxy, etc.

The term "polyoxyethylene" is intended to indicate a group of formula R*(OCH₂CH₂)_S-, wherein R* is hydrogen or alkyl as defined above and s is an integer of 1-200, such as 1- 100, 1-80, 1-70, 1-60, 1-50, 1-40, 1-30, 1-20, 1-10, 1-8, 1-6, 1-5, 1-4 or 1-3.

The term "alkylaryl" is intended to indicate an alkyl group covalently joined to an aryl group.

The term "carbamido" is intended to indicate the group -NHC(O)NH₂.

The term "carbamoyl" is intended to indicate the group -C(O)NH₂. The term "pharmaceutically acceptable salt" is intended to indicate salts prepared by reacting a compound of formula I with a suitable inorganic or organic acid, e.g. hydrochloric, hydrobromic, hydroiodic, sulfuric, nitric, acetic, phosphoric, lactic, maleic, phthalic, citric, propionic, benzoic, glutaric, gluconic, methanesulfonic, salicylic, succinic, tartaric, toluenesulfonic, sulfamic or fumaric acid.

The compounds of formula I may be in the form of E- or Z-isomers, i.e. they may adopt an E or Z configuration about the double bond connecting the 2-indolinone moiety to the phenyl moiety or to the group denoted R₆, or they may be a mixture of the E- and Z- isomers. The E- or Z-isomerism of the compounds of formula I is indicated by a zigzag line.

The term "indolinone compound" (used synonymously with "oxindole compound" herein) is intended to include compounds of formula I and formula I' as shown herein as well as other, structurally related compounds, such as the compounds disclosed in WO

96/40116, US 6,225,335, WO 01/83450, EP 934 931, US 5,834,504, WO 02/02551, WO 00/08202 and WO 98/50356 which are hereby incorporated by reference in their entirety. Methods of preparing the compounds are also disclosed in these publications.

The term "prodrug" is intended to indicate a derivative of an active compound of formula I' which does not, or does not necessarily, exhibit the physiological activity of the active compound, but which may be subjected to enzymatic or other type of cleavage such as hydrolysis in vivo so as to release the active compound of formula I' on administration of the prodrug. In this particular instance, the prodrug comprises the active compound which in itself is highly lipophilic provided with a side chain with predominantly hydrophilic properties imparting improved solubility characteristics to the prodrug, thereby making it more suitable for parenteral administration in the form of a solution or for oral administration to obtain an improved bioavailability, cf. H. Bundgaard, Design of Prodrugs, Elsevier, 1985; H. Bundgaard, Arch. Pharm. Chem. 86(1), 1979, pp. 1-39; D. Fleisher et al., Adv. Drug Delivery Rev. 19(2), 1996, pp. 115-130; H. Bundgaard,

Controlled Drug Delivery 17, 1987, pp. 179-196; Friis and Bundgaard, Eur. J. Pharm. SdA, 1996, pp. 49-59; P. Ettmayer et al., 3. Med. Chem. 47(10), 2004, pp. 2393-2404.

The term "EAE" or "EAE model" is used herein to denote experimentally induced autoimmune encephalomyelitis (EAE) which is generally recognised as an animal model of multiple sclerosis. EAE may be induced by injection of antigenic peptides of myelin such as myelin basic protein, proteolipid protein and myelin oligodendrocyte glycoprotein. EAE is an inflammatory condition of the central nervous system characterised by T-cell infiltration and focal demyelination. EAE can also be induced by transfer of myelin reactive T-cells to normal individuals.

The term "ameliorate" is intended to mean reducing the severity of the neurological symptoms during relapses of multiple sclerosis by administering an effective amount of an active compound whereby it may be possible to reduce or delay permanent disability resulting from neurological damage sustained during relapse, in particular demyelination.

The term "delay the onset of multiple sclerosis" is used to indicate a prophylactic administration of an effective amount of an active compound to prolong the period where no symptoms, or at least no severe symptoms, of multiple sclerosis are observed in susceptible individuals, e.g. in first-degree relatives of multiple sclerosis patients.

The term "reduce the relapse rate in multiple sclerosis" is intended to mean reducing the frequency with which relapses occur or, in other words, prolong the periods of remission. This may make it possible to reduce or delay the accumulation of disabilities resulting from the neurological damage sustained during each relapse, in particular demyelination which eventually leads to increasingly severe disability.

Embodiments of the invention

Currently favoured compounds of formula I are compounds wherein R₂, R₃, R₄ and R₅ are all hydrogen.

In currently favoured compounds of formula I, Ri is -OC(O)R₉, -C(O)NR₁₀R₁₁, - OC(O)NR₁₀R₁₁, -S(O)₂R₉, -OR₁₂, -C(O)R₁₂, -C(O)OR₁₂, -OC(O)OR₁₂, -P(O)(OR₁₀)(OR₁₁), - OP(O)(OR₁₀)(OR₁₁), polyoxyethylene, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl or aryl, said C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl or aryl being substituted with one or more substituents selected from the group consisting of -OR₁₀, -C(O)R₁₀, -C(O)OR₁₀, - OC(O)R₁₀, -OC(O)OR₁₀, -NR₁₀R₁₁, -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, - C(O)NR₁₀R₁₁, -OC(O)NR₁₀R_UrOP(O)(OR₁₀)(OR₁O₇-P(O)(OR₁₀)(OR₁₁), -SR₁₀, -S(O)R₁₀, - S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁, -S(O)₂OR₁₀, polyoxyethylene, heteroaryl and heterocyclyl, said heteroaryl and heterocyclyl being optionally substituted with -OR₁₀, -C(O)R₁₀, -C(O)OR₁₀, OC(O)R₁₀, -OC(O)OR₁₀, -P(O)(OR₁₀)(OR₁₁), -OP(O)(OR₁₀)(OR₁₁), -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, -S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁, - S(O)₂ORi₀ or polyoxyethylene, and Ci_-6 alkyl substituted with -ORi₀, -C(O)Ri₀, - C(O)ORi₀, OC(O)Ri₀, -OC(O)OR₁₀, -NRi₀Rn, -C(O)NRi₀Rn, -P(O)(OR_I0)(OR_H), - OP(0)(ORio)(ORu), -NHC(O)Ri₀, -NHC(O)ORi₀, -NHC(O)NR_I0R_U, -SR_I0, -S(O)Ri₀, - S(O)₂Ri₀, -S(O)₂NRi₀Ri₁ or -S(O)₂ORi₀, wherein R₉, Ri₀, Ru and Ri₂ are as indicated above.

Thus, Ri may be -OC(O)R₉, -0R_i2, -C(O)Ri₂ or Ci_-I0 alkyl, said Ci_-I0 alkyl being substituted with one or more substituents selected from the group consisting of polyoxyethylene, heterocyclyl, -OR_i0, -C(O)Ri₀, -C(O)ORi₀, -OC(O)Ri₀, -OC(O)ORi₀, - NRi₀Ri_I, -C(O)NRi₀R₁₁, -OC(O)NR_1OR_1IrOP(O)(ORi_O)(ORi_I)^P(O)(OR_1O)(OR_1I), NHC(O)Ri₀, -NHC(O)ORi₀, -NHC(O)NR_I0R_U, "SR_I0, -S(O)Ri₀, -S(O)₂Ri₀, -S(O)₂NRi₀Ru, - S(O)₂ORi₀ wherein R₉, Ri₀, Rn and Ri₂ are as indicated above.

The compounds of formula I may be prodrugs, i.e. converted in vivo to compounds of formula I'

r wherein X, R₂, R₃, R₄, R₅, R₆, R_x', R₂', R₃', R₄' and R₅' are as indicated above for formula I. Compounds of formula I' are disclosed in, i.a., WO 96/40116, WO 98/50356 and WO 00/08202 in which they are indicated to be inhibitors of tyrosine kinases and as such useful in the treatment of cancer and other proliferative disorders as well as Parkinson's disease and Alzheimer's disease. It is anticipated that compounds of formula I may also be used for these indications and that they will exhibit improved oral bioavailability compared to the compounds of formula I'. In co-pending international patent application No. PCT2004/DK/000875 filed on 16 December 2004, we have shown that compounds of formula I' show inhibitory activity against EAE induced in mice and are therefore proposed for use in the treatment of multiple sclerosis. However, the activity of compounds of formula I' was low on oral administration when tested in the EAE model. In currently favoured compounds of formula I, Ri is

wherein R₁₀, Rio' and R_u are the same or different and independently selected from the group consisting of hydrogen, Ci_-I0 alkyl, C_2-I0 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein Ri₀ and R_u, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, each of Ci_-I0 alkyl, C_2-I0 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, Ci-₆ alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, -ORi₃, -C(O)Ri₃, -C(O)ORi₃, -OC(O)Ri₃, -OC(O)OR_X3, - NRi₃Ri₄, -C(O)NRi₃R₁₄, -OC(O)NRi₃Ri₄, -NHC(O)Ri₃, -NHC(O)ORi₃, -NHC(O)NRi₃Ri₄, - P(O)(ORi₃)(ORi₄), -OP(O)(ORi₃)(ORi₄), -SRi₃, -S(O)Ri₃, -S(O)₂Ri₃, -S(O)₂NRi₃Ri₄ and - S(O)₂ORi₄, wherein R₁₃ and R₁₄ are as indicated above, provided that Ri₀ and Ru are not both hydrogen.

In other, currently favoured compounds of formula I, Ri is

wherein Ri₀ and R₁₀' are the same or different and independently selected from the group consisting of hydrogen, Ci_-I0 alkyl, C_2-I0 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, each of Ci_-I0 alkyl, C_2-i0 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, Ci_-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, - ORi₃, -C(O)Ri₃, -C(O)OR₁₃, -OC(O)Ri₃, -OC(O)ORi₃, -NRi₃Ri₄, -C(O)NRi₃Ri₄, - OC(O)NRi₃Ri₄, -NHC(O)Ri₃, -NHC(O)ORi₃, -NHC(O)NRi₃Ri₄, -P(O)(OR₁₃)(ORi₄), - OP(O)(ORi₃)(ORi₄), -SR₁₃, -S(O)R₁₃, -S(O)₂R₁₃, -S(O)₂NR₁₃Ri₄ and -S(O)₂OR₁₄, wherein Ri₃ and R_i4 are as indicated above.

In currently favoured compounds of formula I, R₆ is hydrogen. In currently favoured compounds of formula I, wherein R₁', R₂', R₃', R₄' and R₅' are the same or different and independently selected from the group consisting of hydrogen, C_1- io alkyl, Ci_-10 alkoxy, aryl, heteroaryl, aryloxy, Ci_-I0 alkylaryl, Ci_-10 alkylaryloxy, halogen, trihalomethyl, S(O)R₂₀, S(O)₂R₂₀, S(O)₂NR₂₀R_2I, S(O)₂OR₂₀, SR₂₀, NO₂, NR₂₀R₂₁, OR₂₀, CN, CH₂OH, C(O)R₂₀, C(O)OR₂₀, OC(O)R₂₀, NHC(O)R₂₀, -NHC(O)OR₂₀, -NHC(O)NR₂₀R₂₁, (CH₂)_nC(O)₂R₂₀ and C(O)NR₂₀R₂₁, wherein R₂₀ is hydrogen, C_1-6 alkyl, heteroaryl or aryl, said C_1-6 alkyl, heteroaryl or aryl being optionally substituted with hydroxy or NR₂₂R₂₃, wherein R₂₂ and R₂₃ are independently hydrogen or C_1-6 alkyl or, together with the nitrogen atom to which they are attached, form a heteroaryl or heterocyclic ring, R₂₁ is hydrogen, C_1-6 alkyl or aryl, and n is 0-3.

More specifically, R₁', R₂', R₃', R₄' and R₅' may the same or different and independently selected from the group consisting of hydrogen, C_1-6 alkyl, C_1-6 alkoxy and halogen.

Examples of specific compounds of formula I are

3-(2,5-Dimethoxy-benzylidene)-l-hydroxymethyl-l,3-dihydro-indol-2-one 3-(2,5-Dimethoxy-benzylidene)-l-morpholin-4-ylmethyl-l,3-dihydro-indol-2-one 3-(2,5-Dimethoxy-benzylidene)-l-piperidin-l-ylmethyl-l,3-dihydro-indol-2-one 3-(2,5-Dimethoxy-benzylidene)-l-(4-methyl-piperazin-l-ylmethyl)-l,3-dihydro-indol-2- one

3-(2,5-Dimethoxy-benzylidene)-l-(3,3-dimethyl-piperidin-l-ylmethyl)-l,3-dihydro- indol-2-one l-[(Diisobutylamino)-methyl]-3-(2,5-dimethoxy-benzylidene)-l,3-dihydro-indol-2-one l-[l,4']Bipiperidinyl-l'-ylmethyl-3-(2,5-dimethoxy-benzylidene)-l,3-dihydro-indol-2-one 3-(2,5-Dimethyl-benzylidene)-l-morpholin-4-ylmethyl-l,3-dihydro-indol-2-one 3-(2,5-Dimethyl-benzylidene)-l-piperidin-l-ylmethyl-l,3-dihydro-indol-2-one 3-(2,5-Dimethyl-benzylidene)-l-(4-methyl-piperazin-l-ylmethyl)-l,3-dihydro-indol-2- one 3-(2,5-Dimethyl-benzylidene)-l-(3,3-dimethyl-piperidin-l-ylmethyl)-l,3-dihydro-indol- 2-one l-[(Diisobutylamino)-methyl]-3-(2,5-dimethyl-benzylidene)-l,3-dihydro-indol-2-one l-[l,4¹]Bipiperidinyl-l'-ylmethyl-3-(2,5-dimethyl-benzylidene)-l,3-dihydro-indol-2-one 3-[5-Methoxy-2-(2-morpholin-4-yl-ethoxy)-benzylidene]-l-morpholin-4-ylmethyl-l,3- dihydro-indol-2-one; hydrochloride

3-[2-(2-Diethylamino-ethoxy)-5-methoxy-benzylidene]-l-morpholin-4-ylmethyl-l,3- dihydro-indol-2-one; hydrochloride 3-[5-Methoxy-2-(2-piperidin-l-yl-ethoxy)-benzylidene]-l-morpholin-4-ylmethyl-l,3- dihydro-indol-2-one; hydrochloride

Acetic acid l-[3-(2,5-dimethoxy-benzylidene)-2-oxo-2,3-dihydro-indol-l-yl]-ethyl ester Acetic acid [3-(2,5-dimethoxy-benzylidene)-2-oxo-2,3-dihydro-indol-l-yl]-(2,2- dimethyl-[l,3]dioxolan-4-yl)-methyl ester

Acetic acid l-[3-(2,5-dimethyl-benzylidene)-2-oxo-2,3-dihydro-indol-l-yl]-ethyl ester 3-[Hydroxy-(4-nitro-phenyl)-methylene]-2-oxo-2,3-dihydro-indole-l-carboxylic acid phenyl ester

3-[Hydroxy-(4-nitro-phenyl)-methylene]-2-oxo-2,3-dihydro-indole-l-carboxylic acid amide

It is well established that T-cells contribute to the development of several chronic inflammatory and autoimmune diseases. Initially in the disease process, naive T-cells are activated by antigens and produce the proinflammatory cytokine interleukin-2 (IL-2) leading to clonal expansion and production of other inflammatory cytokines involved in the generation of the inflammatory or autoimmune response. Excessive T-cell activity is involved in allergies and immunoinflammatory diseases such as asthma, psoriasis, rheumatoid arthritis and multiple sclerosis. IL-2 has been found to have an important role in promoting the growth of T-cells in that it is a growth factor for both CD4+ and CD8+ T-cells as well as natural killer cells. Furthermore, IL-2 influences the differentiation of T helper cells into ThI and Th2 cells and potentiates the production of cytokines by each cell type. IL-2 appears to be initially produced by activated CD4+ T- cells, inducing proliferation of CD8+ T-cells and production of proinflammatory cytokines such as IL-I, IL-6 and TNF-α.

It is therefore further anticipated that compounds of formula I may be used in the prevention and treatment of other ThI mediated inflammatory diseases and conditions apart from multiple sclerosis, such as inflammatory bowel disease, Crohn's disease, allergies, asthma, arthritis, e.g. gout or rheumatoid arthritis, inflammatory skin diseases, e.g. psoriasis or atopic dermatitis, neuroinflammatory diseases, systemic vasculitis, arteritis, glomerulonephritis, synovitis, osteomyelitis, autoimmune inner ear disease or sepsis and septic conditions.

It is also anticipated that compounds of formula I may be used as modifiers of angiogenesis, e.g. in the treatment pathological conditions and diseases associated with deregulated angiogenesis such as rosacea, atherosclerosis, hemangioma, warts, pyogenic granuloma, scarring, nasal polyps, transplantation, liver regeneration, bone and cartilage destruction, pannus growth, osteophyte formation, endometriosis, dysfunctional uterine bleeding, follicular cysts, ovarian hyperstimulation, thyroiditis, thyroid enlargement, obstructive lung disease, diabetic or ischemic retinopathy, neovascular glaucoma, age-related macular degeneration, acute macular degeneration, retinitis, cytomegalovirus retinitis, macular edema, choroidal neovascularisation, or cerebral ischemia reperfusion injury.

Pharmaceutical compositions

For use in the present invention, the compound or compounds of formula I (the "active ingredient") may be formulated into a pharmaceutical composition together with a pharmaceutically acceptable vehicle and optionally one or more other therapeutic agents. Such other therapeutic agent may be selected from anti-inflammatory drugs such as corticosteroids or non-steroid anti-inflammatory drugs, or immunosuppressive drugs such as methotrexate or cyclophosphamide. For the treatment of multiple sclerosis, it is also envisaged to combine the administration of the compound of formula I with conventional therapeutic agents used in the treatment of multiple sclerosis such as IFN-β or glatiramer acetate.

The vehicle must be "pharmaceutically acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. The formulation may be in a form suitable for oral or parenteral (including subcutaneous, intramuscular, interperitoneal, intraarticular and intravenous) administration.

The formulations may conveniently be presented in dosage unit form and may be prepared by any of the methods well known in the art of pharmacy, e.g. as disclosed in Remington, The Science and Practise of Pharmacy, 20^th Ed., 2000. All methods include the step of bringing the active ingredient into association with the vehicle which constitutes one or more excipients. In general, the formulations are prepared by uniformly and intimately bringing the active ingredient into association with a liquid vehicle or a finely divided solid vehicle or both, and then, if necessary, shaping the product into the desired formulation.

The term "dosage unit" is understood to mean a unitary, i.e. a single dose which is capable of being administered to a patient, and which may be readily handled and packed, remaining as a physically and chemically stable unit dose comprising either the active ingredient as such or a mixture of it with solid or liquid pharmaceutical vehicle materials.

Unlike IFN-β and glatiramer acetate which are peptidic in nature and only suitable for parenteral administration, such by injection, the compounds of formula I are small organic molecules and may therefore be administered orally. This represents a clear benefit for the patient as it permits self-medication and is less painful than for instance injections of IFN-β which often cause pain at the site of injection. Compounds of formula I have surprisingly exhibited an excellent oral bioavailability and EAE inhibitory activity, cf. Table 1 below, and may therefore be suitable for oral administration.

Formulations suitable for oral administration may be in the form of discrete units such as capsules, sachets, tablets or lozenges, each containing a therapeutically effective amount of the active ingredient; in the form of a powder or granules; in the form of a solution or a suspension in an aqueous liquid or non-aqueous liquid, such as ethanol or glycerol; or in the form of an oil-in-water emulsion or a water-in-oil emulsion. Such oils may be edible oils, such as e.g. cottonseed oil, sesame oil, coconut oil or peanut oil. Suitable dispersing or suspending agents for aqueous suspensions include synthetic or natural gums such as tragacanth, alginate, acacia, dextran, sodium carboxymethylcellulose, gelatin, methylcellulose and polyvinylpyrrolidone. The active ingredient may also be administered in the form of a bolus, electuary or paste.

A tablet may be prepared by compressing or moulding the active ingredient optionally with one or more excipients. Compressed tablets may be prepared by compressing, in a suitable machine, the active ingredient(s) in a free-flowing form such as a powder or granules, optionally mixed with a binder, such as lactose, glucose, starch, gelatine, acacia gum, tragacanth gum, sodium alginate, carboxymethylcellulose polyethylene glycol, waxes, hydroxypropylmethylcellulose, microcrystalline cellulose or the like; a lubricant such as sodium oleate, sodium stearate, magnesium stearate, calcium stearate, sodium benzoate, sodium acetate, sodium chloride or the like; a disintegrating agent such as starch, methyl cellulose, agar, bentonite, xanthan gum, croscarmellose, crospovidone, sodium starch glycolate, alginic acid, guar gum, carboxymethylcellulose or the like; or a diluent such as dicalcium phosphate, calcium sulphate, lactose, cellulose, kaolin, mannitol, sorbitol, sucrose or inositol, lactose, mannitol, sorbitol and sucrose also being useful as disintegrating agents. Moulded tablets may be made by moulding, in a suitable machine, a mixture of the powdered active ingredient and suitable carrier moistened with an inert liquid diluent. Capsule formulations may be in the form of hard or soft capsules, e.g. gelatin capsules, containing the active ingredient in a suitable dry or liquid vehicle. Hard capsules typically contain the active ingredient in powder or crystalline form together with one or more dry excipients which may conveniently be selected from the excipients indicated above for use in tablet formulations. Soft capsules typically contain a solution, suspension or emulsion comprising the active ingredient. Suitable excipients for soft capsules include, for instance, suspension vehicles such as vegetable, animal or mineral oils, e.g. triglycerides of lauric, palmitic, stearic, oleic, caproic or linoleic acid, liquid hydrocarbons, or polyethylene glycol; one or more solvents such as ethanol, propylene glycol, dimethylacetamide, lactic acid, glycerol, butanediol or polyvinylpyrrolidone; one or more surfactants such as polyoxyethylene-Ci_2-i8 fatty alcohols, polyoxyethylene stearate, polysorbates, sorbitol esters or monoglycerides. The compounds of formula I may also be included as a solid solution or dispersion, e.g. prepared by dissolving the active ingredient in a molten carrier, filling the molten mixture into the capsules and letting it cool.

In addition to the aforementioned ingredients, the formulations comprising a compound of formula I may include one or more additional ingredients such as buffers, flavouring agents, colourants, thickeners, preservatives, e.g. methyl hydroxy benzoate (including anti-oxidants), emulsifying agents and the like.

Tablets or capsules may optionally be provided with a coating for modifying the release of the active ingredient therefrom. Suitable coating materials comprise, e.g. mixtures of wax with stearic acid, glyceryl monostearate, palmitic acid, glyceryl monopalmitate, cetyl alcohol, shellac, zein, ethylcellulose, acrylic resins or cellulose acetate or diacetate.

For systemic treatment according to the present invention, daily doses of from 0.001-100 mg/kg body weight, preferably from 0.002-15 mg/kg body weight, for example 0.003-10 mg/kg of a compound of formula I are administered, typically corresponding to a daily dose for an adult human of from 0.2 to 750 mg of the active ingredient. Oral compositions are formulated, preferably as tablets, capsules, or drops, containing from 0.05-250 mg, preferably from 0.1-125 mg, of a compound of formula I per dosage unit.

The invention is further described in the following examples which are not in any way intended to limit the scope of the invention as claimed. EXPERIMENTAL

GENERAL SYNTHESIS

The compounds of general formula I can be prepared in a number of ways well known to those skilled in the art of organic synthesis. The compounds of formula I can be synthesised using the methods outlined below, together with methods known in the art of synthetic organic chemistry, or variations thereof as appreciated by those skilled in the art. Preferred methods include, but are not limited to, those described below.

The compounds of formula I can be prepared by techniques and procedures readily available to one of ordinary skill in the art, for example by following the procedures as set forth in the following schemes. The reactions are performed in solvents appropriate to the reagents and materials employed and suitable for the transformations being effected. Also, in the synthetic methods described below, it is to be understood that all proposed reaction conditions, including choice of solvent, reaction atmosphere, reaction temperature, duration of experiment and work-up procedures, are chosen to be conditions of standard for that reaction, which should be readily recognised by one skilled in the art. It is understood by one skilled in the art of organic synthesis that the functionalities present on various portions of the starting molecules in a reaction must be compatible with the reagents and reactions proposed. Not all compounds of formula I falling into a given class may be compatible with some of the reaction conditions required in some of the methods described. Such restrictions to the substituents which are compatible with the reaction conditions will be readily apparent to one skilled in the art and alternative methods can be used.

The schemes described in this section are not intended to limit the scope of the invention in any way. All substituents, unless otherwise indicated, are previously defined. The reagents and starting materials are either available from commercial suppliers or prepared by methods known to one of ordinary skilled in the art following procedures set forth in references such as Fieser and Fieser's Reagents for Organic Synthesis, Volumes 1-22 (John Wiley and Sons, 2004); Rodd's Chemistry of Carbon Compounds, Volumes 1-5 and Supplements (Elsevier Science Publishers, 2000); Organic Reactions, Volumes 1-64 (John Wiley and Sons, 2004); March's Advanced Organic Chemistry (John Wiley and Sons, 5^th Edition) and Larock's Comprehensive Organic Transformations (VCH Publishers Inc., 1999). These schemes are merely illustrative of some methods by which the compounds of this invention can be synthesised, and various modifications to these schemes can be made and will be suggested to one skilled in the art having referred to this disclosure. The starting materials and the intermediates of the reactions may be isolated and purified if desired using conventional techniques, including but not limited to filtration, distillation, crystallisation, chromatography and the like. Such materials may be characterised using conventional means, including physical constants and spectral data.

The introduction of R₁ (R₁ Ψ H) may occur at different stage of the synthesis. The precursor of R₁ may be purchased or synthesised as a reactive species. R₁ may be chemically modified after its introduction on the indolinone, before or after condensation to the benzaldehyde / benzoketone. Some examples for the introduction of R₁ are described in, but not limited to, the following section.

In the same manner eventual protecting groups may be added and removed at different step of the synthesis.

General method A:

Some of the compounds of general formula I can be prepared according to the general method A described below.

Scheme 1

a. Base such as nitrogen base or inorganic base. "Nitrogen bases" are selected from acyclic and cyclic amines. Examples of nitrogen bases include, but are not limited to, ammonia, methylamine, trimethylamine, triethylamine, aniline, 1,8- diazabicyclo-[5.4.1]-undec-7-ene, diisopropyl ethylamine, pyrrolidine, piperidine, morpholine, and pyridine or substituted pyridine (e.g., 2,6-di-te/tbutylpyridine).

"Inorganic bases" are bases that do not contain any carbon atoms. Examples of inorganic bases include, but are not limited to, hydroxide, phosphate, bisulfate, hydrosulfide (SH^"), and amide anions. Preferred nitrogen bases are piperidine and morpholine. Preferred inorganic bases are hydroxide anion, preferably used as its sodium or potassium salt. The reaction generally takes place in a protic solvent such as water or alcohols or in an aprotic solvent such as toluene, CH₂CI₂, THF, DMF. Most preferred solvents are alcohols such as ethanol, and CH₂CI₂.

General method B

Ri may be introduced by alkylation of the nitrogen of the parent compound where Ri =

H as in the following scheme.

Scheme2

b. Alkylation by R₁X in the presence of a base such as NEt₃, NaH, NaOH, KOH, carbonates in an appropriate solvent such as DMF, pyridine, DMSO, CH₃CN, acetone, toluene. X is a leaving group such as chloride, bromide, and iodide.

Alternatively reaction with an alcohol R_xOH may also be considered. Such reaction is run in the presence of a phosphine such as PBu₃, PPh₃ and the like, an azodicarboxylate or an azodicarboxamide in an aprotic solvent, typically THF.

General method C

Some of the compounds of the present disclosure have the general formula II. Scheme 3 illustrates the synthesis of II.

Scheme 3

c. Reaction with an aldehyde such as formaldehyde, acetaldehylde, and the like, and a suitable amine affords a compound of general formula II. The solvent in which the reaction is carried out may be a protic or an aprotic solvent, preferably it is a protic solvent such as an alcohol e.g., methanol or ethanol, or an aqueous alcohol. The reaction may be carried out at temperatures greater than room temperature. The temperature is generally from about 20 ⁰C to about 100 ⁰C, preferably about 40 ⁰C to about 80 ⁰C. By "about" is meant that the temperature range is preferably within 10 degrees Celsius of the indicated temperature, more preferably within 5 degrees Celsius of the indicated temperature, most preferably within 2 degrees Celsius of the indicated temperature. Suitable amines include acyclic and cyclic secondary amines such as diethyl amine, Λ/-te/t-butyl methylamine, diisobutylamine, piperidine, morpholine, Λ/-methyl piperazine, 3,3- dimethylpiperidine and the like.

Alternatively, compounds of general formula II can be prepared via reaction of compounds of general formula I where Ri = H and a preformed Mannich reagent such as an aminal, an /V,O-acetal, a benzotriazole aminal or an iminium salt in the presence of a base such as NEt₃, t-BuOK, n-BuLi. Compounds of general formula II where R₁₀ and R_n combine to form a heteroaryl ring, may be prepared by reacting I where Ri = H with a suitable aldehyde Rio'CHO to yield a C-substituted /V-hydroxymethyl intermediate (see conditions d) and reacting the intermediate with phosphorus oxychloride and a suitable heteroaryl such as pyridine, pyrrole, oxazolyl, imidazolyl, and the like. The reaction may be carried out at temperature less than r.t..

General method D

Scheme 4 is depicting the synthesis of compounds of general formula III. Scheme 4

d. Reaction with an aldehyde of formula Rio'CHO in the presence of an organic base, preferably a tertiary nitrogen base such as NMe₃, NEt₃, pyridine, diisopropylethylamine, l,8-diazabicyclo-[5.4.1]-undec-7-ene, and the like in an aprotic solvent such as acetonitrile, DMF, THF or pyridine. e. To obtain HIa reaction with an acylating agent such as acid anhydride (e.g. acetic anhydride, succinic anhydride), acid halides (e.g. acetyl chloride, propionyl chloride) and carboxylic acid active esters (e.g. p-nitrophenyl ester, pentafluorophenyl ester). The reaction is carried out in an organic base such as pyridine, DMAP and the like. In a similar fashion IHb can be obtained by reaction with chloroformates; IUc may be obtained by reaction with carbamoyl chlorides or icocyanates. Formation of IHd may be achieved by alkylation of the intermediate alcohol following b. f. To obtain Ilia reaction with an aldehyde and a suitable acylating agent as described in d. and e., respectively without isolating the intermediate alcohol. Alternatively R₁ = CHR₁₀OC(O)R₁₀ , R₁ = CHR₁₀OC(O)OR₁₀, Rl = CHR₁₀OC(O)NR₁₀R₁₁ and R₁ = CHR₁₀OR₁₀ may be introduced by alkylation of I where R₁ = H by R₁X following b. A synthesis of R₁CI is exemplified in scheme 5

(for R₁ = CHR₁₀OCOR₁₀, CHR₁₀OCOOR₁₀, CHR₁₀OCONR₁₀R₁₁).

Scheme 5 is an example but one skilled in the art may consider alternative method for the synthesis of such alkylating agent. Scheme 5 exemplifies the synthesis of carboxylic esters, carbonic esters, and carbamates. The chlorine atom may be interchanged with iodide with methods known to those skilled in the art such as NaI in acetone. Scheme 5

g. HCISO₃ in an aprotic solvent such as pentane, CH₂CI₂, CHCI₃, THF at temperature below room temperature (Synth. Communic. 14(9), 857 (1984))

h. Carboxylic acid or carbonic acid or carbamic acid in the presence of a base such as alkali carbonate, alkali bicarbonate, alkali hydroxide and a phase transfer catalyst such as tetra n-butylammonium hydrogensulfate, tetra n- butylammonium chloride in a mixture of water and an aprotic solvent such as

CH₂CI₂, Et₂O, CHCI₃.

i. Reaction with RioOH in the neat alcohol or in a suitable solvent such as CH₂CI₂, CHCI₃, THF, diethyl ether and DMF optionally in the presence of an acid such as hydrochloric acid, trifluoroacetic acid, and the like or a base such as pyridine,

NEt₃, and the like.

j. Reaction with HNRi₀Ru in a suitable solvent such as CH₂CI₂, CHCI₃, THF, diethyl ether, dioxane, DMSO and DMF, eventually with water as a co-solvent, optionally in the presence of a non-nucleophilic base such as pyridine, NaHCO₃, NEt₃.

General method E Compounds of general formula IV may be obtained by either direct acylation (k) or in two steps as shown in the following schemes.

Scheme 6

IVa IVb IVc

k. Acylation may be obtained via reaction with an acylating agent such as acid anhydride (e.g. acetic anhydride, succinic anhydride), acid halides (e.g. acetyl chloride, propionyl chloride), carboxylic acid active esters (e.g. p-nitrophenyl ester, pentafluorophenyl ester), chloroformates and carbamoyl chlorides. The reaction is carried out in an organic base such as pyridine, DMAP and the like or in an aprotic solvent such as CH₃CN, DMF in the presence of a base such as NEt₃, DMAP, pyridine or NaH.

Scheme 7

IVc

I. For examples for LG = Cl, the intermediate can be prepared by treatment of I (Ri = H) with triphosgene in the presence of an organic amine such as NEt₃, pyridine and the like. A compound where LG is imidazol-1-yl can be prepared by treatment of I (R_x = H) with carbonyl diimidazole under conditions well known in the art.

General method F

If Ri = OR₉, derivatisation of l-hydroxy-3-(lH-indol-3-ylmethylene)-l,3-dihydro-indol- 2-one may be used.

Scheme 8

V

The hydroxyl functionality may be alkylated following b using R₉X (X = leaving group) instead of RiX in an appropriate solvent such as DMSO, toluene, DMF or CH₃CN in the presence of a base such as NaOH, KOH, K₂CO₃, Cs₂CO₃ and the like. Reaction with an alcohol R₉OH (instead of R₁OH) may also be considered. (See b)

m. Addition to double bonds may also be considered to afford I where R_x = OR₉. The reaction may be catalysed by bases or by acids such as p-toluene sulfonic acid, pyridinium p-toluenesulfonate, POCI₃, trimethylsilyliodide and the like.

The hydroxyl functionality may be acylated following e.

General method G If Ri contains a carboxylic functionality, it may be modified as described in the following scheme, wherein n is an integer of from 1 to 10.

Scheme 9

n. Standard hydrolysis conditions using first a base such as an aqueous solution of LiOH or NaOH followed by treatment with an acid such as an aqueous solution of HCI. o. Esterification using classical esterification conditions (see March's Advanced Organic Chemistry Reactions, mechanisms, and Structure, 5^th edition, by M. B. Smith and 3. March chapter 10-23 p. 484, chapter 10-26 p. 488 and chapter 10- 28 p. 490) such as acid catalysed esterification (e.g. using sulphuric acid as a catalyst in MeOH), or such as base catalyzed esterification (e.g. using chlorosulfates as reagents under phase transfer conditions, see Synthetic Communications, 14 (9), 857-864, 1984) or such as using diazomethane in a suitable solvent such as Et₂O or EtOH, or using trimethylsilyldiazomethane in a suitable solvent such as toluene, or using an electrophilic reactant (as for example benzyl bromide or methyliodide) in the presence of a base such as

K₂CO₃ or Cs₂CO₃ in a solvent such as DMF or acetonitrile. p. Formation of the amide using cyclic and acyclic amines in the presence of classical coupling agents. Preferred coupling agents include 1,1'- carbonyldiimidazole (CDI), diphenylphoshinic chloride (DPP-CI), benzotriazol- yloxy-tripyrolidinophosphonium hexafluorophosphate (PyBOP), benzotriazol-1- yloxy-tris(dimethylamino)-phosphonium hexafluorophosphate (BOP), N, N'- dicyclohexylcarbodiimide (DCC), or l-ethyl-3-(3-dimethylaminopropyl)- carbodiimide; hydrochloride (EDCI). Preferred bases include diisopropylethylamine, triethylamine, 4-methylmorpholine, or pyridine or a substituted pyridine, for example 4-dimethylaminopyridine or 2,6- dimethylpyridine. Preferred solvents are solvents such as diethylether, dichloromethane, tetrahydrofuran, l-methyl-2-pyrrolidinone, dimethylsulfoxyde or dimethylformamide. The reactions are generally carried out in the presence of a base such as Et₃N or Bu₃N and in the presence of an activator such as HOBt (for example where HOBt is used to improve reactions rates, see Windridge, G. C; Jorgensen, E. C. JACS 1971, 93, 6318), at a temperature between about -78⁰C to about 60⁰C, and are normally complete within about 2 hours to about 5 days.

Examples

For ¹H nuclear magnetic resonance (NMR) spectra (300 MHz) and ¹³C NMR (75.6 MHz) chemical shift values (δ) (in ppm) are quoted for dimethyl-cfe sulfoxide (DMSO-c/₆) or CDCI₃ solutions relative to internal tetramethylsilane (δ = 0) standard. The value of a multiplet, either defined (doublet (d), triplet (t), quartet (q)) or not (m) at the approximate mid point is given unless a range is quoted, (bs) indicates a broad singlet. The ES mass spectra were obtained on a VG Quattro II triple quadrapole mass spectrometer (Micromass, Manchester, UK) operating in either positive or negative electrospray mode with a cone voltage of 30V.

The compounds of the present invention can exist in two isomeric forms: the Z and the E isomeric forms. The NMR data characterize the isomer forms that are present in the solvent used to record the NMR spectrum and determine their molar ratio. For NMR solutions where both the E-isomer and the Z-isomer are present in equal or close to equal amounts, the chemical shifts of both forms are given. For NMR solutions where the equilibrium is shifted in favour of one form, the chemical shifts of the dominating form are given.

The organic solvents used were anhydrous unless otherwise specified. Flash chromatography was performed on silica gel or Sephadex LH-20 from Fluka Chemie GmbH, Switzerland.

The following abbreviations have been used throughout: Ac Acetate or acetyl aq. Aqueous

Brine Saturated aqueous sodium chloride

Boc te/t-Butoxycarbonyl DMAP 4-Dimethylaminopyridine

DMF /^/V-Dimethylformamide

DMSO Dimethylsulfoxide

EDAC l-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride

EDCI l-ethyl-S-CS-dimethylaminopropyO-carbodiimide

EtOAc Ethyl acetate eq. Equivalent

HOBt 1-hydroxybenzotriazole

LG Leaving group

M Molar (mol/L)

NMR Nuclear magnetic resonance

PE Petroleum ether (bp 40-60 ⁰C) r.t. Room temperature sat. Saturated

THF Tetrahydrofuran

TLC Thin layer chromatography

Preparation 1: 3-(2,5-Dimethoxy-benzylidene)-l,3-dihydro-indol-2-one

l,3-Dihydro-indol-2-one (26.0 g, 195.3 mmol), 2,5-dimethoxy benzaldehyde (32.8 g, 195.2 mmol) and piperidine (12 mL) were suspended in ethanol (600 mL). The suspension was heated to reflux for 3 h. Standing in the fridge afforded a thick precipitate which was filtered and washed with petroleum ether. Drying in vacuo afforded 44.3 g (81%) of the title compound as a yellow powder.

Preparation 2: 3-(2,5-Dimethyl-benzylidene)-l,3-dihydro-indol-2-one

l,3-Dihydro-indol-2-one (5 g, 37.5 mmoles), 2,5-dimethyl benzaldehyde (5.3 mL, 37.5 mmol) and piperidine (12 mL) were suspended in ethanol (125 mL). The suspension was heated to reflux overnight before being slightly concentrated in vacuo. Standing in the fridge afforded a thick precipitate which was filtered and washed with ethanol. Drying in vacuo afforded 7.8 g (84%) of the title compound as a yellow powder.

Preparation 3: 3-[5-methoxy-2-(2-morpholin-4-yl-ethoxy)-benzylidene]-l,3-dihydro- indol-2-one; hydrochloride

l,3-Dihydro-indol-2-one (0.74 g, 5.6 mmol), 5-methoxy-2-(2-morpholin-4-yl-ethoxy)- benzaldehyde (1.50 g, 5.6 mmol) and piperidine (0.35 mL) in EtOH (20 mL) were heated to reflux overnight. After concentration in vacuo chromatography afforded 2.04 g (96%) of the title compound (as a free amine) as an orange solid.

To a solution of the free amine (1.15 g, 3 mmol) in EtOAc (10 mL) and MeOH (2 mL) was added a solution of HCI in dioxane (4N, 0.75 mL, 3 mmol). After addition of Et₂O, a precipitate formed which was filtered and washed with Et₂O to afford the title compound (1.11 g, 89% yield).

Preparation 4: 3-[2-(2-diethylamino-ethoxy)-5-methoxy-benzylidene]-l,3-dihydro-indol- 2-one

l,3-Dihydro-indol-2-one (0.8 g, 6 mmol), 2-(2-diethylamino-ethoxy)-5-methoxy- benzaldehyde (1.53 g, 6 mmol) and piperidine (0.35 mL) in EtOH (20 mL) were heated to reflux overnight. After chromatography the title compound as free amine was obtained as a hygroscopic solid (1.54 g, 70% yield).

¹³C-NMR (DMSO-d₆, mixture of the E and Z isomers, signals of the predominant E isomer) δ 168.5, 152.5, 151.1, 142.7, 131.6, 129.8, 127.5, 123.7, 122.4, 121.1, 120.9, 116.8, 114.3, 113.7, 109.9, 67.6, 55.4, 51.0, 47.0, 11.9. To a solution of the free amine (1.5 g, 4 mmol) in EtOAc (3 mL) was added a solution of HCI in dioxane (4N, 2 mL, 8.2 mmol). After addition of Et₂O, a precipitate formed which was filtered and washed with Et₂O to afford the title compound (1.32 g, 80% yield).

Preparation 5: 3-[5-methoxy-2-(2-piperidin-l-yl-ethoxy)-benzylidene]-l,3-dihydro- indol-2-one; hydrochloride

l,3-Dihydro-indol-2-one (0.76 g, 5.7 mmol), 5-methoxy-2-(2-piperidin-l-yl-ethoxy)- benzaldehyde (1.52 g, 5.8 mmol) and piperidine (0.35 mL) in EtOH (20 mL) were heated to reflux overnight. After chromatography the title compound as free amine was obtained as an orange solid (2.07 g, 96% yield). To a solution of the free amine (896 mg, 2.4 mmol) in EtOAc (10 ml_) and MeOH (2 mL) was added a solution of HCI in dioxane (4N, 0.6 mL, 2.4 mmol). After addition of Et₂O, a precipitate formed which was filtered and washed with Et₂O to afford the title compound (759 mg, 76% yield).

Example 1: 3-(2,5-Dimethoxy-benzylidene)-l-hydroxymethyl-l,3-dihydro-indol-2-one

To a suspension of preparation 1 (600 mg, 2.13 mmol) in MeOH (30 mL) was added formaldehyde (13.1 M in water, 1.63 mL, 10 eq.). The mixture was heated to 60 ⁰C overnight and allowed to come to r.t. The solvent was removed. EtOAc was added and the organic phase was washed with sat. aq. NH₄CI until pH was 6. The product was washed further with water (Ix), dried over MgSO₄, filtered, concentrated in vacuo and purified by flash chromatography to afford 315 mg (47%) of the title compound as yellow powder. ¹H NMR (DMSO-CZ₅) δ 7.72 (s,lH), 7.49 (d,lH), 7.32 (t,lH), 7.23 (d,lH), 7.15 (d,lH), 7.12 ~7.07(m,2H), 6.96 (t,lH), 6.26 (t,lH), 5.16 (d,2H), 3.80 (s,3H), 3.74 (s,3H)

General procedure A

To a suspension of 3-(substituted-benzylidene)-l,3-dihydro-indol-2-one in MeOH were added formaldehyde (13.1 M in water, 10 eq.) and an amine (10 eq.). The mixture was heated to 60 ⁰C overnight unless otherwise stated and allowed to come to r.t. The precipitate was filtered and washed with water. If no precipitation occurred, the solvent was removed. EtOAc was added and the organic phase was washed with sat. aq. NH₄CI until pH was 6. The product was washed further with water (Ix), dried over MgSO₄, filtered, concentrated in vacuo and purified by flash chromatography. Example 2: 3-(2,5-Dimethoxy-benzylidene)-l-morpholin-4-ylmethyl-l,3-dihydro-indol- 2-one

General procedure A was followed using preparation 1 (1.0 g, 3.55 mmol), formaldehyde (2.7 ml.) and morpholine (3.09 mL) in MeOH (40 ml_). Flash chromatography afforded 748 mg (53%) of the title compound as an orange foam.

¹H NMR (DMSO-Cf₆) δ 7.72 (s,lH), 7.50 (d,lH), 7.31 (t,lH), 7.23 (d,lH), 7.19 (d,lH), 7.12 - 7.07 (m,2H), 6.95 (t,lH), 4.48 (s,2H), 3.81 (s,3H), 3.74 (s,3H), 3.55 (m,4H), 2.55 (m,4H)

Example 3: 3-(2,5-Dimethoxy-benzylidene)-l-piperidin-l-ylmethyl-l,3-dihydro-indol-2- one

General procedure A was followed using preparation 1 (2.0 g, 7.1 mmol), formaldehyde (5.4 mL) and piperidine (7.02 mL) in MeOH (200 mL). Flash chromatography afforded 1.74 g (65%) of the title compound as yellow oil. ¹H NMR (DMSO-Cf₆) δ 7.71 (s,lH), 7.50 (d,lH), 7.30 (d,lH), 7.25 (d,lH), 7.19 - 7.02 (m,3H), 6.94 (t,lH), 4.45 (s,2H), 3.81 (s,3H), 3.74 (s,3H), 2.52 (m,4H), 1.52 - 1.27 (m,6H)

Example 4: 3-(2,5-Dimethoxy-benzylidene)-l-(4-methyl-piperazin-l-ylmethyl)-l,3- dihydro-indol-2-one

General procedure A was followed using preparation 1 (600 mg, 2.13 mmol), formaldehyde (0.48 mL, 3 eq.) and Λ/-methylpiperazine (0.7 mL, 3 eq.) in MeOH (25 mL). Work-up afforded 800 mg (96%) of the title compound as yellow oil.

¹³C NMR (DMSOd₆) δ 167.8, 152.5, 151.8, 143.7, 132.3, 129.8, 126.2, 123.1, 122.0, 121.6, 120.1, 117.1, 114.3, 112.7, 110.3, 61.0, 55.9, 55.5, 54.4, 50.1, 45.6

Example 5: 3-(2,5-Dimethoxy-benzylidene)-l-(3,3-dimethyl-piperidin-l-ylmethyl)-l,3- dihydro-indol-2-one

General procedure A was followed using preparation 1 (600 mg, 2.13 mmol), formaldehyde (1.63 mL) and 3,3-dimethylpiperidine (2.43 g) in MeOH (30 mL). Flash chromatography afforded 744 mg (86%) of the title compound.

¹H NMR (DMSO-CZ₆) δ 7.70 (s,lH), 7.50 (d,lH), 7.30 (t,lH), 7.24 (d,lH), 7.16 (d,lH),

7.14 - 7.02 (m,2H), 6.94 (t,lH), 4.43 (s,2H), 3.81 (s,3H), 3.74 (s,3H), 2.45 (m,2H),

2.2 (br,2H), 1.47 (m,2H), 1.15 (m,2H), 0.88 (s,6H)

Example 6: l-[(Diisobutylamino)-methyl]-3-(2,5-dimethoxy-benzylidene)-l,3-dihydro- indol-2-one

General procedure A was followed using preparation 1 (600 mg, 2.13 mmol), formaldehyde (1.63 mL) and di-isobutylamine (2.75 g) in MeOH (30 mL). Flash chromatography afforded 260 mg (29%) of the title compound.

¹H NMR (DMSO-Of₆) δ 7.70 (s,lH), 7.49 (d,lH), 7.31 (t,lH), 7.22 (d,lH), 7.14 - 7.02 (m,3H), 6.94 (t,lH), 4.51 (s,2H), 3.81 (s,3H), 3.74 (s,3H), 2.27 (d,4H), 1.79 (m,2H), 0.81 (d,12H)

Example 7: l-[l,4']Bipiperidinyl-l'-ylmethyl-3-(2,5-dimethoxy-benzylidene)-l,3- dihydro-indol-2-one

General procedure A was followed using preparation 1 (700 mg, 2.49 mmol), formaldehyde (1.9 mL) and piperidinopiperidine (4.2 g) in MeOH (60 mL). Flash chromatography afforded 686 mg (60%) of the title compound as a yellow oil. ¹³C NMR (DMSO-CZ₆) δ 167.9, 152.5, 151.8, 143.9, 132.3, 129.8, 126.2, 123.1, 122.0, 121.7, 120.2, 117.1, 114.3, 112.7, 110.2, 61.9, 60.8, 55.9, 55.5, 54.8, 49.7, 49.1, 48.5

Example 8: 3-(2,5-Dimethyl-benzylidene)-l-morpholin-4-ylmethyl-l,3-dihydro-indol-2- one

General procedure A was followed using preparation 2 (700 mg, 2.81 mmol), formaldehyde (2.15 mL) and morpholine (2.45 ml_) in MeOH (60 ml_). Flash chromatography afforded 598 mg (52%) of the title compound as yellow oil.

¹³C NMR (DMSO-de) δ 167.6, 143.7, 135.9, 134.9, 133.5, 133.5, 130.3, 130.1, 129.8, 128.5, 127.0, 121.8, 121.8, 120.3, 110.3, 65.9, 61.3, 50.6, 20.3, 18.9

Example 9: 3-(2,5-Dimethyl-benzylidene)-l-piperidin-l-ylmethyl-l,3-dihydro-indol-2- one

General procedure A was followed using preparation 2 (1.0 g, 4.01 mmol), formaldehyde (3.06 mL) and piperidine (3.96 mL) in MeOH (100 mL) at the difference that the mixture was refluxed for 2 days. Work-up afforded 1.2 g (86%) of the title compound as yellow oil.

¹H NMR (DMSO-CZ₆) δ 7.75 (s,lH), 7.37 (s,lH), 7.32 - 7.10 (m,5H), 6.86 (t,lH), 4.45

(s,2H), 2.53 (m,4H), 2.31 (s,3H), 2.25 (s,3H), 1.47 (m,4H), 1.36 (m,2H)

Example 10: 3-(2,5-Dimethyl-benzylidene)-l-(4-methyl-piperazin-l-ylmethyl)-l,3- dihydro-indol-2-one

General procedure A was followed using preparation 2 (700 mg, 2.81 mmol), formaldehyde (2.15 mL) and Λ/-methylpiperazine (3.12 mL) in MeOH (60 mL). Flash chromatography afforded 360 mg (35%) of the title compound as yellow oil. ¹³C NMR (DMSO-Of₆) δ 167.7, 143.8, 135.9, 134.9, 133.6, 130.4, 130.2, 129.9, 128.6, 127.2, 121.9, 121.8, 120.4, 110.4, 61.1, 54.9, 54.5, 50.2, 45.7, 20.4, 19.0

Example 11: 3-(2,5-Dimethyl-benzylidene)-l-(3,3-dimethyl-piperidin-l-ylmethyl)-l,3- dihydro-indol-2-one

General procedure A was followed using preparation 2 (700 mg, 2.81 mmol), formaldehyde (2.15 mL) and 3,3-dimethylpiperidine (3.21 g) in MeOH (60 mL). Flash chromatography afforded 769 mg (73%) of the title compound as yellow oil. ¹³C NMR (DMSO-CZ₆) δ 167.5, 143.9, 135.7, 134.8, 133.5, 130.3, 130.0, 129.7, 128.5, 127.2, 121.8, 121.6, 120.3, 110.4, 62.8, 62.0, 51.2, 36.7, 30.4, 26.9, 21.8, 20.3, 18.9

Example 12: l-[(Diisobutylamino)-methyl]-3-(2,5-dimethyl-benzylidene)-l,3-dihydro- indol-2-one

General procedure A was followed using preparation 2 (700 mg, 2.81 mmol), formaldehyde (2.15 ml_) and di-isobutylamine (3.62 g) in MeOH (60 ml_). Flash chromatography afforded 605 mg (55%) of the title compound.

¹³C NMR (DMSO-de) δ 167.8, 143.9, 135.6, 134.9, 133.5, 133.4, 130.3, 130.0, 129.6, 128.4, 127.3, 121.8, 121.7, 120.3, 110.0, 61.6, 60.1, 26.1, 20.6, 20.3, 18.8

Example 13: l-[l,4']Bipiperidinyl-l'-ylmethyl-3-(2,5-dimethyl-benzylidene)-l,3-dihydro- indol-2-one

General procedure A was followed using preparation 2 (700 mg, 2.81 mmol), formaldehyde (2.15 mL) and piperidinopiperidine (4.72 ml.) in MeOH (60 ml_) at the difference that the mixture was refluxed for 2 days. After work-up, precipitation occurred upon concentration of the organic layer. Filtration afforded 216 mg (18%) of the title compound as light yellow crystals.

¹³C NMR (DMSO-Gf₆) δ 167.7, 143.8, 135.8, 134.9, 133.5, 130.3, 130.1, 129.8, 128.5, 127.0, 121.8, 120.3, 110.2, 62.1, 60.6, 49.3, 48.7, 25.5, 22.5, 20.3, 18.9

General procedure B

To a suspension of 3-(substituted-benzylidene)-l,3-dihydro-indol-2-one in MeOH were added formaldehyde (13.1 M in water, 10 eq.) and an amine (10 eq.). The mixture was heated to 60 ⁰C for 3 days and allowed to come to r.t. The solvent was removed. The residue was taken in diethyl ether. A precipitate appeared; it was filtered and dried in vacuo.

Example 14: 3-[5-Methoxy-2-(2-morpholin-4-yl-ethoxy)-benzylidene]-l-morpholin-4- ylmethyl-l,3-dihydro-indol-2-one; hydrochloride

General procedure B was followed using preparation 3 (75 mg, 0.18 mmol), formaldehyde (0.14 mL) and morpholine (0.16 ml_) in MeOH (2 ml_). Filtration afforded 34 mg (40%) of the title compound as a yellow solid.

¹³C NMR (DMSO-c/e) δ 167.8, 153.2, 150.2, 143.7, 132.3, 129.9, 126.8, 124.0, 122.1, 121.8, 120.2, 116.9, 114.4, 110.3, 65.9, 63.7, 63.1, 61.3, 55.6, 55.2, 52.0, 50.7, 42.5

Example 15: 3-[2-(2-Diethylamino-ethoxy)-5-methoxy-benzylidene]-l-morpholin-4- ylmethyl-l,3-dihydro-indol-2-one; hydrochloride

CIH

General procedure B was followed using preparation 4 (75 mg, 0.18 mmol), formaldehyde (0.14 ml_) and morpholine (0.16 mL) in MeOH (2 mL). Filtration afforded 13 mg (15%) of the title compound as a yellow solid.

¹³C NMR (DMSO-CZ₆) δ 167.8, 153.1, 150.1, 143.8, 132.2, 129.9, 126.8, 123.7, 122.1, 121.8, 120.2, 116.9, 114.5, 113.7, 110.3, 65.9, 61.3, 55.6, 50.7, 49.6, 47.3, 8.7

Example 16: 3-[5-Methoxy-2-(2-piperidin-l-yl-ethoxy)-benzylidene]-l-morpholin-4- ylmethyl-l,3-dihydro-indol-2-one; hydrochloride

General procedure B was followed using preparation 5 (75 mg, 0.18 mmol), formaldehyde (0.14 mL) and morpholine (0.16 mL) in MeOH (2 mL). Filtration afforded

19 mg (22%) of the title compound as a yellow solid.

¹³C NMR (DMSO-CZ₆) δ 167.9, 153.2, 150.1, 143.8, 132.3, 130.0, 126.9, 123.9, 122.2, 121.9, 120.2, 117.0, 114.5, 114.2, 110.4, 66.0, 61.3, 55.6, 54.9, 52.8, 50.7, 48.3,

22.6, 21.3

General procedure C

To a solution of preparation 1 or preparation 2 in DMF were added an aldehyde, acetic anhydride (2 eq.) and triethylamine (2 eq.). The solution was heated to 80 ⁰C. After reaction completion the solution was concentrated in vacuo and purified by flash chromatography.

Example 17: Acetic acid l-[3-(2,5-dimethoxy-benzylidene)-2-oxo-2,3-dihydro-indol-l- yl]-ethyl ester

General procedure C was followed using preparation 1 (422 mg, 1.5 mmol), acetaldehyde (0.85 mL, 10 eq.), acetic acid anhydride (0.28 ml_) and NEt₃ (0.42 ml_) in DMF (5 mL). The reaction time was 2 h. Flash chromatography afforded 240 mg (44%) of the title compound as a yellow solid.

¹H NMR (DMSO-CZ₆) δ 7.72 (s,lH), 7.52 (d,lH), 7.38 - 7.25 (m,2H), 7.22 (d,lH), 7.15 - 7.04 (m,2H), 6.99 (t,lH), 6.84 (q,lH), 3.80 (s,3H), 3.73 (s,3H), 2.04 (s,3H), 1.72 (d,3H)

Example 18: Acetic acid [3-(2,5-dimethoxy-benzylidene)-2-oxo-2,3-dihydro-indol-l-yl]- (2,2-dimethyl-[l,3]dioxolan-4-yl)-methyl ester

General procedure C was followed using preparation 1 (422 mg, 1.5 mmol), 2,2- dimethyl-[l,3]dioxolane-4-carbaldehyde (399 mg, 2 eq.), acetic acid anhydride (0.28 mL) and NEt₃ (0.42 mL) in DMF (5 mL). The reaction time was 15 h. Flash chromatography afforded 125 mg (18%) of the title compound as a yellow oil. ¹³C NMR (DMSO-CZ₆) δ 168.8, 166.6, 152.5, 151.8, 140.5, 133.1, 129.9, 125.6, 122.9, 122.4, 122.1, 120.5, 117.4, 114.1, 112.7, 111.1, 109.6, 75.2, 71.8, 65.6, 55.9, 55.5, 26.4, 24.8, 20.3 Example 19: Acetic acid l-[3-(2,5-dimethyl-benzylidene)-2-oxo-2,3-dihydro-indol-l-yl]- ethyl ester

General procedure C was followed using preparation 2 (500 mg, 2.0 mmol), acetaldehyde (1.13 ml_, 10 eq.) acetic acid anhydride (0.38 mL) and NEt₃ (0.56 ml_) in DMF (6 mL). The reaction time was 2 h. Flash chromatography afforded 144 mg (21%) of the title compound as an orange oil. ¹³C NMR (DMSO-CZ₆) δ 168.9, 165.6, 140.3, 136.6, 134.9, 133.6, 133.3, 130.3, 130.2, 129.9, 128.4, 126.5, 122.3, 122.1, 120.7, 111.0, 72.8, 20.4, 20.3, 18.9, 17.6

Example 20: 3-[Hydroxy-(4-nitro-phenyl)-methylene]-2-oxo-2,3-dihydro-indole-l- carboxylic acid phenyl ester

To a solution of 2-indolinone (1.33 g, 10 mmol) in THF (30 mL) were added NEt₃ (3 mL, 22 mmol) and phenyl chloroformate (2.6 mL, 22 mmol). Stirring at r.t. was continued for 4 h before addition of water (20 mL) and ice. The precipitate was filtered and washed with water to afford 3.38 g (90%) of 2-phenoxy carbonyloxy-indole-1-carboxylic acid phenyl ester.

To an ice-cold solution of 2-phenoxycarbonyloxy-indole-l-carboxylic acid phenyl ester (3.73 g, 10 mmol) in DMF (50 mL) was added (NhU)₂CO₃ (0.8 g, 8 mmol). Stirring was continued while the ice-bath was melting. After 3 h water (100 mL) was added. The precipitate was filtered and washed with water to afford 2.0 g (79%) of 2-oxo-2,3- dihydro-indole-1-carboxylic acid phenyl ester.

To an ice-cold solution of 2-oxo-2,3-dihydro-indole-l-carboxylic acid phenyl ester (0.51 g , 2 mmol) in DMF (5 mL) were added 4-nitrobenzoylchloride (0.41 g, 2.2 mmol) and

DMAP (0.54 g, 4.4 mmol). Stirring at r.t. was continued for 1.5 h before addition of water (10 mL). The precipitate was filtered and washed with water. This solid was suspended in water (50 mL) and treated with glacial acetic acid (20 mL). The mixture turned from red to yellow. The solid was filtered and washed with water to afford 0.78 g (96%) of the title compound.

¹³C NMR (DMSO-CZ₆) δ 168.5, 164.7, 149.9, 149.3, 148.0, 141.7, 134.7, 130.2, 129.5, 126.1, 125.9, 124.3, 124.2, 123.1, 122.0, 121.6, 113.8, 101.5

Example 21: 3-[Hydroxy-(4-nitro-phenyl)-methylene]-2-oxo-2,3-dihydro-indole-l- carboxylic acid amide

To a solution of 3-[hydroxy-(4-nitro-phenyl)-methylene]-2-oxo-2,3-dihydro-indole-l- carboxylic acid phenyl ester (0.26 g, 0.5 mmol) in DMF (5 mL) was added (NH₄)₂CO₃

(0.08 g, 0.8 mmol).The mixture was heated to 80 ⁰C for 20 h. HCI (37%, 0.13 mL) in water (2.5 mL) was added. The precipitate was filtered and washed with water to afford

0.12 g (75%) of the title compound.

¹³C NMR (DMSO-CZ₆) δ 168.9, 168.3, 152.6, 148.1, 141.7, 135.9, 130.2, 125.7, 123.4, 123.1, 121.6, 114.6, 102.2 Example 22

EAE activity of compounds of formula I on oral administration

Materials and Methods Compounds

Compound A: 3-(2,5-Dimethoxy-benzylidene)-l-piperidin-l-ylmethyl-l,3-dihydro-indol- 2-one;

Compound B: l-[l,4']Bipiperidinyl-l'-ylmethyl-3-(2,5-dimethoxy-benzylidene)-l,3- dihydro-indol-2-one; Compound C: 3-(2,5-dimethoxy-benzylidene)-l,3-dihydro-indol-2-one (compound 110 in PCT/DK2004/000875)

Peptide The following peptide from myelin proteolipid protein was used; PLPi_39-I53 H- HCLGKWLGHPDKFVG-OH. The peptide was synthesized by Fmoc chemistry (Schafer-N, Copenhagen, Denmark). Purity (>95%) was verified by reversed-phase HPLC and integrity by mass spectrometry.

Mice Female SJL/J (H-2^S) inbred mice purchased from Charles River.

Immunization The SJL/J mice (about 8 weeks old) were immunized on day 0 with the PLPi₃₉-i₅₃ peptide (dissolved in sterile NaCI) emulsified 1:1 (vol/vol) in Complete Freund's Adjuvant (5 mg Mycobacterium tuberculosis/ ^'ml) (SSI, Copenhagen, Denmark). Intradermal injections corresponding to 100 μg peptide and 125 μg Mycobacterium tuberculosis were given at the base of the tail in a total volume of 50 μl. The mice were additionally given an i.v. injection with 100 ng pertussis toxin (Sigma) dissolved in sterile NaCI on day 0 and day 2, injection volume was 100 μl.

Compound treatment Groups of 10 mice were dosed daily with compounds (50 mg/kg p.o.) in suspension vehicle (4 g Tween-80, 2 g Carboxy-methyl cellulose 7H4XF, 8 g NaCI, 1 liter H₂O), starting on day 1. Control groups were given either suspension vehicle or dexamethasone (Dexadreson Vet, Intervet, Holland) (1 mg/kg).

Clinical evaluation Mice were weighed and assessed clinically daily from day 5 p.i. according to the following criteria: 0, no disease; 1, tail paralysis; 2, clumsy gait/poor righting ability and limb weakness; 3, moderate or total hind limb paralysis; 4, moribund state or dead. Statistics

Area-under-curve (AUC) of the disease score was calculated for all mice. The medians of AUC of all groups were compared using Kruskal-Wallis test. When P<0.05 in the Kruskal- Wallis test, the Mann-Whitney test was used to compare drug treated groups with the suspension vehicle treated control group (P<0.05). AUC were calculated from day 0 to the termination of the experiment (day 21).

Results

Compounds of formula I were tested orally in the EAE model. The results for two of the compounds are shown in table 1. The inhibitory effects apparent from Table 1 are significant (p<0.05). The compounds exhibit a high activity in the EAE model when dosed orally.

Table 1. Inhibition of EAE on oral administration of compounds I

* Area-under-curve of the disease score. A suspension vehicle group was included in all experiments. AUC of the disease score were calculated for all groups and the Mann- Withney test was used to compare the treated groups with the suspension vehicle group. ** The average (8 experiments) inhibition of EAE after treatment with Compound C.

Claims

1. A compound of general formula I

wherein X is O or S;

R₁ is -OC(O)R₉, -NR₁₀Rn, -C(O)NR₁₀R₁₁, -OC(O)NR₁₀R₁₁, -NHC(O)R₁₀, -NHC(O)OR₁₀, - NHC(O)NR₁₀R₁₁, -S(O)R₉, -S(O)₂R₉, -S(O)₂OR₉, -S(O)₂NR₁₀R₁₁, -OR₁₂, -C(O)R₁₂, - C(O)OR₁₂, -OC(O)OR₁₂, -P(O)(OR₁₀)(OR₁₁), -OP(O)(OR₁₀)(OR₁₁), polyoxyethylene, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, carbocyclyl, heteroaryl or heterocyclyl, said C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, carbocyclyl, heteroaryl or heterocyclyl being substituted with one or more substituents selected from the group consisting of -OR₁₀, - C(O)R₁₀, -C(O)OR₁₀, -OC(O)R₁₀, -OC(O)OR₁₀, -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -OC(O)NR₁₀R₁₁, - OP(O)(OR₁₀)(OR₁₁), -P(O)(OR₁₀)(OR₁₁), -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, - SR₁₀, -S(O)R₁₀, -S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁, -S(O)₂OR₁₀, polyoxyethylene, aryl, heteroaryl, carbocyclyl and heterocyclyl, said aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with -OR₁₀, -C(O)R₁₀, -C(O)OR₁₀, OC(O)R₁₀, -OC(O)OR₁₀, - P(O)(OR₁₀)(OR₁₁), -OP(O)(OR₁₀)(OR₁₁), -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -NHC(O)R₁₀, - NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, -S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁, -S(O)₂OR₁₀ or polyoxyethylene, and C_1-6 alkyl substituted with polyoxyethylene, -OR₁₀, -C(O)R₁₀, - C(O)OR₁₀, OC(O)R₁₀, -OC(O)OR₁₀, -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -P(O)(OR₁₀)(OR₁₁), - OP(O)(OR₁₀)(OR₁₁), -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, - S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁ or -S(O)₂OR₁₀, wherein

R₉ is C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl or heterocyclyl, said C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl or heterocyclyl being optionally substituted with one or more substituents selected from the group consisting of halogen, trihalomethyl, Ci_-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, polyoxyethylene, -ORi₃, -C(O)Ri₃, -C(O)ORi₃, -OC(O)Ri₃, - OC(O)ORi₃, -N₃, -NRi₃Ri₄, -C(O)NRi₃R₁₄, -OC(O)NR₁₃Ri₄, -NHC(O)Ri₃, -NHC(O)ORi₃, - NHC(O)NRi₃Ri₄, -P(O)(OR₁₃)(ORi₄), -OP(O)(ORi₃)(ORi₄), -SRi₃, -S(O)Ri₃, "S(O)₂Ri₃, - S(O)₂NRi₃Ri₄ and -S(O)₂OR₁₃;

R₁₀ and R₁₁ are the same or different and independently selected from the group consisting of hydrogen, polyoxyethylene, C_1-10 alkyl, C_2-10 alkenyl, C_2-I0 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₀ and R₁₁, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, said Ci- _lo alkyl being substituted with one or more, same or different substituents selected from the group consisting of halogen, trihalomethyl, C_x-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, polyoxyethylene, -0R_i3, -C(O)Ri₃, -C(O)ORi₃, -OC(O)Ri₃, - OC(O)OR₁₃, -NR₁₃R₁₄, -C(O)NR₁₃R₁₄, -OC(O)NR₁₃R₁₄, -NHC(O)R₁₃, -NHC(O)OR₁₃, - NHC(O)NR₁₃R₁₄, -P(O)(OR₁₃)(OR₁₄), -OP(O)(OR₁₃)(OR₁₄), -SR₁₃, -S(O)R₁₃, -S(O)₂R₁₃, - S(O)₂NR₁₃R₁₄ and -S(O)₂OR₁₄, said aryl, heteroaryl, carbocyclyl or heterocyclyl being optionally substituted with one or more substituents selected from the group consisting of -OR₁₃, -C(O)R₁₃, -C(O)OR₁₃, -C(O)NR₁₃R₁₄ and polyoxyethylene, and said C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, C_1-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, polyoxyethylene, -OR₁₃, -C(O)R₁₃, -C(O)OR₁₃, -OC(O)R₁₃, -OC(O)OR₁₃, - NR₁₃R₁₄, -C(O)NR₁₃R₁₄, -OC(O)NR₁₃R₁₄, -NHC(O)R₁₃, -NHC(O)OR₁₃, -NHC(O)NR₁₃R₁₄, - P(O)(OR₁₃)(OR₁₄), -OP(O)(ORi₃)(ORi₄), -SR₁₃, -S(O)Ri₃, -S(O)₂Ri₃, -S(O)₂NR₁₃Ri₄ and - S(O)₂OR₁₄, said aryl, heteroaryl, carbocyclyl or heterocyclyl being optionally substituted with one or more substituents selected from the group consisting of -OR₁₃, -C(O)R₁₃, - C(O)ORi₃, -C(O)NR₁₃R₁₄ and polyoxyethylene, provided that R₁₀ and R₁₁ are not both hydrogen;

R₁₂ is C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl or carbocyclyl substituted with hydroxy, amino, cyano, halogen, carboxy, nitro, amido, polyoxyethylene, -OR₁₃, -C(O)R₁₃, - C(O)OR₁₃, -OC(O)R₁₃, -OC(O)OR₁₃, -NR₁₃R₁₄, -C(O)NR₁₃R₁₄, -OC(O)NR₁₃R₁₄, -NHC(O)R₁₃, -NHC(O)OR₁₃, -NHC(O)NRi₃R₁₄, -P(O)(OR₁₃)(OR₁₄), -OP(O)(OR₁₃)(OR₁₄), -SR₁₃, -S(O)R₁₃, -S(O)₂R₁₃, -S(O)₂NR₁₃R₁₄ and -S(O)₂OR₁₄, or heteroaryl or heterocyclyl optionally substituted with hydroxy, amino, cyano, halogen, carboxy, nitro, amido, polyoxyethylene, -ORi₃, -C(O)R₁₃, -C(O)OR₁₃, -OC(O)R₁₃, -OC(O)OR₁₃, -NR₁₃R₁₄, - C(O)NR₁₃R₁₄, -OC(O)NR₁₃R₁₄, -P(O)(OR₁₃)(OR₁₄), -OP(O)(OR₁₃)(OR₁₄), -NHC(O)R₁₃, - NHC(O)OR₁₃, -NHC(O)NR₁₃R₁₄, -SR₁₃, -S(O)R₁₃, -S(O)₂R₁₃, -S(O)₂NR₁₃R₁₄ and - S(O)₂OR₁₄; R₁₃ and R₁₄ are the same or different and independently selected from the group consisting of hydrogen, C_1-6-alkyl, C_2-6-alkenyl, C_4-6-alkadienyl, C_2-6-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₃ and R₁₄, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, each C₁. ₆ alkyl, C_2-6 alkenyl, C_2-6 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl substituent being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, hydroxy, C_1-4 alkyl, C_1-4 alkoxy, nitro, cyano, amino, oxo, halogen, trihalomethyl, C_1-4 alkylthio, C_1-4 alkylamino, C_1-4 alkoxycarbonyl, carboxy, -CONH₂ Or -S(O)₂NH₂;

R₂, R₃, R₄ and R₅ are the same or different and independently selected from the group consisting of hydrogen, halogen, trihalomethyl, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, -OR₁₅, -C(O)R₁₅, -C(O)OR₁₅, -OC(O)R₁₅, -OC(O)OR₁₅, - NR₁₅R₁₆, -C(O)NR₁₅R₁₆, -OC(O)NR₁₅R₁₆, -NHC(O)R₁₅, -NHC(O)OR₁₅, -NHC(O)NR₁₅R₁₆, - SR₁₅, -S(O)R₁₅, -S(O)₂R₁₅, -S(O)₂NR₁₅R₁₆ and -S(O)₂OR₁₅, wherein R₁₅ and R₁₆ are the same or different and independently selected from the group consisting of hydrogen, C_{1- lo} alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₅ and R₁₆, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, said C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, C_1-6-alkyl, C₂.₆-alkenyl, C_2-6-alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, - OR₁₇, -C(O)R₁₇, -C(O)OR₁₇, -OC(O)OR₁₇, -OC(O)R₁₈, -NR₁₇R₁₈, -C(O)NR₁₇R₁₈, - OC(O)NR₁₇R₁₈, -NHC(O)R₁₇, -NHC(O)OR₁₇, -NHC(O)NR₁₇R₁₈, -SR₁₇, -S(O)R₁₇, -S(O)₂R₁₇, - S(O)₂NR₁₇R₁₈ and -S(O)₂OR₁₇, wherein R₁₇ and R₁₈ are the same or different and independently selected from the group consisting of hydrogen, C_1-6-alkyl, C_2-6-alkenyl, C₂.₆-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₇ and R₁₈, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, each C_1-6-alkyl, C_2-6-alkenyl, C₂.₆-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl substituent being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, hydroxy, C^₄ alkyl, Ci_-4 alkoxy, nitro, cyano, amino, oxo, halogen, trihalomethyl, C₁-₄ alkylthio, Ci_-4 alkylamino, C_1-4 alkoxycarbonyl, carboxy, -CONH₂ or -S(O)₂NH₂;

R₆ is hydrogen, C_1-6 alkyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, halogen, -OR₇, - C(O)R₇, -C(O)OR₇, -NR₇R₈, -S(O)₂NR₇R₈, wherein R₇ and R₈ are independently hydrogen, C_1-6 alkyl, aryl or heterocyclyl, said C_1-6 alkyl or heterocyclyl being optionally substituted by heterocyclyl, -OR₇, -C(O)R₇ or -C(O)OR₇;

R₁', R₂' R₃', R₄' and R₅' are the same or different and independently selected from the group consisting of hydrogen, halogen, trihalomethyl, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, -OR₁₅, -C(O)R₁₅, -C(O)OR₁₅, -OC(O)R₁₅, - OC(O)OR₁₅, -NR₁₅R₁₆, -C(O)NR₁₅R₁₆, -OC(O)NR₁₅R₁₆, -NHC(O)R₁₅, -NHC(O)OR₁₅, - NHC(O)NR₁₅R₁₆, -SR₁₅, -S(O)R₁₅, -S(O)₂R₁₅, -S(O)₂NR₁₅R₁₆ and -S(O)₂OR₁₆, wherein R₁₅ and R₁₆ are the same or different and independently selected from the group consisting of hydrogen, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₅ and R₁₆, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, said C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, Ci.₆-alkyl, C_2-6-alkenyl, C_2-6-alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, -OR₁₇, -C(O)R₁₇, -C(O)OR₁₇, -OC(O)R₁₈, -OC(O)OR₁₇, -NR₁₇R₁₈, -C(O)NR₁₇R₁₈, -OC(O)NR₁₇R₁₈, -NHC(O)R₁₇, -NHC(O)OR₁₇, -NHC(O)NR₁₇R₁₈, -SR₁₇, -S(O)R₁₇, -S(O)₂R₁₇, -S(O)₂NR₁₇R₁₈ and -S(O)₂OR₁₇, wherein R₁₇ and R₁₈ are the same or different and independently selected from the group consisting of hydrogen, C_1-6-alkyl, C_2-6-alkenyl, C_2-6-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₇ and R₁₈, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, each C_1-6-alkyl, C_2-6-alkenyl, C_2-6-alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl substituent being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, hydroxy, C_1-4 alkyl, C_1-4 alkoxy, nitro, cyano, amino, oxo, halogen, trihalomethyl, Ci_-4 alkylthio, C_1-4 alkylamino, C_1-4 alkoxycarbonyl, carboxy, -CONH₂ or -S(O)NH₂; or pharmaceutically acceptable salts thereof.

2. A compound according to claim 1, wherein R₂, R₃, R₄ and R₅ are all hydrogen.

3. A compound according to claim 1, wherein R₁ is -OC(O)R₉, -C(O)NRI₀RII, - OC(O)NR₁₀R₁₁, -S(O)₂R₉, -OR₁₂, -C(O)R₁₂, -C(O)OR₁₂, -OC(O)OR₁₂, -P(O)(OR₁₀)(OR₁₁), - OP(O) (OR₁₀) (ORI₁), polyoxyethylene, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl or aryl, said C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl or aryl being substituted with one or more substituents selected from the group consisting of -OR₁₀, -C(O)R₁₀, -C(O)OR₁₀, -

OC(O)R₁₀, -OC(O)OR₁₀, -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -OC(O)NR₁₀R_UrOP(O)(OR₁₀)(OR₁₁),- P(O)(OR₁₀)(OR₁₁), -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, - S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁, -S(O)₂OR₁₀, polyoxyethylene, heteroaryl and heterocyclyl, said heteroaryl and heterocyclyl being optionally substituted with -OR₁₀, -C(O)R₁₀, -C(O)OR₁₀, OC(O)R₁₀, -OC(O)OR₁₀, -P(O)(OR₁₀)(OR₁₁), -OP(O)(OR₁₀)(OR₁₁), -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, -S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁, - S(O)₂OR₁₀ or polyoxyethylene, and C_1-6 alkyl substituted with -OR₁₀, -C(O)R₁₀, - C(O)OR₁₀, OC(O)R₁₀, -OC(O)OR₁₀, -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -P(O)(OR₁₀)(OR₁₁), - OP(O)(OR₁₀)(OR₁₁), -NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, - S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁ or -S(O)₂OR₁₀, wherein R₉, R₁₀, R₁₁ and R₁₂ are as indicated in claim 1.

4. A compound according to claim 3, wherein R₁ is -OC(O)R₉, -OR₁₂, -C(O)R₁₂ or C_1-10 alkyl, said C_1-10 alkyl being substituted with one or more substituents selected from the group consisting of polyoxyethylene, heterocyclyl, -OR₁₀, -C(O)R₁₀, -C(O)OR₁₀, -

OC(O)R₁₀, -OC(O)OR₁₀, -NR₁₀R₁₁, -C(O)NR₁₀R₁₁, -OC(O)NR₁₀R₁₁^OP(O)(OR_1O)(OR₁₁),- P(O)(OR₁₀)(OR₁₁), NHC(O)R₁₀, -NHC(O)OR₁₀, -NHC(O)NR₁₀R₁₁, -SR₁₀, -S(O)R₁₀, - S(O)₂R₁₀, -S(O)₂NR₁₀R₁₁, -S(O)₂OR₁₀ wherein R₉, R₁₀, R₁₁ and R₁₂ are as indicated in claim 1.

5. A compound according to claim 3 or 4, wherein R₁ is

wherein R₁₀, R₁₀' and R₁₁ are the same or different and independently selected from the group consisting of hydrogen, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, or wherein R₁₀ and R₁₁, together with the nitrogen atom to which they are attached form a heterocyclic or heteroaryl ring, each of C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, C_1-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, -OR₁₃, -C(O)R₁₃, -C(O)OR₁₃, -OC(O)R₁₃, -OC(O)OR₁₃, - NR₁₃R₁₄, -C(O)NR₁₃R₁₄, -OC(O)NR₁₃R₁₄, -NHC(O)R₁₃, -NHC(O)OR₁₃, -NHC(O)NR₁₃R₁₄, - P(O)(OR₁₃)(OR₁₄), -OP(O)(OR₁₃)(OR₁₄), -SR₁₃, -S(O)R₁₃, -S(O)₂R₁₃, -S(O)₂NR₁₃R₁₄ and - S(O)₂OR₁₄, wherein R₁₃ and R₁₄ are as indicated in claim 1, provided that R₁₀ and R₁₁ are not both hydrogen.

6. A compound according to claim 3 or 4, wherein R₁ is

wherein R₁₀ and R₁₀' are the same or different and independently selected from the group consisting of hydrogen, C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl, each of C_1-10 alkyl, C_2-10 alkenyl, C_2-10 alkynyl, aryl, heteroaryl, carbocyclyl and heterocyclyl being optionally substituted with one or more, same or different substituents selected from the group consisting of hydrogen, halogen, trihalomethyl, C_1-6 alkyl, C_2-6 alkenyl, C_2-6 alkynyl, hydroxy, carboxy, formyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, amino, carbamoyl, cyano, guanidino, carbamido, - OR₁₃, -C(O)R₁₃, -C(O)OR₁₃, -OC(O)R₁₃, -OC(O)OR₁₃, -NR₁₃R₁₄, -C(O)NR₁₃R₁₄, - OC(O)NR₁₃R₁₄, -NHC(O)R₁₃, -NHC(O)OR₁₃, -NHC(O)NR₁₃R₁₄, -P(O)(OR₁₃)(OR₁₄), -

OP(O)(OR₁₃)(OR₁₄), -SR₁₃, -S(O)R₁₃, -S(O)₂R₁₃, -S(O)₂NR₁₃R₁₄ and -S(O)₂OR₁₄, wherein R₁₃ and R₁₄ are as indicated in claim 1.

7. A compound according to claim 1, wherein R₆ is hydrogen.

8. A compound according to claim 1, wherein R₁', R₃', R₄', R₅' and R₆' are the same or different and independently selected from the group consisting of hydrogen, C_1-10 alkyl, C_1-10 alkoxy, aryl, heteroaryl, aryloxy, C_1-10 alkylaryl, C_1-10 alkylaryloxy, halogen, trihalomethyl, -S(O)R₂₀, -S(O)₂R₂₀, -S(O)₂NR₂₀R₂₁, -S(O)₃R₂₀, -SR₂₀, -NO₂, -NR₂₀R₂₁, - OR₂₀, -CN, -CH₂OH, -C(O)R₂₀, -C(O)OR₂₀, -OC(O)R₂₀, -NHC(O)R₂₀, -NHC(O)OR₂₀, -

NHC(O)NR₂₀R₂₁, -(CH₂)_nC(0)₂R_2o and -C(O)NR₂₀R₂₁, wherein R₂₀ is hydrogen, C_1-6 alkyl, heteroaryl or aryl, said C_1-6 alkyl, heteroaryl or aryl being optionally substituted with hydroxy or -NR₂₂R₂₃, wherein R₂₂ and R₂₃ are independently hydrogen or C_1-6 alkyl or, together with the nitrogen atom to which they are attached, form a heteroaryl or heterocyclic ring, R₂₁ is hydrogen, C_1-6 alkyl or aryl, and n is 0-3.

9. A compound according to claim 8, wherein Ri', R₂', R₃', R₄' and R₅' are the same or different and independently selected from the group consisting of hydrogen, C_1-6 alkyl, C_1-6 alkoxy and halogen.

10. A compound according to claim 1 selected from the group consisting of 3-(2,5-Dimethoxy-benzylidene)-l-hydroxymethyl-l,3-dihydro-indol-2-one 3-(2,5-Dimethoxy-benzylidene)-l-morpholin-4-ylmethyl-l,3-dihydro-indol-2-one 3-(2,5-Dimethoxy-benzylidene)-l-piperidin-l-ylmethyl-l,3-dihydro-indol-2-one 3-(2,5-Dimethoxy-benzylidene)-l-(4-methyl-piperazin-l-ylmethyl)-l,3-dihydro-indol-2- one

3-(2,5-Dimethoxy-benzylidene)-l-(3,3-dimethyl-piperidin-l-ylmethyl)-l,3-dihydro- indol-2-one l-[(Diisobutylamino)-methyl]-3-(2,5-dimethoxy-benzylidene)-l,3-dihydro-indol-2-one l-[l,4']Bipiperidinyl-l'-ylmethyl-3-(2,5-dimethoxy-benzylidene)-l,3-dihydro-indol-2-one

3-(2,5-Dimethyl-benzylidene)-l-morpholin-4-ylmethyl-l,3-dihydro-indol-2-one

3-(2,5-Dimethyl-benzylidene)-l-piperidin-l-ylmethyl-l,3-dihydro-indol-2-one

3-(2,5-Dimethyl-benzylidene)-l-(4-methyl-piperazin-l-ylmethyl)-l,3-dihydro-indol-2- one 3-(2,5-Dimethyl-benzylidene)-l-(3,3-dimethyl-piperidin-l-ylmethyl)-l,3-dihydro-indol-

2-one l-[(Diisobutylamino)-methyl]-3-(2,5-dimethyl-benzylidene)-l,3-dihydro-indol-2-one l-[l,4']Bipiperidinyl-l'-ylmethyl-3-(2,5-dimethyl-benzylidene)-l,3-dihydro-indol-2-one

3-[5-Methoxy-2-(2-morpholin-4-yl-ethoxy)-benzylidene]-l-morpholin-4-ylmethyl-l,3- dihydro-indol-2-one; hydrochloride

3-[2-(2-Diethylamino-ethoxy)-5-methoxy-benzylidene]-l-morpholin-4-ylmethyl-l,3- dihydro-indol-2-one; hydrochloride

3-[5-Methoxy-2-(2-piperidin-l-yl-ethoxy)-benzylidene]-l-morpholin-4-ylmethyl-l,3- dihydro-indol-2-one; hydrochloride Acetic acid l-[3-(2,5-dimethoxy-benzylidene)-2-oxo-2,3-dihydro-indol-l-yl]-ethyl ester

Acetic acid [3-(2,5-dimethoxy-benzylidene)-2-oxo-2,3-dihydro-indol-l-yl]-(2,2- dimethyl-[l,3]dioxolan-4-yl)-methyl ester

Acetic acid l-[3-(2,5-dimethyl-benzylidene)-2-oxo-2,3-dihydro-indol-l-yl]-ethyl ester

3-[Hydroxy-(4-nitro-phenyl)-methylene]-2-oxo-2,3-dihydro-indole-l-carboxylic acid phenyl ester

3-[Hydroxy-(4-nitro-phenyl)-methylene]-2-oxo-2,3-dihydro-indole-l-carboxylic acid amide

11. A pharmaceutical composition comprising one or more compounds of formula I together with a pharmaceutically acceptable excipient or vehicle.

12. Use of a compound of general formula I for the preparation of a medicament for preventing, treating or ameliorating multiple sclerosis, or delaying the onset of or reducing the relapse rate in multiple sclerosis.

13. A method of preventing, treating or ameliorating multiple sclerosis, or delaying the onset of or reducing the relapse rate in multiple sclerosis, the method comprising administering to a patient in need thereof an effective amount of a compound of general formula I.