WO2012022740A1

WO2012022740A1 - Combination treatment of multiple sclerosis

Info

Publication number: WO2012022740A1
Application number: PCT/EP2011/064080
Authority: WO
Inventors: Manolo Beelke; Jérôme Dauvillier; Isabelle Guilleret; Emmanuel Monnet
Original assignee: Merck Serono S.A.
Priority date: 2010-08-18
Filing date: 2011-08-16
Publication date: 2012-02-23

Abstract

The present invention is related to the use of Vitamin D in combination with Interferon-beta for the treatment of multiple sclerosis.

Description

Combination treatment of Multiple Sclerosis

Field of the Invention

The present invention relates to the use of Vitamin D in combination with Interferon-beta for the treatment of multiple sclerosis.

Background of the Invention

Multiple sclerosis (MS) is the most investigated chronic inflammatory demyelinating disease of the central nervous system in humans. The onset of the disease typically occurs during ages 20 to 40. Women are affected approximately twice as often as men.

Over time, MS may result in the accumulation of various neurological disabilities. Clinical disability in MS is presumed to be a result of repeated inflammatory injury with subsequent loss of myelin and axons, leading to tissue atrophy.

MS is manifested in physical symptoms (relapses and disability progression), Central Nervous System (CNS) inflammatory and neurodegenerative processes (as reported on MRI read outs), brain atrophy and cognitive impairment. Presenting symptoms include focal sensory deficits, focal weakness, visual problems, imbalance and fatigue. Sexual impairment and sphincter dysfunction may occur. Approximately half of the patients with MS may experience cognitive impairment or depression.

MS is now considered to be a multi-phasic disease and periods of clinical quiescence (remissions) occur between exacerbations. Remissions vary in length and may last several years but are infrequently permanent.

Four main forms of the disease are described: relapsing-remitting (RR), secondary progressive (SP), primary progressive (PP) and progressive relapsing (PR) multiple sclerosis.

More than 80% of patients with MS have been diagnosed as RR form of the disease with clinical exacerbation of neurological symptoms, followed by a recovery that may or may not be complete (Lublin and Reingold, Neurology, 1996, 46:907-911). In RRMS, accumulation of disability results from progressive inflammatory insults with only partial repair or absence of repair of the damaged area of the CNS. At the early stages of the disease brain plasticity allows a functional recovery where anatomical repair is not possible. Nevertheless, over time the functional areas are reduced, which will clinically express as an incomplete recovery from relapses and a consequently increase of disability (loss of functions). When this stage is reached, patients with RRMS switch to a progressive course, called SPMS. Before the availability of disease modifying drugs (DMDs), it has been described that ten years after disease onset approximately half of the, patients with RRMS at diagnosis converted to SPMS. During the SP phase, worsening of disability results from the accumulation of residual symptoms after exarcerbation but also from insidious progression between exacerbations {Lublin and Reingold above). 10% of MS patients have PPMS at time of diagnosis which is characterized by insidious progression of the symptoms from the disease onset. Less than 5 % of patients have PRMS and are often considered to have the same prognosis as PPMS. It is suggested that distinct pathogenic mechanisms may be involved in different patient sub-groups and have wide-ranging implications for disease classification (Lassmann et al, 2001, Trends Mol. Med., 7, 115-121; Lucchinetti et al, Curr. Opin. Neurol, 2001, 14, 259-269).

MS onset is defined by the occurrence of the first neurological symptoms of CNS dysfunction in addition to at least further signs of dissemination in space and time of CNS lesions (MRI supported or clinically confirmed). Advances in cerebrospinal fluid (CSF) analysis and magnetic resonance imaging (MRI) have simplified the diagnostic process and facilitated early diagnostic. The International Panel on the Diagnosis of MS issued revised criteria facilitating the diagnosis of MS and including MRI together with clinical and para-clinical diagnostic methods (Polman CH, Reingold SC, Edan G, Filippi M, Hartung HP, Kappos L, Lublin FD, Metz LM, McFarland HF, O 'Connor PW, Sandberg-Wollheim M, Thompson A J, Weinshenker BG, Wolinsky JS. Diagnostic criteria for multiple sclerosis: 2005 revisions to the "McDonald Criteria". Ann Neurol. 2005 Dec; 58(6): 840ff).

Current medications for MS which are disease modifying treatments, i.e. modifying the course of MS, modulate or suppress the immune system. There are several FDA approved immunomodulating agents for RRMS including: beta interferons (Betaseron®, Berlex; Avonex®, Biogen; Rebif®, Merck Serono; Extavia®, Novartis), a humanized monoclonal antibody natalizumab (Tysabri®, Biogen and Elan), and Glatiramer Acetate (Copaxone®, Amgen). There is also one FDA approved immunosuppressing drug for worsening MS, Mitoxantrone (Novantrone®, Amgen). Azathioprine is produced by a number of generic manufacturers and as branded name, it is approved for example in Germany. Several other immunosuppressive agents are used, although not FDA approved. Among them, Cladribine, a chlorinated purine analogue 2-chloro- 2'deoxyadenosine analogue (2-CdA), has been suggested to be useful in the treatment of MS (EP 626853B1 and US 5,506,214).

Considering the chronic nature of multiple sclerosis, long-term treatment is required as well as high treatment adherence.

There is a need for improved methods for treating MS. In particular, there is a need to further improve the efficacy of MS treatment using interferons, in particular Rebif®, without negatively affecting safety, tolerability and immunogenicity of the current standard treatment for MS.

Summary of the Invention

The present invention is directed towards a method of treating a subject suffering from multiple sclerosis wherein Vitamin D or an analogue thereof is administered on top of Interferon-beta.

In a preferred embodiment of the present invention Vitamin D or an analogue thereof is administered during a titration period of 4 weeks in a dose of 7Ό00 IU/day and thereafter in a dose of 14Ό00 IU/day.

In a preferred embodiment of the present invention Interferon-beta is Interferon- betala, more preferably Rebif®.

In a preferred embodiment of the present invention Interferon is administered in a dose of 22 or 44 μg three times a week.

In the methods according to the present invention Vitamin D or an analogue thereof is administered orally and Interferon is administered intramuscularly or subcutaneously.

In a preferred embodiment of the present invention the subject has a 25-hydroxy- vitamin D plasma level before treatment below 150 nmol/L.

In a preferred embodiment of the present invention the subject is carrier of a variant of the CYP24A1 gene. Detailed Description of the invention

Four of the currently approved MS therapeutics (Rebif®, Avonex®, Betaseron/Betaferon®, and Copaxone®) are regarded as effective and well tolerated in the treatment of people suffering from MS. As MS is a chronic disease without any cure currently available, the above-mentioned drugs are the only therapeutic agents able to slow down the disease progression of MS. Considering the chronic nature of the disease, long-term treatment is required, as well as high treatment adherence. The currently marketed formulation of Rebif® (recombinant human interferon beta- la) has been evaluated in a series of well-controlled MS studies that have demonstrated that Rebif® significantly reduces clinical attack rate, MRI lesion activity, accumulation of new lesion burden, and disability progression. Long-term data have also demonstrated that Rebif® at either 22 μg or 44 μg tiw maintains these clinical benefits in a dose-related manner (JG Phadke et al, J. Epidemiol Community Health 1987; 41:5-13), and that high-dose, high-frequency administration (44 μg tiw) provides more clinical benefit than 30 μg administered once weekly (Panitch H. et al, Journal of the Neurological Sciences 2005; 239(l):67-74).

It has now been concluded that the efficacy of Interferon-beta therapy of MS may be further improved by adding Vitamin D or an analogue thereof to the treatment regime. Vitamin D or an analogue thereof is administered according to the invention during a titration period in a dose of 7Ό00 IU/day and thereafter in a dose of 14Ό00 IU/day. The titration period has a duration of about 4 weeks. This dosing leads to particularly good results.

Interferon-beta is administered in the methods of the invention according to approved standard treatment regimes, in particular in a dose of 22 or 44 μg three times a week.

The term "interferon-beta (IFN-β)", as used herein, is intended to include fibroblast interferon in particular of human origin, as obtained by isolation from biological fluids or as obtained by DNA recombinant techniques from prokaryotic or eukaryotic host cells, as well as its salts, functional derivatives, variants, analogs and active fragments.

IFN-β suitable in accordance with the present invention is commercially available e.g. as Rebif® (Serono), Avonex® (Biogen) or Betaferon® (Schering). The use of interferons of human origin is also preferred in accordance with the present invention. The term interferon, as used herein, is intended to encompass salts, functional derivatives, variants, analogs and active fragments thereof. Rebif® (recombinant human interferon-β) is the latest development in interferon therapy for multiple sclerosis (MS) and represents a significant advance in treatment. Rebif® is interferon (IFN)-beta la, produced from mammalian cell lines. It was established that interferon beta- la given subcutaneously three times per week is efficacious in the treatment of Relap sing-Remitting Multiple Sclerosis (RRMS). Interferon beta- la can have a positive effect on the long-term course of MS by reducing number and severity of relapses and reducing the burden of the disease and disease activity as measured by MRI.

The dosing of IFN-β in the treatment of relapsing-remitting MS according to the invention depends on the type of IFN-β used.

In accordance with the present invention, where IFN is recombinant IFN- ib produced in E. Coli, commercially available under the trademark Betaseron®, it may be administered sub-cutaneously every second day at a dosage of about of 250 to 300 μg or 8 MIU to 9.6 MIU per person.

In accordance with the present invention, where IFN is recombinant IFN- ia, produced in Chinese Hamster Ovary cells (CHO cells), commercially available under the trademark Avonex®, it may be administered intra-muscularly once a week at a dosage of about of 30μg to 33 μg or 6 MIU to 6.6 MIU per person.

In accordance with the present invention, when IFN is recombinant IFN- ia, produced in Chinese Hamster Ovary cells (CHO cells), commercially available under the trademark Rebif®, it may be administered sub-cutaneously three times a week (TIW) at a dosage of 22 to 44 μg or 6 MIU to 12 MIU per person.

Vitamin D according to the invention is preferably Vitamin D3 (cholecalciferol). Vitamin D according to the invention may also be Vitamin D2 (ergocalciferol). According to the invention it is most preferred to use the cholecalciferol product Vigantol® oil. "Analogues of Vitamin D" according to the invention are structurally modified Vitamin D derivatives which are approximately equal to Vitamin D in lessening a patient's multiple sclerosis symptoms when used in combination with Interferon-beta, while preferably producing fewer side effects. The term analogues of Vitamin D encompasses for example the following compounds: 22- Oxacalcitriol, Paricalcitol, Doxercalciferol, Alfacalcidol, Dihydrotachysterol₂ and Falecalcitriol. 22-oxacalcitriol (OCT) differs from l,25(OH)₂D₃ by virtue of an oxygen atom replacing carbon-22 on the side chain. Paricalcitol is a vitamin D₂ derived sterol lacking the carbon- 19 methylene group found in all natural vitamin D metabolites. Doxercalciferol (lo:-hydroxy vitamin D₂), like alfacalcidol (lis-hydroxy vitamin D₃), is a pro-drug which is hydroxylated in the liver to 1« ,25(OH)₂D₂. Unlike alfacalcidol, doxercalciferol is also 24-hydroxylated to produce le¾,24(S)- (OH)₂D₂, a metabolite with potent pro-differentiation actions and low calcaemic potency. Dihydrotachysterol₂ (DHT₂), hydroxylated in vivo to 25(OH)DHT₂ and l,25(OH)₂DHT₂ is also of interest. The fluorinated calcitriol analogue 26,27-hexafluorocalcitriol (falecalcitriol) may also be used. (S. Steddon et al, Nephrol Dial Transplant, 2001, 16: 1965-1967).

Vitamin D is biologically inactive and is a precursor for hormone-like active substances that influence the regulation of the calcium and phosphate balance. The most important, though most probably not the only, metabolically active metabolite is 1, 25-dihydroxycholecalciferol (1, 25- (OH)₂D₃) or calcitriol. The cellular effect of calcitriol is mediated, like the mechanism of action of other steroid hormones, via binding to a nuclear receptor protein, the vitamin D receptor. Calcitriol regulates the transcription of specific genes. The primary sites of activity for the vitamin D hormones are the bone intestine, kidney, and parathyroid.

Calcitriol plays not only a pivotal role in systemic calcium homeostasis but also in the intracellular calcium homeostasis of various tissues. Vitamin D receptors are present in more than 30 different tissues and calcitriol can be locally produced in tissues that possess vitamin D receptors. The number of genes known to be regulated by the vitamin D hormone is still growing. Effects on other cell systems include modulation of the immune system and inhibition of proliferation of cancer cells. Epidemiological data indicate that a low vitamin D status is correlated with disturbed muscle function, tuberculosis, rheumatoid arthritis, type 1 diabetes, multiple sclerosis (MS), inflammatory bowel diseases, hypertension and specific types of cancer (Barthel et al. Dtsch Med Wochenschr 2003, 128:440-46; Bikle DD, Current Opinion in Rheumatology 2007, 19, 4:383-8; Zittermann A., British Journal of Nutrition 2003, 89:552-572).

The interest in the role of vitamin D in the pathogenesis and severity of MS has been increasing over the last years. Vitamin D₃ decreases the proliferation of pro-inflammatory T lymphocytes and influences the production of cytokines, both of which contribute to the pathogenesis of MS (Cantorna et al. J Immunol 1998, 160 :5314-5319 ; Kimball et al, Am J Clin Nutr 2007, 86:645- 651; Takeuchi et al, J Immunol 1998, 160:209-218 )

Therefore, low serum 25-hydroxyvitamin D (25(OH) D) levels are associated with increased multiple sclerosis prevalence and risk (Burton et al, Multiple Sclerosis 2008, 14 (suppl. 1):34). 25(OH) D levels below 100 nmol/1 are considered as vitamin D deficient

Evidence from large prospective epidemiologic studies suggests that compared with individuals with 25(OH) D levels between 99.2 and 152.9 nmol/1, individuals with lower levels of vitamin D have a significantly higher risk of developing MS (Hiremath et al, Multiple Sclerosis 2009, 15: 735-740 ).

Munger et al. (Neurology 2004, 62:60-65) evaluated the dietary vitamin D intake in relation to risk of MS in two large cohorts of women: the Nurses^' Health Study (NHS, 253 women followed from 1980 to 2000) and Nurses^' Health Study II (NHS II, 310 women followed from 1991 to 2001). The authors found an inverse association between the intake of vitamin D from supplements and the risk of developing MS. Women who used supplemental vitamin D, largely from multivitamins, had a 40% lower risk of MS than women who did not use vitamin D supplements. The RR comparing women with intake of >400 IU/day with women with no supplemental vitamin D intake was 0.59 (95% CI= 0.38 to 0.91; p for trend = 0.006).

According to Hiremath et al (2009) up to 90% of all MS patients are vitamin D deficient.

First clinical data, from studies conducted in subjects with Vitamin D deficiency, however not suffering from multiple sclerosis, suggest that between 7,000 and 14,000 IU of vitamin D₃ daily are needed to reach sufficient 25(OH) D serum levels (Hathcock et al. Am J Clin Nutr 2007, 85:6-18;

Vieth R., Am J Clin Nutr 1999, 69:842-856; Vieth R., Ann Epidemiol 2009 in press ).

There are no clinical studies available on the benefit of administering Vitamin D on top of established therapies for multiple sclerosis. The present invention provides evidence that daily administration of 14 '000 IU Vitamin D in conjunction with standard treatment with Interferon-beta results in particularly good clinical efficacy in multiple sclerosis treatment.

This new add-on therapy is particularly beneficial to subjects with a low Vitamin D status.

Vitamin D status is defined as serum levels of 25 -hydroxy vitamin D (25(OH)D). A low endogenous Vitamin D status according to this invention is defined as a 25 -hydroxy- vitamin D plasma level before treatment below 100 nmol/L.

In individuals, vitamin D status is determined not only by the dietary intake of vitamin D, and exposure of the skin to sun, but also by genetic constitution. In a twin-study, Orton et al. showed that a genetic factor contributes to the 25(OH)D levels which were measured in the sera of their participants (Orton et al. Am J Clin Nutr 2008;88:441-7). A heritability estimate of 0.77 was calculated. As drivers of this effect, polymorphisms within several vitamin D metabolism-related genes have been assessed, including S Ps in the vitamin D receptor (VDR), vitamin D binding protein (VDBP), CYP27B1, and CYP24A1.

Subjects which are carriers of a variant of the CYP24A1 gene may have low Vitamin D status and thus the methods according to this invention would be particularly beneficial to them (Fu et al, Clin. Biochem. 2009 Jul : 42 (10-11): 1174-7. Epub 2009 May 18).

A genetic association study conducted by the applicant, investigating the presence of variants in the CYP24A1 gene showed a statistically significant correlation with absence of relapses in MS patients treated with Interferon-beta during two years (unpublished data).

Table 1 : List of CYP24A1 genetic variants found associated with the absence of relapses.

CYP24A1 Genetic Variant Parameter p-value

rs2762934 Relapse-free 0.0077

rs2296241 Relapse-free 0.00713

rs2762942 Relapse-free 0.04082

Figure 1 shows the distribution of genotype of SNP rs 2762934 in gene CYP24A1 and of the absence of relapses. The total number of subj ects is indicated at the bottom of each patch. The number of responders is indicated at the top of each patch. The dotted line represents the mean response for the total population. P-value is based on Fisher' s test. The Star indicates a p- value<=0.05. The double star indicates a p-value <=0.01.

Figure 2 shows the distribution of genotype of SNP rs2296241 in gene CYP24A1 and of the absence of relapses.

The total number of subjects is indicated at the bottom of each patch. The number of responders is indicated at the top of each patch. The dotted line represents the mean response for the total population. P-value is based on Fisher' s test. The Star indicates a p-value<=0.05. The double star indicates a p-value <=0.01.

Figure 3 shows the distribution of genotype of SNP rs2762942 in gene CYP24A1 and of the absence of relapses.

Total number of subjects is indicated at the bottom of each patch. The number of responders is indicated at the top of each patch. The dotted line represents the mean response for the total population. P-value is based on Fisher's test. The Star indicates a p-value<=0.05.

The observed proportion of patients without relapses during the 2-years treatment is

For rs2762934:

AA: 0%

AG: 36%

GG:74%

For rs2296241 :

AA: 20%

AG: 73%

GG: 71%

For rs2762942:

AA: 51%

AG: 80%

GG: 100% For each of these markers, the proportion of subjects responding to treatment was higher for carriers of the G allele. Based on the observed association, it was concluded that genetic variants involved in vitamin D metabolism could affect the disease course. Therefore supplementing current treatments by adapting vitamin D concentrations based on the genetic background of the patients could enhance the effectiveness of the treatments. For example, carriers of AA genotype might receive a high dose, carriers of AG a lower dose, and carriers of GG an even smaller dose.

Definitions

"A week" refers to a period of time of about 7 days.

The term "treat" or "treating" as used herein is meant to ameliorate, alleviate symptoms, eliminate the causation of the symptoms either on a temporary or permanent basis, or to prevent or slow the appearance of symptoms of the named disorder or condition. The term "treatment" as used herein also encompasses the term "prevention of the disorder", which is, e.g., manifested by delaying the onset of the symptoms of the disorder to a medically significant extent. Treatment of the disorder is, e.g., manifested by a decrease in the symptoms associated with the disorder or an amelioration of the reoccurrence of the symptoms of the disorder.

"Relapses" involve neurologic problems that occur over a relatively short period, which lasts however at least 24 hours.

All of the following criteria must be met for a clinical event to qualify as an MS attack (relapse):

1. Neurological abnormality, either newly appearing or re-appearing, with abnormality specified by both

► Neurological abnormality separated by at least 30 days from onset of a preceding MS attack,

and

► Neurological abnormality lasting for at least 24 hours.

2. Absence of fever or known infection (fever with temperature [axillary, oral, or intrauricular] > 37.5°C/99.5°F).

3. Objective neurological impairment, correlating with the subject's reported symptoms, defined as either

► Increase in at least one of the functional systems of the EDSS, or

► Increase of the total EDSS score.

Sometimes the attack onset occurs over several weeks. A typical MS relapse involves a period of worsening, with development of neurological deficits, then a plateau, in which the patient is not getting any better but also not getting any worse followed by a recovery period. Recovery usually begins within a few weeks.

"Efficacy" of a treatment according to the invention can be measured based on changes in the course of disease in response to a use according to the invention. For example, treatment of MS efficacy can be measured by the frequency of relapses in RRMS and the presence or absence of new lesions in the CNS as detected using methods such as MRI technique {Miller et al, 1996, Neurology, 47(Suppl 4): S217; Evans et al, 1997, Ann. Neurology, 41: 125-132).

The observation of the reduction and/or suppression of MRI Ti gadolinium-enhanced lesions (thought to represent areas of active inflammation) gives a primary efficacy variable.

Secondary efficacy variables include MRI Ti enhanced brain lesion volume, MRI Ti enhanced lesion number, MRI T₂ lesion volume (thought to represent total disease burden, i.e. demyelination, gliosis, inflammation and axon loss), MRI Ti enhanced hypointense lesion volume (thought to represent primarily demyelination and axon loss), time-to-progression of MS, frequency and severity of exacerbations and time-to-exacerbation, Expanded Disability Status Scale score and Scripps Neurologic Rating Scale (SNRS) score (Sipe et al, 1984, Neurology, 34, 1368-1372). Methods of early and accurate diagnosis of multiple sclerosis and of following the disease progression are described mMattson, 2002, Expert Rev. Neurotherapeutics, 319-328.

Degree of disability of MS patients can be for example measured by Kurtzke Expanded Disability Status Scale (EDSS) score (Kurtzke, 1983, Neurology, 33, 1444-1452). Typically a decrease in EDSS score corresponds to an improvement in the disease and conversely, an increase in EDSS score corresponds to a worsening of the disease.

Compositions

Compositions of this invention may further comprise one or more pharmaceutically acceptable additional ingredient(s). Compositions of this invention may be liquid formulations including, but not limited to, aqueous or oily suspensions, solutions, emulsions, syrups, and elixirs. The compositions may also be formulated as a dry product for constitution with water or other suitable vehicle before use.

Patients

Patients according to the invention are patients suffering from multiple sclerosis, preferably RRMS or early SPMS with superimposed relapses.

In an embodiment of the invention, patients are selected from human males or females between 18 and 65 years age.

In another embodiment of the invention, patients had at least one relapse within the prior 12 months of the treatment.

Examples

The efficacy and safety of combined use of Interferon-beta and Vitamin D according to the invention can be assessed for example following the protocol below:

Example 1: Vigantol® oil as add-on to Rebif® in the treatment of RRMS

A three-arm, randomized, double-blinded, placebo-controlled multicenter phase II trial to evaluate the efficacy (by means of anti-inflammatory effects assessed by MS attacks and MRI parameters), safety, and tolerability of Vigantol® oil as add-on to Rebif® 22 or 44 μg three times a week (tiw) in subjects with RRMS according to the revised McDonald criteria is undertaken.

A maximum of 358 eligible subjects are enrolled in this trial: 348 subjects with 25-hydroxy-vitamin D plasma levels below 150 nmol/L and around 10 subjects with 25-hydroxy-vitamin D plasma levels equal or higher than 150 nmol/L. Patients are selected from Male or Female, between 18 and 65 years of age with a history of one recent MS attack or an MRI-based proven disease activity within the last 12 months before screening. Female subjects must be neither pregnant nor breastfeeding and must lack child-bearing potential, as defined by either being post-menopausal or surgically sterile or using a highly effective method of contraception for the entire duration of the trial.

The subjects with 25 -hydroxy- vitamin D plasma levels below 150 nmol/L are randomized to Treatment Group 1 or 2. The subjects with 25 -hydroxy- vitamin D plasma levels equal or higher than 150 nmol/L are automatically assigned to Treatment Group 3.

• Treatment Group 1: Vigantol oil® 14.000 IU/d (350 μg/d) as add-on to Rebif® 22 or 44 μg three times a week in subjects with 25-hydroxy -vitamin D plasma levels below 150 nmol/L (174 subjects)

• Treatment Group 2: Matching placebo daily as add-on to Rebif® 22 or 44 μg three times a week in subjects with 25-hydroxy-vitamin D plasma levels below 150 nmol/L (174 subjects)

• Treatment Group 3: Matching placebo daily as add-on to Rebif® 22 or 44 μg three times a week in subjects with 25-hydroxy-vitamin D plasma levels equal or higher than 150 nmol/L (around 10 subjects)

Assignment to all three Treatment Groups is done by means of interactive voice-response system (IVRS).

Randomization is done only for subjects with 25-hydroxy-vitamin D plasma levels lower than 150 nmol/L by means of IVRS. These subjects are randomly assigned to Vigantol oil® (Treatment Group 1) and placebo treatment (Treatment Group 2) in a 1 : 1 ratio on top of the preexisting treatment with Rebif®. Subjects with 25-hydroxy-vitamin D plasma levels equal or higher than 150 nmol/L are assigned automatically to Treatment Group 3. All three treatment groups are double- blinded for all assessments.

Recruitment is concluded once the requested number of subjects for the Treatment Groups 1 and 2 are included in the study.

There is one Screening (within 14 days prior to Study Day 1 visit [Baseline]) and a Study Day 1 visit, at which time subjects are randomized if the subject is fulfilling all eligibility criteria.

At the Study Day 1 visit, there is a pre-dosing MRI, after which study treatments are initiated. During the Treatment period there are 10 subsequent visits at week 4 and every 12 weeks (i.e. every 84 days) thereafter, up to Week 96. A final follow-up visit occurs at Week 100 (4 weeks after the last dose of trial treatment).

All subjects start the treatment period with a 4 weeks lasting titration period, in which for the first 4 weeks subjects receive 7.000 IU/day (175 μg/d; 1 capsule) of Vigantol® oil or matching placebo, after which they receive full-dose Vigantol® oil (14.000 IU/d [350 μg/d], 2 capsules) or matching placebo for the remaining 92 weeks of the treatment period.

All subjects continue to be treated with Rebif® 22 or 44 μg three times a week which was initiated for at least 60 days before Study Day 1, and continue for the whole duration of this study. Doses of Rebif® are adjusted at the discretion of the treatment physician for the patient's best benefit.

At Study Day 1, pre-dosing baseline assessments are performed for safety (blood sample), MRI and Neurological evaluation. In addition one single blood sample is taken for exploratory PGx analysis at Study Day 1. During the treatment period MRI scans are performed after 48 and 96 weeks from Study Day 1. Blood samples are collected for routine analysis at Weeks 4 and 12, and every 12 weeks thereafter. Neurological assessments are performed at Study Day 1 and every 12 weeks. For this trial double-blinding is ensured for the whole duration of the 96 weeks of the treatment period for all subjects' assessments of all three treatment groups. Double blinding is ensured by means of the Two-Physician concept for the clinical assessments and by means of centralized lab procedures for MRI, PGx, heamatology and blood chemistry analysis. The Two-Physician concept consists of a "Treating Physician" and an "Evaluating Physician". The treating Physician is responsible for all aspects of treatment and clinical management of the subject, including the management of clinical attacks. He has the same blinding as the subject. The Evaluating Physician performs all the standardized neurological examinations needed. He is completely blinded to all aspects related to the subj ect' s treatment, including the assessment of side effects and drug administration problems.

The primary endpoint is a composite endpoint, based on predefined MRI and clinical efficacy outcomes:

1. The primary MRI endpoint is the change from baseline in the mean number of T2-lesions at Week 48;

2. The primary clinical endpoint is the proportion of relapse-free patients at Week 96. Patients in Group 1 have a statistically significant decrease in total volume of T2 brain lesions and relapse rate with respect to Group 2.

The data show that a treatment regimen comprising the combination of Rebif® and Vigantol ® oil is efficient in decreasing brain lesions and the annualized relapse rate, and no severe adverse effect is observed in Group 1 with respect to Group 2. From the literature available, this treatment regime might potentially not only be particularly adapted for patients having low pre-treatment plasma levels of 25-hydroxy-vitamin D, but to all subjects affected by MS.

References:

1. Lublin and Reingold, Neurology, 1996, 46:907-911

2. Lassmann et al, 2001, Trends Mol. Med., 7, 115-121

3. Lucchinetti et al, Curr. Opin. Neurol, 2001, 14, 259-269

4. Polman CH, Reingold SC, Edan G, Filippi M, Hartung HP, Kappos L, Lublin FD, Metz LM, McFarland HF, O'Connor PW, Sandberg-Wollheim M, Thompson AJ, Weinshenker BG, Wolinsky JS. Diagnostic criteria for multiple sclerosis: 2005 revisions to the "McDonald Criteria". Ann Neurol. 2005 Dec; 58(6): 840ff

5. EP 626853B1

6. US 5,506,214

7. JG Phadke et al, J. Epidemiol Community Health 1987; 41 :5-13

8. Panitch H. et al, Journal of the Neurological Sciences 2005; 239(l):67-74

9. S. Steddon et al, Nephrol Dial Transplant, 2001, 16: 1965-1967

10. Barthel et al. Dtsch Med Wochenschr 2003, 128:440-46

11. Bikle DD, Current Opinion in Rheumatology 2007, 19, 4:383-8

12. Zittermann A., British Journal of Nutrition 2003, 89:552-572

13. Cantorna et al. J Immunol 1998, 160 :5314-5319

14. Kimball et al., Am J Clin Nutr 2007, 86:645-651

15. Takeuchi et al, J Immunol 1998, 160:209-218

16. Burton et al, Multiple Sclerosis 2008, 14 (suppl. 1):34

17. Hiremath et al, Multiple Sclerosis 2009, 15:735-740

18. Munger et al. (Neurology 2004, 62:60-65)

19. Hathcock et al. Am J Clin Nutr 2007, 85:6-18

20. Vieth R., Am J Clin Nutr 1999, 69:842-856

21. Vieth R., Ann Epidemiol 2009 in press

22. Orton et al. Am J Clin Nutr 2008;88:441-7

23. Fu et al., Clin. Biochem. 2009 Jul. :42 (10-1 1): 1174-7. Epub 2009 May 18

24. Miller et al, 1996, Neurology, 47(Suppl 4): S217

25. Evans et al, 1997, Ann. Neurology, 41 : 125-132

26. Sipe et al., 1984, Neurology, 34, 1368-1372 27. Mattson, 2002, Expert Rev. Neurotherapeutics, 319-328

28. Kurtzke, 1983, Neurology, 33, 1444-1452

Claims

Claims:

1. Vitamin D or an analogue thereof in combination with Interferon-beta for the treatment of multiple sclerosis, preferably relapsing remitting multiple sclerosis (RRMS).

2. A method of treating a subject suffering from multiple sclerosis wherein a composition comprising Interferon-β and a composition comprising Vitamin D or an analogue thereof are administered.

3. A method according to claim 2 wherein Vitamin D or an analogue thereof is administered during a titration period in a dose of 7Ό00 IU/day and thereafter in a dose of 14Ό00 IU/day.

4. A method according to claim 2 or 3 wherein Interferon-beta is Interferon- betala, preferably Rebif®.

5. A method according to any of claims 2 to 4 wherein Interferon is administered in a dose of 22 or 44 μg three times a week.

6. A method according to any of claims 2 to 5 wherein the titration period lasts about 4 weeks.

7. A method according to any of claims 2 to 6 wherein Vitamin D or an analogue thereof is administered orally.

8. A method according to any of claims 2 to 7 wherein Interferon is administered intramuscularly or subcutaneously.

9. A method according to any of claims 2 to 8 wherein the subject has a 25-hydroxy-vitamin D plasma level before treatment below 150 nmol/L.

10. A method according to any of claims 2 to 9 wherein the subject is carrier of any genetic variant from genes involved in the Vitamin D pathway.