KR20150036398A - Optimum dose regime of an anti-nogo-a antibody in the treatment of amyotrophic lateral sclerosis - Google Patents

Abstract

The present invention relates to a method for the treatment or prevention of neurological diseases, particularly, but not limited to, muscular atrophy sclerosis (ALS), including administration of an anti-Nogo-A antibody.

Description

TECHNICAL FIELD The present invention relates to an anti-NOGOA antibody, and more particularly, to an anti-NOGOA antibody against an anti-NOGOA antibody in the treatment of muscular atrophy sclerosis.

The present invention relates to a method for the treatment or prevention of neurological diseases, in particular, but not limited to, amyotrophic lateral sclerosis (ALS), which comprises administration of an anti-Nogo-A antibody.

Muscle Atrophy Sclerosis (ALS), also known as Lou Gehrig's disease or Maladie de Charcot, is the most common adult-onset motor neuron disease. The main disease feature is the gradual degeneration of gastric and lower motor neurons in surface waterways. Downward motor nerve cell dysfunction (in the brainstem and spinal cord) leads to systemic weakness, atrophy and paralysis. Respiratory muscle dysfunction is a fatal event that usually occurs within 1-5 years of onset.

ALS is the most common motor neuron disease of adults affecting approximately 30,000 people in the United States and 5,000 people in the United Kingdom each year (Leigh & Swash, 1991). Typical onset age is from 50 to 70 years, but sometimes also occurs at a younger age. Most pathologies (90-95%) are classified as sporadic ALS (sALS) and the remainder are inherited and referred to as familial ALS (fALS). The sporadic and familial forms are clinically and pathologically similar, suggesting a common mechanism of onset (Bruijn et al, 2004). However, the exact cause of most pathologies is not yet known, and there is no effective treatment to stop the progression of the disease.

Nogo, also known as Reticulon-4 or neurite proliferation inhibitor, is a protein encoded by the RTN4 gene identified as an inhibitor of neurite proliferation specific to the central nervous system in humans.

Three forms of human Nogo have been identified: Nogo-A has 1192 amino acid residues (GenBank accession number AJ251383); Nogo-B is a splice variant (GenBank Accession No. AJ251384) and a shorter splice variant lacking residues 186-1004 in the putative extracellular domain and Nogo-C also lacks residues 186-1004 and is also a smaller and alternative Amino terminal domain (GenBank Accession No. AJ251385) (Prinjha R et al (2000) Nature 403, 383-384). Inhibition of CNS inhibitory proteins, such as Nogo, can improve neuronal cell damage, promote neuronal cell repair and growth, and provide a therapeutic means of potentially helping recovery from neuronal damage such as persistence in stroke. Examples of such Nogo inhibitors may include small molecules, peptides and antibodies.

It has been reported that the murine monoclonal antibody IN-1 (and subsequently identified as a fragment of Nogo-A) generated against the myelin protein NI-220/250, a potent inhibitor of neurite outgrowth, promotes axonal regeneration Nature 378 498-501 and Thallmair, M (1990) Nature 343 269-272; Bregman, BS et al (1995) Nature 378 498-501 et al (1998) Nature Neuroscience 1 124-131). It has also been reported that Nogo-A is an antigen for IN-1 (Chen et al (2000) Nature 403: 434-439). Administration of IN-1 Fab fragments or humanized IN-1 improved the recovery of rats undergoing spinal cord transection (Fiedler, M et al (2002) Protein Eng 15 931-941; Brosamle, C et al. Neuroscience 20 8061-8068).

Monoclonal antibodies that bind to Nogo are described in WO 2004/052932, WO 2005/028508, WO 2005/061544 and WO 2007/068750. WO 2004/052932 describes a murine antibody 11C7 that binds to a particular form of human Nogo with high affinity. WO 2005/061544 also describes high affinity monoclonal antibodies comprising murine monoclonal antibody 2A10, and generally humanized variants thereof, such as H1 L11 (the sequence for H1 and L11 is shown in SEQ ID NO. 33 And 34 (VH or VL sequences only), respectively. The described antibodies bind to human Nogo-A with high affinity. WO 2007/068750 discloses a number of high affinity humanized monoclonal anti-Nogo antibodies comprising H28L16 (the sequences for H28 and L16 are provided in SEQ ID NOs: 49 and 14, respectively (VH or VL sequences only) . The antibodies described were found useful in the treatment of neurological disorders.

Despite the art of providing high affinity anti-Nogo antibodies for the treatment of neurological disorders, there remains a very desirable goal to optimize therapeutic regimens for such diseases.

Summary of the Invention

According to a first aspect of the present invention there is provided the use of an anti-Nogo-A antibody, or a functional fragment thereof, in an amount sufficient to achieve co-localization of human Nogo-A on the patient's cell membrane in excess of 90% A method of treating or preventing a neurological disorder in a patient, comprising administering to the patient a dose that maintains the neurological condition.

According to a second aspect of the present invention there is provided a method of treating an anti-Nogo-A antibody, or a functional fragment thereof, at a dose that achieves and maintains human Nogo-A on the cell membrane of a patient and a co-localization level of the antibody above 90% Nogo-A < / RTI > antibody for use in the treatment or prevention of a neurological disease in said patient.

According to a third aspect of the present invention there is provided a method of treating an anti-Nogo-A antibody, or a functional fragment thereof, at a dose that achieves and maintains a co-localization level of human Nogo-A on the cell membrane of a patient, There is provided a pharmaceutical composition comprising an anti-Nogo-A antibody, or a functional fragment thereof, for use in the treatment or prevention of a neurological disease in said patient.

Brief Description of Drawings

Figure 1: A laser scanning cell count (LSC) well scan was placed in each section to identify tissue sections (gray image) and four areas (cyan box). A high resolution field scan produced images of the positions of three dyes. Individual proteins were stained with blue (gamma sarcoglycan), red (Nogo A) and green (H28L16).

Figure 2: Gating of event groups (pre-dose biopsy). Based on the fluorescence intensity, a scatter plot was generated to isolate the pixels of the dye for the feature.

Figure 3: Gating of event groups (pre-dose biopsy). This figure is a field image showing the process of "back gating" used to display gated events for an organization section.

Figure 4: Gating of event group (biopsy after administration). As in Figure 2, however, a scatter plot was generated after administration of 15 mg / kg of H28L16.

Figure 5: Gating of event groups (biopsies after administration). As in FIG. 3, however, backgating was performed after administration of 15 mg / kg of H28L16.

Figure 6: NogoA / H28L16 expression (gamma sarcoglycan channels not shown). Individual fluorescence channels are shown. The top row shows only the target (Nogo A), the second row shows only H28L16, and the third row shows Nogo A and H28L16.

Figure 7: Percentage of Nogo A co-localized with H28L16 on the membrane. Percentage of Nogo A on H28L16 and co-localized muscle fiber membranes. The highlighted "15 mg / kg" box represents the largest amount of co-localization achieved after administration of 15 mg / kg of H28L16. Each bar represents one section.

Figure 8: Co-localized membrane Nogo-A median percentage with H28L16. Data from repeated doses of 2.5 mg / kg (left panel), 15 mg / kg of repeated doses (3 panels at the center) and 15 mg / kg of single doses (right panel). Median of triplicate sections derived from biopsies from a repeat-dose cohort, with a single section derived from a biopsy from a single dose cohort.

Figure 9: Model estimation relationship (Emax) between plasma H28L16 concentration and H28L16 and percentage of co-localized membrane Nogo-A.

Figure 10: Predicted median (5 ^th and 95 ^th percentile) H28L16 plasma concentration time profile for 15 mg / kg of H28L16 administered every 4 weeks.

Figure 11: Predicted median (5 ^th and 95 ^th percentile) H28L16 plasma concentration time profile for 15 mg / kg of H28L16 administered every two weeks.

Figure 12: Variable heavy and variable light chain regions of the H28L16 antibody.

DETAILED DESCRIPTION OF THE INVENTION

According to a first aspect of the present invention there is provided a method of treating or preventing an anti-Nogo-A antibody, or a functional fragment thereof, in a dose sufficient to achieve and maintain a co-localization level of human Nogo-A on the patient ' There is provided a method of treating or preventing a neurological disorder in a patient, comprising administering to the patient.

It will be understood that reference to "co-localization" herein refers to the rate at which the bound Nogo-A is localized or associated with or associated with the anti-Nogo-A antibody. For example, a co-localization value of 100% is equal to that of all membrane Nogo-A co-localized with anti-Nogo-A antibody. Those skilled in the art will appreciate that although co-localization is not equal to the target occupancy, it provides a very useful measure of perhaps the epidemiological effect to confirm optimal dosing regimens.

Data are provided herein that demonstrate that co-localization of human Nogo-A and anti-Nogo-A antibodies on cell membranes at levels greater than 90% achieved maximal pharmacodynamic effects.

The term "anti-Nogo-A antibody " as used herein refers to any antibody or variant form thereof, including, but not limited to, antibody fragments, domain antibodies or single chain antibodies capable of binding to Nogo-A. The anti-Nogo-A antibody may be a neutralizing anti-Nogo-A antibody. Anti-Nogo-A can be murine, chimeric, humanized, or fully human antibodies or fragments thereof. "Neutralization" and its grammatical variants refer to the inhibition of any Nogo function, more particularly any Nogo-A function, whether wholly or in part.

As used herein, reference to "treatment" refers to the treatment of muscular atrophy, including gradual degeneration of neurons in surface conduits, denervation of muscular fibers and / or muscle weakness and / Means reducing or eliminating the symptoms of the disease and / or its cause.

Reference to "prevention ", as used herein, refers to progressive degeneration of neurons in surface conduits, delamination of muscular fibers and / or muscle weakness and / or delayed stiffness, prevention or minimization of muscle stiffness, Preventing, or minimizing the symptoms of the disease associated therewith.

It will be appreciated that the amount and frequency of anti-Nogo-A antibody administered will be selected to ensure that co-localization with Nogo-A greater than 90% on the cell membrane is achieved and maintained.

WO 2010/004031 discloses that a suitable dose of an anti-Nogo-A antibody for the treatment of ALS or other neurological diseases in humans is in the range of 0.1 mg / kg to 300 mg / kg, generally about 2 mg / kg to about 40 mg / kg. < / RTI > Thus, in one embodiment, the dose of the anti-Nogo-A antibody is from 0.1 mg / kg to 300 mg / kg. In a further embodiment, the dose of the anti-Nogo-A antibody is from 2 mg / kg to 40 mg / kg.

WO 2007/068750 also discloses that a suitable dose of an anti-Nogo-A antibody for the treatment of stroke and other neurological disorders in humans is in the range of 700 mg to 3500 mg for a 70 kg body weight. Thus, in one embodiment, the dose of the anti-Nogo-A antibody is from 10 mg / kg to 50 mg / kg.

In one particular embodiment of the invention, the dose of the anti-Nogo-A antibody is 15 mg / kg. Here we present data showing surprisingly beneficial results for a dose of 15 mg / kg. In particular, these doses are adjusted to a particularly convenient dosage frequency to achieve a higher threshold than the present invention threshold of Nogo-A and 90% co-localization on the cell membrane.

Here we present data for a particularly advantageous dose of 15 mg / kg of the present invention, wherein the data show that co-localization with Nogo-A on the cell membrane has increased by more than 90% over a period of 8 days after administration, And falls to about 70% between 22 and 27 days. Thus, in one embodiment, when the dose is 15 mg / kg, the dosing interval suitable for maintaining more than 90% co-localization is 8 to 22 days. In a further embodiment, when the dose is 15 mg / kg, a dosage interval suitable for maintaining a co-localization of greater than 90% is 10 to 18 days.

In one particular embodiment of the invention, when the dose is 15 mg / kg, the administration interval suitable for maintaining more than 90% co-localization is 14 days. It will be appreciated that slight variations in the 14-day dosing interval may be tolerated and that administration may occur within 14 days +/- 3 days before and after the 14-day optimal dosing interval. Thus, in one embodiment, a dosage interval that is suitable to maintain more than 90% co-localization when the dose is 15 mg / kg is 11 to 17 days (i.e., ). In alternative embodiments, a dosage interval that is suitable to maintain more than 90% co-localization when the dose is 15 mg / kg is 12 to 16 days (i.e., two days before and after the 14 day optimal dose interval) . In alternative embodiments, a dosage interval that is suitable to maintain greater than 90% co-localization when the dose is 15 mg / kg is 13 to 15 days (i. E., Around 1 day before and / .

According to a further aspect of the present invention there is provided a method of treating or preventing a neurological disease in a patient comprising administering to the patient an anti-Nogo-A antibody at a dose of 15 mg / kg every 14 days (+/- 3 days) Method is provided.

Examples of anti-Nogo-A antibodies that may be used in accordance with the present invention are described in WO 2004/052932, WO 2005/028508, WO 2005/061544 and WO 2007/068750, Incorporated herein by reference.

In one embodiment, the anti-Nogo-A antibody is a monoclonal antibody.

In one embodiment, the anti-Nogo-A antibody is a humanized antibody. The term "humanized antibody" refers to a engineered antibody of the type having its CDRs derived from a non-human donor immunoglobulin, wherein the remaining immunoglobulin-derived portion of the molecule comprises one (or more) human immunoglobulin ). Also, the framework support residues can be modified to preserve binding affinity (see, for example, Queen et al., Proc. Natl Acad Sci USA, 86: 10029-10032 (1989), Hodgson et al., Bio / Technology , 9: 421 (1991)). Suitable human receptor antibodies may be selected from conventional databases, such as the KABAT® database, the Los Alamos database, and the Swiss Protein database, through identity with the nucleotide and amino acid sequences of the donor antibody (in this case murine donor antibody 2A10) . Human antibodies characterized by identity with the framework region of the donor antibody (on an amino acid basis) may be suitable to provide a heavy chain constant region and / or a heavy chain variable framework region for insertion of a donor CDR. Suitable receptor antibodies capable of donating light chain constant or variable framework regions can be selected in a similar manner. It should be noted that the heavy and light chains of the receptor antibody need not be derived from the same receptor antibody. Particularly EP-A-0239400 and EP-A-054951 describe several methods for producing such humanized antibodies.

The term "donor antibody" refers to a non-human antibody that provides a humanized antibody whose amino acid sequence of a variable region, CDR, or other functional fragment or analogue thereof contributes to a humanized antibody and that is characterized by antigen specificity and neutralizing activity of the donor antibody do.

The term "receptor antibody" refers to an antibody heterologous to a donor antibody that provides the humanized antibody with its heavy and / or light chain framework region and / or its heavy chain and / or light chain constant region. Receptor antibodies can be derived from any mammal, provided that they are non-immunogenic in humans. In one embodiment, the receptor antibody is a human antibody.

Alternatively, humanization can be accomplished through a "veneering " method. Statistical analysis of the immunoglobulin heavy and heavy chain variable regions of unique human and murine reveals that the precise patterns of exposed residues are different in human and murine antibodies and that most individual surface positions strongly select a small number of different residues (See Padlan EA et al. (1991) Mol. Lmmunol.28, 489-498 and Pedersen JT et al (1994) J. Mol. Biol. 235: 959-973). Thus, it is possible to reduce the immunogenicity of non-human Fv by substituting the exposed residues of the framework regions that are different from those generally identified in human antibodies. Substitution of surface residues may be sufficient to " invisible " the variable region of the mouse to the human immune system, since antigenicity of the protein may correlate with surface accessibility (see also Mark GE et al 1994) in Handbook of Experimental Pharmacology vol.113: The pharmacology of monoclonal Antibodies, Springer-Verlag, pp 105-134). This process of humanisation is referred to as "veneering " because only the surface of the antibody is modified so that the support residues remain unstoppable. A further alternative approach is described in WO 2004/006955.

"CDR" is defined as the amino acid sequence of the complementarity determining region of an antibody which is a hypervariable region of an immunoglobulin heavy chain and a light chain. See, for example, Kabat et al., Sequences of Proteins of Immunological Interest, 4th Ed., U.S. Pat. National Institutes of Health (1987)]. There are three heavy chains and three light chain CDRs (or CDR regions) in the variable region of the immunoglobulin. Thus, "CDR" as used herein refers to all three heavy chain CDRs, all three light chain CDRs (or, where appropriate, both heavy and light chain CDRs). Structure and protein folding of the antibody may mean that other residues are considered to be part of the antigen binding region and will be understood by those skilled in the art. For example, Chothia et al. (1989) Conformations of immunoglobulin hypervariable regions; Nature 342, p. 877-883.

In one embodiment, the anti-Nogo-A antibody is a neutralizing anti-Nogo-A antibody or fragment thereof such as murine antibodies 2A10, 15C3 and 2C4 (WO 2005/016544, the contents of which is incorporated herein by reference in its entirety) ).

In one embodiment, the anti-Nogo-A antibody is any of the antibodies disclosed in WO 2004/052932 (the entire contents of which are incorporated herein by reference). Examples of antibodies described in WO 2004/052932 include 11C7 and humanized variants thereof.

In one embodiment, the anti-Nogo-A antibody is any human anti-Nogo-A antibody described in WO 2005/028508 and WO 2009/056509, the contents of which are incorporated herein by reference. In a further embodiment, the anti-Nogo-A antibody is the human anti-Nogo-A antibody 6A3 described in WO 2009/056509.

In one embodiment, the anti-Nogo-A antibody comprises the anti-Nogo-A antibody described in WO 2007/068750. In a further embodiment, the anti-Nogo-A antibody is a humanized antibody, such as a humanized variant of 2A10, such as H20L16, H28L16, H28L13 and H27L16 (the disclosure of which is incorporated herein by reference in WO 2007/068750) ), Human antibodies, or fragments thereof.

In a further embodiment, the anti-Nogo-A antibody is H27L16 (SEQ ID NO: 48 and SEQ ID NO: 14 of WO 2007/068750), H28L13 (SEQ ID NO: 49 of WO 2007/068750 and SEQ ID NO: 13 ) Or H28L16 (SEQ ID NO: 49 and SEQ ID NO: 14 of WO 2007/068750). In yet further embodiments, the anti-Nogo-A antibody comprises H28L16 (SEQ ID NO: 49 and SEQ ID NO: 14 of WO 2007/068750, SEQ ID NO: 1 and SEQ ID NO: 2, respectively, herein) do.

In a further embodiment, the anti-Nogo-A antibody binds to H27FL L16FL (SEQ ID NO: 54 and SEQ ID NO: 18 of WO 2007/068750), H28FL L13FL (SEQ ID NO: 55 of WO 2007/068750 and SEQ ID NO: : 17) or H28FL L16FL (SEQ ID NO: 55 and SEQ ID NO: 18 of WO 2007/068750). In yet further embodiments, the anti-Nogo-A antibody comprises H28FL L16FL (SEQ ID NO: 55 of WO 2007/068750 and SEQ ID NO: 18).

In a further embodiment, the anti-Nogo-A antibody is selected from the group consisting of H28L16 (SEQ ID NO: 49 and SEQ ID NO: 14 of WO 2007/068750) or H28FL L16FL (SEQ ID NO: 55 of WO 2007/068750 and SEQ ID NO: 18). In still further embodiments, the anti-Nogo-A antibody comprises H28L16 (SEQ ID NO: 49 and SEQ ID NO: 14 of WO 2007/068750).

In a further embodiment, the anti-Nogo-A antibody or fragment thereof comprises one or more of 2A10, H28L16 or 6A3, optionally six CDRs. In a further embodiment, the anti-Nogo-A antibody or fragment thereof binds to the same human Nogo-A epitope as H28L16 (human Nogo-A 610-621aa comprising SEQ ID NO: 6 from WO 2010/004031) Is an antibody that competes with H28L16 for binding to Nogo-A.

Examples of neurological disorders that can be treated according to the administration regimen of the present invention include stroke (ischemia or hemorrhage), traumatic brain injury, spinal cord injury, Alzheimer ' s disease, Frontotemporal dementias (tauopathies), peripheral neuropathy, Parkinson ' s disease, Creutzfeldt-Jakob disease (CJD), schizophrenia Schizophrenia), muscular atrophy sclerosis (ALS), multiple sclerosis, Huntington's disease, multiple sclerosis, inclusion body myositis, polymyositis, dermatomyositis, morphology Morphologically nonspecific myopathies, congestive heart failure, and neuropathic pain.

In one embodiment, the neurological disorder is muscular atrophy sclerosis (ALS).

The mode of administration of the anti-Nogo-A antibody or fragment thereof of the present invention may be any suitable route for delivering the agent to the host. The antibodies and pharmaceutical compositions of the present invention are particularly useful for parenteral administration, i.e., subcutaneous (sc), intrathecal, intraperitoneal (ip), intramuscular (im), intravenous (iv), or intranasal . In one embodiment, the anti-Nogo-A antibody or fragment thereof is administered intravenously. In an alternative embodiment, the anti-Nogo-A antibody or fragment thereof is administered subcutaneously.

According to a second aspect of the present invention there is provided a method of treating an anti-Nogo-A antibody, or a functional fragment thereof, at a dose that achieves and maintains human Nogo-A on the cell membrane of a patient and a co-localization level of the antibody above 90% Nogo-A < / RTI > antibody for use in the treatment or prevention of a neurological disease in said patient.

According to a third aspect of the present invention there is provided a method of treating an anti-Nogo-A antibody, or a functional fragment thereof, at a dose that achieves and maintains a co-localization level of human Nogo-A on the cell membrane of a patient, There is provided a pharmaceutical composition comprising an anti-Nogo-A antibody, or a functional fragment thereof, for use in the treatment or prevention of a neurological disease in said patient.

The pharmaceutical composition of the present invention typically contains an effective amount of the antibody of the present invention as an active ingredient in a pharmaceutically acceptable carrier. In the prophylactic agonists of the present invention, aqueous suspensions or solutions containing the engineered antibodies, typically buffered at physiological pH in a form ready for injection, are preferred. Compositions for parenteral administration will generally comprise a solution of a pharmaceutically acceptable carrier, typically an antibody of the invention or its cocktail dissolved in an aqueous carrier. For example, various aqueous carriers such as 0.9% saline, 0.3% glycine and the like can be used. These solutions are sterile and generally free of particulate matter. Such solutions can be sterilized by well-known conventional sterilization techniques (e.g., filtration). The composition may contain pharmaceutically acceptable auxiliary substances necessary to approximate physiological conditions such as pH adjusting agents and buffers. In such pharmaceutical formulations, the concentration of the antibody of the invention can vary greatly, i.e., from less than about 0.5%, generally about 1%, or at least about 1% to as much as 15 or 20% Depending on the mode of administration, it will be selected primarily based on fluid volume, viscosity, and the like.

Thus, the pharmaceutical compositions for intramedullary injection of the present invention comprise 1 mL of sterile buffered water and from about 1 ng to about 100 mg, such as from about 50 ng to about 30 mg or more, such as from about 5 mg to about 25 mg of Can be prepared to contain the antibody of the present invention. Similarly, a pharmaceutical composition of the invention for intravenous infusion contains about 250 ml of a 0.9% saline (sodium chloride) solution, and about 1 to about 30 and typically 5 mg per 1 ml of 0.9% saline (sodium chloride) To about 25 mg of the engineered antibody of the present invention. Actual methods of preparing compositions that can be administered parenterally are well known or will be apparent to those skilled in the art and are described in more detail in, for example, Remington's Pharmaceutical Science, 15th ed., Mack Publishing Company, Easton, Pennsylvania have. For the preparation of intravenously administrable antibody formulations of the invention, see Lasmar U and Parkins D "The formulation of Biopharmaceutical products ", Pharma. Sci.Tech.today, page 129-137, Vol.3 (3rd April 2000), Wang, "Instability, stabilization and formulation of liquid protein pharmaceuticals ", Int. J. Pharm 185 (1999) 129-188, Stability of Protein Pharmaceuticals Part A and B ed Ahem T. J., Manning M. C, New York, NY: Plenum Press (1992), Akers.M.J. J Pharm Sci 92 (2003), " Effects of Types of Sugar on Stabilization of Protein in the Dried State ", J. Pharm Sci 91 (2002) 2283-2300, Imamura, K et al., &Quot; Excipient-Drug Interactions in Parenteral Formulations & 266-274, lzutsu, Kkojima, S. "Excipient crystalinity and its protein-structure-stabilizing effect during freeze-drying", J Pharm. Pharmacol., 54 (2002) 1033-1039, Johnson, R., "Mannitol-sucrose mixtures-versatile formulations for protein lyophilization", J. Pharm. Sci, 91 (2002) 914-922. Ha, E Wang W, Wang YJ. &Quot; Peroxide Formation in Polysorbate 80 and Protein Stability ", J. Pharm Sci., 91, 2252-2264 (2002), the entire contents of which are incorporated herein by reference and the reader concretely references this).

The antibodies described herein can be lyophilized for storage and reconstituted in a suitable carrier before use. This technique has been found to be effective with conventional immunoglobulins and the known-lyophilization and reconstitution techniques can be used.

In one embodiment, the anti-Nogo-A antibody is administered with an additional therapeutic agent. In a further embodiment, the additional therapeutic agent is an agonist that is effective against muscle atrophic lateral sclerosis (ALS). In yet a further embodiment, the agonist effective in muscle atrophic lateral sclerosis (ALS) is dexpramipexole. (R) -N ⁶ -propyl-4,5,6,7-tetrahydrobenzo [d] thiazole-2,6-diamine) was developed by Knopp Biosciences and Biogen It is a drug. Phase II clinical trials involving 102 patients showed a slowing of ALS disease progression and phase III studies are currently underway.

In one embodiment, the anti-Nogo-A antibody is administered with a compound having anti-glutamate activity. In certain embodiments, the compound having anti-glutamate activity is riluzole. In another embodiment, the compound having anti-glutamate activity is an antagonist of an AMPA receptor, such as a 2,3-benzodiazepine compound, especially talampanel. In another embodiment, the compound having anti-glutamate activity is TRO 19622 or ceftriaxone. The anti-Nogo-A antibody and the compound having anti-glutamate activity can be administered to a patient simultaneously, sequentially or separately. When the compound having anti-glutamate activity is riluzole, about 50 mg to about 150 or 200 mg of riluzole can be administered daily to the patient, and typically 100 mg of riluzole is administered daily to the patient. Riluzole is typically administered orally. When the compound having anti-glutamate activity is a Thalamphanel, Thalampanel is administered at about 10 mg to about 250 mg, typically 1 to 5 times a day, orally. In one embodiment, Thalamand paln is administered at a dose of 25 mg or 50 mg once to 5 times a day, optionally 3 times a day.

The present invention is illustrated with reference to the following studies:

Example  One: Nogo  A and H28L16 The co- Localization  analysis

Laser scanning cell counting (LSC) enables the measurement of expression at that location by generating an image of the dye position in the cell and tissue sections using a laser to stimulate the fluorescent dye. The dye is attached to an antibody that binds the targeted protein. A commercially available anti-Nogo A antibody attached to Alexa 647 dye was used to identify targets for H28L16. The use of commercially available Nogo A antibodies allows the measurement of targets in tissue sections. It is also desirable to know how many targets (Nogo A) are located in the muscle fiber membrane. This was accomplished by staining a section with antibodies against muscle fiber membrane protein anti-gamma sarcomoglycan adhering to Alexa 488 dye.

This muscle fiber membrane and co-makes it possible to determine and measure the amount of non-target localization (Nogo A) - and membrane localization and co. Next, it is necessary to quantitate co-localization with the amount of H28L16 and its target (Nogo A) in the administered muscle section. To provide this quantification of H28L16, the muscle sections were labeled using a non-neutralizing anti-individual specific antibody against H28L16 labeled with Alexa 555 dye. Once the three components are labeled with three different dyes, each feature can be separated and measured using LSC software.

A well scan was performed to check the tissue section (gray image) and four areas (cyan box) were placed in each section. A high resolution field scan produced images of the positions of three dyes.

Figure 1 shows an LSC area image with color assigned to each channel. For example, individual proteins were stained with blue (gamma sarcoglycan), red (Nogo A) and green (H28L16). The rectangle boxes summarized in white are the individual field images that make up the area image. Each region image contained 10 field im- ages.

A scatter plot was generated to separate the pixels of the dye for characterization based on fluorescence intensity. The pixel was a threshold using a process known as "gating " (the background noise was separated from the specific signal). Gating is indicated as colored lines on the scatterplots and gates are labeled R1, R2, R3, etc. according to the order in which the gates were created (Fig. 2). The scatter diagram of Figure 2 was generated by plotting two channels with respect to each other. By plotting "green max pixel" for "green intensity", the membrane and non-membrane portions of the tissue were separated. The brightest pixel in R1 is on the membrane labeled with gamma sarcohoglycan antibody. The darker pixels at R5 are unlabeled fibers. The long red intensity value was plotted against the membrane value (green max pixel) to obtain the amount of Nogo A co-localized with membrane R2. To obtain the amount of drug on the film, the green max pixel was plotted against the yellow intensity. This is a pre-dose biopsy, so if present, there is no event at R3 containing the drug on the membrane. The final plot represents the drug to be co-localized with Nogo A on the membrane.

A group of pixels within a gated population may be displayed on the image as a color box. This process is known as "back gating. &Quot; This is a quality control operation to demonstrate the most accurate position of the gate (Figure 3). The group gated in the scatter diagram of Fig. 2 is shown as an image in Fig. It should be noted from FIG. 3 that this data is from patients receiving vehicle only, with minimal background staining of anti-individual specific antibodies against H28L16 and little evidence of Nogo-A and co-localization Red box).

4 and 5, which show data from patients receiving 15 mg / kg of H28L16, backgating was performed using a green box showing the location of H28L16 that was not co-localized with Nogo-A, and a co- it should be noted as showing a red box shows the localization. Thus, Figures 4 and 5 illustrate the positive drug event in R3 and the co-localization of the drug and Nogo A.

Figure 6 shows a separate fluorescence channel. Administration - The pre-biopsy is red and there is no orange / yellow fluorescence compared to post-biopsy showing co-localization of H28L16 and Nogo A and an excess of H28L16 that is not co-localized with Nogo A. High co-localization occurred along the fiber membrane.

Figure 7 is a graph showing the percentage of Nogo A on co-localized muscle fiber membranes with H28L16. Three sections were cut and evaluated for each patient's biopsy (before and after the drug or placebo was administered). Some low levels of co-localization appeared in biopsies from untreated subjects (administration-transfer or placebo administration) showing nonspecific background staining seen in anti-individual specific antibodies co-localized with Nogo-A. Higher levels of anti-individual specific staining and co-localization were consistent in the post-drug administration samples, and co-localization showed a dose-dependent increase. The highlighted section shows the highest level of co-localization of H28L16 and Nogo A on the muscle membrane, which appeared in biopsies collected from the subject 8 days after administration of 15 mg / kg of H28L16.

Example  2: Muscular atrophy sclerosis  ( ALS ) For H28L16 Lt; RTI ID = 0.0 > co- Localization  Use of experiments

Capacity-selectivity is determined using co-localization of Nogo-A and H28L16 at the target site on the membrane of skeletal muscle cells as described in Example 1, without direct measurement of the pharmacodynamic response to H28L16 in muscle. For example, in a "First Time in Human" (FTiH) study of muscle biopsies to estimate the biodistribution of H28L16 to muscle level and skeletal muscle of Nogo-A, Lt; / RTI > A frozen biopsy (mainly 0.5 mg / kg, 2.5 mg / kg and 15 mg / kg cohort) of sufficient quality to support immunohistochemical analysis was performed using an antibody against Nogo-A, an anti-individual specific antibody against H28L16 And stained with anti-gamma sarcomoglycan (to define the muscle membrane). Expression was visualized using a secondary antibody labeled with fluorescence and quantified using laser scanning cell count (LSC). Using this approach, expression of Nogo-A on the muscle membrane could be assessed and differentiated from intracellular Nogo-A expression by co-localization with gamma sarcoglycans. An estimate of the level of H28L16 in the muscle was also measured and an estimate of the degree of co-localization of Nogo-A and H28L16 on the muscle membrane was derived.

A dose-dependent increase in drug staining was observed when the H28L16 level was assessed in the muscle using anti-individual specific staining. No staining was detectable at 0.5 mg / kg, and the highest staining in the biopsy was 15 mg / kg < / RTI > on the 22-27 day after administration (consistent with immunoassay measurements at the drug level in muscle). By varying the time of biopsy collection at the 15 mg / kg repeat-dose cohort (cohort 8), the time course of the drug biological distribution to the muscle after administration can be assessed. These data showed a low level of initial H28L16 in muscle on day 1 and slightly decreased again on day 23-24 after increasing to day 8.

Additional measurements of membrane Nogo-A levels in these biopsies allow the assessment of co-localization of Nogo-A and H28L16 at the target site after administration. Figure 8 shows the median percentage of H28L16 and co-localized Nogo-A in myocardial membranes in post-treatment biopsies taken from the 2.5 mg / kg and 15 mg / kg cohorts, where 100% It would be the same as co-localized with H28L16).

Higher levels were observed in biopsies from subjects treated with 15 mg / kg than in biopsies from the 2.5 mg / kg cohort, with measured levels of H28L16 closely following muscle co-localization. At a 15 mg / kg repeated dose cohort, co-localization was again associated with H28L16 levels, and at approximately 50% central co-localization on day 1, co-localized Nogo-A with the study drug on the muscle membrane after 8 days of administration Increased to> 90%, and then decreased again to about 70% at 22-27 days.

Although co-localization is not equal to the target share,> 90% co-localization of muscle fibers Nogo-A and H28L16 is considered to be required throughout the entire administration interval to achieve maximal pharmacodynamic effect. Based on the LSC data, a dose level of 15 mg / kg of H28L16 is required to achieve > 90% co-localization of the membranes Nogo-A and H28L16 8 days after administration, but this co- It is not maintained. An exploratory PK-PD modeling for investigating the potential relationship between the estimated plasma H28L16 concentrations (using the population PK model) at the level of co-localization biopsy is shown in FIG. The E _max model was applied to these data to estimate an EC ₅₀ of about 18 μg / mL and an EC ₉₀ of about 160 μg / mL.

Based on the population PK modeling and subsequent simulation to investigate the dosage regimen that maintains plasma H28L16 concentrations in excess of the EC _90, the dosage regimen used in the FTiH study (4 weeks every single 15 mg / kg), the average , It is clear that predictions will not be sufficient to maintain plasma concentrations above 160 μg / mL during the dosing interval. However, increasing the dosing frequency to 15 mg / kg once every two weeks was expected to, on average, maintain a concentration of greater than 160 μg / mL during the entire dose interval. Predicted plasma H28L16 concentration time profiles for 15 mg / kg administered every 2 and 4 weeks are presented below in Figures 10 and 11, respectively. Predicted plasma exposure (C _max and AUC _{(0- tau )} ) of H28L16 in relation to the dosing regimen of 15 mg / kg every two weeks as proposed is presented in Table 1.

Table 1: 15 administered every 2 weeks mg / kg of H28L16 The predicted center (5 ^th  And 95 ^th  Percentile) H28L16  plasma Cmax  And AUC (O- tau )

Predicted human systemic exposures at steady state after repeated administration of 15 mg / kg once every two weeks are approximately twice that after a single 15 mg / kg dose.

Experience from the FTiH study in ALS patients showed that H28L16 at a dose of 15 mg / kg once every four weeks was well tolerated. In the long-term 12-month toxicity study in Cynomolgus monkeys administered intravenously at a maximum of 500 mg / kg every 14 days, no significant toxicity was observed. The monkey-to-human exposure limit for H28L16 compared to the predicted human plasma exposure in the steady state of the proposed two phase study is based on (i) the dose ratio based on the dose of 15 mg / kg administered every two weeks 33 times, (ii) 10 times based on the steady-state AUC ratio, and (iii) 27 times based on the steady-state _Cmax ratio.

Safety and systemic exposure limits from non-clinical studies; Clinical safety from FTiH studies, PK-co-localization relationship data; And population PK modeling, the optimal dose for H28L16 appears to be 15 mg / kg once every two weeks. These doses are expected to exceed the levels of exposure in clinical studies achieved so far, but are well supported by non-clinical safety data and maximize the opportunity for delivery of efficacy in ALS patients.

                                SEQUENCE LISTING <110> Glaxo Group Limited       Bulllman, Jonathan       Krull, David <120> NOVEL TREATMENT    <130> AGS / PB65206 WO <140> US 61/668134 <141> 2012-07-05 <160> 2 <170> FastSEQ for Windows Version 4.0 <210> 1 <211> 113 <212> PRT <213> Artificial Sequence <220> Polypeptide sequence <400> 1 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala  1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr             20 25 30 Trp Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile         35 40 45 Gly Asn Ile Asn Pro Ser Asn Gly Gly Thr Asn Tyr Asn Glu Lys Phe     50 55 60 Lys Ser Lys Ala Thr Met Thr Arg Asp Thr Ser Thr Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys                 85 90 95 Glu Leu Met Gln Gly Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser             100 105 110 Ser      <210> 2 <211> 112 <212> PRT <213> Artificial Sequence <220> Polypeptide sequence <400> 2 Asp Ile Val Met Thr Gln Ser Pro Leu Ser Asn Pro Val Thr Leu Gly  1 5 10 15 Gln Pro Val Ser Ile Ser Cys Arg Ser Ser Ser Ser Leu Leu Tyr Lys             20 25 30 Asp Gly Lys Thr Tyr Leu Asn Trp Phe Gln Gln Arg Pro Gly Gln Ser         35 40 45 Pro Gln Leu Leu Ile Tyr Leu Met Ser Thr Arg Ala Ser Gly Val Pro     50 55 60 Asp Arg Phe Ser Gly Gly Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Gln Gln Leu                 85 90 95 Val Glu Tyr Pro Leu Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys             100 105 110

Claims (20)

Comprising administering to said patient an anti-Nogo-A antibody, or a functional fragment thereof, at a dose that achieves and sustains a co-localization level of human Nogo-A on the patient &apos; s cell membrane in excess of 90% A method for treating or preventing a neurological disorder in a patient. The method according to claim 1, wherein the dose of the anti-Nogo-A antibody is 0.1 mg / kg to 300 mg / kg. 3. The method of claim 2, wherein the dose of the anti-Nogo-A antibody is 15 mg / kg. 4. The method of claim 3, wherein greater than 90% of the co-localization is maintained for 8 to 22 days. 5. The method of claim 4, wherein greater than 90% co-localization is maintained for 10 to 18 days. 6. The method of claim 5, wherein greater than 90% co-localization is maintained for 11 to 17 days. 7. The method of claim 6, wherein greater than 90% co-localization is maintained for 12-16 days. 7. The method of claim 6, wherein said greater than 90% co-localization is maintained for between 13 and 15 days. 9. The method of claim 8, wherein the co-localization of greater than 90% is maintained for 14 days. A method of treating or preventing a neurological disease in a patient, the method comprising administering to the patient an anti-Nogo-A antibody, or a functional fragment thereof, at a dose of 15 mg / kg every 14 days +/- 3 days. 11. The method according to any one of claims 1 to 10, wherein the anti-Nogo-A antibody is a monoclonal antibody. 12. The method according to any one of claims 1 to 11, wherein the anti-Nogo-A antibody is a humanized antibody. 13. The method according to any one of claims 1 to 12, wherein the anti-Nogo-A antibody comprises H28L16. 14. A method according to any one of claims 1 to 13, wherein the neurological disorder is selected from the group consisting of stroke (ischemia or hemorrhage), traumatic brain injury, spinal cord injury, Alzheimer &apos; s disease, frontotemporal dementia (tauopathy), peripheral neuropathy, (CJD), schizophrenia, muscular atrophy (ALS), multiple sclerosis, Huntington's disease, multiple sclerosis, inclusive myositis, multiple myelitis, dermatomyositis, morphologic nonspecific myopathy, congestive heart failure and neuropathic &Lt; / RTI &gt; 15. The method according to claim 14, wherein the neurological disorder is muscle atrophic sclerosis (ALS). 16. The method according to any one of claims 1 to 15, further comprising administering with an additional therapeutic agent. 17. The method of claim 16, wherein the additional therapeutic agent is an agonist effective against muscle atrophy cirrhosis (ALS). 18. The method of claim 17, wherein the agent that is effective for muscle atrophy sclerosis (ALS) is dexpramipexole. 17. The method of claim 16, wherein the additional therapeutic agent is a compound having an anti-glutamate activity. 20. The method of claim 19, wherein the compound having anti-glutamate activity is rilsuzole.

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