Humanized antibody

• US 20100080800A1
• Filed: 07/23/2009
• Published: 04/01/2010
• Est. Priority Date: 07/14/2006
• Status: Active Grant

• Current Assignee: AC Immune SA, Genentech, Inc. (Roche Holding AG)
• Original Assignee: AC Immune SA, Genentech, Inc. (Roche Holding AG)
• Inventors: Pihlgren, Maria, Muhs, Andreas, Watts, Ryan, Pfeifer, Andrea
• Granted Patent: US 8,246,954 B2
• Time in Patent Office: Days
• US Class Current: 424/133.100
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/16 Anti-Parkinson drugs
• CPC Class Codes: A61P 25/28 for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• CPC Class Codes: C07K 16/18 against material from animals or humans
• CPC Class Codes: C07K 16/28 against receptors, cell surface antigens or cell surface determinants
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/34 Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues
• CPC Class Codes: C07K 2317/56 variable (Fv) region, i.e. VH and/or VL
• CPC Class Codes: C07K 2317/565 Complementarity determining region [CDR]
• CPC Class Codes: G01N 33/6896 Neurological disorders, e.g. Alzheimer's disease

Abstract

The present invention is related to chimeric and humanized antibody and to methods and compositions for the therapeutic and diagnostic use in the treatment of amyloidosis, a group of disorders and abnormalities associated with amyloid protein such as Alzheimer'"'"'s disease.

Claims

1. A chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof, which specifically binds to at least one epitope on the β

-amyloid protein wherein the epitope comprises at least two consecutive amino acid residues predominantly involved in binding to the antibody, wherein the at least two consecutive amino, acid residues are -Lys-Leu- embedded within the following core sequence (SEQ ID NO;

     10);

3. A chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof according to claim 1, which specifically binds to at least two epitopes on the β

-amyloid protein wherein said at least two epitopes each comprise at least two consecutive amino acid residues predominantly involved in the binding of the antibody, wherein the at least two consecutive amino acids are separated by at least one amino acid residue not involved in antibody binding or to a significantly smaller extent than said consecutive amino acid residues, which are -Phe-Phe- and -Lys-Leu-, respectively, embedded within the following core sequence;

4. A chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof according to claim 1, wherein

Xaa_l is an amino acid selected from the group consisting of His and Arg;

Xaa₂ is an amino acid selected from the group consisting of Asn and Gln;

Xaa₃ is an amino acid selected from the group consisting of Val and Leu,

Xaa₄ is an amino acid selected from the group consisting of Ala and Val;

Xaa₅ is an amino acid selected from the group consisting of Glu and Asp, and

Xaa₆ is an amino acid selected from the group consisting of Glu and Asp.

5. A chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof according to claim 4, wherein

Xaa_l is His

Xaa₂ is Gln;

Xaa₃ is Val;

Xaa₄ is Ala;

Xaa₅ is Glu; and

Xaa₆ is Asp.

6. A chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof according to claim 1, which specifically binds to at least two epitopes on the 13-amyloid protein wherein said at least two epitopes each comprise at least two consecutive amino acid residues predominantly involved in the binding of the antibody, which are -Phe-Phe- and -Lys-Leu-, respectively, and wherein said at least two distinct binding sites exhibit amino acid sequence -Val-Phe-Phe-Ala-Glu-Asp- shown in SEQ ID NO:

7 and amino acid sequence His-Gln-Lys-Leu-Val- shown in SEQ ID NO;

8, respectively.

8. A humanized antibody or a fragment thereof according to claim 1, wherein a CDR of the antibody or fragment thereof is of non-human origin and is obtained from a donor antibody raised against an antigen fragment which does not contain said epitope.

12. A humanized antibody or a fragment thereof according to claim 1, wherein the humanized antibody comprises variable regions with human- or primate-derived framework regions and at least two CDRs with an amino acid sequence selected from the group of sequences consisting of SEQ ID NO:

1 representing CDR1, SEQ ID NO;

2 representing CDR2 and SEQ ID NO;

3 representing CDR3 of the Heavy Chain Variable Region (HCVR) and SEQ ID NO;

4 representing CDR1, SEQ ID NO;

5 representing CDR2 and SEQ ID NO;

6 representing CDR3 of the Light Chain Variable Region (LCVR).

45. A chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof antibody according to claim 1, wherein the Heavy Chain Variable Region (HCVR) has an amino acid sequence that is 90%, particularly 95%, more particularly 98% identical to the sequence given in SEQ ID NO:

15.

46. A chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof according to claim 1, wherein the Light Chain Variable Region (LCVR) has an amino acid sequence that is 90%, particularly 95%, more particularly 98% identical to the sequence given in SEQ ID NO:

12.

61. A nucleic acid molecule comprising a nucleotide sequence encoding a chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof according to claim 1.

70. An expression vector comprising the nucleotide sequence of any one of claims 61 to 63.

71. A cell comprising an expression vector comprising the nucleotide sequence of any one of claims 61 to 63 or an expression vector of claim 70.

72. A composition comprising the antibody according to claim 1 including any functionally equivalent antibody or functional parts thereof in a therapeutically effective amount.

75. A mixture comprising a chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof according to claim 1 including any functionally equivalent antibody or functional parts thereof in a therapeutically effective amount and, optionally, a further biologically active substance and/or a pharmaceutically acceptable carrier and/or a diluent and/or an excipient.

81. Use of a chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof and/or a functional part thereof according to claim 1 and/or a composition according to claim 72, or a mixture according to claim 75 for the preparation of a medicament for treating or alleviating the effects of amyloidosis, a group of diseases and disorders associated with amyloid plaque formation including secondary amyloidosis and age-related amyloidosis such as diseases including, but not limited to neurological disorders such as Alzheimer'"'"'s Disease (AD), Lewy body dementia, Down'"'"'s syndrome, hereditary cerebral hemorrhage with amyloidosis (Dutch type);

the Guam Parkinson-Dementia complex;

as well as other diseases which are based on or associated with amyloid-like proteins such as progressive supranuclear palsy. multiple sclerosis;

Creutzfeld Jacob disease, Parkinson'"'"'s disease, HIV-related dementia, ALS (amyotropic lateral sclerosis), Adult Onset Diabetes;

senile cardiac amyloidosis;

endocrine tumors, and others, including macular degeneration.

86. A method for the preparation of a composition according to claim 72, or a mixture according to claim 75 for use in a method of preventing, treating or alleviating the effects of amyloidosis, a group of diseases and disorders associated with amyloid plaque formation including secondary amyloidosis and age-related amyloidosis such as diseases including. but not limited to, neurological disorders such as Alzheimer'"'"'s Disease (AD), Lewy body dementia, Down'"'"'s syndrome, hereditary cerebral hemorrhage with amyloidosis (Dutch type);

the Guam Parkinson- Dementia complex;

as well as other diseases which are based on or associated with amyloid-like proteins such as progressive supranuclear palsy, multiple sclerosis;

Creutzfeld Jacob disease, Parkinson'"'"'s disease, HIV-related dementia, ALS (amyotropic lateral sclerosis), Adult Onset Diabetes;

senile cardiac amyloidosis;

endocrine tumors, and others, including macular degeneration comprising formulating an antibody according to the invention in a pharmaceutically acceptable form.

93. A method for preventing, treating or alleviating the effects of amyloidosis, a group of diseases and disorders associated with amyloid plaque formation including secondary amyloidosis and age-related amyloidosis such as diseases including, but not limited to, neurological disorders such as Alzheimer'"'"'s Disease (AD). Lewy body dementia, Dawn'"'"'syndrome, hereditary cerebral hemorrhage with amyloidosis (Dutch type);

the Guam Parkinson-Dementia complex;

as well as other diseases which are based on or associated with amyloid-like proteins such as progressive supranuclear palsy, multiple sclerosis;

Creutzfeld Jacob disease, Parkinson'"'"'s disease, HIV-related dementia, ALS (amyotropic lateral sclerosis), Adult Onset Diabetes;

senile cardiac amyloidosis;

endocrine tumors, and others, including macular degeneration by administering a chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof and/or a functional part thereof according to claim 1 and/or a pharmaceutical composition according to claim 72, or a mixture according to claim 75, to a animal or a human affected by such a disorder comprising administering the antibody in a therapeutically effective amount.

99. Method of diagnosis of an amyloid-associated disease or condition in a patient comprising

(a) bringing the sample or a specific body part or body area suspected to contain the amyloid protein into contact with an antibody according to claim 1;

(b) allowing the antibody to bind to the amyloid protein;

(c) detecting the antibody bound to the protein; and

(d) correlating the presence or absence of antibody binding with the presence or absence of amyloid protein in the sample or specific body part or area.

100. Method of determining the extent of amyloidogenic plaque burden in a tissue and/or body fluids comprising

(a) obtaining a sample representative of the tissue and/or body fluids under investigation;

(b) testing said sample for the presence of amyloid protein with an antibody according to claim 1;

(c) determining the amount of antibody bound to the protein; and

(d) calculating the plaque burden in the tissue and/or body fluids.

101. Test kits for detection and diagnosis of amyloid-associated diseases and conditions comprising an antibody according to claim 1.

121. A chimeric antibody or a fragment thereof, or a humanized antibody or a fragment thereof according to claim 1, which antibody substantially binds Aβ

monomers but, preferably, does not show any significant cross-reactivity with amyloid precursor protein (APP).

151. A method of preventing Abeta-induced neuron degradation comprising treating neurons with an effective amount of a humanized antibody or a fragment thereof according to claim 1.

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62. A nucleic acid molecule comprising a nucleotide sequence encoding a humanized light chain variable region comprising SEQ ID NO:

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63. A nucleic acid molecule comprising a nucleotide sequence encoding a humanized heavy chain variable region comprising SEQ ID NO:

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105. Heavy Chain Variable Region (HCVR) comprising human- or primate-derived heavy chain framework regions and at least two CDRs with an amino acid sequence selected from the group of sequences consisting of SEQ ID NO:

1 representing CDR1, SEQ ID NO;

2 representing CDR2 and SEQ ID NO;

3 representing CDR3 of the Heavy Chain Variable Region (HCVR).

106. Light Chain Variable Region (LCVR),comprising human- or primate-derived light chain framework regions and at least two CDRs with an amino acid sequence selected from the group of sequences consisting of SEQ ID NO:

4 representing CDR1, SEQ ID NO;

5 representing CDR2 and SEQ ID NO;

6 representing CDR3 of the Light Chain Variable Region (LCVR).

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113. A humanized antibody or a fragment thereof, which comprises the Light Chain Variable Region (LCVR) of SEQ ID NO:

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119. A humanized antibody or a fragment thereof, which comprises the Heavy Chain Variable Region (HCVR) of SEQ ID NO:

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120. A humanized antibody or a fragment thereof, which comprises the Heavy Chain of SEQ ID NO:

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149. A method for disaggregating preformed beta-amyloid fibers, comprising interacting an hC2 antibody with preformed beta-amyloid fibers.

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