JP2016047839A - Treatment of amyloidogenic disease - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a novel therapy and reagent for the treatment of Alzheimer disease.SOLUTION: The present invention provides a preferable dosage range, maximum concentration, average concentration and monitoring method for use in the treatment of Alzheimer disease using an antibody to Aβ. The present invention also provides a monitoring method capable of evaluating a change in patient's symptom or indication after the treatment. Optionally, this humanized antibody is a humanized type of murine antibody 3D6 expressed by hybridoma deposited with ATCC under the number of PTA-5130.SELECTED DRAWING: None

Description

BACKGROUND OF THE INVENTION Alzheimer's disease (AD) is a progressive disease that results in senile dementia. In general, Se
lkoe, TINS 16: 403 (1993); Hardy et al., WO 92/13069.
Selkoe, J .; Neuropathol. Exp. Neurol. 53: 438 (
1994); Duff et al., Nature 373: 476 (1995); Games et al.,
See Nature 373: 523 (1995). Broadly speaking, the disease can be divided into two categories: late onset that occurs in old age (65 years and older) and early onset before old age, that is, between 35 and 60 years old. Divided. In both types of disease, the pathology is the same, but abnormalities tend to be more severe and widespread when they begin at an early age. This disease is characterized by at least two types of lesions in the brain: neurofibrillary tangles and senile plaques. A neurofibrillary tangle is an intracellular deposit of microtubule-associated tau protein consisting of two filaments wound together in pairs. Senile plaques (ie, amyloid plaques) are areas of a disturbed neural network up to 150 μm across, with extracellular amyloid deposits in the center, which are visible by microscopic analysis of brain tissue sections. Accumulation of amyloid plaques in the brain is also associated with Down syndrome and other cognitive disorders.

The main component of the plaque is a peptide called Aβ or β-amyloid peptide. The Aβ peptide is a 39-43 amino acid 4-kDa internal fragment of a larger transmembrane glycoprotein, termed a protein called amyloid precursor protein (APP). As a result of proteolytic processing of APP by various secretase enzymes, A
β is mainly found in both the short form with a length of 40 amino acids and the long form with a length in the range of 42-43 amino acids. Part of the hydrophobic transmembrane domain of APP is found at the carboxy terminus of Aβ, and in particular in the long form, can contribute to Aβ's ability to aggregate and become plaques. Accumulation of amyloid plaques in the brain eventually leads to neuronal cell death. Physical symptoms associated with this type of neurodegeneration characterize Alzheimer's disease.

Several mutations within the APP protein have been correlated with the presence of Alzheimer's disease. For example, Goate et al., Nature 349: 704 (1991) (valine ⁷¹⁷ to isoleucine); Charter Harlan et al., Nature 3
53: 844 (1991)) (valine ⁷¹⁷ to glycine); Murrell et al., Sci.
ence 254: 97 (1991) (valine ⁷¹⁷ to phenylalanine); Mull
an et al., Nature Genet. 1: 345 (1992) (double mutation changing from lysine ⁵⁹⁵ -methionine ⁵⁹⁶ to asparagine ⁵⁹⁵ -leucine ⁵⁹⁶ ). Such mutations may cause Alzheimer's disease by increased or altered processing of APP to Aβ, particularly by processing APP to increased amounts of long forms of Aβ (ie, Aβ1-42 and Aβ1-43). Conceivable. Mutations in other genes, such as the presenilin genes PS1 and PS2, are thought to produce increased amounts of long form Aβ by indirectly affecting APP processing (Hardy, TINS 20). 154 (1997)).

A mouse model has been used successfully to determine the importance of amyloid plaques in Alzheimer's disease (Games et al., Supra, Johnson-Wood et al., Proc. Nat.
l. Acad. Sci. USA 94: 1550 (1997)). In particular, when a PDAPP transgenic mouse (which expresses a mutated form of human APP and develops Alzheimer's disease at an early age) is injected with a long form of Aβ, the mouse has a reduced progression of Alzheimer's disease and Aβ Shows both increased antibody titers against peptides (Schenk et al., Nat
ure 400, 173 (1999)). The above observations show that Aβ in its long form
Is a causative factor of Alzheimer's disease.

McMichael, EP 526511, proposes to administer a homeopathic dose (less than or equal to 10 ⁻² mg / day) of Aβ to patients with pre-proven AD. In a typical human having about 5 liters of plasma, even this upper dose limit would be expected to produce a concentration of 2 pg / ml or less. A in human plasma
The normal concentration of β is generally in the range of 50-200 pg / ml (Seuber
t et al., Nature 359: 325 (1992)). Because the proposed dose of EP526511 seems to change little the level of endogenous circulating Aβ, and EP52
In 6511, it is unlikely that any therapeutic benefit would be provided because the use of adjuvants is not recommended as an immunostimulant.

Thus, there is a need for new therapies and reagents for the treatment of Alzheimer's disease, particularly those that can provide therapeutic benefits at physiological (eg, non-toxic) doses.

Cross-reference to related applications U.S. Application No. 60/648631 filed Jan. 28, 2005;
U.S. Publication No. US 20060193850 Al published on May 31; International Publication No. WO 06/083589 published on August 10, 2006; U.S. Application No. 60/622525 filed on October 26, 2004. No. and US Publication No. US20060160161Al, published July 20, 2006, are related applications, all of which are hereby incorporated by reference in their entirety for all purposes.

SUMMARY OF THE INVENTION The present invention provides a method for therapeutically treating Alzheimer's disease. The method includes administering to a patient suffering from the disease a dose of antibody in the range of about 0.5 mg / kg to less than 5 mg / kg by intravenous infusion. This antibody is at least 10 ⁷ M ⁻
Specifically binds to the N-terminal fragment of β-amyloid peptide (Aβ) with a binding affinity of ¹ , thereby treating the patient therapeutically. In some cases, the antibody is a humanized antibody. In some cases, the humanized antibody is a humanized form of murine antibody 3D6 expressed by a hybridoma deposited with the ATCC under number PTA-5130. In some cases, the humanized antibody comprises (i) a light chain comprising three complementarity determining regions (CDRs) from the immunological light chain variable region of murine antibody 3D6, and (ii) immunity of murine antibody 3D6. A heavy chain comprising three complementarity determining regions (CDRs) derived from a biological heavy chain variable region. In some cases, the humanized antibody comprises (i) a variable light chain region having the sequence shown in SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5, and (ii) SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 A variable heavy chain region having the sequence shown in FIG. In some cases, the humanized antibody comprises (i) a variable light chain region having the sequence shown in SEQ ID NO: 1 or SEQ ID NO: 5, and (ii) a variable heavy having the sequence shown in SEQ ID NO: 2 or SEQ ID NO: 6. Contains the chain region. In some cases, the humanized antibody is bapineuzumab. In some cases, the antibody has the number PTA-51.
Mouse antibody 10D5 expressed by the hybridoma deposited with ATCC at 29
Is a humanized version of In some cases, the humanized antibody comprises (i) a light chain comprising three complementarity determining regions (CDRs) from the immunological light chain variable region of murine antibody 10D5, and (ii)
) Three complementarity determining regions (CDRs) derived from the immunological heavy chain variable region of mouse antibody 10D5
Containing heavy chains. In some cases, the humanized antibody comprises (i) a variable light chain region having the sequence set forth in SEQ ID NO: 7 or SEQ ID NO: 7 set forth in US Patent Publication No. 20050142131, and (ii) SEQ ID NO: 8 or US It includes a variable heavy chain region having the sequence shown in SEQ ID NO: 8 shown in Patent Publication No. 20050142131. In some cases, humanized antibodies have been deposited by the hybridoma deposited with the American Type Culture Collection (ATCC), Manassas, VA 20108, USA, on April 8, 2003, in accordance with the terms of the Budapest Treaty and having deposit number PTA-7271. Humanized form of mouse antibody 12A11 expressed.

In some cases, the humanized antibody comprises (i) a light chain comprising three complementarity determining regions (CDRs) from the immunological light chain variable region of mouse antibody 12A11, and (ii) mouse antibody 1
It contains a heavy chain comprising three complementarity determining regions (CDRs) derived from the immunological heavy chain variable region of 2A11. In some cases, the humanized antibody is (i) US Patent Publication No. 200501865.
A variable light chain region having the sequence shown in SEQ ID NO: 2 shown in No. 1 and (ii) a variable heavy chain region having the sequence shown in SEQ ID NO: 4 shown in US Patent Publication No. 200501118651. In some methods, the dosage is about 0.5 mg / kg. In some methods, the dosage is about 1.5 mg / kg. In some methods, the dosage is 0.
5 to 3 mg / kg. In some methods, the dosage is 0.5 to 1.5 mg / k
g. In some methods, this dosage is administered on multiple occasions, such as every 13 weeks.

Some methods include the Minimental State Test (MMSE), the Alzheimer's Disease Rating Scale-Cognition (ADAS-COG), and clinician interview-base.
d impression) (CIBI), neurological test battery
) (NTB), disability assessment for dementia (DAD)
, Clinical dementia assessment-sum of boxes (CDR-SOB), neuropsychiatric inventory (NPI), positron emission tomography (PET imaging)
The method further includes monitoring the patient with at least one assessment selected from the group consisting of scans, magnetic resonance imaging (MRI) scans, and blood pressure measurements. In some methods, the type of assessment is MMSE, which can be used on multiple occasions, such as before dose administration, 4 weeks, 16 weeks, 6 months, 1 year after dose administration, etc. To be applied. In some methods, the MMSE score measured after administration is higher than the previously evaluated MMSE score.

The invention further provides a method of therapeutically treating Alzheimer's disease, wherein a patient suffering from this disease is administered intravenously with a dose of antibody in the range of about 0.5 mg / kg to less than 5 mg / kg. Administering by injection, wherein the antibody specifically binds to β-amyloid peptide (Aβ) with a binding affinity of at least 10 ⁷ M ⁻¹ , posterior reversible encephalopathy syndrome (PRES) or monitoring the patient for angioedema. In some cases, the monitoring step may be FLAIR (water suppression inversion recovery method: Fluid Attenuat
ed Inversion Recovery, including sequence imaging, and performing an MRI scan. In some methods, the monitoring step causes the PRE to
At least one clinical symptom related to S, for example, headache, nausea, vomiting, confusion, seizures, visual abnormalities, changes in mental function, ataxia, frontal symptom, parietal symptoms (par
ietal symptom), stupor, or local neurological signs. In some ways,
Dosage is reduced or suspended based on MRI scan results indicating PRES or angioedema. In some methods, the dosage is FLAI indicating PRES or angioedema.
Based on R-sequence imaging results, reduction or suspension. In some ways,
The dosage is reduced or suspended based on the identification of at least one clinical symptom associated with PRES. In some methods, MRI scans are every 3 months, every 6 months, or every year. In some methods, FLAIR sequence imaging is every 3 months, every 6 months, or every year.

Some of the methods further include determining the presence or absence of hypertension in the patient, and if the patient has hypertension, the method further includes administering an antihypertensive drug. In some cases, the antihypertensive agent is selected from the group consisting of hydrochlorothiazide, angiotensin converting enzyme (ACE) inhibitor, angiotensin II receptor blocker (ARE), beta blocker, and calcium channel blocker.

Some methods further comprise treating PRES or angioedema by administering a steroid to the patient. In some cases, the steroid is dexamethasone or methylprednisolone.

Some methods are based on MRI scan results showing PRES or blood vessels, reducing or suspending dosage, and at least one clinical symptom associated with PRES or angioedema, such as headache, nausea, vomiting Further including identifying confusion, seizures, visual abnormalities, changes in mental function, ataxia, frontal lobe symptoms, parietal lobe symptoms, stupor, or local neurological signs. Some methods include reducing or suspending dosage based on FLAIR sequence imaging results indicating PRES or angioedema, and PRES
Or at least one clinical symptom associated with angioedema, such as headache, nausea, vomiting,
The method further includes identifying confusion, seizures, visual abnormalities, changes in mental function, ataxia, frontal lobe symptoms, parietal lobe symptoms, stupor, or local neurological signs.

In some methods, the monitoring step includes PRES at a first time point after administration.
Or the presence of PRES or angioedema is indicated at the second time point after the first time point, indicating that there is no PRES or angioedema, and the step of monitoring is present A second dose is administered or administered after the step of administering and monitoring the patient at the first dose is detected that PRES or angioedema is present. Otherwise, after the monitoring step detects that PRES or angioedema is not present, the third dose is administered, the first and third doses being greater than the second dose.

In some of the above methods, the antibody is a humanized antibody. In some cases, humanized antibodies are deposited by the hybridoma deposited with the American Type Culture Collection (ATCC), Manassas, VA 20108, USA, on April 8, 2003, according to the terms of the Budapest Treaty and having deposit number PTA-5130. Mouse antibody 3 expressed
It is a humanized form of D6.

In some cases, the humanized antibody comprises (i) a light chain comprising three complementarity determining regions (CDRs) from the immunological light chain variable region of murine antibody 3D6, and (ii) murine antibody 3D6.
A heavy chain comprising three complementarity determining regions (CDRs) derived from the immunological heavy chain variable region. In some cases, the humanized antibody comprises (i) a variable light chain region having the sequence shown in SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5, and (ii) SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 A variable heavy chain region having the sequence shown in FIG. In some cases, the humanized antibody is
(I) a variable light chain region having the sequence shown in SEQ ID NO: 1 or SEQ ID NO: 5, and (ii)
) Comprising a variable heavy chain region having the sequence shown in SEQ ID NO: 2 or SEQ ID NO: 6. In some cases, the humanized antibody is bapineuzumab.

In some of the above methods, the dosage is about 0.5 mg / kg. In some methods, the dosage is about 1.5 mg / kg. In some methods, the dosage is 0.5
To 3 mg / kg. In some methods, the dosage is 0.5 to 1.5 mg / kg
It is. In some methods, this dosage is administered on multiple occasions, such as every 13 weeks.

In some of the above methods, bapineuzumab is prematurely treated with PRES or angioedema.
Administered at the first dose before being determined from I-scan, and PRES or angioedema is MRI
A second dose is administered after being determined from the scan, and the second dose is less than the first dose. In some cases, the first dose is 3-5 mg / kg and the second dose is 0
. 5 to 3 mg / kg. In some cases, the second dose is half of the first dose. In some cases, bapineuzumab is administered at a first frequency before MRI shows PRES or angioedema, and at a second frequency after MRI shows PRES or angioedema, The frequency is less than the first frequency.

In some of the above methods, the type of evaluation is blood pressure, and the presence or absence of hypertension is determined. In some cases, if the patient has hypertension, the method further comprises administering an antihypertensive drug. In some cases, the antihypertensive agent is a hydrochlorothiazide, an angiotensin converting enzyme (ACE) inhibitor, an angiotensin II receptor blocker (ARB).
), Beta blockers, and calcium channel blockers.

The present invention further provides a therapeutic product. This product includes a glass vial and instructions for use. The glass vial contains about 10 mg to about 250 mg of humanized anti-Aβ antibody, about 4%
Mannitol or about 150 mM NaCl, about 5 mM to about 10 mM histidine,
And a formulation containing about 10 mM methionine. Instructions for monitoring patients to whom the formulation is administered for PRES and / or angioedema are included in the product.

The present invention relates to a method of treating Alzheimer's disease, wherein a patient having this disease is treated with Aβ.
Administering subcutaneously an antibody that specifically binds to an N-terminal fragment of the antibody, wherein the antibody is administered at a dose of 0.01-0.6 mg / kg and at a frequency between weekly and monthly. A method comprising: In some cases, the antibody is administered at a dose of 0.05 to 0.5 mg / kg. In some cases, the antibody is administered at a dose of 0.05 to 0.25 mg / kg. In some cases, the antibody is administered from weekly to biweekly at a dose of 0.015-0.2 mg / kg. In some cases, the antibody is administered weekly to biweekly at a dose of 0.05 to 0.15 mg / kg. In some cases, this antibody is 0.05-0.07 m.
Administered weekly at a dose of g / kg. In some cases, the antibody is administered weekly at a dose of 0.06 mg / kg. In some cases, the antibody is administered every other week at a dose of 0.1-0.15 mg / kg. In some cases, the antibody is administered monthly at a dose of 0.1-0.3 mg / kg. In some cases, the antibody is administered monthly at a dose of 0.2 mg / kg.

The present invention relates to a method of treating Alzheimer's disease, wherein a patient having this disease is treated with Aβ.
An antibody that specifically binds to the N-terminal fragment of is administered subcutaneously, wherein the antibody is administered at a dose of 1-40 mg and at a frequency between weekly and monthly. . In some cases, the antibody is administered at a dose of 5-25 mg. In some cases, the antibody is administered at a dose of 2.5-15 mg. In some cases, this antibody is
Administered weekly to biweekly at a dose of 1-12 mg. In some cases, this antibody is 2.5
Administered weekly to biweekly at a dose of -10 mg. In some cases, the antibody is 2.5 to
It is administered weekly at a dose of 5 mg. In some cases, the antibody is administered weekly at a dose of 4-5 mg. In some cases, the antibody is administered every other week at a dose of 7-10 mg.

The present invention relates to a method of treating Alzheimer's disease, wherein a patient having this disease is treated with Aβ.
Administering an antibody that specifically binds to an N-terminal fragment of an antibody in a regimen sufficient to maintain a maximum serum concentration of the antibody in the patient, less than about 28 μg of antibody per ml of serum, thereby treating the patient A method of including is provided. In some cases, the maximum serum concentration is in the range of about 4-28 μg antibody per ml of serum. In some cases, the maximum serum concentration is
Within the range of about 4-18 μg of antibody per ml of serum. In some cases, the average serum concentration of antibody in a patient is less than about 7 μg of antibody per ml of serum. In some cases, the average serum concentration is in the range of about 2-7 μg of antibody per ml of serum. In some cases,
The average serum concentration is about 5 μg of antibody per ml of serum. In some cases, the antibody is administered intravenously. In some cases, the antibody is administered subcutaneously. In some cases, a dose of 0.1-1.0 mg / kg is administered monthly. In some cases, 0.5-1.
A dose of 0 mg / kg is administered monthly. In some cases, the antibody is administered at a frequency between weekly and monthly. In some cases, the antibody is administered weekly or biweekly. Some methods further include measuring the concentration of antibody in the serum and adjusting the regimen if the measured concentration is out of range. In some cases, the antibody is a humanized antibody. In some cases, the humanized antibody is ATCC with the number PTA-5130.
Is a humanized version of mouse antibody 3D6 expressed by the hybridoma deposited at In some cases, the humanized antibody is bapineuzumab. In some cases, the humanized antibody is a humanized form of murine antibody 10D5 expressed by a hybridoma deposited with the ATCC under number PTA-5129. In some cases, the humanized antibody is a humanized form of murine antibody 12A11 expressed by a hybridoma deposited with the ATCC under the number PTA-7271.

The present invention relates to a method of treating Alzheimer's disease, wherein a patient having this disease is treated with Aβ.
Administering an antibody that specifically binds to an N-terminal fragment of an antibody in a regimen sufficient to maintain an average serum concentration of antibody in the patient of less than about 7 μg of antibody per ml of serum, thereby treating the patient A method of including is provided. In some cases, the average serum concentration is serum 1
Within the range of about 2-7 μg antibody per ml. In some cases, the average serum concentration is about 5 μg of antibody per ml of serum. In some cases, the antibody is administered intravenously.
In some cases, the antibody is administered subcutaneously. In some cases, 0.1-1.0 mg
A dose of / kg is administered monthly. In some cases, a dose of 0.5-1.0 mg / kg is administered monthly. In some cases, the antibody is administered at a frequency between weekly and monthly. In some cases, the antibody is administered weekly or biweekly. Some methods further include measuring the concentration of antibody in the serum and adjusting the regimen if the measured concentration is out of range. In some cases, the antibody is a humanized antibody. In some cases, the humanized antibody is a humanized form of murine antibody 3D6 expressed by a hybridoma deposited with the ATCC under number PTA-5130. In some cases, the humanized antibody is bapineuzumab. In some cases, the humanized antibody has the number PTA-
Mouse antibody 10 expressed by a hybridoma deposited with the ATCC at 5129
It is a humanized form of D5. In some cases, the humanized antibody is designated A, with the number PTA-7271.
Humanized form of murine antibody 12A11 expressed by a hybridoma deposited with TCC.

FIG. 1 shows the light and heavy chain amino acid sequences of bapineuzumab deduced from the DNA sequence of the expression construct. FIG. 2 shows the average MMSE change from baseline. Figure 3 shows the change from baseline MMSE by cohort at 4 months. FIG. 4 shows the mean, median and standard deviation of MMSE change from baseline at 4 months. FIG. 5 shows the statistical test results of MMSE change from baseline at 4 months. FIG. 6 shows simulated antibody steady state serum concentrations from various subcutaneous regimens assuming 70% bioavailability. FIG. 7 shows the steady state concentration of antibody after subcutaneous administration of AAB-001 at a dose of 0.05 or 0.06 mg / kg assuming 70% or 100% bioavailability. FIG. 8 shows plasma Aβ levels after administration of AAB-001 at a dose of 0.15, 0.5, or 1.0 mg / kg. FIG. 9 shows the pharmacokinetic parameters after intravenous administration of AAB-001 at doses of 0.15, 0.5, 1.0, and 2.0 mg / kg. FIG. 10 shows the mean serum AAB-001 concentration versus time characteristics after intravenous administration of AAB-001 at doses of 0.15, 0.5, 1.0, and 2.0 mg / kg. FIG. 11 shows plasma Aβ levels after intravenous administration of AAB-001 at doses of 0.15, 0.5, and 1.0 mg / kg.

This application is intended to maximize therapeutic benefit against the occurrence of side effects, particularly angiogenic edema.
Administration of antibodies to the N-terminal fragment of Aβ is provided at a preferred dosage and frequency. A series of experiments using various regimens of murine 3D6 antibody in transgenic PDAPP mice identified a steady state average serum concentration of approximately 3.7 μg / ml antibody for reduction of amyloid accumulation. This data is 0.5m administered intravenously every 13 weeks.
It provides evidence that doses of g / kg and 1.5 mg / kg were effective in inhibiting cognitive decline in Alzheimer's disease patients. These regimens result in an average serum concentration of antibody that includes an effective dose of 3.7 μg / ml in mice. For example, 13
It was found that a dose of 0.5 mg / kg administered weekly produced an average serum concentration of about 3 μg / ml. A dose of 1 mg / kg administered every 13 weeks was found to produce an average serum concentration of about 5.5 μg / ml, administered every 13 weeks.
A dose of 2 mg / kg was expected to produce an average serum concentration of 9.4 μg / ml. Thus, the data show that serum antibodies of the same magnitude that are effective in mice are effective in humans. These data represent full-length Aβ1-4
Further supported by clinical trials with immunotherapy at 2. In these studies, antibody responders were found to be statistically significantly inhibited from cognitive decline. The antibody responder has an ELI of at least 1 of 2200 antibodies corresponding to a serum titer of about 1 μg / ml.
Has SA titer.

This data shows that for higher doses of antibody, especially 5 mg / kg, 0.5-1.5 mg / kg
Only achieves a therapeutic benefit that is not (possibly smaller) than lower doses in the range, but also provides evidence that some patients also experience significant side effects, particularly vasogenic edema. While carrying out the present invention does not depend on an understanding of the mechanism, side effects are believed to be due to its high maximum concentration after administration of the antibody.

In general, these data have shown that the therapeutic benefit is effective in mice.
It shows that it can be obtained with a relatively modest dose of antibody designed to produce an average serum concentration similar to that encompassing this value, which would be effective in humans at 7 μg / ml. This data shows that for regimens that give an equivalent area under the curve with respect to the serum concentration of antibody as a function of time, the smaller doses that are administered more frequently have the side effect profile than the larger doses that are administered less frequently. The former regimen also indicates that the former regimen avoids antibody concentration spikes associated with the administration of large doses in the latter regimen. In the study of this example, administration was given intravenously every 13 weeks. The interval between doses can be reduced approximately every month with corresponding reductions in individual doses, but further increases in frequency every week or every other week (every other week) usually require a visit to an infusion center High risk of non-compliance due to inconvenience of intravenous administration. However, weekly or biweekly dosing is practical for subcutaneous administration, which can be easily self-administered,
Or it can be administered by a caregiver who has not received medical training. By subcutaneous delivery
Slower delivery to the blood is also provided, and concentration spikes are avoided. The bioavailability as measured by the area under the curve of the antibody in plasma for subcutaneous delivery compared to intravenous delivery is about 70-100%.

Thus, a preferred regimen for administering an antibody that specifically binds to the N-terminal fragment of Aβ achieves an average serum concentration of administered antibody of 1-15 μg / ml in the patient.
This range covers the effective concentrations demonstrated in mice and humans, allowing some margin for measurement error and individual patient variability. Serum concentrations are determined by actual measurement or based on the amount of antibody administered, frequency of administration, route of administration and half-life of the antibody (eg WinNonline Version 4
. 0.1 (Pharsight Corporation, Cary, USA)). Mean antibody concentration in serum is 1-10, 1-5 or 2-4 μg / m
It is preferable to be within the range of l.

The data show that antibodies that specifically bind to the N-terminal fragment of Aβ are approximately 2 antibodies per ml of serum.
There is also evidence that administration of a regimen sufficient to maintain the highest serum concentration of antibody in a patient, less than 8 μg, maximizes the therapeutic benefit against possible side effects, particularly the occurrence of angioedema. provide. A preferred maximum serum concentration is about 4 to 4 antibodies per ml of serum.
It is in the range of 28 μg. The highest serum with less than about 28 μg of antibody per ml of serum and 1 m of serum
A combination with an average serum concentration of antibody in the patient of less than about 7 μg antibody per liter is particularly beneficial. See FIGS. 9 and 10.

This data shows that an antibody that specifically binds to the N-terminal fragment of Aβ is approximately 7 antibodies per ml of serum.
Also provides evidence that administration of a regimen sufficient to maintain an average serum concentration of antibody of less than μg maximizes the therapeutic benefit against possible side effects, particularly the occurrence of angioedema . A preferred average concentration is in the range of about 2-7 μg of antibody per ml of serum.

If the antibody is administered intravenously, it is inconvenient if it needs to be administered more frequently than about every month, as discussed above. Preferred antibody doses for monthly intravenous administration are 0.1-1.
Occurs within the range of 0 mg / kg antibody, or preferably 0.5-1.0 mg / kg antibody.

For more frequent administration, eg weekly to monthly dosing, subcutaneous administration is preferred. The dose used for subcutaneous administration is usually 0.1 to 0.6 mg / kg or 0.01 to 0.35
mg / kg, preferably in the range of 0.05 to 0.25 mg / kg. For weekly or biweekly dosing, the dose is 0.015-0.2 mg / kg, or 0.05-0.1
It is preferably within the range of 5 mg / kg. For weekly dosing, the dose is 0.05
To 0.07 mg / kg, for example about 0.06 mg / kg. For biweekly dosing, the dose is preferably 0.1 to 0.15 mg / kg. For monthly dosing, the dosage is preferably 0.1 to 0.3 mg / kg, or about 2 mg / kg. Monthly administration includes dosing by calendar month or lunar month (ie every 4 weeks).

FIG. 6 shows the results from various subcutaneous regimens assuming 70% bioavailability.
The steady state serum concentration of the simulated antibody is shown. It can be seen that a dose of 0.1 mg / kg results in an average serum concentration that is very close to 3.7 μg / ml found to be effective in mice and has only a slight peak-to-trough variation. Figure 7 shows 70% or 10
The steady state concentration of the antibody is shown after subcutaneous administration at doses of 0.05 and 0.06 mg / kg, assuming 0% bioavailability. At 70% bioavailability, the 0.05 and 0.06 mg / kg doses were just below and directly above the 3.7 μg / ml dose found to be effective in mice, with only slight peak-to-trough variations I understand that I do not have it.

The treatment regimen is as described above, wherein the average serum concentration of the antibody is at least 6 months or 1 year,
And in some cases it is usually continued to be maintained for a lifetime. Serum concentrations can be measured at any time during the treatment, and if the average concentration falls below the target range, the dose and / or frequency of administration can be increased and the average concentration is within the target range. If so, the dose and / or frequency can be reduced.

Determining the optimal plasma concentration of an antibody is useful for determining a dosage regimen or optimizing the dosage in an individual patient, but in practice, mg / kg or m
Once an effective dosage regimen and dosage frequency in g has been determined, the same dosage regimen can be used for many other patients without having to calculate or measure the patient titer in detail. Thus, any of the above dosages and treatment regimens can be used regardless of whether titers are measured or expected in a particular patient. For example, one suitable regimen is 0.1-1.0 mg / kg antibody, or preferably 0.5-1.0 m.
Intravenous administration at monthly intervals, with doses ranging from g / kg antibody. For subcutaneous dosing, the dose used is usually 0.01-0.6 mg / kg or 0.01-0.35
mg / kg, preferably in the range of 0.05 to 0.25 mg / kg. For weekly or biweekly dosing, the dose is 0.015-0.2 mg / kg, or 0.05-0.1
It is preferably within the range of 5 mg / kg. For weekly dosing, the dose is 0.05
It is preferable that it is -0.07 mg / kg, for example, 0.06 mg / kg. For biweekly dosing, the dose is preferably 0.1 to 0.15 mg / kg. For monthly dosing, the dose is preferably 0.1 to 0.3 mg / kg or 2 mg / kg.

As elsewhere in this application, the dose expressed in mg / kg is given here by multiplying the general patient mass (eg 70 or 75 kg) and generally rounding to the nearest whole number. Can be converted to a quantity. When expressed in absolute mass, antibodies are usually administered at a dose between 1 and 40 mg, with a frequency between weekly and monthly. A preferred range is 5-25 mg or 2.5-15 mg, with a weekly to monthly frequency. For weekly to biweekly administration, the dose is often 1-12 mg or 2.5-10 mg. For weekly administration, the dose is often 2.5 to 5 mg or 4 to 5 mg. For biweekly administration, the dose can be 7-10 mg. The mass of antibody packed for administration in unit dose is an integer, eg 1, 5, 10, 20, 30, 40, 50, 75 or 100 m.
Usually rounded off to g.

The present invention is for use in treating Alzheimer's disease with an antibody against Aβ.
It is provided to monitor preferred dosage ranges and regimens. This method presupposes to some extent the clinical test results described in the examples. A preferred dosage range for antibodies that bind to the N-terminal fragment of Aβ is about 0.5 to 5 mg antibody per kg patient body weight. A preferred dosage is less than 5 mg / kg. 0.5 to 3 mg / kg, 0.5
To 1.5 mg / kg and 1.5 mg / kg dosages are particularly preferred.

The present invention also provides for monitoring a regimen that can assess changes in the patient's symptoms or signs after treatment. This symptom or sign may be related to Alzheimer's disease itself and / or side effects of treatment. The dose of the drug or the frequency of its administration can be adjusted based on the monitoring results. Alternatively, or in addition, additional agents can be administered to treat any side effects. For example, MRI and / or F
By using LAIR sequencing imaging monitoring, PRES or angioedema or its signs or symptoms can be detected. Presence of PRES or angioedema indicates that doses should be reduced or discontinued or dose intervals should be increased. Alternatively, or in addition, steroids can be administered to the patient to treat PRES or angioedema. After dose reduction or suspension, or after increasing the interval between doses and / or after steroid administration,
Continued monitoring may indicate that PRES or angioedema has disappeared, in which case the original dose and / or dosing interval can be resumed. Administration of steroids may or may not be continued as a preventive measure at this point. As another example,
Blood pressure monitoring may indicate the development of hypertension. Similarly, administration can be reduced in dosage or suspended, and / or intervals between administrations can be increased, and / or antihypertensive drugs can be administered. If further monitoring indicates that hypertension has disappeared, the original amount and / or dosing interval can be resumed. Administration of antihypertensive drugs may or may not continue at this point as a preventive measure.

Before describing the present invention, it will be helpful in understanding the present invention to provide definitions of certain terms used below.

The term “immunoglobulin” or “antibody” (used interchangeably herein) refers to antigen binding having a basic four polypeptide chain structure consisting of two heavy chains and two light chains. Refers to a sex protein, the chain being stabilized by, for example, an interchain disulfide bond, which has the ability to specifically bind antigen. Both heavy and light chains are folded into domains. The term “domain” includes, for example, peptide loops stabilized by β-pleated sheets and / or intrachain disulfide bonds (eg, 3-4
The globular region of the heavy chain or light chain. The domain
Referred to herein as “constant” or “variable”, which is the relative lack of sequence variation within the domains of the various class members in the case of the “constant” domain, or the various in the case of the “variable” domain. Based on significant mutations within class member domains. The “constant” domains on the light chain are referred to interchangeably as “light chain constant regions”, “light chain constant domains”, “CL” regions or “CL” domains. “Constant” domains on the heavy chain are referred to interchangeably as “heavy chain constant regions”, “heavy chain constant domains”, “CH” regions or “CH” domains. A “variable” domain on a light chain is referred to interchangeably as a “light chain variable region”, “light chain variable domain”, “VL” region or “VL” domain. “Variable” domains on the heavy chain are referred to interchangeably as “heavy chain constant regions”, “heavy chain constant domains”, “CH” regions or “CH” domains.

The term “region” refers to a portion or portion of an antibody chain and includes constant or variable domains as defined herein, as well as more discrete portions or portions of said domains.
For example, a light chain variable domain or region includes “complementarity determining regions” or “CDRs” interspersed within “framework regions” or “FRs” as defined herein.

The immunoglobulin or antibody can exist in monomeric or polymeric form. The term “antigen-binding fragment” refers to an antibody that binds or is intact with an antigen (ie,
It refers to immunoglobulin or antibody polypeptide fragments that compete for antigen binding (ie, specific binding) with the intact antibody from which they are derived. The term “conformation”
Is a protein or polypeptide (eg, antibody, antibody chain, domain or region thereof)
The tertiary structure of For example, the phrase “light chain (or heavy chain) conformation” refers to the tertiary structure of the light chain (or heavy chain) variable region, and the phrases “antibody conformation” or “antibody fragment conformation” Refers to the tertiary structure of an antibody or fragment thereof.

“Specific binding” of an antibody means that the antibody exhibits appreciable affinity for the antigen or preferred epitope, and preferably does not exhibit significant cross-reactivity. “Sensitive” or preferred binding is at least 10 ⁶ , 10 ⁷ , 10 ⁸ , 10 ⁹ M
^-1, or a bond with the affinity of ¹⁰ 10 ^{M -1.} Affinities greater than 10 ⁷ M ⁻¹ , preferably greater than 10 ⁸ M ⁻¹ are more preferred. These intermediate values presented herein are also intended to be within the scope of the present invention, and preferred binding affinities are, for example, 10 ⁶ to 1
0 ¹⁰ M ⁻¹ , preferably 10 ⁷ to 10 ¹⁰ M ⁻¹ , more preferably 10 ⁸ to 1
It can be shown as an affinity range of 0 ¹⁰ M ⁻¹ . “No significant cross-reactivity”
An antibody is an antibody that does not appreciably bind to an unwanted entity (eg, an unwanted proteinaceous entity). For example, an antibody that specifically binds Aβ binds Aβ appreciably but does not react significantly with non-Aβ proteins or peptides (eg, non-Aβ proteins or peptides contained in plaques). Antibodies specific for the preferred epitope are:
For example, it does not significantly cross-react with distant epitopes on the same protein or peptide.
Specific binding can be determined by any means recognized in the art for determining such binding. Preferably, specific binding is determined by Scatchard analysis and / or competitive binding assays.

Binding fragments are generated by recombinant DNA techniques or by enzymatic or chemical cleavage of intact immunoglobulins. Binding fragments include Fab, Fab ′, F (ab ′)
₂ , Fabc, Fv, single chain, and single chain antibodies are included. Other than “bispecific” or “bifunctional” immunoglobulins or antibodies, an immunoglobulin or antibody is understood to have each of its binding sites identical. "Bispecific" or "bifunctional antibody" is 2
An artificial hybrid antibody with two different heavy / light chain pairs and two different binding sites. Bispecific antibodies can be produced by a variety of methods including fusion of hybridomas or linking of Fab ′ fragments. For example, Songsivillai & Lac
hmann, Clin. Exp. Immunol. 79: 315-321 (1990);
Koselny et al. Immunol. 148, 1547-1553 (1992).

The term “humanized immunoglobulin” or “humanized antibody” includes an immunoglobulin comprising at least one humanized immunoglobulin chain or humanized antibody chain (ie, at least one humanized light chain or heavy chain). Or refers to an antibody. The term “humanized immunoglobulin chain” or “
A “humanized antibody chain” (ie, “humanized immunoglobulin light chain” or “humanized immunoglobulin heavy chain”) is a variable framework region substantially derived from a human immunoglobulin or human antibody, and a non-human immunoglobulin. Or a complementarity determining region (CDR) substantially derived from a non-human antibody (eg, at least one CDR, preferably two CDRs, more preferably three CDRs) and further a constant region (eg, a light chain An immunoglobulin chain or an antibody chain (ie light chain or heavy chain, respectively) having a variable region further comprising at least one constant region or portion thereof in the case, and preferably three constant regions in the case of the heavy chain).
Point to. The term “humanized variable region” (eg, “humanized light chain variable region” or “humanized heavy chain variable region”) refers to variable framework regions and non-human immunity substantially derived from human immunoglobulins or human antibodies. Refers to a variable region comprising a complementarity determining region (CDR) substantially derived from a globulin or non-human antibody.

The phrase “substantially derived from a human immunoglobulin or human antibody” or “substantially human”, when aligned with the amino acid sequence of a human immunoglobulin or human antibody for comparison purposes, Shares at least 80-90%, preferably 90-95%, more preferably 95-99% identity (ie local sequence identity) with the sequence of the constant region, eg, conservative substitutions, consensus sequences It means to allow substitution, germline substitution, backmutation and the like. Introduction of conservative substitutions, consensus sequence substitutions, germline substitutions, back mutations, etc. is often referred to as “optimization” of a humanized antibody or chain. The phrase “substantially derived from a non-human immunoglobulin or non-human antibody” or “substantially non-human” refers to the sequence of a non-human organism, such as a non-human mammal, at least 80-95%, preferably 90 Meaning having an immunoglobulin or antibody sequence that is -95%, more preferably 96%, 97%, 98%, or 99% identical.

Thus, except possibly for CDRs, all regions or residues of a humanized immunoglobulin or humanized antibody, or humanized immunoglobulin chain or humanized antibody chain are the corresponding regions of one or more natural human immunoglobulin sequences. Or substantially the same as the corresponding residue. The term “corresponding region” or “corresponding residue” is the same (ie equivalent) as a region or residue on the first amino acid or nucleotide sequence when the first and second sequences are optimally aligned for comparison purposes. N) refers to a region or residue on the second amino acid or nucleotide sequence that occupies a position.

The term “humanized immunoglobulin” or “humanized antibody” is not intended to encompass chimeric immunoglobulins or antibodies as defined below. Humanized immunoglobulins or humanized antibodies are chimeric in their configuration (ie, contain regions from more than one species of protein), but they are chimeric immunoglobulins or chimeric antibodies as defined herein. Additional features not found in (ie, variable regions comprising donor CDR residues and acceptor framework residues).

The term “chimeric immunoglobulin” or antibody refers to an immunoglobulin or antibody whose variable region is derived from a first species and whose constant region is derived from a second species. Chimeric immunoglobulins or chimeric antibodies can be constructed from immunoglobulin gene segments belonging to different species, eg, by genetic engineering.

An “antigen” is an entity (eg, proteinaceous entity or peptide) to which an antibody specifically binds.

The term “epitope” or “antigenic determinant” refers to a site on an antigen to which an immunoglobulin or antibody (or antigen-binding fragment thereof) specifically binds. Epitopes can be formed from both contiguous amino acids or non-adjacent amino acids that are flanked by tertiary folds of the protein. Epitopes formed from adjacent amino acids are generally retained when exposed to denaturing solvents, whereas epitopes formed by tertiary fold structures are generally lost upon treatment with denaturing solvents. An epitope generally comprises at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 amino acids in a unique spatial conformation. Methods for determining the spatial conformation of an epitope include, for example, X-ray crystallography and two-dimensional nuclear magnetic resonance. For example, Epitope Mapping
Protocols in Methods in Molecular Biolo
gy, 66, G.G. E. See Morris ed. (1996).

Antibodies that recognize the same epitope can be identified in a simple immunoassay, ie, a competitive binding assay, that shows the ability of one antibody to block another antibody from binding to the target antigen. Competitive binding occurs when the immunoglobulin under test is a common antigen of the reference antibody, eg, A
It is determined by an assay that inhibits specific binding to β or the like. Numerous types of competitive binding assays such as solid phase direct or indirect radioimmunoassay (RIA), solid phase direct or indirect enzyme immunoassay (EIA), sandwich competitive assays (Stahli et al., Meth).
ods in Enzymology 9: 242 (1983)), solid phase direct biotin-avidin EIA (Kirkland et al., J. Immunol. 137: 3).
614 (1986)); solid phase direct labeling assay, solid phase direct labeling sandwich assay (Harlow and Lane, Antibodies: A Laborat).
ory Manual, Cold Spring Harbor Press (1988)
); Solid phase direct labeling RIA using the I-125 label (Morel et al., Mo.
l. Immunol. 25 (1): 7 (1988)); solid phase direct biotin-avidin EIA (Cheung et al., Virology 176: 546 (1990)).
And directly labeled RIA (Moldenhauer et al., Scand. J. Immunol
. 32:77 (1990)). In general, such assays involve the use of purified antigen bound to a solid surface, or cells having any of these, unlabeled test immunoglobulin and labeled reference immunoglobulin. Competitive inhibition is measured by determining the amount of label bound to the solid surface or cells in the presence of the test immunoglobulin. Usually the test immunoglobulin is present in excess. Usually, when a competing antibody is present in excess, it will have at least 50-55% specific binding of the reference antibody to the common antigen,
55-60%, 60-65%, 65-70%, 70-75% or more will be inhibited.

Epitopes are also recognized by immune cells such as B cells and / or T cells. Epitope cell recognition is determined by in vitro assays that measure antigen-dependent proliferation as determined by ³ H-thymidine incorporation, cytokine secretion, antibody secretion, or antigen-dependent killing (cytotoxic T lymphocyte assay). can do.

Exemplary epitopes or antigenic determinants are human amyloid precursor protein (APP)
Although it can be found within, it is preferably found within the Aβ peptide of APP. There are multiple APP isoforms, such as APP ⁶⁹⁵ , APP ⁷⁵¹ , and APP ⁷⁷⁰ . The amino acids in APP are in the number specified by the sequence of APP ⁷⁷⁰ isoform (see, eg, GenBank accession number P05067, also shown in SEQ ID NO: 38). Aβ (also referred to herein as β-amyloid peptide and A-β) peptide is the 39-43 amino acids of APP (Aβ39, Aβ40, Aβ41, A
An internal fragment of β42 and Aβ43) of about 4 kDa. Aβ40 consists of residues 672-711 of APP, for example, and Aβ42 consists of residues 673-713 of APP. in
As a result of proteolytic processing of APP by various secretase enzymes in vivo or in situ, Aβ is a “short form” 40 amino acids in length and a “long” in the range 42-43 amino acids in length. Found both in "form". As described herein, preferred epitopes or antigenic determinants are located within the N-terminus of the Aβ peptide and are within amino acids 1-10 of Aβ, preferably residues 1-3, 1-4 of Aβ42, 1-5,
Includes residues from 1-6, 1-7, or 3-7. Additional referenced epitopes or antigenic determinants are Aβ residues 2-4, 5, 6, 7 or 8, Aβ residues 3-5, 6, 7, 8 or 9, or Aβ42 residue 4 -7, 8, 9 or 10.

The term “amyloidogenic disease” includes any disease associated with (or resulting from) the formation or deposition of insoluble amyloid fibrils. Exemplary amyloidogenic diseases include, but are not limited to, systemic amyloidosis, Alzheimer's disease, mature onset diabetes, Parkinson's disease, Huntington's disease, frontotemporal dementia, and prion-related infectious sponges Encephalopathy (Kool and Creutzfeldt-Jakob disease in humans, and scrapie and BSE in sheep and cattle, respectively). Depending on the nature of the polypeptide component of the deposited fibrils, various amyloidogenic diseases are defined or characterized. For example, in a subject or patient with Alzheimer's disease, β-amyloid protein (eg, wild-type, mutant, or truncated β-amyloid protein) is a characteristic polypeptide component of amyloid deposits. Therefore,
Alzheimer's disease is an example of a “disease characterized by Aβ deposition” or a “disease associated with Aβ deposition” in the brain of a subject or patient, for example. The terms “β-amyloid protein”, “β-amyloid peptide”, “β-amyloid”, “Aβ” and “Aβ peptide” are used interchangeably herein.

The term “effective dose” or “effective dose” is defined as an amount sufficient to achieve, or at least to some extent, achieve the desired effect. The term “therapeutically effective dose” is defined as an amount sufficient to cure or at least partially inhibit the disease and its complications in a patient already suffering from the disease. Effective amounts for this use will depend on the severity of the infection and the general condition of the patient's own immune system.

The term “patient” includes human and other mammalian subjects that receive either prophylactic or therapeutic treatment.

“Soluble” or “dissociated” Aβ refers to non-aggregated or disaggregated Aβ polypeptide. “Insoluble” Aβ refers to an aggregated Aβ polypeptide, eg, Aβ held together by non-covalent bonds. Aβ (eg, Aβ42) is believed to aggregate at least to some extent due to the presence of hydrophobic residues at the C-terminus of the peptide (part of the transmembrane domain of APP). One method for preparing soluble Aβ is to dissolve the lyophilized peptide in pure DMSO while sonicating. The obtained solution is centrifuged to remove any insoluble fine particles.

The term “effector function” refers to an activity that belongs to the Fc region of an antibody (eg, an IgG antibody) and includes, for example, the ability of an antibody to bind effector molecules, such as complement and / or Fc receptors. Can control several immune functions of antibodies, such as effector cell activity, lysis, complement-mediated activity, antibody clearance, and antibody half-life.

The term “effector molecule” refers to a molecule capable of binding to the Fc region of an antibody (eg, an IgG antibody), including but not limited to complement proteins or Fc receptors.

The term “effector cell” refers to a cell that is capable of binding to the Fc portion of an antibody (eg, an IgG antibody) generally, through Fc receptors expressed on the surface of effector cells, including but not limited to lymphatic cells. Includes spheres such as antigen presenting cells and T cells.

The term “Fc region” refers to the C-terminal region of an IgG antibody, particularly the heavy chain (s) of the IgG antibody.
The C-terminal region of Although the boundaries of the Fc region of an IgG heavy chain can vary slightly, an Fc region is generally defined as extending from about amino acid residue Cys226 to the carboxyl terminus of the IgG heavy chain (s).

Unless otherwise stated, the term “Kabat numbering” is specifically used by Kabat et al. (Sequences of Pro, which is incorporated herein by reference.
teins of Immunological Interest, 5th edition Public
c Health Service, National Institutes of
Health, Bethesda, Md. (1991)) defined as numbering of residues in, for example, an IgG heavy chain antibody using the EU index.

The term “Fc receptor” or “FcR” refers to a receptor that binds to the Fc region of an antibody. Common Fc receptors that bind to the Fc region of antibodies (eg, IgG antibodies) include, but are not limited to, FcγRI, FcγRII, and FcγRIII subclass receptors (
Allelic variants and alternative spliced forms of these receptors). Fc receptors are described in Ravetch and Kinet, Annu. Rev. Im
munol 9: 457-92 (1991); Capel et al., Immunomethod.
s 4: 25-34 (1994); and de Haas et al. Lab. Clin.
Med. 126: 330-41 (1995).

Immunological and therapeutic reagents The immunological and therapeutic reagents of the present invention comprise or are derived from an immunogen or antibody, or a functional or antigen-binding fragment thereof, as defined herein. Become. The basic antibody structural unit is known to comprise a tetramer of subunits. Each tetramer consists of two identical pairs of polypeptide chains, each pair consisting of one “light” chain (approximately 25 kDa)
And one “heavy” chain (about 50-70 kDa). The amino-terminal portion of each chain contains a variable region of about 100 to 110 or more amino acids that is primarily responsible for antigen recognition. The carboxy-terminal portion of each chain defines a constant region primarily responsible for effector functions.

Antibody As used herein, the term “antibody” refers to immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, ie, molecules that contain an antigen binding site that specifically binds (recognizes) an antigen. Point to. Examples of immunologically active portions of immunoglobulin molecules include F (ab
) And F (ab ′) 2 fragments, which can be produced by treating the antibody with an enzyme such as pepsin, or can be produced by art-recognized recombinant engineering techniques. it can. Aspects of the invention that also relate to antibody stabilization include, for example, polyclonal and monoclonal antibodies that bind a therapeutic target antigen such as an antigen, eg, Aβ. As used herein, the term “monoclonal antibody” or “monoclonal antibody composition” is the only type capable of recognizing and binding to a specific epitope of a target antigen, eg, the epitope (s) of Aβ. Refers to a population of antibody molecules containing the antigen binding sites of Thus, a monoclonal antibody composition generally exhibits a single binding specificity and affinity for a particular target antigen with which it immunoreacts.

Polyclonal antibodies Polyclonal antibodies can be prepared as described above by immunizing a suitable subject with an immunogen. The antibody titer of the immunized subject can be monitored over time by standard techniques, such as an enzyme linked immunosorbent assay (ELISA) using immobilized target antigen. Optionally, antibody molecules directed against the target antigen can be isolated from the mammal (eg, from blood) and further purified by well-known techniques, such as protein A sepharose chromatography, For example, an IgG fraction can be obtained. At an appropriate time after immunization, for example when the anti-antigen antibody titer is highest, antibody producing cells are obtained from the subject and standard techniques such as Kohler and Milste are used.
The hybridoma technique (Brown et al. (1981) J. Immunol. 127: 539-46) first described by in (1975) Nature 256: 495-497.
Brown et al. (1980) J .; Biol. Chem. 255: 4980-83; Yeh
(1976) Proc. Natl Acad. Sci. USA 76: 2927-31
And Yeh et al. (1982) Int. J. et al. Cancer 29: 269-75), etc.) can be used to prepare monoclonal antibodies. For the preparation of chimeric polyclonal antibodies, see Buechler et al. US Pat. No. 6,420,113.

Monoclonal antibodies Any of a number of well-known protocols used for the fusion of lymphocytes with immortalized cell lines can be applied for the purpose of generating monoclonal antibodies (eg G
. Galfred et al. (1977) Nature 266: 55052; Gefter et al.
Somatic Cell Genet. Cited above; Lerner, Yale J .;
Biol. Med. Cited above; Kenneth, Monoclonal Antib
odies, cited above). Moreover, those skilled in the art will appreciate that many variations of such methods exist and are also useful. In general, immortal cell lines (
For example, myeloma cell lines) are obtained from the same mammalian species as the lymphocytes. For example, murine hybridomas can be made by fusing lymphocytes from a mouse immunized with the immunogenic preparation of the present invention with an immortalized mouse cell line. Preferred immortal cell lines are mouse myeloma cell lines that are sensitive to medium containing hypoxanthine, aminopterin and thymidine (“HAT medium”). Any number of myeloma cell lines, eg, P3-NS1 / 1-Ag4-1, P3-x63-Ag8.653 or Sp2 / O-A
The g14 myeloma line can be used as a fusion partner by standard techniques. These myeloma lines are available from the ATCC. Generally, HAT-sensitive mouse myeloma cells are fused to mouse splenocytes using polyethylene glycol (“PEG”). The hybridoma cells resulting from the fusion are then selected using HAT medium that kills unfused and non-productively fused myeloma cells (
Unfused splenocytes are not transformed and die after a few days). Hybridoma cells producing the monoclonal antibodies of the invention can be obtained using standard ELISA assays.
It is detected by screening hybridoma culture supernatants for antibodies that bind the target antigen, eg, Aβ.

Recombinant antibodies As an alternative to preparing monoclonal antibody-secreting hybridomas, monoclonal antibodies are identified and isolated by screening a recombinant combinatorial immunoglobulin library (eg, antibody phage display library) with the target antigen. Thereby, immunoglobulin library members that bind the target antigen can be isolated. Kits for generating and screening phage display libraries are commercially available (eg, Pharmacia Recombinant
Page Antibody System, catalog number 27-9400-01; and Stratagene SurfZAP ™ Page Display Ki.
t, catalog number 240612). In addition, examples of methods and reagents that are particularly applicable for use in generating and screening antibody display libraries include, for example, Ladner et al., US Pat. No. 5,223,409; Kang et al., PCT International Publication No. W
O92 / 18619; Dower et al., PCT International Publication No. WO 91/17271; Wi
nter et al., PCT International Publication No. WO 92/20791; Markland et al., PCT International Publication No. WO 92/15679; Breitling et al., PCT International Publication No. WO 93 /
No. 01288; McCafferty et al., PCT International Publication No. WO 92/01047; G
Arrrard et al., PCT International Publication No. WO 92/09690; Ladner et al., PCT International Publication No. WO 90/02809; Fuchs et al. (1991) Bio / Technol.
ogy 9: 1370-1372; Hay et al. (1992) Hum. Antibody. H
bridomas 3: 81-85; Huse et al. (1989) Science 24.
6: 1275-1281; Griffiths et al. (1993) EMBO J 12: 7.
25-734; Hawkins et al. (1992) J. MoI. Mol. Biol. 226: 889
Clarkson et al. (1991) Nature 352: 624-628;
Gram et al. (1992) Proc. Natl. Acad. Sci. USA 89:35
76-3580; Garrad et al. (1991) Bio / Technology 9: 1
373-1377; Hoogenboom et al. (1991) Nuc. Acid Res.
19: 4133-4137; Barbas et al. (1991) Proc. Natl. Aca
d. Sci. USA 88: 7978-7982; and McCafferty et al., Na.
true (1990) 348: 552-554.

Chimeric and humanized antibodies In addition, recombinant antibodies, such as chimeric and humanized monoclonal antibodies, contain both human and non-human portions and can be made using standard recombinant DNA techniques. While within the scope of the present invention.

The term “humanized immunoglobulin” or “humanized antibody” refers to at least one humanized immunoglobulin chain or humanized antibody chain (ie, at least one humanized light chain or heavy chain).
An immunoglobulin or antibody comprising The term “humanized immunoglobulin chain” or “humanized antibody chain” (ie, “humanized immunoglobulin light chain” or “humanized immunoglobulin heavy chain”) is substantially derived from a human immunoglobulin or human antibody. Variable framework regions and complementarity determining regions substantially derived from non-human immunoglobulin or non-human antibodies (
CDR) (eg, at least one CDR, preferably two CDRs, more preferably three CDRs), and in the case of a constant region (eg, at least one constant region or portion thereof in the case of light chain and heavy chain) Refers to an immunoglobulin or antibody chain (ie, light or heavy chain, respectively) having a variable region further comprising three constant regions). The term “humanized variable region” (eg, “humanized light chain variable region” or “humanized heavy chain variable region”) refers to variable framework regions and non-human immunity substantially derived from human immunoglobulins or human antibodies. Refers to a variable region comprising a complementarity determining region (CDR) substantially derived from a globulin or non-human antibody.

The phrase “substantially derived from a human immunoglobulin or human antibody” or “substantially human”, when aligned with the amino acid sequence of a human immunoglobulin or human antibody for comparison purposes, Constant region sequence and at least 80-90%, 90
Means ~ 95%, or 95-99% identity (ie, local sequence identity), allowing for example conservative substitutions, consensus sequence substitutions, germline substitutions, backmutations, etc. To do. Introduction of conservative substitutions, consensus sequence substitutions, germline substitutions, back mutations, etc. is often referred to as “optimization” of a humanized antibody or chain. The phrase “substantially derived from a non-human immunoglobulin or non-human antibody” or “substantially non-human” refers to a sequence of a non-human organism, such as a non-human mammal, at least 80-95%, preferably at least Meaning having an immunoglobulin or antibody sequence that is 90-95%, more preferably 96%, 97%, 98%, or 99% identical.

Thus, with the exception of the CDRs, all regions or residues of a humanized immunoglobulin or humanized antibody, or humanized immunoglobulin chain or humanized antibody chain are the corresponding regions of one or more natural human immunoglobulin sequences or It is substantially identical to the corresponding residue. The term “corresponding region” or “corresponding residue” is the same (ie equivalent) as a region or residue on the first amino acid or nucleotide sequence when the first and second sequences are optimally aligned for comparison purposes. N) refers to a region or residue on the second amino acid or nucleotide sequence that occupies a position.

The term “significant identity” refers to two polypeptide sequences, the default gap weight (ga
these sequences have at least 50-60% sequence identity, preferably at least 60-70% sequence identity, more preferably when optimally aligned, such as by GAP or BESTFIT of programs using p weight) At least 70-80% sequence identity,
More preferably at least 80-90% sequence identity, even more preferably at least 90-95% sequence identity, and even more preferably at least 95% sequence identity (eg, 99% sequence identity or more) Means sharing. The term “substantial identity” refers to two polypeptide sequences, the G of the program using default gap weights.
When optimally aligned, such as by AP or BESTFIT, these sequences have at least 80-90% sequence identity, preferably at least 90-95% sequence identity, and more preferably at least 95% sequence or more. It means sharing identity (for example, 99% or more sequence identity). With respect to sequence comparison, typically one sequence acts as a reference sequence, to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are entered into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. The sequence comparison algorithm then calculates the percent sequence identity for the test sequence (s) relative to the reference sequence, based on the designated program parameters.

Optimal alignment of sequences for comparison is described, for example, in Smith & Waterman, Adv.
Appl. Math. 2: 482 (1981) Local homology algorithm, Need
leman & Wunsch, J.M. Mol. Biol. 48: 443 (1970) homology alignment algorithm, Pearson & Lipman, Proc. Nat '
l. Acad. Sci. USA 85: 2444 (1988) similarity search, computerization of the implementation of these algorithms (Wisconsin Genetics So)
ftware Package, Genetics Computer Group, 5
75 Science Dr. , Madison, WI GAP, BESTFIT
, FASTA, and TFASTA), or visual inspection (generally Ausubel et al., C
current Protocols in Molecular Biology). One example of an algorithm suitable for determining percent sequence identity and sequence similarity is the BLAST algorithm, which is the Alt
schul et al. Mol. Biol. 215: 403 (1990).
Software for performing BLAST analyzes is available at the National Center for Biotechnology Information (publicly accessible through the National Institutes of Health NCBI Internet Server)
Publicly available through In general, default program parameters can be used to perform the sequence comparison, but customized parameters can also be used. Regarding the amino acid sequence, the BLASTP program has a word length (W) of 3;
Ten predictions (E) and the BLOSUM62 scoring matrix are used as defaults (Henikoff & Henikoff, Proc. Natl. Acad.
Sci. USA 89: 10915 (1989)).

Preferably, residue positions that are not identical differ by conservative amino acid substitutions. For the purpose of classifying amino acid substitutions as conservative or non-conservative, amino acids are as follows: group I (hydrophobic side chain): leu, met, ala, val, leu, ile; group II (neutral Hydrophilic side chain): cys, ser, thr; Group III (acidic side chain)
: Asp, glu; group IV (basic side chain): asn, gln, his, lys, a
rg; group V (residues that affect chain orientation): gly, pro; and group V
I (aromatic side chain): grouped into trp, tyr, phe. Conservative substitutions involve substitutions between amino acids within the same class. Non-conservative substitutions constitute exchanging one member of these classes for another member.

Preferably, the humanized immunoglobulin or humanized antibody binds the antigen with an affinity that is within 3, 4, or 5 times the affinity of the corresponding non-humanized antibody. For example, a non-humanized antibody
A humanized antibody having a binding affinity of 10 ⁻⁹ M is at least 3 × 10 ⁻⁸ M, 4 ×
It will have a binding affinity of 10 ⁻⁸ M, 5 × 10 ⁻⁸ M, or 10 ⁻⁹ M. When describing the binding properties of an immunoglobulin or antibody chain, this chain is described as “direct antigen (eg,
It can be described on the basis of “Aβ) binding” ability. A chain is said to be “direct antigen binding” if it imparts specific binding properties or affinity to an intact immunoglobulin or antibody (or antigen-binding fragment thereof). An antibody comprising an equivalent chain that lacks said mutation in the binding affinity of an intact immunoglobulin or antibody (or antigen-binding fragment thereof) comprising said chain (or back mutation), or A mutation substantially affects the ability of a heavy or light chain to direct antigen binding if it affects (eg, reduces) at least an order of magnitude compared to the binding affinity of the antigen binding fragment). It is said that. Of an antibody (or antigen-binding fragment thereof) comprising an equivalent chain lacking said mutation in the binding affinity of an intact immunoglobulin or antibody (or antigen-binding fragment thereof) comprising said chain. A mutation "does not substantially affect (eg does not reduce) the ability of the chain to bind directly to antigen" if it only affects (eg, does not reduce) the binding affinity by 2, 3, or 4 times.

The term “chimeric immunoglobulin” or antibody refers to an immunoglobulin or antibody whose variable region is derived from a first species and whose constant region is derived from a second species. Chimeric immunoglobulins or chimeric antibodies can be constructed from immunoglobulin gene segments belonging to different species, eg, by genetic engineering. The term “humanized immunoglobulin” or “humanized antibody” is not intended to encompass chimeric immunoglobulins or antibodies as defined below. Humanized immunoglobulins or humanized antibodies are chimeric in their configuration (ie, contain regions from more than one species of protein), but they are chimeric immunoglobulins or chimeric antibodies as defined herein. Additional features not found in (ie, variable regions comprising donor CDR residues and acceptor framework residues).

Such chimeric and humanized monoclonal antibodies can be obtained by recombinant DNA techniques known in the art, for example, Robinson et al., International Application No. P
CT / US86 / 02269; Akira et al., European Patent Application No. 184187; Tan
iguchi, M .; Morrison et al., European Patent Application No. 173494; Neuberger et al., PCT International Publication No. WO 86/01533; Cabilly et al., US Pat. No. 4,816,567; (1988) Science 240: 1041-1
043; Liu et al. (1987) Proc. Natl. Acad. Sci. USA 84
: 3439-3443; Liu et al. (1987) J. MoI. Immunol. 139: 3521
-3526; Sun et al. (1987) Proc. Natl. Acad. Sci. USA
84: 214-218; Nishimura et al. (1987) Canc. Res. 47:
999-1005; Wood et al. (1985) Nature 314: 446-449;
And Shaw et al. (1988) J. MoI. Natl. Cancer Inst. 80: 155
3-15559; Morrison, S .; L. (1985) Science 229: 1
202-1207; Oi et al. (1986) BioTechniques 4: 214; W
inter, US Pat. No. 5,225,539; Jones et al. (1986) Nature.
321: 552-525; Verhoeyan et al. (1988) Science 239
: 1534; and Beidler et al. (1988) J. MoI. Immunol. 141: 40
It can be generated using the method described in 53-4060.

Transgenic animals and human antibodies derived from phage display Alternatively, immunization creates a transgenic animal (eg, a mouse) that can produce the entire repertoire of human antibodies in the absence of endogenous immunoglobulin production. Is currently possible. For example, it has been described that homozygous deletion of the antibody heavy chain joining region (J _H ) gene in chimeric and germ-line mutant mice results in complete inhibition of endogenous antibody production. Transfer of a human germline immunoglobulin gene array in such germline mutant mice results in the production of human antibodies upon antigen challenge.
For example, US Pat. Nos. 6,150,584; 6,114,598; and 5,770,42.
See No. 9.

All human antibodies can also be obtained from phage display libraries (Hoog
enroom et al. Mol. Biol. 227: 381 (1991); Marks et al., J. Biol. Mol. Biol. 222: 581-597 (1991)). Chimeric polyclonal antibodies can also be obtained from phage display libraries (Buechle).
r et al., US Pat. No. 6,420,113).

Bispecific antibodies, antibody fusion polypeptides, and single chain antibodies Bispecific antibodies (BsAbs) are antibodies that have binding specificities for at least two different epitopes. Such antibodies include full-length antibodies or antibody fragments (eg F (ab
) '2 bispecific antibody). Methods for making bispecific antibodies are known in the art. Traditional production of full length bispecific antibodies is based on the co-expression of two immunoglobulin heavy chain-light chain pairs, where the two chains have different specificities (M
illstein et al., Nature, 305: 537-539 (1983)). Because of the random combination of immunoglobulin heavy and light chains, these hybridomas (quadromas) produce a potential mixture of various antibody molecules (WO 93/08829 and Traunecker et al., EMBO J., 10: 3655). ~ 3659 (1991)).

Bispecific antibodies also include cross-linked or “heteroconjugate” antibodies. For example, one of the antibodies in the heterozygote binds to avidin and the other binds biotin or other payload (pa
yload). Heterozygous antibodies can be made using any convenient crosslinking method. Suitable crosslinking agents are well known in the art and are disclosed in US Pat. No. 4,676,980, along with several crosslinking techniques.

In yet another aspect, the antibody may generate an antibody fusion polypeptide by chemically or genetically fusing with a payload, such as a reactive moiety, a detectable moiety, or a functional moiety, such as an immunotoxin. it can. Such payloads include, for example, immunotoxins, chemotherapeutic agents, and radioisotopes, all of which are well known in the art.

Single chain antibodies are also suitable for stabilization according to the present invention. The fragment includes a heavy chain variable domain (VH) linked to a light chain variable domain (VL) by a linker, which allows each variable region to associate with each other and reproduces the antigen-binding pocket of the parent antibody. VL from there
And a VH region. Gruber et al. Immunol. 152: 53
68 (1994).

NANOBODY
Nanobodies are antibody-derived therapeutic proteins that contain the properties of naturally-derived heavy chain antibodies. Nanobodies can function as a single, relatively small, functional antigen-binding structural unit, domain or protein. Nanobody ™ technology (Ably
nx N.R. V. ) Was originally developed after the discovery that camelids (camel and llama) have fully functional antibodies that lack light chains. These heavy chain antibodies contain one variable domain (
VHH) and two constant domains (CH2 and CH3). VHH is used to distinguish these from the heavy chain variable domains (referred to as “VH domains”) present in normal 4-chain antibodies. Cloned and isolated VHH domains are stable polypeptides that have the full antigen-binding ability of the original heavy chain antibody. VHH domains and Nanobodies can also be manipulated into multivalent and multi-selective formats. A Nanobody having an amino acid sequence corresponding to the amino acid sequence of a naturally-occurring VHH domain, ie, one in the amino acid sequence of the naturally-occurring VHH sequence (and particularly in the framework sequence).
Humanization by substituting one or more amino acid residues with one or more amino acid residues present in the corresponding position (s) in a VH domain derived from a normal human four-chain antibody can do. For details, US20050130266, US20040, each of which is incorporated herein by reference for all purposes.
253638, WO / 2006/040153, US20050214857, WO / 2
See 006/079372 or WO / 2006/122825. Antibodies against Aβ can also be generated by the Nanobody ™ method.

It will be understood that any of the aforementioned polypeptide molecules, alone or in combination, are suitable for preparation as a stabilized formulation according to the present invention.

Anti-Aβ Antibodies In general, the formulations of the present invention comprise various antibodies for treating amyloidogenic diseases, particularly Alzheimer's disease, by targeting Aβ peptides.

The terms “Aβ antibody”, “anti-Aβ antibody” and “anti-Aβ” refer to an antibody that binds to one or more epitopes or antigenic determinants of human amyloid precursor protein (APP), Aβ protein, or both. Therefore, they are used interchangeably herein. Exemplary epitopes or antigenic determinants can be found in APP, but are preferably found in the Aβ peptide of APP. APP isoforms such as APP ⁶⁹⁵ , APP ^75
1 and APP ⁷⁷⁰ are present. The number of amino acids in APP is the number specified by the sequence of APP ⁷⁷⁰ isoform (eg, GenBank accession number P05067).
See). An example of a specific isoform of APP currently known to exist in humans is the 695 amino acid polypeptide described by Kang et al. (1987) Nature 325: 733-736 and named “normal” APP; Ponte Et al. (198
8) Nature 331: 525-527 (1988) and Tanzi et al. (1988)
) Nature 331: 751-amino acid polypeptide described by 528-530; and Kitaguchi et al. (1988) Nature 331: 530-53.
770-amino acid polypeptide described by 2. in vivo or in
As a result of proteolytic processing of APP by various secretase enzymes in situ, Aβ is a “short form” 40 amino acids in length, and amino acids 42-4 in length.
It is found in both of the three ranges of “long form”. The short form of Aβ ₄₀ consists of residues 672-711 of APP. The long form, eg Aβ ₄₂ or Aβ _43, is the residue 6
72-713 or 672-714. Part of the hydrophobic domain of APP is found at the carboxy terminus of Aβ, and in particular in the long form, can contribute to the ability of Aβ to aggregate.
Aβ peptides can be found in or purified from body fluids of humans or other mammals, including normal individuals and individuals suffering from amyloidogenic disorders, such as cerebrospinal fluid.

The terms “β-amyloid protein”, “β-amyloid peptide”, “β-amyloid”, “Aβ” and “Aβ peptide” are used interchangeably herein. Aβ peptides (eg Aβ39, Aβ40, Aβ41, Aβ42 and Aβ43) are
An internal fragment of about 4 kDa of ˜43 amino acids. Aβ40 consists of residues 672-711 of APP, for example, and Aβ42 consists of residues 672-713 of APP. Aβ peptides include peptides resulting from secretase cleavage of APP and synthetic peptides having the same or essentially the same sequence as the cleavage products. Aβ peptides can be obtained from a variety of sources, eg, tissues, cell lines, or body fluids (eg, serum or cerebrospinal fluid). For example, Aβ is, for example, Walsh et al. (2002), Nature 416,
As described on pages 535-539, it can be obtained from APP-expressing cells such as Chinese hamster ovary (CHO) cells stably transfected with APP _{717V → F.} Aβ preparations can be obtained from tissue sources using previously described methods (eg, Johnson-Wood et al. (1997), Proc. Natl. Acad. Scr.
. USA 94: 1550). Alternatively, Aβ peptide can be synthesized using methods well known to those skilled in the art. For example, Fields et al., Synthetic
Peptides: A User's Guide, Grant ed. H. Freem
an & Co. New York, NY, 1992, p. 77. Thus, the peptide can be, for example, Applied Biosystems Peptide Sy.
Using an automated Merrifield method for solid-phase synthesis with an α-amino group protected by either t-Boc or F-moc chemistry using a side chain protected amino acid on an nthesizer Model 430A or 431 Can be synthesized. Longer peptide antigens can be synthesized using well-known recombinant DNA techniques. For example, a polynucleotide encoding a peptide or fusion peptide can be synthesized or molecularly cloned and inserted into a suitable expression vector for transfection and heterologous expression by a suitable host cell. Aβ peptide also refers to the related Aβ sequence resulting from a mutation in the Aβ region of a normal gene.

The terms “Aβ-derived diffusible ligand” and “ADDL” are small soluble Aβ42 oligomers, primarily trimers and tetramers, but also higher order species (eg, each in its entirety for all purposes). Lambert, MP, et al., Incorporated by reference to
(1998) Proc. Natl. Acad. Sci. USA, 95, 6448-64.
53; Chromy, B .; A. Et al. (2000) Soc. Neurosci. Abst
r. 26, page 1284, WO 2004/031400).

The term “anti-ADDL antibody” refers to an antibody produced and selected for its ability to specifically bind ADDLs without binding to Aβ monomers or amyloid fibrils. For example,
WO 2004/03, which is incorporated by reference in its entirety for all purposes
See 1400.

The term “epitope” or “antigenic determinant” refers to a site on an antigen to which an immunoglobulin or antibody (or antigen-binding fragment thereof) specifically binds. Exemplary epitopes or antigenic determinants to which the Aβ antibody binds can be found in the human amyloid precursor protein (APP), but are preferably found in the Aβ peptide of APP. Exemplary epitopes or antigenic determinants within Aβ are located within the N-terminus, central region, or C-terminus of Aβ. An “N-terminal epitope” is an epitope or antigenic determinant located within the N-terminus of an Aβ peptide. Exemplary N-terminal epitopes include amino acids 1-10 or 1-1 of Aβ
2, preferably, residues 1-3, 1-4, 1-5, 1-6, 1-7, 2-6 of Aβ42,
Residues from 2-7, 3-6, or 3-7 are included. Another exemplary N-terminal epitope begins at residues 1-3 of Aβ and ends at residues 7-11. Additional exemplary N-terminal epitopes include residues 2-4, 5, 6, 7 or 8 of Aβ, residues 3-5, 6, 7, 8, or 9 of Aβ, or residues 4-4 of Aβ42 7, 8, 9 or 10 are included. A “central epitope” is an epitope or antigenic determinant that includes residues located within the middle or middle portion of an Aβ peptide. Exemplary central epitopes include residues from amino acids 13-28 of Aβ, preferably from residues 14-27, 15-26, 16-25, 17-24, 18-23, or 19-22 of Aβ. A group is included. Another exemplary central epitope includes amino acid 1 of Aβ
6-24, 16-23, 16-22, 16-21, 18-21, 19-21, 19-22
, 19-23, or 19-24 residues. The “C-terminal” epitope or antigenic determinant is located within the C-terminus of the Aβ peptide, and amino acids 33-40, 33-41 of Aβ,
Or residues within 33-42 are included. A “C-terminal epitope” includes residues located within the C-terminus of an Aβ peptide (eg, within about amino acids 30-40 or 30-42 of Aβ),
Epitope or antigenic determinant. Additional exemplary C-terminal epitopes or antigenic determinants include residues 33-40 or 33-42 of Aβ.

When an antibody is said to bind to an epitope within a particular residue, such as Aβ3-7, the antibody specifically binds to a polypeptide containing a particular residue (ie, Aβ3-7 in this example) It means to do. Such antibodies do not necessarily contact every residue within Aβ3-7. In addition, substitution or deletion of any one amino acid in Aβ3 to 7 does not necessarily significantly affect the binding affinity.

In various aspects, the Aβ antibody is end-specific. As used herein, the term “terminal specific” specifically binds to an N-terminal or C-terminal residue of an Aβ peptide, but is present within a longer Aβ species containing that residue, or within an APP. When referring to antibodies that do not recognize the same residue. In various aspects, the Aβ antibody is “C-terminal specific”. As used herein, the term “C-terminal specific” means that the antibody specifically recognizes the free C-terminus of the Aβ peptide. As an example of a C-terminal specific Aβ antibody, one that recognizes an Aβ peptide that ends with residue 40 but does not recognize an Aβ peptide that ends with residues 41, 42, and / or 43, residue 4
Recognizes Aβ peptides ending in 2, but ending in residues 40, 41, and / or 43
Examples include those that do not recognize β-peptides.

In one aspect, the Aβ antibody can be a 3D6 antibody or variant thereof, or a 10D5 antibody or variant thereof, both of which are disclosed in US Patent Publication No. 20030165496A.
No. 1, US Patent Publication No. 20040087777A1, International Patent Publication No. WO 02/462
37A3, and International Patent Publication No. WO 04/080419 A2. 3
Descriptions of D6 and 10D5 antibodies can be found, for example, in International Patent Publication No. WO 02 / 088306A2.
And International Patent Publication No. WO 02 / 088307A2.
The 10D5 antibody is also described in US Patent Publication No. 20050142131. Additional 3D6 antibodies are described in US Patent Application No. 11/303478 and International Application No. PCT / US05.
/ 45614. 3D6 is an N-terminal epitope located in the human β-amyloid peptide, in particular a monoclonal antibody (mAb) that specifically binds to residues 1-5
It is. In comparison, 10D5 is a mAb that specifically binds to an N-terminal epitope located in the human β-amyloid peptide, particularly residues 3-6. A cell line producing 3D6 monoclonal antibody (RB96 3D6.33.2.2.4) was introduced on April 8, 2003, according to the terms of the Budapest Treaty, American Type Culture Collection (ATCC), M
deposited at Anaassas, VA 20108, USA and has the deposit number PTA-5130. A cell line producing 10D5 monoclonal antibody (RB44 10D5.19.21
) Was deposited with the ATCC on April 8, 2003 in accordance with the terms of the Budapest Treaty and has the deposit number PTA-5129.

Bapineuzumab means a humanized 3D6 antibody comprising a light chain having a mature variable region having the amino acid sequence specified in SEQ ID NO: 1 and a heavy chain having a mature variable region having the amino acid sequence specified in SEQ ID NO: 2. To do.

Humanized 3D6 light chain variable region

Humanized 3D6 heavy chain variable region

Bapineuzumab is also known as AAB-001. Figure 1 shows the amino acid sequences of the light and heavy chain variable regions of bapineuzumab deduced from the DNA sequence of the expression construct. AAB-001 light and heavy chain amino acid sequences deduced from the DNA sequence of the expression construct. The CDR region is underlined. Cysteines that are expected to form intermolecular disulfide bonds are underlined, indicating binding. The N-linked glycosylation consensus site is bold italic. The expected heavy chain COOH-terminated lysine is shown in parentheses.

A humanized 3D comprising a light chain having a mature variable region having the amino acid sequence specified in SEQ ID NO: 3, and a heavy chain having a mature variable region having the amino acid sequence specified in SEQ ID NO: 4
A second version of 6 antibodies is shown below.

Humanized 3D6 light chain variable region

Humanized 3D6 heavy chain variable region

A third version of the humanized 3D6 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 5 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 6
US 2005/0090649 Al, published on May 28, which is hereby incorporated by reference for all purposes.

Humanized 3D6 light chain

Humanized 3D6 heavy chain

A humanized 10 comprising a light chain having a mature variable region having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having a mature variable region having the amino acid sequence specified in SEQ ID NO: 8.
The D5 antibody version is shown below.

Humanized 10D5 light chain variable region

Humanized 10D5 heavy chain variable region

In another aspect, the antibody can be a 12B4 antibody or variant thereof, as described in US Patent Publication No. 20040082762A1 and International Patent Publication No. WO03 / 0777858A2. 12B4 is an mAb that specifically binds to an N-terminal epitope located in the human β-amyloid peptide, particularly residues 3-7.

12A11 or a chimeric or humanized or Nanobody form thereof is a preferred antibody. The 12A11 antibody or a variant thereof is described in US Patent Publication No. 200501118651, US Patent Publication No. 20060198851, International Patent Publication No. WO04 / 108895, and International Patent Publication No. WO06 / 0666089. All in all are hereby incorporated by reference for all purposes. 12A
11 is an N-terminal epitope located in the human β-amyloid peptide, in particular residues 3-7
MAb that specifically binds to. Cell lines producing 12A11 monoclonal antibody are:
December 12, 2005, ATCC (American Type Culture Collection, 10
801 University Boulevard, Manassas, VA 201
10-2209) and has ATCC accession number PTA-7271.

A first version of a humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain (version 1) having the amino acid sequence specified in SEQ ID NO: 8 was published in June 2005. US 20050118651 Al, published on the 2nd, is incorporated herein by reference for all purposes.

Humanized 12A11 light chain

Humanized 12A11 heavy chain (version 1)

A second version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain (version 2) having the amino acid sequence specified in SEQ ID NO: 9 was published in June 2005. US 20050118651 Al, published on the 2nd, is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 2)

A third version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 10 (version 2.1) was published in 2005. No. US20050118651 Al published on June 2, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 2.1)

A fourth version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain (version 3) having the amino acid sequence specified in SEQ ID NO: 11 was published in June 2005. US 20050118651 Al, published on the 2nd, is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 3)

A fifth version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 12 (version 4.1) No. US20050118651 Al published on June 2, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 4.1)

A sixth version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 13 (version 4.2) No. US20050118651 Al published on June 2, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 4.2)

A seventh version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain (version 4.3) having the amino acid sequence specified in SEQ ID NO: 14 No. US20050118651 Al published on June 2, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 4.3)

An eighth version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain (version 4.4) having the amino acid sequence specified in SEQ ID NO: 15 No. US20050118651 Al published on June 2, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 4.4)

A ninth version of a humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain (version 5.1) having the amino acid sequence specified in SEQ ID NO: 16 No. US20050118651 Al published on June 2, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 5.1)

The 10th humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 17 (version 5.2).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 5.2)

The eleventh of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 18 (version 5.3).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 5.3)

The 12th humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 19 (version 5.4).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 5.4)

The thirteenth of a humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7, and a heavy chain having the amino acid sequence specified in SEQ ID NO: 20 (version 5.5).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 5.5)

The 14th humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 21 (version 5.6).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 5.6)

The 15th of a humanized 12A11 antibody comprising a light chain having the amino acid sequence specified by SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified by SEQ ID NO: 22 (version 6.1).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 6.1)

The 16th humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified in SEQ ID NO: 23 (version 6.2).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 6.2)

The 17th of a humanized 12A11 antibody comprising a light chain having the amino acid sequence designated SEQ ID NO: 7 and a heavy chain having the amino acid sequence designated SEQ ID NO: 24 (version 6.3).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 6.3)

The 18th humanized 12A11 antibody comprising a light chain having the amino acid sequence specified by SEQ ID NO: 7 and a heavy chain having the amino acid sequence specified by SEQ ID NO: 25 (version 6.4).
Is described in US20050118651 Al, published June 2, 2005, which is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 6.4)

A nineteenth version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain (version 7) having the amino acid sequence specified in SEQ ID NO: 26 was published in June 2005. US 20050118651 Al, published on the 2nd, is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 7)

The 20th version of the humanized 12A11 antibody comprising a light chain having the amino acid sequence specified in SEQ ID NO: 7 and a heavy chain (version 8) having the amino acid sequence specified in SEQ ID NO: 27 was published in June 2005. US 20050118651 Al, published on the 2nd, is incorporated herein by reference for all purposes.

Humanized 12A11 heavy chain (version 8)

In yet another aspect, the antibody is “Humanized Antibodies tha.
It can be the 6C6 antibody or variant thereof described in US Patent Publication No. US20060166582 and International Patent Publication No. WO06 / 06604, titled “t Recognize Beta Amyloid Peptide”. 6C6 is a mAb that specifically binds to an N-terminal epitope located in the human β-amyloid peptide, particularly residues 3-7. Cell lines producing antibody 6C6 are in accordance with the terms of the Budapest Treaty, 200
On November 1, 5 years, it was deposited with ATCC and assigned accession number PTA-7200.

In yet another aspect, the antibody can be a 2H3 antibody as described in US Patent Publication No. US20060257396. 2H3 is a mAb that specifically binds to an N-terminal epitope located in the human β-amyloid peptide, particularly residues 2-7. A cell line producing the antibody 2H3 was established on December 13, 2005 in accordance with the terms of the Budapest Treaty.
And assigned accession number PTA-7267.

In yet another aspect, the antibody can be the 3A3 antibody described in US Patent Publication US20060257396. 3A3 is a mAb that specifically binds to an N-terminal epitope located in the human β-amyloid peptide, particularly residues 3-7. A cell line producing antibody 3A3 was introduced on December 13, 2005 in accordance with the terms of the Budapest Treaty.
And assigned accession number PTA-7269.

In yet another aspect, the antibody can be 2B1, 1C2, or 9G8. Cell lines producing antibodies 2B1, 1C2 and 9G8 are in accordance with the terms of the Budapest Treaty, 20
Deposited with ATCC on November 1, 2005, accession number PTA-72, respectively.
02, PTA-7199 and PTA-7201 were assigned.

Antibodies for use in the present invention can be produced recombinantly or synthetically. For example, antibodies can be obtained from, for example, CHO cells, NIH 3T3 cells, PER. It can be produced by recombinant cell culture methods using C6® cells, NS0 cells, VERO cells, chicken embryo fibroblasts, or BHK cells. In addition, antibodies with minor modifications that retain the main functional properties of binding Aβ peptide are contemplated by the present invention. In a particular embodiment, the antibody is a humanized anti-Aβ peptide 3D6 antibody that selectively binds Aβ peptide. More specifically, the humanized anti-Aβ peptide 3D6 antibody is an NH ₂ terminal epitope, eg, human beta-amyloid 1 found in plaque deposits in the brain (eg, in patients suffering from Alzheimer's disease). Amino acid residue 1 located in the ~ 40 or 1-42 peptide
Designed to specifically bind to ~ 5.

Prophylactic and therapeutic methods The present invention relates to therapeutic responses beneficial in a patient (e.g. inducing phagocytosis of Aβ, reducing plaque burden in the patient, e.g. for the prevention or treatment of amyloidogenic diseases, among others) Therapeutic immunologic against specific epitopes within Aβ under conditions that result in inhibition of plaque formation, reduction of neuritic dystrophy, improvement of cognitive function, and / or reversal of cognitive decline, treatment or prevention It relates to the treatment of Alzheimer's disease and other amyloidogenic diseases by administering reagents (eg, humanized immunoglobulins) to patients. The invention also relates to the use of the disclosed immunological reagents (eg, humanized immunoglobulins) in the manufacture of a medicament for the treatment or prevention of amyloidogenic diseases.

As used herein, the term "treatment" cures, heals, alleviates, reduces, changes, corrects, restores, improves, or improves a disease, symptoms of the disease or predisposition to the disease. As an application or administration of a therapeutic agent to a patient having a disease, a symptom of the disease or a predisposition to the disease with an influencing purpose, or as an application or administration of the therapeutic agent to an isolated tissue or cell line from the patient Defined.

In one aspect, the present invention provides a method for preventing or treating a disease associated with amyloid deposits of Aβ in a patient's brain. Such diseases include Alzheimer's disease, Down's syndrome, and cognitive impairment. The latter can occur with or without other features of amyloidogenic disease. Some methods of the invention involve administering to a patient an effective dose of an antibody that specifically binds to a component of the amyloid deposit. Such methods are particularly useful for preventing or treating Alzheimer's disease in human patients. An exemplary method involves administering an effective dose of an antibody that binds to Aβ. The preferred method is
An effective dose of an antibody that specifically binds to an epitope within residues 1-10 of Aβ, eg, Aβ
Antibody that specifically binds to an epitope within residues 1 to 3, an antibody that specifically binds to an epitope within residues 1 to 4 of Aβ, and specifically binds to an epitope within residues 1 to 5 of Aβ Antibody, A
Specific to an antibody that specifically binds to an epitope within residues 1-6 of β, an antibody that specifically binds to an epitope within residues 1-7 of Aβ, or specific to an epitope within residues 3-7 of Aβ Administering an antibody that binds to. In yet another aspect, the invention features administering an antibody that binds to an epitope comprising a free N-terminal residue of Aβ. In yet another aspect, the invention features administering an antibody that binds to an epitope within residues 1-10 of Aβ, wherein residues 1 and / or 7 of Aβ is aspartic acid. . In yet another aspect,
The present invention is characterized by administering an antibody that specifically binds to Aβ peptide without binding to full-length amyloid precursor protein (APP). In yet another aspect, the antibody isotype is human IgG1.

In yet another aspect, the invention features administering an antibody that binds to a amyloid deposit in a patient and elicits a clearing response to the amyloid deposit.
For example, such a withdrawal response can be caused by Fc receptor-mediated phagocytosis.

The therapeutic agents of the present invention are generally not substantially contaminated with unwanted contaminants. This means that the agent is generally at least about 50% w / w (weight / weight) purity and is substantially free of interfering proteins and contaminants. In some cases, the agent is at least about 80% w / w, and more preferably at least 90 or about 95% w / w purity. However, using conventional protein purification techniques, at least 9
A 9% w / w homogeneous peptide can be obtained.

This method can be used for both asymptomatic patients and patients currently showing symptoms of the disease. The antibodies used in such methods can be human, humanized, chimeric or non-human antibodies, or fragments thereof (eg, antigen-binding fragments), as described herein, and monoclonal Or it can be polyclonal. In yet another aspect, the invention features administering an antibody prepared from a human immunized with an Aβ peptide, which can be a patient treated with the antibody.

In another aspect, the invention features administering an antibody with a pharmaceutical carrier as a pharmaceutical composition. Alternatively, the antibody can be administered to a patient by administering a polynucleotide encoding at least one antibody chain. This polynucleotide produces an antibody chain by being expressed in a patient. In some cases, the polynucleotide encodes the heavy and light chains of the antibody. The polynucleotide is expressed in a patient to produce heavy and light chains. In other embodiments, the patient is monitored for the level of antibody administered in the patient's blood.

Thus, the present invention fulfills the long-standing need for therapeutics to prevent or ameliorate neuropathology and, in some patients, cognitive impairment associated with Alzheimer's disease.

Patients applicable to treatment Patients applicable to treatment include individuals who are at risk for disease but who are not showing symptoms, as well as patients who are currently showing symptoms. In the case of Alzheimer's disease, if a person lives long, virtually everyone is at risk of suffering from Alzheimer's disease. Thus, the method can be applied prophylactically to the general population without requiring any assessment of the subject patient's risk. The method is particularly useful for individuals with a known genetic risk of Alzheimer's disease. Such individuals include those with relatives who have experienced the disease and those whose risk is determined by analysis of genetic or biochemical markers. Genetic markers of risk for Alzheimer's disease include mutations in the APP gene, particularly mutations at positions 717 and 670 and 671, referred to as Hardy mutation and Swedish mutation, respectively (see Hardy, supra). Wanna) Other markers of risk are the presenilin genes PS1 and PS2, and ApoE4 (family history of AD, hypercholesterolemia or atherosclerosis)
Mutation. Individuals currently suffering from Alzheimer's disease have characteristic dementia,
In addition, it can be recognized from the existence of the risk factors described above. In addition, several diagnostic tests are available to identify individuals with AD. These include measurements of CSFτ levels and Aβ42 levels. The increase in τ and the decrease in Aβ42 levels are
Indicates the presence of AD. Individuals suffering from Alzheimer's disease, as discussed in the Examples section,
Diagnosis can also be made according to ADRDA diagnostic criteria.

In asymptomatic patients, treatment can begin at any age (eg, 10, 20, 30). However, it is usually not necessary to begin treatment until the patient reaches 40, 50, 60 or 70. Treatment generally involves multiple administrations over a period of time. Treatment can be monitored by assaying antibody levels over time. If response decreases, booster administration is indicated. In patients with latent Down's syndrome, treatment can be initiated by administering the therapeutic agent to the mother prior to childbirth or by administering the therapeutic agent immediately after birth.

Patients applicable for treatment include those aged 50 to 87 years, patients suffering from mild to moderate Alzheimer's disease, patients having an MMSE score of 14 to 26, Neurolo
gical and Communicative Disorders and St
roke-Alzheimer's disease Related Diode
Patients with a presumed diagnosis of Alzheimer's disease based on rs (NINCDS-ADRDA) diagnostic criteria and / or patients with a Rosen Modified Hachinski Ischemic score less than or equal to 4 Can be mentioned. Some patients have MRI scans consistent with the diagnosis of Alzheimer's disease, ie other abnormalities that may be due to other diseases (eg, stroke, traumatic brain injury, arachnoid cyst, tumor, etc.) on MRI Applicable to treatment.

The method of the present invention comprises the following: encephalitis; clinically evident stroke; clinically significant carotid or vertebral floor stenosis or plaque; seizures excluding childhood febrile seizures; any significant It is particularly applicable to patients who do not have any history or evidence of autoimmune disease or immune system disorder; and / or clinically significant kidney damage. The methods of the invention are particularly applicable to patients who did not have a clinically significant infection, such as chronic persistent or acute infection, within 30 days prior to initiating treatment. The method of the present invention provides for immunosuppressive drug therapy (eg, systemic corticosteroids) (for local and nasal corticosteroids and corticosteroids for asthma within 90 days prior to initiating treatment. Inhalation is acceptable) or is particularly applicable to patients who have not been treated with chemotherapeutic agents for malignant tumors within 3 years prior to initiating treatment. The method of the present invention provides for patients who do not have clinically significant abnormalities (eg, atrial fibrillation) on physical examination, neurological examination, clinical examination or EKG examination where the abnormality can be harmful to the patient This is particularly applicable to the above. The method of the invention applies to patients who are not using anticonvulsants for seizures, anti-Parkinson's, anticoagulants (excluding use of 325 mg / day or less of aspirin), or narcotic therapy. Is also particularly applicable.

Treatment regimens and dosages For prophylactic use, the pharmaceutical composition or drug may be used to treat the biochemical, histological and / or behavioral illness of patients susceptible to Alzheimer's disease or otherwise at risk for Alzheimer's disease. In an amount sufficient to eliminate or reduce the risk, reduce the severity, or delay the development of the disease, including the symptoms, its complications, and the intermediate pathological phenotype shown during the onset of the disease Be administered. For therapeutic use, the composition or drug may have an intermediate pathological phenotype in the onset of the complications and disease in patients suspected of such disease or who already suffer from such disease. Disease symptoms including (biochemical, histological and / or
Or behavioral) is administered in an amount sufficient to cure, or at least to some extent, inhibit.

In some methods, administration of the agent reduces or eliminates myocognitive impairment in patients who have not yet developed characteristic Alzheimer's disease pathology. An amount adequate to accomplish therapeutic or prophylactic treatment is defined as a therapeutically or prophylactically effective dose. In both prophylactic and therapeutic regimes, the agent is usually administered in several doses until a sufficient immune response is achieved. The term “immune response” or “immunological response” includes humoral (antibody-mediated) responses to antigens and / or cellular (antigen-specific T cells or their secreted products) within a recipient subject. A) Response occurrence is included. Such a response is elicited by an active response, i.e., a response elicited by administration of an immunogen, or a passive response, i.e., an immunoglobulin or antibody, or a primed T cell. Response.

An “immunogen” or “immunogen” can elicit an immunological response against itself when administered to a mammal, optionally with an adjuvant. Generally, the immune response is monitored and repeated administrations are given when the immune response begins to decline.

Effective doses of the compositions of the present invention for the treatment of the above-mentioned medical conditions include the means of administration, target site, patient physiological condition, whether the patient is a human or animal, and whether other therapies are being administered. , And depending on many different factors, including whether the treatment is prophylactic or therapeutic. Usually, the patient is a human but non-human mammals, including transgenic mammals, can also be treated. Treatment doses need to be optimized for safety and efficacy by titration.

For passive immunization, the dosage is about 0.0001 to 10 per kg of host body weight.
0 mg, and more usually in the range of 0.01 to 5 mg. For example, the dosage range can be less than 20 mg / kg body weight or less than 10 mg / kg body weight, or within the range of 1.0 to 7 mg / kg. For passive immunization, preferred dosages range from 0.5 to less than 5 mg, and more usually from 0.5 to 3 mg per kg of host body weight. For example, the dosage is less than 5 mg / kg body weight or 1 kg body weight
Per mg, or in the range of 0.5 to 1.5 mg / kg, preferably at least 1.5 mg / kg. Subjects can be dosed daily, every other day, weekly, or according to any other schedule determined by empirical analysis. An exemplary treatment involves administration over multiple periods, eg, multiple doses of at least 6 months. Additional exemplary treatment regimens are once every two weeks, or once a month, or 3
1 dose every 6 months.

Exemplary passive dosing schedules include 1.5-3 mg / kg or 1.5 mg / kg every 13 weeks. The agents of the present invention are usually administered on multiple occasions. The interval between single doses can be weekly, monthly, every 13 weeks, or yearly. The interval is
It may be irregular if indicated by measuring blood levels of antibodies to Aβ in the patient.

In some methods, dosage is adjusted to achieve a plasma antibody concentration of 1-1000 μg / ml, and in some methods 25-300 μg / ml. Alternatively, the antibody can be administered as a sustained release formulation, in which case less frequent administration is required. Dosage and frequency will vary depending on the half-life of the antibody in the patient. In general, human antibodies show the longest half life, followed by humanized antibodies, chimeric antibodies, and nonhuman antibodies.

The dosage and frequency of administration will vary depending on whether the treatment is prophylactic or therapeutic. For prophylactic use, a composition containing the antibody or a cocktail thereof increases patient tolerance by administration to a patient who is not yet in a disease state. Such an amount is defined to be a “prophylactic effective dose”. For this use, the exact amount will again depend on the patient's health and systemic immunity, but is generally 0.1 to 25 mg per dose, especially 0.5 to 2.5 mg per dose. A relatively low dose is administered at relatively infrequent intervals over a long period of time. Some patients continue to receive treatment for the rest of their lives.

In therapeutic applications, relatively high doses (eg, 10 per dose) at relatively short intervals.
From 2 to 250 mg antibody, and a dose of 5 to 25 mg is more commonly used), but preferably the patient exhibits partial or complete recovery of disease symptoms until disease progression is reduced or stopped Until needed occasionally. Thereafter, the patient can be administered a prophylactic regime.

The therapeutic agent can be administered by parenteral, topical, intravenous, oral, subcutaneous, intraarterial, intracranial, intraperitoneal, intranasal or intramuscular means for prophylactic and / or therapeutic treatment. . The most common route of administration of passive immunogens is intravenous infusion, although other routes can be equally effective. The next most common route is intramuscular injection. This type of injection is most commonly performed in the arm or leg muscles. In some methods, the agent is injected directly into a particular tissue where deposits have accumulated, for example, intracranial injection. Intramuscular injection or intravenous infusion is preferred for administration of the antibody. In some methods, a particular therapeutic antibody is
It is injected directly into the skull. In some methods, the antibody is administered as a sustained release composition or device, such as a Medipad ™ device.

In some cases, the agents of the present invention can be administered in combination with other agents that are at least partially effective in treating amyloidogenic diseases. In the case of Alzheimer's disease and Down's syndrome, where amyloid deposits occur in the brain, the agent of the invention should be administered together with other agents that increase the passage of the agent of the invention across the blood brain barrier. You can also.

Pharmaceutical Compositions Agents of the invention are often administered as a pharmaceutical composition comprising an active therapeutic agent, ie, a pharmaceutical composition that also includes various other pharmaceutically acceptable ingredients. Remington's
Pharmaceutical Science (15th edition, Mack Publishing)
ng Company, Easton, Pennsylvania (1980)). The preferred form depends on the intended mode of administration and therapeutic application. Depending on the desired formulation, the composition can also include a pharmaceutically acceptable non-toxic carrier or diluent, which is commonly used to formulate a pharmaceutical composition for animal or human administration. Defined as vehicle. The diluent is selected so as not to affect the biological activity of the combination.
Examples of such diluents are distilled water, physiological phosphate buffered saline, Ringer's solution, dextrose solution, and Hank's solution. In addition, the pharmaceutical composition or formulation can also include other carriers, adjuvants, or nontoxic, nontherapeutic, nonimmunogenic stabilizers and the like.

Pharmaceutical compositions are large, slowly metabolized macromolecules such as proteins, polysaccharides such as chitosan, polylactic acid, polyglycolic acid and copolymers (latex functional Sepharose ™, agarose, cellulose, etc.), polymeric amino acids, Amino acid copolymers,
And lipid aggregates (such as oil droplets or liposomes) and the like. In addition, these carriers can function as immunostimulatory agents (ie, adjuvants).

For parenteral administration, an agent of the invention is in a physiologically acceptable diluent, including a pharmaceutical carrier, which can be a sterile liquid, such as water oil, saline, glycerol, or ethanol. It can be administered as an injectable dose solution or suspension of the substance. In addition, auxiliary substances such as wetting or emulsifying agents, surfactants, pH buffering substances and the like may be present in the composition. Other ingredients of the pharmaceutical composition are of petroleum, animal, vegetable, or synthetic origin, such as peanut oil, soybean oil, and mineral oil. In general, glycols such as propylene glycol or polyethylene glycol are preferred liquid carriers, particularly for injectable solutions. The antibodies can be administered in the form of depot injections or implant preparations, which can be formulated in such a manner as to allow sustained release of the active ingredient. An exemplary composition is 50 mM L-histidine, 150 mM NaCl.
5 mg / mL formulated in an aqueous buffer consisting of and adjusted to pH 6.0 with HCl
Including monoclonal antibodies.

In general, the compositions are prepared as injections, either as liquid solutions or suspensions. Solid forms suitable for solution or suspension in liquid vehicle prior to injection can also be prepared. The preparation can also be emulsified or encapsulated in liposomes or microparticles, such as polylactides, polyglycolides, or copolymers, etc. (Langer, Science 249) for enhanced adjuvant effect as described above. : 1527
(1990) and Hanes, Advanced Drug Delivery Re
views 28:97 (1997)). The agents of the present invention can be administered in the form of depot injections or implant preparations, which can be formulated in such a manner as to allow sustained or pulsed release of the active ingredient.

Additional formulations suitable for other modes of administration include oral, intranasal, and pulmonary formulations, suppositories, and transdermal applications. For suppositories, binders and carriers include, for example, polyalkylene glycols or triglycerides, and such suppositories contain active ingredient in the range of 0.5% to 10%, preferably 1% to 2%. It can be formed from the mixture it contains. Oral formulations include excipients such as pharmaceutical grade mannitol, lactose,
Contains starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, and the like. These compositions take the form of solutions, suspensions, tablets, pills, capsules, sustained release formulations, or powders, 10% to 95% active ingredient, preferably 25% to 70%.
%.

Topical application can result in transdermal or intradermal delivery. Topical administration can be facilitated by co-administering the agent with cholera toxin, or a detoxified derivative or subunit thereof, or other similar bacterial toxin (Glenn et al., Nature
391, 851 (1998)). Co-administration can be accomplished by using the components as a mixture or as a binding molecule obtained by chemical cross-linking or expression as a fusion protein.

Alternatively, transdermal delivery can be achieved using the skin path or using the transferosome (Paul et al., Eur.
. J. et al. Immunol. 25: 3521 (1995); Cevc et al., Biochem. B
iophys. Acta 1368: 201-15 (1998)).

Monitoring the progress of treatment The present invention provides a method for monitoring treatment in patients suffering from or susceptible to Alzheimer's disease, ie, monitoring the course of treatment being administered to a patient. This method can be used to monitor both therapeutic treatment to symptomatic patients and prophylactic treatment to asymptomatic patients. In particular, this method is useful for monitoring passive immunization (eg, measuring the level of antibody administered).

Some methods include, for example, determining a baseline value of an antibody level or characteristic in a patient, and comparing it with a post-treatment characteristic or level value before administering a dose of the agent, for example. Accompanied by. Significant increase in level or characteristic value (ie,
A positive treatment result (ie, administration of an agent desired), expressed as one standard deviation from the mean of repeated measurements of the same sample, is greater than the general margin of experimental error in such measurements. The response was achieved). If the value for the immune response does not change significantly or decreases, a negative treatment outcome is suggested.

In other methods, level or characteristic control values (ie, mean and standard deviation) are determined for a control population. In general, individuals in the control population have not received prior treatment.
The measured level or characteristic in the patient after administration of the therapeutic agent is then compared to a control value. A significant increase (eg, greater than one standard deviation from the mean value) relative to the control value indicates a positive or satisfactory treatment outcome. A lack or decrease in significant increase indicates a negative or insufficient treatment outcome. Administration of the agent is generally continued while the level is increased relative to the control value. As mentioned above, reaching a plateau compared to the control value indicates that the treatment application can be interrupted or reduced in dosage and / or frequency.

In other methods, level or characteristic control values (eg, mean and standard deviation) are determined from a control population of individuals who have been treated with a therapeutic agent and whose level or characteristic has reached a plateau in response to treatment. The Level or characteristic measurements in patients are compared to control values.
If the measured level in the patient is not significantly different from the control value (eg, greater than one standard deviation), the treatment can be discontinued. If the level in the patient is significantly below the control value, continued administration of the agent is guaranteed. If the level in the patient persists below the control value, a treatment change may be indicated.

In other methods, patients who are not currently receiving treatment, but who have previously undergone the course of treatment, are monitored for antibody levels or characteristics to determine if treatment needs to be resumed. The measured level or characteristic in the patient can be compared to the value previously reached in the patient after the previous course of treatment. A significant decrease compared to previous measurements (ie, greater than the general margin of error in repeated measurements of the same sample) indicates that treatment can be resumed. Alternatively, the value measured in the patient can be compared to control values (mean value and standard deviation) determined in the patient population after undergoing the course of treatment. Or
Measurements in patients are a prophylactically treated population of patients who still have no disease symptoms,
Or it can be compared to a control value in a population of therapeutically treated patients showing recovery of disease characteristics. In all of these cases, a significant decrease (ie, greater than the standard deviation) compared to the control level indicates that the patient should resume treatment.

The tissue sample for analysis is generally blood, plasma, serum, mucus fluid, or cerebrospinal fluid from a patient. The sample is analyzed, for example, for the level or characteristics of antibodies against Aβ peptide, eg, the level or characteristics of humanized antibodies. ELI to detect antibodies specific for Aβ
The SA method is described in the Examples section. In some methods, the level or characteristic of an administered antibody is determined using, for example, in vitro, as described herein, using a clearance assay.
Determined by ro phagocytosis assay. In such a method, a tissue sample from the patient being tested is contacted with amyloid deposits (eg, derived from PDAPP mice) and phagocytic cells having Fc receptors. The subsequent removal of amyloid deposits is then monitored. The presence and extent of the ablation response provides an indication of the presence and level of antibodies effective to eliminate Aβ in the tissue sample of the patient under test.

Antibody properties after passive immunization typically show an immediate peak of antibody concentration followed by an exponential decay. In the absence of further administration, this decay approaches pre-treatment levels within a period of days to months, depending on the half-life of the administered antibody. For example, the half-life of some human antibodies is on the order of 20 days.

In some methods, a baseline measurement of antibodies to Aβ in a patient is made prior to administration, followed immediately by a second measurement to determine peak antibody levels, one or more times at intervals. The antibody level decay is monitored by making further measurements of. If the level of antibody drops to a predetermined percentage of the peak (eg, 50%, 25% or 10%) below baseline or below baseline, an additional dose of antibody is administered. Some methods constitute a beneficial prophylactic or therapeutic treatment regimen in other patients by comparing subsequent measurement levels below the peak or background to previously determined reference levels. If the measured antibody level is significantly lower than the reference level (eg, less than one standard deviation minus the mean of the reference values in the population of patients benefiting from treatment) Administration is indicated.

Additional methods include amyloidogenicity by monitoring any physiological symptoms (eg, physical or psychological symptoms) recognized in the technical field periodically during the course of treatment, relying on a researcher or physician. Diagnosing or monitoring a sex disorder (eg, Alzheimer's disease). For example, cognitive impairment can be monitored. The latter is a symptom of Alzheimer's disease and Down's syndrome, but can occur without the other features of any of these diseases. For example, cognitive impairment can be monitored by determining a patient's score in a customary mini-mental state test throughout the course of treatment.

Patients have a Mini-Mental State Test (MMSE), Alzheimer's Disease Rating Scale-Cognition (
ADAS-COG), Clinician interview impression (CIBl), Neurological test battery (NTB), Dementia disability assessment (DAD), Clinical dementia assessment-Box total (CDR-
At least one selected from the group consisting of SOB), neuropsychiatric examination slip (NPI), positron emission tomography (PET imaging) scanning, magnetic resonance imaging (MRI) scanning, EKG and blood pressure measurement Can be monitored by evaluation. This type of evaluation can be made on multiple occasions. For example, MMSE should be performed before administering a dose of immunogen and at 4, 6, 16, 6 months, and 1 year after administering that dose of immunogen. Can do. In some patients, MMSE may be performed before administering a single dose of the immunogenic agent, and at weeks 6 and 16. MRI scans can be performed every 3 months, every 6 months, or annually.

Patients can be monitored for posterior reversible encephalopathy syndrome (PRES) or angioedema after administering an antibody in the range of about 0.5 mg / kg to less than 5 mg / kg, the antibody comprising at least 10 It specifically binds to β-amyloid peptide (Aβ) with a binding affinity of ⁷ M- ¹ . PRES classically consists of reversible angiogenic edema in the posterior circulation region, but a conversion to irreversible cytotoxic edema has been described. PRES is typically characterized by headache, nausea, vomiting, confusion, seizures, visual abnormalities, changes in mental function, ataxia, frontal symptoms, parietal symptoms, stupor, and local neurological signs. In addition to the aforementioned clinical symptoms, the presence of PRES can be shown by using MRI scanning or water-suppressed reversal recovery (FLAIR) sequence imaging (Pediatric Neurology, 20 (3): 24
1-243; AJNR, 26: 825-830; NEJM, 334 (8): 494-50
0; Pediatr Nephrol, 18: 1161-1166; Internal
Medicine Journal, 35: 83-90; JNNP, 68: 790-79.
1; AJNR, 23: 1038-1048; Pak J Med Sci, 21 (2):
149-154, and AJNR, 21: 1199-1209).

Patients can be monitored for PRES or angioedema every month, every 4 months, every 6 months, or yearly. Patients can be monitored for at least one clinical symptom associated with PRES or angioedema. This monitoring may include performing an MRI scan. This monitoring may further include performing a FLAIR sequence imaging method. The results of monitoring can affect the dosing method. For example, if PRES or angioedema is detected, dosing can be suspended, the dosage can be reduced, or the interval between doses can be increased.

Patients with PRES or angioedema can have their blood pressure measured for hypertension. If hypertension is detected in a patient, the patient can treat hypertension by administering an antihypertensive drug. The antihypertensive agent can be selected from the group consisting of hydrochlorothiazide, angiotensin converting enzyme (ACE) inhibitor, angiotensin II receptor blocker (ARB), beta blocker, and calcium channel blocker. Patients with PRES or angioedema can be treated with steroids such as dexamethasone or methyprednisol.

C. Kits The present invention further provides therapeutic products. This product includes a glass vial and instructions for use. The glass vial contains about 10 mg to about 250 mg of humanized anti-Aβ antibody, about 4%
Mannitol or about 150 mM NaCl, about 5 mM to about 10 mM histidine,
And a formulation containing about 10 mM methionine. Instructions for monitoring patients to whom the formulation is administered for PRES and / or angioedema are included in the product. In some therapeutic products, glass vials contain about 10 mg of humanized anti-Aβ antibody, about 10 mM methionine, about 4% mannitol, and about 0.00% in about 10 mM histidine at a pH of about 6.0. Contains a formulation containing 005% polysorbate-80 (plant-derived). Instructions for monitoring patients to whom the formulation is administered for PRES and / or angioedema are included in the product.

Example I
Prevention and Treatment of Human Subjects Bapineuzumab (AAB-001) is a humanized monoclonal antibody against Aβ. The purpose of this study is to determine the safety and tolerability of a single dose of bapineuzumab in AD.

Method: Randomization of bapineuzumab infusion in patients with mild to moderate AD,
Double-blind, placebo-controlled single incremental dose trial. Patients enrolled in this trial met all of the following criteria:
1. National Institute of Neurological an
d Communicative Disorders and Stroke-Alz
heimer's disease and Related Disorders (N
INCDS-ADRDA) Diagnosis presumed to be Alzheimer's disease (AD) by diagnostic criteria.
2. Age between 50 and 87 years (inclusive).
3. Mini-mental state test (MMSE) score of 14-26.
Less than 4.4 Lowsen improved Hachinsky ischemic score.
5). Live at home with an appropriate caregiver who can accompany the patient at all hospital visits, or can accompany the patient at all hospital visits, approximately 5 per week for the duration of the study Living in a community living with a caregiver who can visit a patient.
6). A visitor's magnetic resonance imaging (MRI) scan that is consistent with the diagnosis of AD, ie other that may belong to other diseases (eg, stroke, traumatic brain injury, arachnoid cyst, tumor, etc.) Screening for scans where no anomalies are present on MRI.
7). Surgically sterile or 2 years after menopause.
8). Evidence of English fluency and sufficient intellectual function before onset. Patients must have sufficient visual and auditory capabilities to perform all aspects of cognitive and functional assessment.
9. Receiving stable doses of drug therapy (s) for treatment of non-exclusive medical condition (s) for at least 30 days prior to screening. If the patient is taking an acetylcholinesterase inhibitor or memantine, these drug therapy (s) must be maintained at a stable dosage regimen for at least 60 days prior to the screening evaluation.
Patients with any of the following criteria were excluded from enrollment in trials.
1. Significant neurological disease other than AD that can affect cognition.
2. Diagnostic and Statistical Manual of
Mental Disorders, 4th edition (DSM-IV) criteria, clinically significant major psychiatric disorders (eg major depressive disorder) or symptoms that may affect the patient's ability to complete the study ( For example, hallucinations) currently exist.
3. Current clinically significant systemic illness that is prone to worsening of the patient's condition or impacting patient safety during the study.
4). Encephalitis; clinically evident stroke; clinically significant carotid or vertebral base stenosis or plaque; seizures excluding childhood febrile seizures; any clinically significant autoimmune disease or immune system Disability; history or evidence of any clinically significant renal impairment.
5). Clinically significant infection (eg, chronic persistent or acute infection) in the last 30 days.
6). Immunosuppressive pharmacotherapy within the last 90 days (eg systemic corticosteroids) (inhalation of local and nasal corticosteroids and corticosteroids for asthma is acceptable) or within the last 3 years Treatment with chemotherapeutic agents for malignant tumors.
7). Myocardial infarction within the last 2 years.
8). Cancer history within the last 5 years, excluding non-metastatic basal cell carcinoma and squamous cell carcinoma of the skin.
9. Other clinically significant abnormalities (eg, atrial fibrillation) in physical, neurological, clinical or ECG tests that could harm the study or could be harmful to the patient ).
Hemoglobin less than 10.11 g / dL.
11. History of alcohol or drug addiction or abuse within the last 2 years.
12.12 Hamilton Depression Psychiatric Rating Scale (Hamilton psychiatric rating sc
ale for depression) (HAM-D) (17 items) score.
13. Current use of anticonvulsants, antiparkinsonian drugs, anticoagulants (excluding use of 325 mg / day or less aspirin), or narcotic therapy for seizures.
14 Current use of prescription or non-prescription drugs for cognitive enhancement other than cholinesterase inhibitors and memantine. The current use of cholinesterase inhibitors and memantine is prohibited unless the following conditions are met: (a) Maintain a stable dosage regimen for at least 60 days prior to screening; (b) There are no clinically significant side effects that can be attributed to the drug; and (C) Except for unforeseen circumstances, the patient and caregiver agree to continue the same regimen for the duration of the trial.
15. Any other medications with the potential to affect cognition other than those described in # 18 (but not limited to) unless screened at a stable dosage regimen for at least 30 days prior to screening Anxiolytics, sedatives, hypnotics, antipsychotics, antidepressants,
Commercially available (OTC) sleep aids, sedative antiallergic drugs, vitamin E, thyroid supplements,
And vitamin B12 supplement by injection).
16. Patients who have discontinued cholinesterase inhibitors, memantine, cognitive enhancers, or drugs that potentially affect cognition within 60 days prior to screening.
17. 30 days prior to screening of experimental drug therapy (compound) or appliance for AD, or any other investigational drug therapy or appliance for indicators other than treatment of AD,
Or use within the 5 half-life of the use of such medication, whichever is longer.
18. Any prior experimental treatment with AN1792, AAB-001, ACC-001, or other experimental immunotherapy or vaccine for AD.
19. Optional pre-treatment with biological products other than those mentioned in # 18 for the treatment of AD within the last 3 years.
20. Patients who donated blood (regular blood donation) within 90 days before screening.
21. Any known hypersensitivity to any excipients included in the study drug formulation.
22. Presence of pacemakers, aneurysm clips, prosthetic heart valves, ear implants, eyes, skin, or foreign metal pieces in the body that may be contraindicated in brain MRI scans.
Body weight greater than 23.120 kg (264 lb).

Results: 30 patients with 0.5 mg / kg (6 active, 2 placebo), 1
. Bapineuzumab was administered at doses of 5 mg / kg (6 active, 2 placebo) and 5 mg / kg (10 active, 4 placebo). 5 mg / kg 3
/ 10 patients with MRI mainly due to high signal abnormalities in FLAIR sequence
Abnormalities occurred and the study did not continue beyond that dose. In two patients, these abnormalities were seen in regular surveillance scans without clinical symptoms, while the third patient experienced increased confusion. MRI FLAIR abnormalities resolved in 12 weeks after administration in all three cases. As part of the safety assessment, the MMSE is baseline, week 4,
Conducted at 6 weeks, 6 months and 1 year. FIG. 2 shows the average MMSE change from baseline. Figure 3 shows the change from baseline MMSE by cohort at 4 months. At 16 weeks, the pre-designated main time point for the analysis, treatment differences compared to placebo favored treatment groups at a dose of 0.5 mg / kg (2.0 treatment vs. placebo difference). , P = .152), statistical significance was reached at a dose of 1.5 mg / kg (2.5 treatment vs. placebo difference, p = 0.047). There was no significant difference in MMSE changes compared to placebo for the 5.0 mg / kg group. FIG. 4 shows the mean, median and standard deviation of MMSE change from baseline at 4 months. FIG. 5 shows the statistical test results of MMSE change from baseline at 4 months. No correlation was found between MRI FLAIR abnormalities and differences in MMSE changes.

Plasma Aβ rose from baseline levels in a dose-dependent manner and peaked approximately 24 hours after infusion. Pharmacokinetic analysis showed a half-life of 22-28 days and supported a 13-week dosing interval in multiple dosing.

Conclusion: In this small study, MM with a dose of 1.5 mg / kg bapineuzumab
SE was statistically significantly improved compared to placebo. The highest single infusion dose of 5 mg / kg was accompanied by MRI FLAIR abnormalities, which disappeared.

Although the foregoing invention has been described in detail for purposes of clarity of understanding, it will be apparent that certain modifications may be practiced within the scope of the appended claims. All publications and patent literature cited in this specification, as well as the text listed in the figures, are in whole for all purposes, as if individually indicated as such. , Which is incorporated herein by reference.

Example II
Pharmacokinetic Study of AAB-001 The purpose of this study is to determine human pharmacokinetics (PK) after intravenous administration of AAB-001.

Methods: Randomized, double-blind, placebo-controlled, multiple incremental dose trial of AAB-001 administered intravenously. Six doses of AAB-001 were administered intravenously once every 13 weeks. 0.1
There were four dose cohorts that were 5, 0.5, 1.0, and 2.0 mg / kg.

Results: PK was predictable and dose independent. PK-CL 0.05-0.06 mL / h / kg across all dose levels. Dose proportional exposure. At all dose levels, it is a pre-infusion concentration that can be quantified, but very little accumulation with intravenous dosing once every 13 weeks. t1 / 2 ranged from 21 to 34 hours. At steady state, about 3.3 μg / m
L, Cavg after 0.5 mg / kg AAB-001 (ie about 35 mg). FIG.
Please refer to. About 3.3 μg / mL Cav after 0.5 mg / kg AAB-001
g is close to a concentration of 3.7 μg / mL found to be effective in PDAPP mice. FIG. 10 shows AAB-0 at doses of 0.15, 0.5, 1.0, and 2.0 mg / kg.
2 shows mean serum AAB-001 concentration versus time characteristics after intravenous administration of 01.

Plasma Aβ concentrations tend to reflect AAB-001 concentrations. Plasma Aβ levels above the baseline level and above the placebo group at a dose level of AAB-001 of 0.5 mg / kg or higher at the end of the 13 week dosing interval. Please refer to FIG. t
The max ranged from 14 to 48 hours.

Serum anti-AAB-001 antibody levels were undetectable in any sample (0
. (Up to # 6 before injection for a cohort of 5 mg / kg AAB-001).

From the foregoing, it will be apparent that the invention provides several uses. For example, the present invention relates to the use of any of the above-mentioned antibodies against Aβ in the treatment, prevention or diagnosis of amyloidogenic diseases, or the manufacture of a drug or diagnostic composition for use in the treatment, prevention or diagnosis of amyloidogenic diseases. In any of the above mentioned antibodies against Aβ.

Claims (115)

A method of therapeutically treating Alzheimer's disease, comprising administering to a patient suffering from said disease a dose of antibody in the range of about 0.5 mg / kg to less than 5 mg / kg by intravenous infusion. Wherein the antibody specifically binds to β-amyloid peptide (Aβ) with a binding affinity of at least 10 ⁷ M ⁻¹ , thereby treating the patient therapeutically. A method of therapeutically treating Alzheimer's disease, comprising administering to a patient suffering from said disease a dose of antibody in the range of about 0.5 mg / kg to less than 5 mg / kg by intravenous infusion. Wherein the antibody specifically binds to an N-terminal fragment of β-amyloid peptide (Aβ) with a binding affinity of at least 10 ⁷ M ⁻¹ , thereby treating the patient therapeutically. A method for therapeutic treatment of Alzheimer's disease, which specifically binds to an N-terminal fragment of β-amyloid peptide (Aβ) with a binding affinity of at least 10 ⁷ M ⁻¹ to a patient suffering from said disease Administering one dose by intravenous infusion and monitoring said patient for posterior reversible encephalopathy syndrome (PRES) or angioedema. A method of therapeutically treating Alzheimer's disease, comprising administering to a patient suffering from said disease a dose of antibody in the range of about 0.5 mg / kg to less than 5 mg / kg by intravenous infusion. The antibody specifically binds to an N-terminal fragment of β-amyloid peptide (Aβ) with a binding affinity of at least 10 ⁷ M ⁻¹ , posterior reversible encephalopathy syndrome (PRES) or angioedema Monitoring said patient for.   5. The method of claim 1, 2, 3 or 4 wherein the antibody is a humanized antibody. 6. The method of claim 5, wherein the humanized antibody is a humanized form of mouse antibody 3D6 expressed by a hybridoma deposited with the ATCC under number PTA-5130.   7. The method of claim 6, wherein the humanized antibody is bapineuzumab. 5. The method of claims 1-4, wherein the humanized antibody is a humanized form of murine antibody 10D5 expressed by a hybridoma deposited with the ATCC under the number PTA-5129. 6. The method of claim 5, wherein the humanized antibody is a humanized form of murine antibody 12A11 expressed by a hybridoma deposited with the ATCC under the number PTA-7271. 10. The method according to any of claims 1 to 9, wherein the dose is about 0.5 to 3 mg / kg. 10. The method according to any of claims 1 to 9, wherein the dose is about 0.5 to 1.5 mg / kg.   10. The method according to any one of claims 1 to 9, wherein the dosage is about 0.5 mg / kg.   10. The method according to any one of claims 1 to 9, wherein the dosage is about 1.5 mg / kg.   10. A method according to any preceding claim, wherein the dosage is administered on multiple occasions.   15. The method of claim 14, wherein the dose is administered every 8 to 16 weeks.   16. The method of claim 15, wherein the dose is administered every 10 to 14 weeks.   17. The method of claim 16, wherein the dosage is administered every 13 weeks. Mini-mental state test (MMSE), Alzheimer's disease rating scale-cognition (ADAS)
-COG), Clinician interview impression (CIBI), Neurological test battery (NT
B), Dementia disability assessment (DAD), Clinical dementia assessment-Box total (CDR-SOB)
By at least one evaluation selected from the group consisting of: neuropsychiatric examination sheet (NPI), positron emission tomography (PET imaging) scanning, magnetic resonance imaging (MRI) scanning, EKG and blood pressure measurement 5. The method of claim 1, 2, 3 or 4 further comprising the step of monitoring the patient. 19. The method of claim 18, wherein the type of evaluation is Alzheimer's Disease Rating Scale-Cognition (ADAS-COG).   The method of claim 19, wherein the ADAS-COG is applied on multiple occasions. The method of claim 18, wherein the evaluation type is a neurological test battery (NTB).   The method of claim 21, wherein the NTB is applied on multiple occasions.   The method of claim 18, wherein the evaluation type is MMSE.   24. The method of claim 23, wherein the MMSE is applied on multiple occasions. Before the MMSE administers the dose, as well as 4 weeks after the administration of the dose,
25. The method of claim 24, performed after 16 weeks, 6 months, and 1 year. 25. The method of claim 24, wherein the MMSE score measured after administration is higher than the previously assessed MMSE score. The monitoring step comprises performing an MRI scan.
The method described in 1. 28. The method of claim 27, wherein the monitoring step further comprises performing a FLAIR (Water Suppression Inversion Recovery Method) sequence imaging method. 28. The method of claim 27, wherein the monitoring step comprises identifying at least one clinical symptom associated with PRES or angioedema. The at least one clinical symptom is selected from the group consisting of headache, nausea, vomiting, confusion, seizures, visual abnormalities, changes in mental function, ataxia, frontal symptoms, parietal symptoms, stupor, and local neurological signs; 30. The method of claim 29. 28. The method of claim 27, further comprising reducing or suspending the dosage based on the results of an MRI scan indicating PRES or angioedema. 29. The method of claim 28, further comprising reducing or suspending the dosage based on FLAIR sequence imaging results indicating PRES or angioedema. 32. The method of claim 30, further comprising reducing or suspending the dosage based on the identification of at least one clinical symptom associated with PRES or angioedema. 28. The method of claim 27, wherein the MRI scan is every 3 months, every 6 months, or every year. 29. The method of claim 28, wherein the FLAIR sequence imaging method is every 3 months, every 6 months, or every year. 34. The method of any one of claims 31 to 33, further comprising determining the presence or absence of hypertension in the patient, and further comprising administering an antihypertensive drug if the patient has hypertension. . 37. The antihypertensive agent is selected from the group consisting of hydrochlorothiazide, angiotensin converting enzyme (ACE) inhibitor, angiotensin II receptor blocker (ARB), beta blocker, and calcium channel blocker. the method of. 34. The method of any one of claims 31 to 33, further comprising treating PRES or angioedema by administering a steroid to the patient.   40. The method of claim 38, wherein the steroid is dexamethasone.   40. The method of claim 38, wherein the steroid is methylprednisolone. MRI scan results showing PRES and identification of at least one clinical symptom related to PRES, or MRI scan results showing angioedema and at least one related to angioedema
28. The method of claim 27, further comprising reducing or suspending the dosage based on the identification of one clinical symptom. The at least one clinical symptom is selected from the group consisting of headache, nausea, vomiting, confusion, seizures, visual abnormalities, changes in mental function, ataxia, frontal symptoms, parietal symptoms, stupor, and local neurological signs; 42. The method of claim 41. To identify the results of FLAIR sequencing imaging showing PRES and at least one clinical symptom related to PRES, or the results of FLAIR sequencing to show angioedema and at least one clinical symptom related to angioedema 29. The method of claim 28, further comprising reducing or suspending the dosage based on. The clinical symptoms are selected from the group consisting of headache, nausea, vomiting, confusion, seizures, visual abnormalities, changes in mental function, ataxia, frontal lobe symptoms, parietal symptoms, stupor, and local neurological signs;
44. The method of claim 43. The monitoring step indicates the presence of PRES or angioedema at a first time point after administration and the absence of PRES or angioedema at a second time point after the first time point. Before the step of monitoring indicates that PRES or angioedema is present, the patient is administered a first dose and the step of monitoring presents PRES or angioedema A second dose is administered or not administered, and the monitoring step detects that no PRES or angioedema is present, and then the third dose is 5. The method of claim 3 or 4, wherein the first and third doses are administered and greater than the second dose.   46. The method of claim 45, wherein the first and third doses are the same.   The method according to claim 3 or 4, wherein the antibody is a humanized antibody. 5. The method of claim 3 or 4, wherein the humanized antibody is a humanized form of mouse antibody 3D6 expressed by a hybridoma deposited with the ATCC under the number PTA-5130.   49. The method of claim 48, wherein the humanized antibody is bapineuzumab. The antibody is administered at a first dose before PRES or angioedema is determined from the MRI scan, and is administered at a second dose after PRES or angioedema is determined from the MRI scan. The method of claim 3 or 4, wherein the second dose is less than the first dose. The first dose is 3-5 mg / kg and the second dose is 0.5-3 mg / kg.
51. The method of claim 50, wherein the weight is kg.   52. The method of claim 51, wherein the second dose is half of the first dose. The bapineuzumab is administered at a first dose before PRES or angioedema is determined from the MRI scan, and at a second dose after PRES or angioedema is determined from the MRI scan. Wherein the second dose is less than the first dose,
50. The method of claim 49. The first dose is 3-5 mg / kg and the second dose is 0.5-3 mg / kg.
54. The method of claim 53, wherein the method is kg.   55. The method of claim 54, wherein the second dose is half of the first dose. a. i. From about 10 mg to about 250 mg of a humanized anti-Aβ antibody,
ii. About 4% mannitol or about 150 mM NaCl,
iii. From about 5 mM to about 10 mM histidine, and iv. A glass vial containing a formulation comprising about 10 mM methionine, and b. A kit for the treatment of Alzheimer's disease comprising instructions for monitoring patients to whom the formulation is administered for PRES and / or angioedema. A method for treating Alzheimer's disease, comprising:
For patients with the disease, an antibody that specifically binds to the N-terminal fragment of Aβ is sufficient to maintain an average serum concentration of the antibody in the patient in the range of 1-15 μg antibody per ml of serum. Administering in a regimen and thereby treating said patient. 6. The mean serum concentration is in the range of 1-10 [mu] g antibody per ml of serum.
8. The method according to 7. 58. The average serum concentration is in the range of 1-5 [mu] g antibody per ml of serum.
The method described in 1. 58. The average serum concentration is in the range of 2-4 [mu] g antibody per ml of serum.
The method described in 1.   58. The method of claim 57, wherein the antibody is administered intravenously.   62. The method of claim 61, wherein a dose of 0.1 to 1.0 mg / kg is administered monthly.   62. The method of claim 61, wherein a dose of 0.5-1.0 mg / kg is administered monthly.   58. The method of claim 57, wherein the antibody is administered subcutaneously.   58. The method of claim 57, wherein the antibody is administered at a frequency between weekly and monthly.   58. The method of claim 57, wherein the antibody is administered weekly or biweekly. 58. The method of claim 57, wherein the antibody is administered at a dose of 0.01 to 0.35 mg / kg. 58. The method of claim 57, wherein the antibody is administered at a dose of 0.05 to 0.25 mg / kg. 58. The method of claim 57, wherein the antibody is administered from weekly to biweekly at a dose of 0.015-0.2 mg / kg. 58. The method of claim 57, wherein the antibody is administered weekly to every other week at a dose of 0.05 to 0.15 mg / kg. 58. The method of claim 57, wherein the antibody is administered weekly at a dose of 0.05 to 0.07 mg / kg.   58. The method of claim 57, wherein the antibody is administered weekly at a dose of 0.06 mg / kg. 58. The method of claim 57, wherein the antibody is administered every other week at a dose of 0.1 to 0.15 mg / kg. 58. The method of claim 57, wherein the average serum concentration of the antibody is maintained for at least 6 months. 58. The method of claim 57, wherein the average serum concentration of the antibody is maintained for at least 1 year. 58. The method of claim 57, further comprising measuring the concentration of antibody in the serum and adjusting the regimen if the measured concentration is outside the range. A method for treating Alzheimer's disease, comprising:
Administering to the patient having the disease subcutaneously an antibody that specifically binds to the N-terminal fragment of Aβ, wherein the antibody is administered at a dose of 0.01-0.6 mg / kg and weekly and monthly. Comprising the step of being administered at a frequency of between. 78. The method of claim 77, wherein the antibody is administered at a dose of 0.05 to 0.5 mg / kg. 78. The method of claim 77, wherein the antibody is administered at a dose of 0.05 to 0.25 mg / kg. 78. The method of claim 77, wherein the antibody is administered from weekly to biweekly at a dose of 0.015-0.2 mg / kg. 78. The method of claim 77, wherein the antibody is administered weekly to every other week at a dose of 0.05 to 0.15 mg / kg. 78. The method of claim 77, wherein the antibody is administered weekly at a dose of 0.05 to 0.07 mg / kg.   78. The method of claim 77, wherein the antibody is administered weekly at a dose of 0.06 mg / kg. 78. The method of claim 77, wherein the antibody is administered every other week at a dose of 0.1 to 0.15 mg / kg. 78. The method of claim 77, wherein the antibody is administered monthly at a dose of 0.1 to 0.3 mg / kg.   78. The method of claim 77, wherein the antibody is administered monthly at a dose of 0.2 mg / kg. A method for treating Alzheimer's disease, comprising:
Administering to the patient having the disease subcutaneously an antibody that specifically binds to an N-terminal fragment of Aβ, wherein the antibody is administered at a dose of 1-40 mg and at a frequency between weekly and monthly. Including the steps of:   90. The method of claim 87, wherein the antibody is administered at a dose of 5-25 mg.   90. The method of claim 87, wherein the antibody is administered at a dose of 2.5-15 mg. 88. The method of claim 87, wherein the antibody is administered weekly to biweekly at a dose of 1-12 mg. 90. The method of claim 87, wherein the antibody is administered weekly to biweekly at a dose of 2.5-10 mg.   90. The method of claim 87, wherein the antibody is administered weekly at a dose of 2.5-5 mg.   90. The method of claim 87, wherein the antibody is administered weekly at a dose of 4-5 mg.   88. The method of claim 87, wherein the antibody is administered every other week at a dose of 7-10 mg. A method for treating Alzheimer's disease, comprising:
Administering to a patient with the disease an antibody that specifically binds to the N-terminal fragment of Aβ in a regimen sufficient to maintain a maximum serum concentration of the antibody in the patient of less than about 28 μg antibody per ml of serum. And thereby treating the patient. 96. The method of claim 95, wherein said maximum serum concentration is in the range of about 4-28 [mu] g antibody per ml of serum. 99. The method of claim 96, wherein said maximum serum concentration is in the range of about 4-18 [mu] g antibody per ml of serum. 98. The method of any one of claims 95 to 97, wherein the mean serum concentration of the antibody in the patient is less than about 7 [mu] g antibody per ml serum. 10. The average serum concentration is in the range of about 2-7 [mu] g antibody per ml of serum.
9. The method according to 8. 99. The method of claim 99, wherein the average serum concentration is about 5 [mu] g of antibody per ml of serum. A method for treating Alzheimer's disease, comprising:
Administering to the patient with the disease an antibody that specifically binds to the N-terminal fragment of Aβ in a regimen sufficient to maintain an average serum concentration of the antibody in the patient of less than about 7 μg of antibody per ml of serum. And thereby treating the patient. 2. The average serum concentration is in the range of about 2-7 [mu] g antibody per ml of serum.
The method according to 01. 103. The method of claim 102, wherein the average serum concentration is about 5 [mu] g antibody per ml of serum.   104. The method of any one of claims 95 to 103, wherein the antibody is administered intravenously.   104. The method of any one of claims 95 to 103, wherein the antibody is administered subcutaneously. 106. The method according to any one of claims 95 to 105, wherein a dose of 0.1 to 1.0 mg / kg is administered monthly. 106. The method according to any one of claims 95 to 105, wherein a dose of 0.5 to 1.0 mg / kg is administered monthly.   108. The method of claim 107, wherein the antibody is administered at a frequency between weekly and monthly.   109. The method of claim 108, wherein the antibody is administered weekly or biweekly. 99. The method further comprising: measuring the concentration of antibody in the serum; and adjusting the regimen if the measured concentration is outside the range.
3. The method according to any one of 2.   108. The method of claim 95 or 107, wherein the antibody is a humanized antibody. 112. The method of claim 111, wherein said humanized antibody is a humanized form of murine antibody 3D6 expressed by a hybridoma deposited with the ATCC under number PTA-5130.   113. The method of claim 112, wherein the humanized antibody is bapineuzumab. 111. The method of claim 111, wherein the humanized antibody is a humanized form of murine antibody 10D5 expressed by a hybridoma deposited with the ATCC under number PTA-5129. 111. The method of claim 111, wherein the humanized antibody is a humanized form of murine antibody 12A11 expressed by a hybridoma deposited with the ATCC under the number PTA-7271.

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