Methods for safe administration of tau phosphopeptide conjugates

Abstract

A method for inducing anti-phosphorylated tau antibodies in humans without inducing serious adverse events is described, comprising administering to a subject an effective amount of a composition containing a tau phosphopeptide conjugated to an immunogenic carrier.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application is an international application claiming priority to U.S. Provisional Patent Application No. 63 / 261,793, filed September 29, 2021, the disclosure of which is incorporated herein by reference in its entirety.

[0002] Electronic Sequence Listing Reference The contents of the electronic sequence listing (065794_8WO1.xml; size: 35,700 bytes; and creation date: August 29, 2022) are incorporated herein by reference in their entirety.

[0003] FIELD OF THEINVENTION The present application is in the field of medicine. In particular, the present application relates to a method for inducing an immune response against tau protein in a subject suffering from a neurodegenerative disease, disorder or condition, using a composition comprising a phosphorylated tau (p-tau) peptide conjugated to an immunogenic carrier.

[0004] background Alzheimer's disease (AD) is a progressive, debilitating neurodegenerative disorder that affects an estimated 44 million people worldwide (Alzheimers.net). Current commercialized AD therapies aim to affect clinical symptoms but do not target the pathogenic processes underlying the disease (disease-modifying effects). Unfortunately, current therapies are only minimally effective, and therefore there is an urgent need to develop and test additional preventive and therapeutic measures.

[0005] The pathology characteristic of Alzheimer's disease is the accumulation of extracellular plaques containing grossly aggregated amyloid beta protein, and intracellular "tangles" or aggregates of hyperphosphorylated tau protein. The molecular events that cause the accumulation of these proteins are poorly characterized. With regard to amyloid, it has been hypothesized that abnormal cleavage of the amyloid precursor protein leads to the accumulation of an aggregation-prone fragment containing amino acids 1-42. With regard to tau, it has been hypothesized that dysregulation of kinases, phosphatases, or both, leads to the abnormal phosphorylation of tau. When tau becomes hyperphosphorylated, it loses its ability to effectively bind and stabilize microtubules, and instead accumulates in the cytoplasm of affected neurons. Unbound hyperphosphorylated tau appears to form first oligomers and then higher order aggregates, the presence of which appears to negatively affect the function of the neurons in which they are formed, possibly via interruption of normal axonal transport.

[0006] In developed countries, individuals diagnosed with Alzheimer's disease or other dementing tauopathies are commonly treated with cholinesterase inhibitors (e.g., Aricept®) or memantine (e.g., Namenda™). These drugs are reasonably well tolerated but have very modest efficacy. For example, Aricept® delays the worsening of symptoms by 6-12 months in approximately 50% of treated individuals. The remaining treatments are non-pharmacological and focus on enabling patients to better handle everyday tasks while their cognitive abilities decline.

[0007] Immunotherapies for the prevention and treatment of AD are currently under development. Active immunization with antigens associated with AD can potentially stimulate both antibody- and cell-mediated immune responses against AD. However, evaluation of the first widely tested human anti-amyloid beta vaccine was halted in 2002. Meningoencephalitis, a potentially fatal form of central nervous system inflammation, was observed in clinical studies of AN-1792, an active immunotherapy targeting Aβ, in AD patients (Orgogozo et al., 2003). The encephalitic reaction, which occurred in 6% of patients exposed to AN-1792, is thought to be induced by unwanted Aβ-specific T cell activation.

[0008] To date, very few studies have been performed with agents that specifically target tau pathology. Currently, tau immunotherapy is moving into clinical trials, but the field is still in its infancy and a mechanistic understanding of the efficacy and safety of the various approaches is not well established (Sigurdsson, Neurodegener Dis. 2016; 16(0): 34-38). Encephalitis, i.e., brain inflammation, has also been reported in mice immunized against full-length tau protein. However, no adverse effects were reported from animals immunized with a single injection of phosphorylated tau peptide in a CNS proinflammatory environment (Rosenmann H., 2013. Curr. Alzheimer Res. 10, 217-228).

[0009] The long-term safety profile of a non-tau phosphopeptide vaccine (AADvac1) in human patients with mild to moderate Alzheimer's disease has been published (Novak et al., Alzheimer's Research & Therapy (2018) 10:108). The vaccine contains a synthetic peptide derived from amino acids 294-305 of the tau sequence coupled to keyhole limpet hemocyanin (KLH) via an N-terminal cysteine. The vaccine was administered with aluminum hydroxide adjuvant (containing 0.5 mg of Al3+) in a dose of 40 μg of peptide (CKDNIKHVPGGGS, SEQ ID NO: 13) coupled to KLH in a volume of 0.3 ml phosphate buffer. Adverse events (AEs) observed from the 26 patients enrolled in the study and associated with AADvac1 treatment in the Phase 1 study (FUNDAMANT study) were injection site reactions (erythema, swelling, warmth, pruritus, pain, nodules). One or more of these AEs were observed in 50% of AADvac1-treated patients. Injection site reactions were reversible and mostly mild in manifestation. Six serious adverse events (SAEs) were observed (abdominal strangulation hernia, dehydration, acute psychosis, behavioral and psychiatric symptoms of dementia, second degree atrioventricular block, and sinus bradycardia). None of the SAEs were deemed by the investigators to be related to AADvac1 treatment. No allergic or anaphylactic reactions were observed. No safety signals emerged in clinical laboratory evaluations (coagulation, blood biochemistry, hematology, and urinalysis), in vital sign evaluations, or in neurological and physical examinations. No safety signals were detected by MRI evaluation. No edematous changes occurred. Meningeal changes and meningoencephalitis were not observed. New microhemorrhages were observed in one ApoE4 homozygote and brain surface hemosiderin was detected in one ApoE4 heterozygote, both events being clinically silent and considered consistent with the background incidence of such lesions in the AD patient population.

[0010] However, the safety profile of tau phosphopeptide conjugates in human patients has not been reported. There is a need for safe and effective treatments for neuronal degenerative diseases such as Alzheimer's disease. Summary of the Invention

[0011] In one general aspect, the present application provides a method of inducing antibodies against at least one of tau, preferably phosphorylated tau and enriched paired helical filaments (ePHF), in a human subject in need thereof, comprising administering to the human subject a composition comprising 5 μg to 200 μg per dose of a conjugate, the conjugate being represented by formula (I):

[0012] [ka] or having the structure of formula (II):

[0013] [ka] The structure is (In the formula, x is an integer from 0 to 10, preferably from 2 to 6, and most preferably 3; n is an integer of 3 to 15, preferably 3 to 12. the carrier represents an immunogenic carrier selected from the group consisting of keyhole limpet hemocyanin (KLH), tetanus toxoid, CRM197 and outer membrane protein mixture (OMP) from Neisseria meningitidis, or derivatives thereof; The tau peptide refers to a tau phosphopeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1 to SEQ ID NO:3 and SEQ ID NO:5 to SEQ ID NO:12.

[0014] In one general aspect, the present application provides a composition for inducing antibodies against tau, preferably phosphorylated tau and / or enriched paired helical filaments (ePHF), in a human subject in need thereof, comprising 5 μg to 200 μg of a conjugate per dose, the conjugate having a structure represented by formula (I):

[0015] [ka] or having the structure of formula (II):

[0016] [ka] The structure is (In the formula, x is an integer from 0 to 10, preferably from 2 to 6, and most preferably 3; n is an integer of 3 to 15, preferably 3 to 12. The carrier represents an immunogenic carrier selected from the group consisting of keyhole limpet hemocyanin (KLH), tetanus toxoid, CRM197 and outer membrane protein mixture (OMP) from Neisseria meningitidis, or derivatives thereof; The tau peptide relates to a composition that represents a tau phosphopeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1 to SEQ ID NO:3 and SEQ ID NO:5 to SEQ ID NO:12.

[0017] In one embodiment, the composition further comprises a pharma- ceutically acceptable carrier.

[0018] In one embodiment of the present application, the conjugate comprises a tau phosphopeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1 to SEQ ID NO:3 or SEQ ID NO:5 to SEQ ID NO:12 conjugated to CRM197 via a linker. Preferably, the tau peptide is a tau phosphopeptide having the amino acid sequence of SEQ ID NO:2. Multiple tau phosphopeptides, for example 2, 3, 4, 5, 6, 7, 8, 9, 10 or more tau phosphopeptides, can be conjugated to one carrier protein. More preferably, the conjugate comprises

[0019] [ka] (In the formula, n is an integer from 3 to 7, and VYKS(p)PVVSGDTS(p)PRHL-CONH2 contains the phospho-tau peptide of SEQ ID NO:2. It has the structure:

[0020] In an embodiment of the present application, the composition further comprises at least one adjuvant. For example, the at least one adjuvant may comprise a TLR9 agonist, such as a CpG oligonucleotide having a nucleotide sequence selected from the group consisting of SEQ ID NO: 14 to SEQ ID NO: 18. In one embodiment, the composition further comprises a CpG oligonucleotide having a nucleotide sequence of SEQ ID NO: 14. In another embodiment, the composition further comprises aluminum hydroxide. In yet another embodiment, the composition further comprises a CpG oligonucleotide having a nucleotide sequence selected from the group consisting of SEQ ID NO: 14 to SEQ ID NO: 18, and aluminum hydroxide.

[0021] In an embodiment of the present application, a method of inducing antibodies against at least one of tau, preferably phosphorylated tau and paired helical fibrils (PHFs), in a human subject in need thereof comprises administering to the human subject a composition comprising a pharma- ceutically acceptable carrier, aluminum hydroxide, a CpG oligonucleotide having the nucleotide sequence of SEQ ID NO: 14, and 5 μg to 200 μg of a conjugate per dose, wherein the conjugate is

[0022] [ka] (wherein n is an integer from 3 to 7, and VYKS(p)PVVSGDTS(p)PRHL-CONH2 comprises the phospho-tau peptide of SEQ ID NO:2). It has the structure:

[0023] In certain embodiments, the methods of the present application comprise administering to a human subject a composition comprising 5 μg, 10 μg, 15 μg, 20 μg, 25 μg, 30 μg, 35 μg, 40 μg, 45 μg, 50 μg, 60 μg, 70 μg, 80 μg, 90 μg, 100 μg, 110 μg, 120 μg, 130 μg, 140 μg, 150 μg, 160 μg, 170 μg, 180 μg, 190 μg, 200 μg, or any value therebetween, of a conjugate described herein per dose.

[0024] In certain embodiments, the composition is administered intramuscularly. In other embodiments, the composition is administered subcutaneously.

[0025] In certain embodiments, the antibodies comprise IgG antibodies against phosphorylated tau (pTau), preferably with anti-pTau IgG titers at least 50, 60, 70, 80, 90, 100 or more times higher than that of a placebo control.

[0026] In certain embodiments, the antibodies comprise IgG antibodies to non-phosphorylated tau, preferably with anti-tau IgG titers at least 50, 60, 70, 80, 90, 100 or more times higher than that of a placebo control.

[0027] In certain embodiments, the antibodies comprise IgG antibodies against enriched paired helical fibrils (ePHF), preferably having anti-ePHF IgG titers at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more times higher than that of a placebo control.

[0028] In certain embodiments, the methods of the present application further include administering to the subject a second dose of a composition comprising a pharma- ceutically acceptable carrier and 5 μg to 200 μg, e.g., 15 μg or 60 μg, of the conjugate per dose, 4 to 12 weeks, e.g., 8 weeks, after the initial administration of the composition.

[0029] In certain embodiments, administration of the second dose of the composition can boost an antibody response induced by the composition, including, for example, an anti-pTau IgG response and / or an anti-ePHF IgG response, preferably the antibody response is increased by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% or more when measured at least two weeks after administration of the second dose of the composition.

[0030] In certain embodiments, the methods of the present application further include administering to the subject a third dose of a composition comprising a pharma- ceutically acceptable carrier and 5 μg to 200 μg, e.g., 15 μg or 60 μg, of the conjugate per dose, 20 to 28 weeks, e.g., 24 weeks, after the initial administration of the composition.

[0031] In certain embodiments, administration of the third dose of the composition can boost an antibody response induced by the composition, including, for example, an anti-pTau IgG response and / or an anti-ePHF IgG response, preferably the antibody response is increased by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% or more when measured at least two weeks after administration of the third dose of the composition.

[0032] In certain embodiments, the methods of the present application further include administering to the subject a fourth dose of a composition comprising a pharma- ceutically acceptable carrier and 5 μg to 200 μg, e.g., 15 μg or 60 μg, of the conjugate per dose, 44 to 52 weeks, e.g., 48 weeks, after the initial administration of the composition.

[0033] In certain embodiments, the fourth dose of the composition can boost an antibody response induced by the composition, including, for example, an anti-pTau IgG response and / or an anti-ePHF IgG response, preferably the antibody response is increased by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% or more when measured at least two weeks after administration of the fourth dose of the composition.

[0034] In one general aspect, the present application provides a method of inducing a sustained immune response against phosphorylated tau protein (pTau) in a human subject in need thereof, comprising: i. administering intramuscularly to a subject a priming vaccine comprising an effective amount of a conjugate; and ii. administering to the subject intramuscularly a first booster vaccine containing an effective amount of the conjugate 6-10 weeks after administration of the priming vaccine; Including, a sustained immune response is produced for at least about 20 weeks following administration of the priming vaccine; The conjugate has the formula (I):

[0035] [ka] or having the structure of formula (II):

[0036] [ka] The structure is (In the formula, x is an integer from 0 to 10, preferably from 2 to 6, and most preferably 3; n is an integer of 3 to 15, preferably 3 to 12. The carrier represents an immunogenic carrier selected from the group consisting of keyhole limpet hemocyanin (KLH), tetanus toxoid, CRM197 and outer membrane protein mixture (OMP) from Neisseria meningitidis, or derivatives thereof; The tau peptide represents a tau phosphopeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1 to SEQ ID NO:3 and SEQ ID NO:5 to SEQ ID NO:12; The method relates to a method wherein the effective amount of the conjugate comprises between 5 μg and 200 μg of the conjugate per dose.

[0037] In certain embodiments, the carrier is CRM197.

[0038] In certain embodiments, the effective amount of the conjugate comprises 15 μg of the conjugate per dose.

[0039] In certain embodiments, the effective amount of the conjugate comprises 60 μg of the conjugate per dose.

[0040] In certain embodiments, the methods of the present application further include intramuscularly administering to the subject a second booster vaccine composition comprising an effective amount of the conjugate 20 to 26 weeks after administration of the priming vaccine, wherein the sustained immune response persists for at least about 36 weeks after administration of the priming vaccine.

[0041] In certain embodiments, the second booster vaccine composition is administered 24 weeks after administration of the priming vaccine, and the sustained immune response persists for at least about 48 weeks after administration of the priming vaccine.

[0042] In certain embodiments, the methods of the present application further comprise intramuscularly administering to the subject a third booster vaccine composition comprising an effective amount of the conjugate 45 to 50 weeks after administration of the priming vaccine, wherein the sustained immune response persists for at least about 67 weeks after administration of the priming vaccine.

[0043] In certain embodiments, the third booster vaccine composition is administered 48 weeks after administration of the priming vaccine, and the sustained immune response persists for at least about 74 weeks after administration of the priming vaccine.

[0044] In certain embodiments, the sustained immune response comprises an IgG response to phosphorylated tau (pTau), preferably with anti-pTau IgG titers at least 50, 60, 70, 80, 90, 100 or more times higher than that of a placebo control.

[0045] In certain embodiments, the sustained immune response comprises an IgG response to non-phosphorylated tau, preferably with anti-tau IgG titers at least 50, 60, 70, 80, 90, 100 or more times higher than that of a placebo control.

[0046] In certain embodiments, the sustained immune response comprises an IgG response to concentrated paired helical fibrils (ePHF), preferably having an anti-ePHF IgG titer that is at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more times higher than that of a placebo control.

[0047] In certain embodiments, the subject is in need of clearance of tau aggregates.

[0048] In certain embodiments, the subject needs to treat a neurodegenerative disease or disorder caused by or associated with the formation of neurofibrillary lesions.Preferably, the human subject needs to treat Alzheimer's disease, such as early Alzheimer's disease, mild cognitive impairment (MCI) due to Alzheimer's disease, mild Alzheimer's disease, or mild to moderate Alzheimer's disease.In certain embodiments, the subject is amyloid positive in the brain but has not yet shown significant cognitive impairment.In other embodiments, the subject has abnormal levels of cerebrospinal fluid (CSF) Abeta amyloid 42 (Aβ42) that are consistent with AD pathology.In another embodiment, the subject needs to treat a neurodegenerative disease or disorder caused by or associated with the formation of neurofibrillary lesions.

[0049] Further aspects, features, and advantages of the present invention will be better understood from a reading of the following detailed description of the invention and the appended claims. [Brief description of the drawings]

[0050] The foregoing summary and the following detailed description of the invention will be better understood when read in conjunction with the accompanying drawings. For the purpose of illustrating the invention, there are shown in the drawings embodiments which are presently preferred. It should be understood, however, that the invention is not limited to the precise embodiments shown in the drawings.

[0051] [Figure 1] Graph of the geometric mean (±95% confidence interval) of anti-p-tau IgG response over time directed against phosphorylated tau peptide (p-tau) in subcohort 2.1 following treatment with either JACI-35.054 (15 μg or 60 μg) or placebo (ITT analysis set). [Diagram 2] Graph of the geometric mean (±95% confidence interval) of anti-tau IgG responses over time directed against non-phosphorylated tau peptides in subcohort 2.1 following treatment with either JACI-35.054 (15 μg or 60 μg) or placebo (ITT analysis set). [Diagram 3]Graph of the geometric mean (±95% confidence interval) of anti-ePHF (enriched vs. helical fibrils) IgG titers over time in subcohort 2.1 after treatment with either JACI-35.054 (15 μg or 60 μg) or placebo (ITT analysis set). [Figure 4-1] 4A and 4B are graphs of epitope recognition profiles of antibodies in 8 AD patients induced by vaccination with JACI-35.054 (15 μg) as determined by epitope mapping ELISA for short 8-mer overlapping peptides covering phospho-peptides T3.30 (SEQ ID NO: 19) and T3.85 (SEQ ID NO: 21) and non-phospho-peptides T3.56 (SEQ ID NO: 20) and T3.86 (SEQ ID NO: 22). Figure 4A shows the epitope recognition profile of anti-phosphorylated tau antibodies in subcohort 2.1. Figure 4B shows the epitope recognition profile of anti-tau antibodies in subcohort 2.1. OD = optical density. [Figure 4-2] (As stated above.)

[0052] Detailed Description of the Invention Various publications, articles, patents, and patent applications are cited or described in the Background and throughout this specification, and each of these references is incorporated herein by reference in its entirety. Any discussion of documents, acts, materials, devices, articles, and the like which has been included in this specification is for the purpose of providing a context for the present invention. Such discussion is not an admission that any or all of these matters form part of the prior art with respect to any invention(s) disclosed or claimed.

[0053] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Otherwise, certain terms used herein have the meanings described herein.

[0054] It must be noted that as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0055] Unless otherwise stated, any numerical value, e.g., concentration or concentration range, described herein should be understood to be modified in all cases by the term "about". Thus, numerical values ​​typically include ±10% of the indicated value. For example, a concentration of 1 mg / mL includes 0.9 mg / mL to 1.1 mg / mL. Similarly, a concentration range of 1% to 10% (w / v) includes 0.9% (w / v) to 11% (w / v). As used herein, the use of numerical ranges expressly includes all possible subranges, i.e., all individual numerical values ​​within such ranges, including integers within such ranges and fractional parts of the values, unless the context clearly indicates otherwise.

[0056] Unless otherwise indicated, the term "at least" preceding a series of elements should be understood to refer to every element in the series. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. Such equivalents are intended to be encompassed by the present invention.

[0057] As used herein, the terms "comprises," "comprising," "includes," "including," "has," "having," "contains," or "containing," or any other variations thereof, can be understood to imply the inclusion of a stated integer or group of integers but not the exclusion of any other integer or group of integers, and are intended to be non-exclusive or non-limiting. For example, a composition, mixture, process, method, article, or device that comprises listed elements is not necessarily limited to only those elements, but can include other elements not expressly listed or inherent to such composition, mixture, process, method, article, or device. Further, unless expressly stated to the contrary, "or" refers to a non-exclusive "or" rather than an exclusive "or." For example, condition 1 or 2 is satisfied by any one of the following: 1 is true (or exists) and 2 is false (or does not exist), 1 is false (or does not exist) and 2 is true (or exists), and both 1 and 2 are true (or exist).

[0058] It should also be understood that the terms "about," "approximately," "generally," "substantially," and similar terms used herein when referring to dimensions or characteristics of preferred inventive components indicate that the described dimensions / characteristics are not precise boundaries or parameters, as would be understood by one of ordinary skill in the art, but do not exclude minor variations therefrom that are functionally the same or similar. At a minimum, such references including numerical parameters may include variations without altering the least significant digit using mathematical and industrial principles accepted in the art (e.g., rounding, measurement or other systematic errors, manufacturing tolerances, etc.).

[0059] The present invention provides a method for inducing antibodies against tau, preferably phosphorylated tau and / or condensed paired helical filaments (ePHF), in a human subject in need thereof without inducing serious adverse events such as encephalitis. In certain embodiments, the method comprises administering to the subject an effective amount of a conjugate comprising a tau phosphopeptide covalently linked to an immunogenic carrier, either directly or via a linker.

[0060] As used herein, the term "anti-phosphorylated tau antibody" refers to an antibody that binds to tau in which an amino acid residue at one or more positions in the amino acid sequence of tau is phosphorylated. The phosphorylated amino acid residue may be, for example, serine (Ser), threonine (Thr), or tyrosine (Tyr). The site of phosphorylated tau to which the anti-phosphorylated tau antibody binds is preferably a site that is specifically phosphorylated in neurodegenerative diseases such as Alzheimer's disease. Examples of sites of phosphorylated tau to which the anti-phosphorylated tau antibody binds include, for example, Tyr18, Ser199, Ser202, Thr205, Thr212, Ser214, Ser396, Ser404, Ser409, Ser422, and Thr427. As used throughout this application, amino acid positions are obtained with reference to the sequence of human microtubule-associated protein tau isoform 2, whose amino acid sequence is represented in GenBank Accession No. NP_005901.2.

[0061] Whether anti-phosphorylated tau antibodies can be induced upon administration can be determined by testing a biological sample from a subject (e.g., blood, plasma, serum, PBMC, urine, saliva, feces, interstitial fluid (ISF), CSF or lymph) for the presence of antibodies, e.g., IgG or IgM antibodies, directed against the immunogenic tau peptide administered in the pharmaceutical composition (see, e.g., Harlow, 1989, Antibodies, Cold Spring Harbor Press). For example, the titer of antibodies produced in response to administration of a composition providing an immunogen can be measured by enzyme-linked immunosorbent assay (ELISA), other ELISA-based assays (e.g., MSD-Meso Scale Discovery), dot blots, SDS-PAGE gels, ELISPOT, or antibody-dependent cellular phagocytosis (ADCP) assays.

[0062] As used herein, the term "adverse event" (AE) refers to any untoward medical occurrence in a patient to whom a pharmaceutical formulation is administered, not necessarily having a causal relationship with treatment. In an embodiment of the present invention, AEs are rated on a three-level scale of increasing severity, using the following definitions: mild (Grade 1), referring to an AE that is easily tolerated by the subject, causes minimal discomfort, and does not interfere with daily activities; moderate (Grade 2), referring to an AE that is sufficiently unpleasant to interfere with normal daily activities and may require intervention; severe (Grade 3), referring to an AE that interferes with normal daily activities and usually requires treatment or other intervention. A serious AE (SAE) may be any AE occurring at any dose that results in any of the following outcomes: death, which is an outcome, not an event; life-threatening, which refers to an event where the patient is at risk of death at the time of the event's occurrence; life-threatening does not refer to an event that may have caused death if it had been more severe; hospitalization for treatment, e.g., an unplanned overnight hospitalization, or an extension of an existing hospitalization; permanent or significant impairment or substantial destruction of the ability to perform normal life functions; congenital anomaly / birth defect; a significant medical event (as determined by the investigator) that may endanger the patient or require medical or surgical intervention to prevent one of the other outcomes listed above (e.g., emergency room or intensive care at home for allergic bronchospasm, or blood disorders or seizures that do not result in hospitalization). Hospitalization is a formal admission to a hospital. Hospitalization or extended hospitalization constitutes the criteria for an AE to be serious, but is not considered an SAE in itself. In the absence of an AE, hospitalization or extended hospital stay should not be reported as an SAE by the participating investigator. This may be the case in the following situations: the hospitalization or extended hospital stay is necessary for a procedure required by the protocol; or the hospitalization or extended hospital stay is part of a routine procedure followed by the central site (e.g., stent removal after a surgical procedure). This should be documented in the study file. Hospitalization for elective procedures for a pre-existing condition that did not worsen during the study is not considered an AE.

[0063] A complication occurring during hospitalization is an AE. If the complication prolongs the hospital stay or meets any of the other SAE criteria, the event is an SAE.

[0064] As used herein, the term "encephalitis" refers to inflammation of the brain, which may be due to infectious and non-infectious causes.As used herein, the term "meningoencephalitis" refers to a condition characterized by infection or inflammation of the brain meninges and brain.The diagnosis of encephalitis or meningoencephalitis can be determined by techniques known to those skilled in the art in view of the present disclosure, for example, clinical, neurological, and psychiatric examinations, biological sample collection, including blood and CSF sample collection, MRI scanning, and electroencephalography (EEG).

[0065] As used herein, the term "tau" or "tau protein", also known as microtubule-associated protein tau, MAPT, neurofibrillary tangle protein, paired helical fibril-tau, PHF-tau, MAPTL, MTBT1, refers to an abundant central and peripheral nervous system protein with multiple isoforms. In the human central nervous system (CNS), six major tau isoforms exist, ranging in size from 352 to 441 amino acids long, due to alternative splicing (Hanger et al., Trends Mol Med. 15:112-9, 2009). Examples of tau include, but are not limited to, tau isoforms in the CNS, such as the longest tau isoform of 441 amino acids (4R2N), also named microtubule-associated protein tau isoform 2, with four repeats and two inserts, such as human tau isoform 2, whose amino acid sequence is represented in GenBank accession number NP_005901.2. Other examples of tau include the shortest (fetal) isoform (3R0N) of 352 amino acids long, also named microtubule-associated protein tau isoform 4, with three repeats and no insertion, such as human tau isoform 4, whose amino acid sequence is represented in GenBank Accession No. NP_058525.1. Examples of tau also include the "big tau" isoform expressed in peripheral nerves, which contains 300 additional residues (exon 4a). Friedhoff et al., Biochimica et Biophysica Acta 1502 (2000) 122-132. Examples of tau include human big tau, a 758 amino acid long protein encoded by a 6762 nucleotide long mRNA transcript (NM_016835.4), or its isoforms. The amino acid sequence of the exemplary human big tau is represented in GenBank Accession No. NP_058519.3. As used herein, the term "tau" includes homologs of tau from non-human species, such as Macaca Fascicularis (cynomolgus monkey), rhesus monkey, or Pan troglodytes (chimpanzee).As used herein, the term "tau" includes proteins that include mutations of full-length wild-type tau, such as point mutations, fragments, insertions, deletions, and splice variants. The term "tau" also includes post-translational modifications of the tau amino acid sequence. Post-translational modifications include, but are not limited to, phosphorylation.

[0066] As used herein, the term "peptide" or "polypeptide" refers to a polymer composed of amino acid residues linked via peptide bonds, related naturally occurring structural variants, and non-naturally occurring synthetic analogs thereof. The term refers to peptides of any size, structure, or function. Typically, peptides are at least three amino acids in length. Peptides can be naturally occurring, recombinant, or synthetic peptides, or any combination thereof. Synthetic peptides can be synthesized, for example, using an automated polypeptide synthesizer. Examples of tau peptides include any peptide of tau protein that is about 5 to about 30 amino acids in length, preferably about 10 to about 25 amino acids in length, and more preferably about 16 to about 21 amino acids in length. In this disclosure, peptides are listed from N-terminus to C-terminus using standard three- or one-letter amino acid abbreviations, with the phosphate residue indicated by "p". Examples of tau peptides useful in the present invention include, but are not limited to, tau peptides comprising any of the amino acid sequences of SEQ ID NOs: 1-12, or tau peptides having an amino acid sequence that is at least 75%, 80%, 85%, 90%, or 95% identical to any of the amino acid sequences of SEQ ID NOs: 1-12.

[0067] The avidity of an antibody can be measured by an avidity index using methods known in the art in view of the present disclosure. The titer of an antibody against a particular antigen is measured at two different concentrations of coated antigen: one at a saturation concentration where all antibodies can bind to the antigen, and the other at a low concentration where only antibodies with very high binding capacity can bind to the antigen. As used herein, "avidity index" refers to the ratio of the levels of antibody titer measured at low and high density coatings of antigen. For example, the avidity of an antibody against an antigen, such as ePHF or pTau, can be measured at multiple different time points after one immunization or after multiple immunizations to assess whether the avidity (as measured by the avidity index) increases over time. As used herein, an antibody with "increased avidity" or "increased binding avidity" against an antigen refers to an antibody with an increased avidity index against the antigen over time during the course of treatment or immunization. An increase in avidity suggests possible affinity maturation of the antibody.

[0068] As used herein, the term "phosphopeptide" or "phospho-epitope" refers to a peptide that is phosphorylated at one or more amino acid residues. Examples of tau phosphopeptides include any tau peptide that includes one or more phosphorylated amino acid residues. Any suitable tau phosphopeptide known to one of skill in the art can be used in the conjugate in light of the present disclosure. In certain embodiments, the one or more tau phosphopeptides include an amino acid sequence of one of SEQ ID NOs: 1-3 or 5-12, or an amino acid sequence that is at least 75%, 80%, 85%, 90%, or 95% identical to an amino acid sequence of one of SEQ ID NOs: 1-3 or 5-12, and one or more of the indicated amino acid residues are phosphorylated. Preferably, the tau phosphopeptide includes an amino acid sequence of one of SEQ ID NOs: 1-3. Abnormally phosphorylated tau readily aggregates into insoluble oligomers, which are neurotoxic and contribute to neurodegeneration (Goedert et al, 1991). The oligomers progress to so-called tangles of paired helical filaments (PHFs) (Alonso et al., 2001).The extent of neurofibrillary tangle pathology has consistently been shown to correlate with the degree of dementia in AD subjects (Bierer et al., 1995; Braak and Braak, 1991; Delacourte, 2001).

[0069] Tau peptides useful in the present invention can be synthesized by solid phase peptide synthesis or recombinant expression systems. Automated peptide synthesizers are commercially available from a number of suppliers, such as Applied Biosystems (Foster City, Calif.). Recombinant expression systems can include bacteria, such as E. coli, yeast, insect cells, or mammalian cells. Recombinant expression procedures are described by Sambrook et al., Molecular Cloning: A Laboratory Manual (CSHP Press, NY 2d ed., 1989).

[0070] Conjugates Examples of conjugates useful in the present invention include, but are not limited to, the tau phosphopeptide conjugates described in U.S. Patent Application Publication No. 2019 / 0119341, the disclosure of each of which is incorporated by reference in its entirety.

[0071] In certain embodiments, the conjugate has the following structure:

[0072] [ka] or having the formula (II):

[0073] [ka] The structure is (In the formula, x is an integer from 0 to 10; n is an integer of 2 to 15, preferably 3 to 11. Carrier refers to an immunogenic carrier, Tau peptide refers to tau phosphopeptide.

[0074] As used herein, the term "immunogenic carrier" refers to an immunogenic substance that can be coupled to a tau peptide. The immunogenic moiety coupled to the tau peptide can induce an immune response to induce the production of antibodies that can specifically bind to the tau peptide. An immunogenic moiety is an operative moiety, including proteins, polypeptides, glycoproteins, complex polysaccharides, particles, nucleic acids, polynucleotides, etc., that is recognized as foreign and thereby induces an immunological response from the host. Any suitable immunogenic carrier known to those of skill in the art can be used in the present invention in view of the present disclosure. In certain embodiments, the immunogenic carrier is keyhole limpet hemocyanin (KLH), tetanus toxoid, CRM197 (a non-toxic form of diphtheria toxin), outer membrane protein mixture (OMP) from Neisseria meningitidis, or derivatives thereof. In certain embodiments, the immunogenic carrier is CRM197.

[0075] In certain embodiments, the tau peptide is conjugated to the immunogenic carrier via a linker. As used herein, the term "linker" refers to a chemical moiety that connects the immunogenic carrier to the tau peptide. Any suitable linker known to those skilled in the art can be used in the present invention in view of the present disclosure. The linker can be, for example, a single covalent bond, a substituted or unsubstituted alkyl, a substituted or unsubstituted heteroalkyl moiety, a polyethylene glycol (PEG) linker, a peptide linker, a sugar-based linker, or a cleavable linker, such as a disulfide linkage or a protease cleavage site, or an amino acid, or a combination thereof. Examples of linkers can include one or more of polyethylene glycol (PEG), succinimidyl 3-(bromoacetamido)propionate (SBAP), m-maleimidobenzoyl-N-hydroxysuccinimidyl ester (MBS), or one or more amino acids, such as Cys, Lys, or sometimes Ser or Thr, or a combination thereof.

[0076] In certain embodiments, x is an integer from 1 to 10, from 2 to 9, from 2 to 8, from 2 to 7, from 2 to 6, from 2 to 5, from 2 to 4, or from 2 to 3. In certain embodiments, x is 3.

[0077] In certain embodiments, multiple tau phosphopeptides may be conjugated to one immunogenic carrier. In some embodiments, n is 2-15, 3-11, 3-9, 3-8, or 3-7.

[0078] In certain embodiments, the conjugate comprises one or more tau peptides. In certain embodiments, the tau peptides of the conjugate can be the same or different.

[0079] In certain embodiments, the tau phosphopeptide consists of the amino acid sequence of one of SEQ ID NOs: 1-3.

[0080] In certain embodiments, the linker comprises (C2H4O)x-cysteine-acetamidepropionamide or m-maleimidobenzoyl-N-hydroxysuccinimide ester-cysteine-(C2H4O)x, where x is an integer between 0 and 10, e.g., 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[0081] In certain embodiments, the carrier is covalently linked to the N-terminus of the tau phosphopeptide via a linker.

[0082] In other specific embodiments, the carrier is covalently linked to the C-terminus of the tau peptide via a linker.

[0083] In certain embodiments, the conjugate comprises:

[0084] [ka] (In the formula, n is an integer from 2 to 15, preferably from 3 to 11, more preferably from 3 to 7, and VYKS(p)PVVSGDTS(p)PRHL-CONH2 contains the phospho-tau peptide of SEQ ID NO: 2. It has the structure:

[0085] The conjugates of the present invention can be made by methods known in the art in view of the present disclosure. For example, the conjugates shown above can be made by reacting succinimidyl-3-(bromoacetamido)propionate (SBAP):

[0086] [ka] with the amino group of CRM197 to form an amide linkage. This CRM197 precursor can then be reacted with a tau peptide (e.g., the tau phosphopeptide of SEQ ID NO:2) conjugated at its N-terminus or its C-terminus to form a tau phosphopeptide conjugate.

[0087] Pharmaceutical Compositions Pharmaceutical compositions comprising an effective amount of the conjugate useful in the present invention together with pharma- ceutically acceptable excipients and / or carriers can be made using methods known in the art in view of the present disclosure. The optimal ratio of each component in the composition can be determined by techniques well known to those skilled in the art in view of the present disclosure.

[0088] Pharmaceutically acceptable excipients and / or carriers are well known in the art (see Remington's Pharmaceutical Science (15th ed.), Mack Publishing Company, Easton, Pa., 1980). The preferred formulation of a pharmaceutical composition depends on the intended method of administration and therapeutic application. The composition may contain a pharma- ceutically acceptable non-toxic carrier or diluent, defined as a vehicle commonly used to formulate pharmaceutical compositions for animal or human administration. 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 may also include other carriers, adjuvants, or non-toxic, non-therapeutic, and non-immunogenic stabilizers, and the like. It will be understood that the characteristics of the carrier, excipient, or diluent may depend on the route of administration for a particular application.

[0089] The pharmaceutical composition can contain a mixture of the conjugate and the same immunogenic tau peptide, or alternatively, the pharmaceutical composition can contain a mixture of the conjugate and a different immunogenic tau peptide of the invention.

[0090] In certain embodiments, the conjugate can be administered in combination with a suitable adjuvant to achieve a desired immune response in a subject.The suitable adjuvant can be administered before, after, or simultaneously with the administration of the conjugate of the present invention.A preferred adjuvant enhances the intrinsic response to an immunogen without causing conformational changes in the immunogen that affect the qualitative form of the response.

[0091] In one embodiment, adjuvants useful in the methods of the present application are aluminum salts (alum), such as aluminum hydroxide, aluminum phosphate, and aluminum sulfate.

[0092] In another embodiment, the adjuvant useful in the method of the present application is a TLR agonist, such as a CpG oligonucleotide. As used herein, the term "CpG oligonucleotide", "CpG oligodeoxynucleotide", or "CpG ODN" refers to an oligonucleotide that contains at least one CpG motif. As used herein, "oligonucleotide", "oligodeoxynucleotide" or "ODN" refers to a polynucleotide formed from a plurality of linked nucleotide units. Such oligonucleotides can be obtained from existing nucleic acid sources or can be produced by synthetic methods. As used herein, the term "CpG motif" refers to a nucleotide sequence that contains unmethylated cytosine-phosphate-guanine (CpG) dinucleotides (i.e., cytosine (C) followed by guanine (G)) linked by phosphate bonds or phosphodiester backbones or other internucleotide linkages, such as phosphorothioate (ps), phosphorodithioate (ps2), methylphosphonate (mp), or methylphosphorothioate (rp). Phosphorothioate, phosphorodithioate, methylphosphonate and methylphosphorothioate are stabilized internucleotide linkages, while phosphodiester is a naturally occurring internucleotide linkage. Oligonucleotide phosphorothioates are typically synthesized as a random racemic mixture of Rp and Sp phosphorothioate linkages. In view of the present disclosure, any suitable CpG oligonucleotide known to those skilled in the art can be used in the present invention. Examples of such CpG oligonucleotides include, but are not limited to, CpG2006 (also known as CpG 7909), CpG 1018, CpG2395, CpG2216 or CpG2336.

[0093] In certain embodiments, the CpG oligonucleotide is lipidated, i.e., conjugated (covalently linked) with a lipid moiety. As used herein, "lipid moiety" refers to a moiety containing a lipophilic structure. Lipid moieties, such as alkyl groups, fatty acids, triglycerides, diglycerides, steroids, sphingolipids, glycolipids, or phospholipids, especially sterols such as cholesterol or fatty acids, when bound to highly hydrophilic molecules, such as nucleic acids, can substantially increase plasma protein binding and, as a result, the circulatory half-life of the hydrophilic molecule. In addition, binding to certain plasma proteins, such as lipoproteins, has been shown to increase uptake into certain tissues expressing the corresponding lipoprotein receptors (e.g., LDL receptor, HDL receptor, or scavenger receptor SR-B1). In particular, lipid moieties conjugated to phosphopeptides and / or CpG oligonucleotides allow the peptides and / or oligonucleotides to be anchored to the membrane of liposomes via the hydrophobic moiety.

[0094] Such adjuvants can be used with or without other specific immunostimulants, such as MPLAs (3 De-O-acylated monophosphoryl lipid A (MPL™), monophosphoryl hexaacyl lipid A 3-deacylated synthetic (3D-(6-acyl)PHAD™, PHAD™, PHAD™-504, 3D-PHAD™) lipid A), polymeric or monomeric amino acids such as polyglutamic acid or polylysine. Such adjuvants can be used with or without other specific immunostimulants, such as muramyl peptides (e.g., N-acetylmuramyl-L-threonyl-D-isoglutamine (thr-MDP), N-acetyl-normuramyl-L-alanyl-D-isoglutamine (nor-MDP), N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1'-2'dipalmitoyl-sn-glycero-3-hydroxyphosphoryloxy)-ethylamine (MTP-PE), N-acetylglucosaminyl-N-acetylmuramyl-L-Al-D-isoglu-L-Ala-dipalmitoxypropylamide (DTP-DPP) Theramide™), or other bacterial cell wall components.Oil-in-water emulsions include MF59, containing 5% squalene, 0.5% Tween 80, and 0.5% Span 85 (optionally containing various amounts of MTP-PE), which is formulated into submicrometer particles using a microfluidizer (see WO 90 / 14837); SAF, containing 10% squalene, 0.4% Tween 80, 5% pluronic block polymer L121, and thr-MDP, which is microfluidized into a submicrometer emulsion or vortexed to generate a larger particle size emulsion; and Ribi™ Adjuvant System (RAS) (Ribi ImmunoChem, Hamilton, Mont.) 0.2% Tween 80, and one or more bacterial cell wall components selected from the group consisting of monophosphoryl lipid A (MPL™), trehalose dimycolate (TDM), and cell wall skeleton (CWS), preferably MPL™+CWS (Detox™). Other adjuvants include complete Freund's adjuvant (CFA), and cytokines such as interleukins (IL-1, IL-2, and IL-12), macrophage colony stimulating factor (M-CSF), and tumor necrosis factor (TNF).

[0095] In certain embodiments, the pharmaceutical compositions useful in the methods of the present application further comprise one or more suitable adjuvants as described herein, such as an aluminum salt, e.g., aluminum hydroxide, aluminum phosphate, and / or aluminum sulfate, and / or aCpG, e.g., CpG2006 (also known as CpG 7909), CpG 1018, pG2395, CpG2216, or CpG2336. In one embodiment, the pharmaceutical composition comprises a pharma- ceutically acceptable carrier, aluminum hydroxide, CpG 7909, and a conjugate of a tau phosphopeptide covalently linked to CRM197 via a linker.

[0096] In other embodiments, the pharmaceutical composition comprises a conjugate as described herein, one or more adjuvants, a buffer containing one or more amino acids, such as histidine or glycine, one or more carbohydrates, such as glucose or sucrose, and / or a surfactant, such as polysorbate 80, polysorbate 20, etc.

[0097] How to use One general aspect of the present application relates to a method for safely inducing an immune response against tau protein in a human subject suffering from a neurodegenerative disease, disorder or condition, comprising administering to the subject a pharmaceutical composition comprising an effective amount of a phosphorylated tau conjugate. In a particular embodiment, the immune response is induced against tau protein, preferably phosphorylated tau protein, more preferably ePHF.

[0098] As used herein, the term "effective amount" refers to the amount of an active ingredient or component that induces a desired biological or medical response in a subject. The selection of a specific effective dose can be determined (e.g., through clinical trials) by a person skilled in the art based on the consideration of several factors, including the disease to be treated or prevented, the symptoms involved, the patient's weight, the patient's immune status, and other factors known by a person skilled in the art. The exact dose to be used in the formulation may also depend on the method of administration, the route of administration, the target site, the physiological state of the patient, other medications administered, and the severity of the disease, and should be determined according to the judgment of the practitioner and the circumstances of each patient. For example, the effective amount of tau phosphopeptide conjugated with an immunogenic carrier protein also depends on whether an adjuvant is also administered, with higher dosages being required in the absence of an adjuvant. Effective doses can be extrapolated from dose-response curves derived from in vitro or animal model test systems.

[0099] Since one or more tau phosphopeptides can be conjugated to an immunogenic carrier, an effective amount of the conjugate includes the total weight of the immunogenic carrier protein, the one or more tau phosphopeptides conjugated thereto, and one or more linkers (if used) in the conjugate. In an embodiment of the present application, the effective amount of the conjugate is about 5 μg to about 200 μg per dose, preferably about 15 μg to about 150 μg per dose, such as 5 μg, 10 μg, 15 μg, 20 μg, 25 μg, 30 μg, 35 μg, 40 μg, 45 μg, 50 μg, 60 μg, 70 μg, 80 μg, 90 μg, 100 μg, 110 μg, 120 μg, 130 μg, 140 μg, 150 μg, 175 μg, 200 μg per dose, or any value therebetween of the immunogenic carrier. Preferably, the effective dose is 15 μg per dose, up to 60 μg, for example 45 μg, 50 μg, 55 μg, 60 μg, or any value therebetween, or up to 150 μg, for example 120 μg, 125 μg, 130 μg, 135 μg, 140 μg, 145 μg, 150 μg, or any value therebetween.

[0100] As used herein, the terms "induce" and "stimulate" and variations thereof refer to any measurable increase in cellular activity. Induction of immune response can include, for example, activation, proliferation, or maturation of a population of immune cells, increasing cytokine production, and / or other indicators of increased immune function. In certain embodiments, induction of immune response can include increasing B cell proliferation, producing antigen-specific antibodies, increasing antigen-specific T cell proliferation, improving dendritic cell antigen presentation, and / or increasing the expression of certain cytokines, chemokines, and costimulatory markers.

[0101] The ability to induce or activate an anti-tau immune response upon administration in an animal or human organism can be assessed either in vitro or in vivo using a variety of assays standard in the art. For a general description of techniques available for assessing immune response initiation and activation, see, for example, Coligan et al. (1992 and 1994, Current Protocols in Immunology; ed. J Wiley & Sons Inc, National Institute of Health). Measurement of cellular immunity can be readily performed by methods known in the art, for example, by measuring the cytokine profile secreted by activated effector cells, including those derived from CD4+ and CD8+ T cells (e.g., quantification of IL-4 or IFNγ producing cells by ELISPOT), by determining the activation state of immune effector cells (e.g., T cell proliferation assay by classical [3H] thymidine incorporation), by assaying for antigen-specific T lymphocytes in sensitized subjects (e.g., peptide-specific lysis in cytotoxicity assays, etc.).

[0102] The ability to stimulate a cellular and / or humoral response can be determined by testing a biological sample (e.g., blood, plasma, serum, PBMC, urine, saliva, feces, CSF, or lymph) from a subject for the presence of antibodies directed to the immunogenic tau peptide administered in the pharmaceutical composition (see, e.g., Harlow, 1989, Antibodies, Cold Spring Harbor Press). For example, the titer of antibodies produced in response to administration of a composition providing an immunogen can be measured by enzyme-linked immunosorbent assay (ELISA), dot blot, SDS-PAGE gel, ELISPOT, or antibody-dependent cellular phagocytosis (ADCP) assay.

[0103] The conjugate composition can be administered parenterally, topically, intravenously, orally, subcutaneously, intraarterially, intracranially, intraperitoneally, intradermally, intranasally, or intramuscularly for preventive and / or therapeutic treatment. The most typical administration route of the immunogenic substance is subcutaneous or intramuscular injection. This latter type of injection is most typically performed in the arm or leg muscle.

[0104] It is easily understood by those skilled in the art that the regimen for priming and boosting administration can be adjusted based on the immune response measured after administration.For example, the boosting composition is generally administered several weeks or months after the administration of the priming composition, for example, about 1 week, or 2 weeks, or 3 weeks, or 4 weeks, or 8 weeks, or 16 weeks, or 20 weeks, or 24 weeks, or 28 weeks, or 32 weeks, or 36 weeks, or 40 weeks, or 44 weeks, or 48 weeks, or 52 weeks, or 56 weeks, or 60 weeks, or 64 weeks, or 68 weeks, or 72 weeks, or 76 weeks, or 1 to 2 years after the administration of the priming composition.

[0105] In certain embodiments, one or more boosts may be administered. The antigens in each priming and boosting composition need not be identical, no matter how many boosting compositions are used, but should have common antigenic determinants or be substantially similar to each other.

[0106] As known to those skilled in the art, immunogenicity, boostability and sustainability are important considerations for the efficacy of a vaccine. It has been discovered in the present invention that administration of an effective amount of the conjugate described herein can induce a strong antibody response against p-tau in a patient in need thereof, for example, a patient in need of treating Alzheimer's disease (e.g., mild to moderate Alzheimer's disease or early Alzheimer's disease) or mild cognitive impairment (MCI) due to Alzheimer's disease. The antibody response is sustainable, for example, lasting for at least 6 weeks. The antibody response is also boosted by one or more subsequent boosting doses. As used herein, "boosted" in the context of an antibody response refers to an antibody response that is maintained or strengthened after a subsequent dose, when measured at least 2 weeks after the administration of the subsequent dose. For example, an antibody response is "boosted" by a subsequent dose if the antibody titer measured 2 weeks after the subsequent dose is increased compared to the antibody titer before the subsequent dose.

[0107] In certain embodiments, the human subject is in need of treatment for a neurodegenerative disease, disorder, or condition.

[0108] As used herein, "neurodegenerative disease, disorder, or condition" includes any neurodegenerative disease, disorder, or condition known to one of skill in the art in light of the present disclosure. Examples of neurodegenerative diseases, disorders, or conditions include neurodegenerative diseases or disorders caused by or associated with the formation of neurofibrillary lesions, such as tau-associated diseases, disorders, or conditions referred to as tauopathies. In certain embodiments, the neurodegenerative disease, disorder, or condition includes Alzheimer's disease, Parkinson's disease, Creutzfeldt-Jakob disease, Pugilist dementia, Down's syndrome, Gerstmann-Straussler-Scheinker disease, inclusion body myositis, prion protein cerebral amyloid angiopathy, traumatic brain injury, amyotrophic lateral sclerosis, Parkinson's dementia complex of Guam, non-Guam motor neuron disease with neurofibrillary tangles, argyrophilic grain dementia, corticobasal degeneration, amyotrophic lateral sclerosis with Lewy dementia, and the like. The neurodegenerative disease, disorder, or condition may be any disease or disorder that exhibits coexistence of tau and amyloid pathology, including, but not limited to, cerebrospinal fluid (CSF) sclerosis, diffuse neurofibrillary tangles with calcification, frontotemporal dementia, preferably frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17), frontotemporal lobar dementia, Hallervorden-Spatz disease, multiple system atrophy, Niemann-Pick disease type C, Pick's disease, progressive subcortical gliosis, progressive supranuclear palsy, subacute sclerosing panencephalitis, tangle only dementia, postencephalitic parkinsonism, myotonic dystrophy, chronic traumatic encephalopathy (CTE), primary age-related tauopathy (PART), cerebrovascular disease, or dementia with Lewy bodies (LBD). In certain embodiments, the neurodegenerative disease, disorder, or condition is Alzheimer's disease or another tauopathy. In a preferred embodiment, the neurodegenerative disease, disorder, or condition is Alzheimer's disease.

[0109] The clinical course of Alzheimer's disease can be divided into multiple stages according to the progression pattern of cognitive and functional impairment.The stages can be defined using rating scales known in the art, including, for example, the NIA-AA research framework (see, for example, Dubois et al., Alzheimer's & Dementia 12 (2016) 292-323, Dubois et al., Lancet Neurol 2014; 13: 614-29, Jack et al., Alzheimer's & Dementia 14 (2018) 535-562) and Clinical Dementia Rating Scale (CDR) (see, for example, Berg L. Clinical Dementia Rating (CDR). Psychopharmacol Bull. 1988; 24 (4): 637-639), the entire contents of each of which are incorporated herein by reference.

[0110] For example, the National Institute on Aging-Alzheimer's Association (NIA-AA) Research Framework defines AD biologically by neuropathological changes or biomarkers, and treats cognitive impairment as a symptom / sign of the disease rather than the definition of the disease (see, e.g., Clifford RJ, NIA-AA Research Framework: Toward a biological definition of Alzheimer's disease. Alzheimer's & Dementia 14 (2018) 535-562, the contents of which are incorporated herein by reference). According to the NIA-AA definition, individuals with biomarker findings of Aβ deposits only (abnormal amyloid PET scan, or low Aβ42 or Aβ42 / Aβ40 ratio in CSF) and normal biomarkers of pathological tau are assigned the label "Alzheimer's pathological changes," and the term "Alzheimer's disease" is applied when biomarker findings of both Aβ and pathological tau are present. The NIA-AA has also developed a system for staging the severity of AD. Specifically, the NIA-AA definition (adapted from Clifford RJ, 2018, Text Box 2 above) is as follows: definition A: Aβ biomarkers determine whether an individual is on the Alzheimer's continuum. T: Pathological tau biomarkers determine whether someone on the Alzheimer's continuum has Alzheimer's disease. Grading of severity: (N): Neurodegeneration / neuronal injury biomarkers (C): Cognitive symptoms A and T indicate the specific neuropathological changes that define Alzheimer's disease, whereas (N) and (C) are not specific for Alzheimer's disease and are therefore placed in parentheses.

[0111] In a preferred embodiment, the neurodegenerative disease, disorder, or condition is early Alzheimer's disease, mild cognitive impairment (MCI) due to Alzheimer's disease, or mild Alzheimer's disease.

[0112] In some embodiments, the neurodegenerative disease, disorder, or condition is mild to moderate Alzheimer's disease.

[0113] In some embodiments, the subject in need of treatment is amyloid positive in the brain, but has not yet shown significant cognitive impairment. Amyloid deposits in the brain can be detected using methods known in the art, such as PET scanning, immunoprecipitation mass spectrometry, or other methods (e.g., using CSF biomarkers) (Clifford RJ, NIA-AA Research Framework: Toward a biological definition of Alzheimer's disease. Alzheimer's & Dementia 14 (2018) 535-562).

[0114] In other embodiments, the human subject in need of treatment has abnormal levels of CSF Abeta amyloid 42 (Aβ42) consistent with AD pathology. For example, the subject may have low levels of CSF Aβ42 or a low Aβ42 / Aβ40 ratio consistent with AD pathology (see, e.g., Clifford RJ, 2018, supra and references therein, the contents of each of which are incorporated herein by reference in their entirety).

[0115] In certain embodiments, one or more additional treatments can be administered in combination with the tau phosphopeptide conjugate.The additional treatments can include administration of tau antigen before, after or simultaneously with administration of the conjugate.The antigens in the additional compositions do not need to be identical, but should have a shared antigenic determinant with the tau phosphopeptide of the conjugate or be substantially similar to each other.

[0116] Thus, in certain embodiments, the method of the present application further comprises administering to a subject liposomes comprising tau phosphopeptides presented on the surface of the liposomes.Examples of tau liposomes useful in the present invention include, but are not limited to, the tau liposomes described in U.S. Patent Nos. 8,647,631 and 9,687,447, and U.S. Patent Publication No. 2019 / 0119341, the disclosures of each of which are incorporated herein by reference in their entirety.For example, the liposomes useful in the present application can comprise an adjuvant that contains tau phosphopeptides; helper T cell epitopes; lipidated CpG oligonucleotides; and toll-like receptor 4 ligands, and the tau phosphopeptides are presented on the surface of the liposomes.

[0117] In certain embodiments, administration of an effective amount of a conjugate of the present application to a subject results in an anti-pTau IgG or anti-Tau IgG (non-phosphorylated Tau peptide) response for at least 20 weeks, e.g., at least 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 weeks. In other embodiments, administration of an effective amount of a conjugate of the present application to a subject results in an IgG response that recognizes pathological ePHF Tau derived from human AD brain, and the response is sustained for at least 20 weeks, e.g., at least 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 weeks.

[0118] As used herein, "in combination" in the context of administration of two or more therapeutic agents to a subject refers to the use of more than one therapy. The use of the term "in combination" does not restrict the order in which the therapies are administered to a subject.

[0119] Composition can be provided in a kit, container or dispenser that can contain one or more unit dosage forms containing active ingredient if desired.Kit can contain metal or plastic foil, such as blister pack.Kit, container or dispenser can be provided with instructions for administration.

[0120] In certain embodiments, the kit comprises at least one of a pharmaceutical composition comprising a liposome in accordance with an embodiment of the present invention, and a pharmaceutical composition comprising a conjugate in accordance with an embodiment of the present invention. EXAMPLES

[0121] [Example 1] Preparation of conjugate vaccines Peptides and adjuvants The tau phosphopeptide (SEQ ID NO:2) used in this study was synthetically produced with the addition of phospho-residues during synthesis (Pepscan, NL). A conjugate comprising a tau phosphopeptide having the amino acid sequence of SEQ ID NO:2 covalently linked to a CRM carrier via a linker is referred to herein as JACI-35.054.

[0122] The vaccine peptide was conjugated to the carrier protein CRM197 via a polyethylene glycol (PEG)-cysteine-acetamide propionamide linker. Tau phosphopeptide with the amino acid sequence of SEQ ID NO:2 was synthetically produced with the addition of a phospho residue and a PEG3 spacer during synthesis (Polypeptide Laboratories SAS). JACI-35.054 was produced by conjugating the carrier protein CRM197 to the cysteine ​​on the N-terminus of the peptide via a succinimidyl 3-(bromoacetamido)propionate (SBAP) linker. SBAP was linked to the CRM197 protein primary amine (-NH2) via NHS ester reaction chemistry. Excess SBAP linker was removed using ultrafiltration and diafiltration (UF / DF). The CRM197-SBAP intermediate was conjugated to the tau phosphopeptide, and the reaction was stopped by adding an excess of L-cystine when the reaction was complete, terminating the conjugation reaction. The crude CRM197-peptide conjugated product was purified using a Capto Q ImpRes (GE Healthcare) chromatography column and eluted using an isocratic salt method. The purified CRM197-peptide product was then formulated in a buffer containing Tris and sucrose, e.g., 20 mM Tris, 250 mM sucrose, at pH 8.1, using UF / DF. A final concentration of 0.01% PS80 was reached by adding polysorbate 80 (PS80) stock buffer, e.g., 10% PS80 stock buffer, to generate a CRM197-tau peptide drug substance (DS) stock solution. The solution was mixed thoroughly before filtration. Prior to injection, the stock solution was diluted with PBS and CpG / alum, e.g., to a first concentration of 0.8 mg / mL CRM197-tau peptide, and then further diluted with PBS and CpG / alum to a final concentration of 30 ug / mL CRM197-tau peptide for injection. Alternatively, the CRM197-tau peptide stock solution was maintained at a concentration of 3.1 mg / mL in 10 mM PBS (pH 7.3) and further diluted with PBS to reach the desired working concentration.CpG oligonucleotides, alum and PBS were then added to reach a final concentration of 30ug / mL CRM197-pTau peptide base and the final formulation was mixed thoroughly prior to injection.

[0123] One concern with targeting CNS antigens with active vaccines is that non-specific or off-target inflammation may cause undesirable neuropathological changes.To investigate this, whole brains were taken from mice immunized with the conjugate composition and stained to visualize perivascular or other cellular infiltration.No signs of neuroinflammation, cellular infiltration, or other undesirable neuropathological changes were observed in any of the immunized mice (data not shown).This suggested that the vaccine-induced antibodies and natural immune response to vaccination did not cause neuropathological changes in mice.

[0124] [Example 2] A 6-Month Intramuscular Toxicity Study of JACI-35.054 in Rhesus Monkeys the purpose The objective of the overall study was to determine the toxicity of JACI-35.054 (JACI-35.054) following seven intramuscular (im) injections administered over a period of approximately 6 months to naive male and female rhesus monkeys and to assess the reversibility of changes following a 4-week recovery period.

[0125] Test and reference articles were administered by im injection. Control animals were dosed on the same treatment schedule as the dosing group receiving the compound with the active moiety removed in the final formulation.

[0126] design The study protocol applied was as follows (Table 1).

[0127] [Table 1]

[0128] The test article JACI-35.054 composition (containing 15, 50 and 150 μg of JACI-35.054 at low, medium and high dose levels, respectively, as well as 500 μg of CpG7909 and 562.5 μg of aluminum hydroxide suspension) and the reference article were administered by intramuscular injection on days 1, 29, 57, 85, 113, 141 and 169. The control group received the reference article, which was a combination of Tris buffer (in place of the active moiety), CpG7909 and aluminum hydroxide suspension.

[0129] Body weight was evaluated weekly from the acclimation period until the end of the study. Ophthalmoscopy was performed once before the study and 5 days after the 4th and last dose. Electrocardiograms (limbs and height leads) were recorded for each animal once before the study and after the 4th and last dose. Blood and urine samples were taken from all animals for clinical pathology (hematology, coagulation, clinical chemistry and urinalysis) once before treatment, on days 90 and 174 (main and recovery studies) and on day 207 (recovery study) for all surviving animals. Blood samples for serum for determination of anti-p-tau, anti-CRM197 by ELISA were taken on days -14, 8, 22, 36, 50, 64, 78, 92, 99, 106, 120, 134, 148, 162, 176, 183 (main and recovery studies) and on days 190, 204 and 211 (recovery study). CSF was collected once pre-dosing and before necropsy. Blood for immunophenotyping was collected pre-dosing, on days 169 and 211 (end of recovery study). PMBC were collected (ELISpot for T cell responses) on days -14 and 183 (main and recovery studies) and on day 211 (recovery study). Potential binding activity of JACI-35.054-induced antibodies (serum sampled pre-study and on day 183 from 150 μg dosed animals) to a panel of 42 frozen human tissues from three unrelated individuals was assessed using immunohistochemistry (IHC). After the completion of the 6-month dosing period and 4-week recovery study period, main and recovery study animals were euthanized and necropsied, and organ weights and gross findings were recorded. Histopathological examination was performed on the brain, injection site, and lymph nodes of all main and recovery study animals.

[0130] JACI-35.054 induced anti-pTau IgG titers in all treated monkeys at all doses tested (15, 50 and 150ug / mL).

[0131] result Results from the draft audited report indicate that after seven im injections on days 1, 29, 57, 85, 113, 141 and 169, JACI-35.054 at all dose levels was well tolerated by rhesus monkeys, there were no unexpected deaths, and there were no JACI-35.054-related clinical signs or effects on body weight, ophthalmology, electrocardiography, immunophenotyping, clinical pathology or cerebrospinal fluid parameters, organ weights, or gross / microscopic findings.

[0132] For all antibody panels tested, no significant differences in immune cell population size were observed between study groups. The concentrations of the various cell populations of interest measured in the test article-treated groups (Groups 2 to 4) were comparable to the control group, and the variation in immune cell population size was within the biological variability for most populations.

[0133] Findings observed in some animals treated with JACI-35.054 were limited to very mild to moderate skin sensitivity (erythema and edema). However, the transient nature of the findings, as well as the lack of persistence and dose-relatedness, did not suggest a trend towards overt skin sensitivity to JACI-35.054. All gross / microscopic changes were considered to be adjuvant-related and confounded by the experimental procedures, and not due to the presence of JACI-35.054 in the injected dose. The slightly lower incidence of microscopic changes in the recovery phase and their tendency to be confined to the muscle layer may suggest that the inflammatory and degenerative / necrotic processes had resolved to some extent after the 4-week recovery period.

[0134] Immunohistochemical studies performed on 42 frozen human tissues from three unrelated individuals clearly demonstrated that monkey antibodies induced by JACI-35.054, tested at 1 / 300 and 1 / 100 dilutions, did not produce off-target staining in any of the tissues tested. conclusion

[0135] Overall, there were no obvious test substance-related changes after seven intramuscular injections on days 1, 29, 57, 85, 113, 141 and 169, and therefore the highest dose level of JACI-35.054 (150 μg) was considered the no observed effect level (NOEL) for this study.

[0136] [Example 3] Three-month repeated subcutaneous dose toxicity study in mice the purpose The objective of this study was to evaluate potential toxicity following seven subcutaneous (SC) injections over a three-month period in CD1 mice of the test articles CRM197-pTau and CpG-7909, and the JACI-35.054 composition, an adjuvanted vaccine formulated with aluminum hydroxide as the adjuvant. After completion of the treatment period, designated animals were euthanized either two weeks after the final injection (early euthanasia) or after an additional two-week treatment-free period (late euthanasia) and assessed for reversibility of findings or potential delayed effects. design

[0137] The study protocol applied was as follows (Table 2).

[0138] [Table 2]

[0139] The experimental design consisted of Swiss CD1 mice (60 males and 60 females) assigned to four groups injected SC with JACI-35.054 (1.7, 5 and 15 μg / dose; expressed as dose of CRM197-pTau [groups 2, 3 and 4]) or placebo / control substances (Tris buffer, and CpG7909 and aluminum hydroxide adjuvant [group 1]) in the interscapular region (days 1, 15, 29, 43, 57, 71 and 85). At the end of the treatment period, the first available 12 mice / sex / group were euthanized 2 weeks after the last dose (early euthanasia) and the last 6 mice / sex of groups 1 and 4 were euthanized after a 2-week treatment-free period (i.e., 4 weeks after the last injection of test or control). Toxicity parameters and endpoints evaluated included morbidity / mortality, clinical observations, local injection reactions, body weight, food consumption, ophthalmology, hematology, blood biochemistry and anatomical pathology evaluation (including organ weights). A complete necropsy was performed on all animals and gross abnormalities were recorded and microscopic examination was performed on all tissues (including potential target organs in animals from groups 1, 2, 3 and 4 euthanized at the end of the treatment period and in the group euthanized at the end of the non-treatment dosing period). Blood was collected from all animals before, during and during the non-treatment period for determination of immunogenicity as measured by the production of anti-CRM197 IgG and anti-p-tau IgG.

[0140] result The test article JACI-35.054 (1.7, 5, and 15 μg / dose) and placebo were generally well tolerated throughout the study, with no notable clinical signs or effects on body weight, food consumption, ophthalmology, hematology, or organ weights observed in the majority of animals following SC administration. Two animal deaths were noted during the study (one female in Group 1 (control) on Day 78 [Week 12] and one male in Group 3 (5 μg) on ​​Day 43 (Week 7)). These deaths were considered accidental and unrelated to JACI-35.054 administration, as one animal belonged to the control group and no unique clinical, in vivo, or microscopic findings were associated or identified with the deceased male in Group 3. Furthermore, no other animals in the JACI-35.054-treated groups died or underwent humane euthanasia during the study.

[0141] Following JACI-35.054 administration, anti-pTau IgG titers increased in a generally dose-dependent manner, confirming the expected vaccine-associated immunogenicity, and anti-CRM197 IgG titers were also induced. Additional CRM197-pTau-associated changes at the end of treatment included an increased incidence of moderate (grade 3) granulomatous inflammation in females and males receiving 5 μg / dose or higher, as well as mild increases in total protein concentrations (+3.6% to +8.1%) and moderate decreases in albumin to globulin ratios (A / G; -7.9% to -21.2%), likely reflecting increases in globulin concentrations induced by CRM197-pTau due to antigenic stimulation.

[0142] Additional notable findings related to the general SC injection procedure or co-administered adjuvants (particularly aluminum hydroxide suspension) in all JACI-35.054 treatment and control groups included: 1) at least one clinical observation of erythema, thickening, and / or swelling during the study (generally observed with greater frequency and / or severity in JACI-35.054-treated animals compared to controls, with a trend toward dose-dependent progression); 2) thickening and white masses at necropsy in the majority of animals, often correlating with microscopic observations of granulomatous inflammation characterized by granulomas and / or inflammatory pseudocysts with necrotic / caseating centers at the injection site; and 3) lymphocytic hyperplasia and foamy macrophage infiltration in the axillary lymph nodes.

[0143] After a recovery period, clinical signs of hyperplasia and swelling were fully reversible at 1.7 μg / mouse and above, and granulomatous inflammation was fully reversible at 5 μg / mouse and above. In females, a reduction in A / G was still observed, and no recovery was observed at the injection site in any group. There was partial recovery of adjuvant-related findings in the axillary lymph nodes (reduced incidence / severity of foamy macrophages).

[0144] conclusion In conclusion, JACI-35.054 (1.7, 5, and 15 μg / dose) formulated as an adjuvanted vaccine of CRM197-pTau with CpG7909 and aluminum hydroxide was well tolerated when administered SC to CD1 mice seven times every two weeks. Consistent with expected immunogenicity, JACI-35.054-associated anti-pTau IgG titers and anti-CRM197 IgG titers were induced in a largely dose-dependent manner. Other CRM197-pTau-associated findings were limited to granulomatous inflammation, mild elevations in total protein concentrations, and moderate reductions in albumin to globulin ratios, which were partially reversible, at doses of 5 μg / mouse / dose and above. As all findings were considered non-adverse, the maximum dose level of JACI-35.054, 15 μg, was considered the no observed adverse effect level (NOAEL) for this study.

[0145] [Example 4] Safety and Efficacy of JACI-35.054 in Humans A multicenter, prospective, placebo-controlled, double-blind, randomized study to evaluate treatment with a tau-targeted vaccine compared to placebo over a 50-week (i.e., 12-month) period in subjects with early Alzheimer's disease. The study population will be individuals (male and female) aged 50-75 years who have been diagnosed with mild AD or MCI due to AD according to National Institute on Aging-Alzheimer's Association (NIA-AA) criteria. Immunizations will be performed at month 0 (week 0), month 2 (week 8), month 6 (week 24), and month 12 (week 48). Based on safety and immunogenicity results, the protocol may be revised to test additional regimens.

[0146] the purpose Primary Objectives: To assess the safety and tolerability of the study vaccine; and to assess the immunogenicity (induction of IgG titers against p-tau in serum) of the study vaccine.

[0147] Secondary objectives: to further evaluate the immunogenicity of the study vaccine (induction of IgG titers to tau and IgM titers to p-tau and tau in serum); and to evaluate the avidity of antibodies induced by immunization.

[0148] Exploratory Objectives: To examine the effect of the study vaccine on putative biomarkers of AD progression, i.e., blood and / or CSF concentrations of total tau and p-tau protein; to examine the effect of the study vaccine on T cell activation in the blood; to examine the activity of the study vaccine on inflammatory cytokines in the blood (e.g., IL-1B, IL-2, IL-6, IL-8, IL-10, IFN-γ, and TNF-α); to further examine the effect of the study vaccine on immune responses (e.g., antibodies to vaccine components, functional capacity of antibodies induced by the vaccine); and to examine the effect of the study vaccine on behavior, cognition, and functional activities.

[0149] treatment JACI-35.054 administered by the intramuscular route at up to three dose levels will be studied in up to three subcohorts. The study is currently ongoing and is being tested in 2.1 subcohorts.

[0150] Subcohort 2.1 (8 subjects): 6 subjects received JACI-35.054 at 15 μg / dose and 2 subjects received placebo. Safety and tolerability data after all subjects received the second injection in subcohort 2.1 allow for dose escalation following review by the Data Safety Monitoring Board (DSMB).

[0151] Subcohort 2.2 (8 subjects) (optional): 6 subjects received JACI-35.054 at 60 μg / dose and 2 subjects received placebo. This subcohort is currently being conducted based on the good safety and tolerability observed in subcohort 2.1 and based on the expectation that antibody responses in this previous subcohort will be optimized at the 60 μg dose.

[0152] Subcohort 2.3 (8 subjects) (optional): JACI-35.054 can be administered at up to 150 μg / dose to 6 subjects and placebo can be administered to 2 subjects. This subcohort is optional and may be performed based on the good safety and tolerability observed in subcohort 2.2 and if the antibody response in this previous subcohort is expected to be optimized at higher doses.

[0153] Subcohort Expansion: Optional recruitment of up to 16 additional subjects (12 active treatment and 4 placebo) may be considered in a given subcohort of each cohort. The goal is to collect additional data at the dose expected to show the most favorable profile in terms of immunogenicity, safety and tolerability. The decision to expand a given subcohort is based on the cumulative safety / tolerability and immunogenicity data obtained from the respective cohort.

[0154] The vaccine or placebo will be administered four times at weeks 0, 8, 24 and 48, respectively, with, for example, 8, 16 and 24 weeks between each dose. The treatment period is expected to be 50 weeks (12 months), followed by a safety follow-up period of 24 weeks (6 months). The overall subject participation period will be up to approximately 80 weeks from the first screening assessment to the final safety follow-up visit.

[0155] Safety Tracking All subjects will be under clinical observation for 24 hours after the first dose of study vaccine and for 4 hours after each subsequent dose of study vaccine. Subsequent safety assessments will also be conducted by telephone for all subjects 48-72 hours after each immunization. In each subcohort, the first dose of the first 4 subjects should be administered after the previous subject's 48-72 hour safety assessment has been conducted. Safety test samples will be collected at baseline, before each injection, and 2-4 weeks after each injection. All treated subjects will have a 24-week (6-month) safety follow-up period after the end of the treatment period. During this period, subjects will be asked to attend their first follow-up visit 19 weeks after the last dose and a final visit at the end of the follow-up period (26 weeks after the last dose). Participant safety will be monitored throughout the study by regular review of safety data by the Data Safety Monitoring Board (DSMB).

[0156] Interim analysis (IA) Interim analyses of safety, tolerability and immunogenicity data can be performed in each subcohort as follows: - Once all subjects in the subcohort have completed Visit 4 (Week 10), i.e., 2-4 weeks after the second injection - After all subjects in the subcohort have completed Visit 6 (Week 26), i.e., 2-4 weeks after the third injection - After all subjects in the subcohort have completed Visit 9 (Week 50), i.e., 2-4 weeks after the last injection at Week 48 - Once all subjects in the subcohort have completed Visit 11 (Week 74), i.e., at the end of safety.

[0157] Follow-up period Available biomarker data can also be examined during any of these IAs. The above IAs can also be performed on expanded subcohorts.

[0158] Additional IAs to examine the sustainability of immune response data can be performed between weeks 26 and 50 and between weeks 50 and 74.

[0159] Study population The study population was 50–75 years of age (male and female) diagnosed with mild AD or MCI due to AD according to the National Institute on Aging-Alzheimer's Association (NIA-AA) criteria and with a Clinical Dementia Rating Scale (CDR) global score of 0.5 or 1.

[0160] The selection criteria were as follows: 1. Male or female between 50 and 75 years of age. 2. Mild cognitive impairment (MCI) due to AD or mild AD according to NIA-AA criteria and a Clinical Dementia Rating (CDR) global score of 0.5 or 1, respectively. Mini-Mental State Examination (MMSE) score of 3.22 or higher. 4. Abnormal levels of CSF Abeta amyloid 42 (Aβ42) at screening consistent with AD pathology. In cases that are borderline with respect to CSF ​​Aβ42 levels, other results that help determine amyloid positivity may be considered, such as Aβ42 / Aβ40 ratio, and a case-by-case history of a positive amyloid PET scan or positive CSF Aβ42 level. Results of CSF sampling performed within 6 months prior to screening are acceptable on a case-by-case basis, provided that they are consistent with the presence of amyloid pathology and that the corresponding CSF sample is available for study use for testing. 5. Subjects not receiving any marketed treatment for AD or receiving stable doses of acetylcholinesterase inhibitors and / or memantine for at least 3 months prior to baseline. 6. Subject cared for by a reliable informant or caregiver who ensures compliance, assists in clinical evaluations, and reports safety issues. 7. Women must be at least 1 year postmenopausal and / or surgically sterilized. Women of childbearing potential or who are not postmenopausal must have a negative blood pregnancy test at screening (blood drawn -14 days prior to baseline through -3 days prior to baseline) and be willing to use highly effective contraception from the screening visit through the end of participation. Urine pregnancy tests will be performed throughout the treatment period to determine whether subjects can continue to receive the study vaccine. Male participants in the study who have female partners of childbearing potential will be required to use barrier contraception (condoms with spermicide) throughout the study in addition to the contraceptive measures used by their female partners. 8. Subjects who, in the investigator's opinion, are able to understand and give written informed consent. 9. Subjects and informants or caregivers must be fluent in one of the study languages ​​and able to comply with all study procedures, including lumbar puncture.

[0161] The exclusion criteria were as follows: 1. Participation in previous clinical trials for AD and / or neuropathy using active immunization, except where subjects were treated with placebo only and there is documented evidence that the placebo vaccine is not expected to induce any specific immune response. 2. Participation within the past 12 months prior to screening in a previous clinical trial for AD and / or neuropathy using any passive immunization, unless subjects were treated with placebo only and there is documented evidence that the placebo is not expected to induce any specific immune response. 3. Participation in a previous clinical trial for AD and / or neurological disorders using any small molecule drug, including a BACE-1 inhibitor, within the past 3 months prior to screening. 4. Concurrent participation in any other clinical trial using an experimental or approved drug or therapy. 5. The presence of positive antinuclear antibody (ANA) titers at a dilution of at least 1:160 in subjects without clinical symptoms of autoimmune disease. 6. Current or past history of autoimmune disease, or clinical symptoms consistent with the presence of an autoimmune disease. 7. Immunosuppression, including but not limited to use of immunosuppressants or systemic steroids, unless temporarily prescribed for >3 months prior to screening. 8. History of severe allergic reactions (e.g., anaphylaxis), including but not limited to severe allergic reactions to previous vaccines and / or medications. 9. Prior history of clinically significant hypoglycemic episodes. 10. Drug or alcohol abuse or dependence currently met or met within the past 5 years according to Diagnostic and Statistical Manual of Mental Disorders V (DSM-V) criteria for any condition. 11. Any clinically significant medical condition likely to interfere with the assessment of safety and tolerability of study treatment and / or compliance with all scheduled study visits. 12. Any clinically significant medical condition likely to affect the immune system (e.g., any history of acquired or innate immune system disorders). 13. Use of hydralazine, procainamide, quinidine, isoniazid, TNF inhibitors, or minocycline within the 12 months immediately prior to screening. 14. Use of diltiazem for at least 3 months prior to screening, except on a stable dose. 15. Overt suicide risk, defined as subject responding "yes" to suicidal ideation questions 4 or 5 or "yes" to suicidal behavior within the past 12 months using the Columbia Suicide Severity Scale. 16. Concurrent psychiatric or neurological disorders other than those considered related to AD (e.g., head injury with loss of consciousness, symptomatic stroke, Parkinson's disease, severe carotid artery occlusive disease, TIA). 17. History or presence of poorly controlled epileptic seizures. In case of a history of epileptic seizures, the seizures must be well controlled such that no seizure occurrences have been present within 2 years prior to screening. Use of antiepileptic medications is acceptable if on a stable dose for at least 3 months prior to screening. 18. History of meningoencephalitis within the past 10 years prior to screening. 19. Subject with a history of hemorrhagic and / or non-hemorrhagic stroke. 20. Presence or history of peripheral neuropathy. 21. History of inflammatory neuropathy with possible CNS involvement. 22. Screening MRI scan showing structural evidence of an alternative pathology inconsistent with AD that could be causing the subject's symptoms: benign meningioma less than 1 cm in diameter, 3 or more lacunar infarcts or a single infarct greater than 1 cm in diameter, or evidence of a space-occupying lesion other than any single area of ​​superficial hemosiderosis, or evidence of a previous major hemorrhage greater than 10 mm. T2 * Microbleeds on MRI can be seen up to 10 regardless of location. 23. Unable to undergo MRI examination for any reason including but not limited to metal implants and / or severe claustrophobia that contraindicate MRI studies. 24. Significant hearing or vision impairment, or other problems judged by the investigator to be relevant to preventing following the protocol and conducting outcome measures. 25. Clinically significant infection or major surgical procedure within 3 months prior to screening. Planned surgical procedures anticipated to be performed during participation in the study must be reviewed and approved by the medical monitor at the time of screening. 26. Any vaccine administered within the past 2 weeks prior to screening, including influenza vaccine. 27. Clinically significant arrhythmias or other clinically significant abnormalities on ECG at screening. 28. Myocardial infarction, unstable angina, or significant coronary artery disease within 1 year prior to baseline. 29. History of cancer within the past 5 years other than treated squamous cell carcinoma, basal cell carcinoma, and intraepidermal melanoma, or noninvasive prostate cancer or noninvasive breast cancer that has been completely removed and is considered to have been treated. 30. Clinically significant deviations from normal values ​​for hematological parameters, liver function tests, and other biochemical laboratory values ​​that, in the opinion of the investigator, are judged to be clinically significant. 31. Subject is pregnant as confirmed by blood test at screening, or is planning pregnancy or breastfeeding. 32. Subjects receiving any anticoagulant or antiplatelet medication except aspirin at a dose of 100 mg or less daily (to avoid the risk of bleeding during scheduled or unscheduled lumbar puncture). 33.Subjects receiving antipsychotics, except for stable low doses for the treatment of insomnia. 34.Has donated blood or blood products within 30 days prior to screening or plans to donate blood while participating in the study. 35. A positive Venereal Disease Research Laboratory (VDRL) value consistent with active syphilis at the time of screening. 36. Positive HIV test at screening. 37. Laboratory or clinical evidence of active hepatitis B and / or C at screening. 38. Serum creatinine >1.5 times the upper limit of normal, abnormal thyroid function test results, or clinically significant decreases in serum B12 or folate levels (Note: all oral doses of thyroid replacement medications, B12 or folate must be stable for at least 3 months prior to screening).

[0162] Study endpoints The following primary endpoints regarding safety and tolerability will be evaluated: adverse events, immediate and delayed reactogenicity (e.g., local and systemic reactogenicity including anaphylaxis, immune complex disease); suicidal ideation (C-SSRS); behavior (NPI); cognitive and functional assessments to evaluate safety (RBANS, CDR-SB); vital signs; MRI imaging; electrocardiogram; routine hematology and biochemistry assessments in blood and urine; assessment of autoimmune antibodies including anti-dsDNA antibodies in blood; inflammatory markers in blood and CSF.

[0163] The following primary endpoints of immune response (i.e., immunogenicity) will also be assessed: anti-pTau IgG titers in serum (geometric mean, change from baseline, responder rate, peak and area under the curve).

[0164] The following secondary endpoints related to immune response (i.e., immunogenicity) will be evaluated: serum anti-tau IgG, anti-p-tau, anti-ePHF IgG and anti-tau IgM titers (geometric mean, change from baseline, responder rate, peak and area under the curve), and determination of the IgG response profile by avidity testing.

[0165] The following exploratory endpoints will be evaluated: change from baseline in titers of putative AD biomarkers in blood and / or CSF (e.g., total tau, p-tau), change from baseline in measured T cell activation levels in blood, change from baseline in inflammatory cytokine titers in blood, change from baseline in antibody titers in blood, change from baseline in behavioral (NPI), cognitive and functional activities (RBANS, CDR-SB) scores.

[0166] Results / Conclusion: The following primary endpoints were / will be assessed: · Safety and tolerability - adverse events, immediate and delayed reactogenicity (e.g. local and systemic reactogenicity including anaphylaxis, pain, redness, immune complex disease, swelling, fever); global assessment of tolerability; suicidal ideation (C-SSRS); behavior (NPI); cognitive and functional assessments to evaluate safety (RBANS, CDR-SB); vital signs; MRI imaging; ECG; routine haematology and biochemistry assessments of blood and urine; assessment of autoimmune antibodies including anti-DNA antibodies in blood; inflammatory markers in blood and CSF. · Immune response - anti-p-tau IgG titers in serum (geometric mean, change from baseline, responder rate, peak, and area under the curve).

[0167] The following secondary endpoints were / will be assessed: Immune response: Serum anti-tau IgG, anti-p-tau, anti-ePHF IgG and anti-tau IgM titers (geometric mean, change from baseline, responder rate, peak and area under the curve), determination of IgG response profile by avidity testing.

[0168] The following exploratory endpoints were / will be evaluated: · Change from baseline in biomarker titers in blood and / or CSF (e.g. total tau and p-tau protein), change from baseline in T cell activation levels in blood, change from baseline in inflammatory cytokine (e.g. IL-1B, IL-2, IL-6, IL-8, IL-10, IFN-γ, and TNF-α) titers in blood, change from baseline in suicidal ideation (C-SSRS), behavior (NPI), cognitive and functional activity (RBANS, CDR-SB) scores.

[0169] The study is ongoing. To date, one subcohort has received JACI-35.054 at a dosage level of 15 μg of conjugate ("15 μg dose"), and placebo (phosphate buffered saline (PBS)), as in Table 3. Study subjects in subcohort 2.2 are receiving JACI-35.054 at a dosage level of 60 μg of conjugate ("60 μg dose").

[0170] [Table 3]

[0171] Interim results through week 74 (2 weeks after the 4th injection) for the 15 μg dose and through week 26 (2 weeks after the 3rd injection) for the 60 μg dose indicate that JACI-35.054 is safe and well tolerated, and no clinically relevant safety concerns associated with the study vaccine were observed. Increases in anti-p-tau specific IgG titers relative to baseline were observed in the sera of active-treated subjects who were all responders after the second dose of JACI-35.054 at 15 μg dose at week 10, and at subsequent weeks 24, 26, 36, 48, 50, 67, and 74. Increases in anti-p-tau specific IgG titers relative to baseline were observed in the sera of active-treated subjects who were also all responders after the second dose of JACI-35.054 at 60 μg dose at week 10, and at subsequent weeks 15, 20, 24, and 26. Increases in anti-tau specific IgG titers (response to non-phosphopeptide tau peptides containing the amino acid sequence of SEQ ID NO: 4) were also observed in all study subjects treated with the active drug who were all responders at weeks 10, 24, 26, 36, 48, 50, 67 and 74 at the 15 μg dose, and in all study subjects treated with the active drug who were all responders at weeks 10, 15, 20, 24 and 26 at the 60 μg dose. Increases in anti-ePHF IgG titers against pathological p-tau were observed in early AD subjects treated with the active drug, of which 83.3% were responders at weeks 26, 36 and 50 at the 15 μg dose, and 83.3% and 100% of those at the 60 μg dose were responders at weeks 10 and 26, respectively. No antibody responses were observed in subjects receiving placebo.

[0172] Anti-p-tau IgG response of JACI-35.054 in humans Specific IgG antibody responses directed against phosphorylated tau peptide (p-tau) induced by the JACI-35.054 vaccine in the subcohorts of Table 3 were measured by MSD. Figure 1 shows anti-p-tau IgG titers after immunization with either 15 μg and 60 μg doses of JACI-35.054 or placebo. As shown by the results in Figure 1, immunization with either 15 μg or 60 μg doses of JACI-35.054 induced a strong anti-p-tau IgG response against a biotinylated phosphorylated tau peptide having the amino acid sequence of SEQ ID NO: 19, which contains a biotin linked to the N-terminus of the phosphorylated tau peptide comprising the amino acid sequence of SEQ ID NO: 2.

[0173] Immunization at the 15 μg dose level at weeks 8, 24 and 48 results in boosting of the anti-pTau IgG response when measured two weeks later at weeks 10, 26 and 50, respectively. Similarly, immunization at the 60 μg dose level at weeks 8 and 24 results in boosting of the anti-pTau IgG response when measured two weeks later at weeks 10 and 26, respectively.

[0174] Table 4 shows the anti-pTau IgG responder rates (ITT population) following immunization with either the 15 μg dose, the 60 μg dose of JACI-35.054, or placebo.

[0175] [Table 4]

[0176] As shown by the results in Table 4, 50% of subjects treated with JACI-35.054 at the 15 μg dose level were responders at week 2 and 66.7% were responders at week 8. All subjects treated with JACI-35.054 at the 15 μg dose level were responders from week 10 to week 74. As shown by the results in Table 4, 66.7% of subjects treated with JACI-35.054 at the 60 μg dose level were responders at week 2 and 83.3% were responders at week 8. All subjects treated with JACI-35.054 at the 60 μg dose level were responders from week 10 to at least week 26. None of the subjects treated with placebo developed an anti-p-tau IgG response.

[0177] Anti-tau IgG response of JACI-35.054 in humans The specific IgG antibody response directed against non-phosphorylated tau peptide induced by JACI-35.054 vaccine in the subcohort of Table 3 was measured by MSD. Figure 2 shows the anti-tau IgG titers after immunization with either JACI-35.054 at a dose of 15 μg or 60 μg, or placebo. As shown by the results in Figure 2, immunization with JACI-35.054 at a dose of either 15 μg or 60 μg induced an IgG antibody response against biotinylated non-p-tau peptide having an amino acid sequence of SEQ ID NO:20, which contains a biotin linked to the N-terminus of the non-phosphorylated tau peptide comprising the amino acid sequence of SEQ ID NO:4. Thus, immunization with JACI-35.054 at a dose of 15 μg or 60 μg induced an IgG antibody response that recognizes the non-p-tau peptide having the amino acid sequence of SEQ ID NO:4 in addition to the p-tau peptide having the amino acid sequence of SEQ ID NO:2.

[0178] Immunization at the 15 μg dose level at weeks 8, 24 and 48 resulted in boosting of anti-tau IgG responses when measured two weeks later at weeks 10, 26 and 50, respectively. Immunization at the 60 μg dose level at weeks 8 and 24 resulted in boosting of anti-tau IgG responses when measured two weeks later at weeks 10 and 26, respectively.

[0179] Table 5 shows the anti-tau IgG responder rate (ITT population) after immunization with either JACI-35.054 at a dose of 15 μg or 60 μg or placebo. As shown by the results in Table 5, 66.7% of subjects treated with JACI-35.054 at the 15 μg dose level were responders at week 2 and 83.3% were responders at week 8. All subjects treated with JACI-35.054 at the 15 μg dose level were responders from week 10 to week 74. 66.7% of subjects treated with JACI-35.054 at the 60 μg dose level were responders at weeks 2 and 8. All subjects treated with JACI-35.054 at the 60 μg dose level were responders from week 10 to at least week 26. No subjects treated with placebo developed an anti-tau IgG response.

[0180] [Table 5]

[0181] Recognition of pathological p-tau (enriched paired helical filaments - ePHF) from human AD brains The ability of IgG polyclonal antibodies induced by immunization with JACI-35.054 in the subcohorts of Table 3 to bind to ePHFs derived from human AD brains was measured over time by MSD. Figure 3 shows anti-ePHF IgG titers (ITT population) following immunization with either 15 μg or 60 μg doses of JACI-35.054 or placebo. The results in Figure 3 show that immunization with JACI-35.054 at 15 μg or 60 μg doses induces an IgG antibody response that recognizes pathological ePHF tau derived from human AD brains.

[0182] Immunization at the 15 μg dose level at weeks 8, 24, and 48 resulted in boosting of anti-ePHF IgG responses when measured two weeks later at weeks 10, 26, and 50, respectively. Similarly, immunization at the 60 μg dose level at weeks 8 and 24 resulted in boosting of anti-ePHF IgG responses when measured two weeks later at weeks 10 and 26, respectively.

[0183] Table 6 shows the anti-ePHF IgG responder rates (ITT population) following immunization with either JACI-35.054 or placebo at the 15 μg or 60 μg dose levels. The anti-ePHF IgG responder rate in six subjects treated with JACI-35.054 at the 15 μg dose increased from 0% at week 2 (i.e., 2 weeks after the first injection) to 83.3% at week 26 (i.e., 2 weeks after the third injection), with subsequent responder rates ranging from 66.7% (weeks 48, 67, and 74) to 83.3% (weeks 36 and 50). The anti-ePHF IgG responder rate in six subjects treated with JACI-35.054 at the 60 μg dose increased from 16.7% at week 2 (i.e., 2 weeks after the first injection) to 100% at week 26 (i.e., 2 weeks after the third injection).

[0184] [Table 6]

[0185] It is understood that the examples and embodiments described herein are for illustrative purposes only, and that modifications may be made to the above-described embodiments without departing from the broad inventive concept thereof. It is therefore understood that the invention is not limited to the particular embodiments disclosed, but is intended to cover modifications that are within the spirit and scope of the invention as defined by the appended claims.

[0186] [Example 5] Vaccination with JACI-35.054 induces antibodies with a high degree of heterogeneity in epitope recognition To further profile the antibody response for breadth and selectivity against pathological p-tau, epitope mapping was performed on sera from human subjects with short p-tau and non-p-tau amino acid sequences. A study was conducted to determine the epitope recognition profile of antibodies induced by JACI-35.054 in human subjects. Eight AD patients were intramuscularly immunized with 15ug per dose of JACI-35.054 or placebo at weeks 0, 8, 24, and 48. The epitope recognition profile of antibodies was determined by epitope mapping ELISA using a library of N-terminally biotinylated 8-mer peptides with a one amino acid shift and covering the entire sequence of phospho-tau peptide T3.30 (SEQ ID NO: 19) as well as the sequence of tau peptide T3.56 (SEQ ID NO: 20) before the first immunization (V1, week 0) and after the third immunization (V6, week 26). In addition, binding of the antibodies to full-length phosphotau peptide T3.30 (sequence number 19) and tau peptide T3.56 (sequence number 20), as well as phosphotau peptide T3.85 (sequence number 21) and tau peptide T3.86 (sequence number 22) (with additional C-terminal amino acids) was also determined.

[0187] Data are presented as pre-subtracted optical density (OD) values ​​(OD obtained before the first immunization (V1: week 0) subtracted from OD obtained after the third immunization (V6, week 26)) for each peptide and each patient. Negative values ​​after subtraction were set to 0.000.

[0188] Table 7, Table 8 and Figure 4 show the epitope recognition profile of antibodies induced by vaccination with JACI-35.054 as determined by epitope mapping ELISA on short 8-mer overlapping peptides covering phosphopeptide T3.30 (sequence number 19) and non-phosphopeptide T3.56 (sequence number 20).

[0189] [Table 7]

[0190] Table 7 and Figure 4A show that two AD patients did not produce IgG antibodies against the sequences of phosphotau peptides T3.30 (SEQ ID NO: 19) and T3.85 (SEQ ID NO: 21) (patients #1 and #2). Six AD patients produced IgG antibodies against the sequences of phosphotau peptides T3.30 (SEQ ID NO: 19) and T3.85 (SEQ ID NO: 21), with at least four AD patients showing an overall low binding to the sequence of phosphotau peptide T3.85 (SEQ ID NO: 21). The OD values ​​obtained with the 8-mer peptide show that in all six AD patients, the IgG antibodies induced after three immunizations bound to the C-terminal part of the sequence of phosphotau peptide T3.30 (SEQ ID NO: 19). In addition, some binding to the N-terminal part of the sequence of phosphotau peptide T3.30 (SEQ ID NO: 19) was found in four AD patients.

[0191] [Table 8]

[0192] Table 8 and FIG. 4B show that two AD patients did not produce IgG antibodies against the sequences of tau peptides T3.56 (SEQ ID NO: 20) and T3.86 (SEQ ID NO: 22) (patients #1 and #2). Six AD patients developed IgG antibodies against the sequences of tau peptides T3.56 (SEQ ID NO: 20) and T3.86 (SEQ ID NO: 22), with overall low binding to the sequence of tau peptide T3.86 (SEQ ID NO: 22). The OD values ​​obtained with the 8-mer peptide show that in all six AD patients, the IgG antibodies induced after three immunizations bound to the C-terminal part of the sequence of tau peptide T3.56 (SEQ ID NO: 20). In addition, in three AD patients, binding to tau peptides Tau393-400 and Tau396-403 was found.

[0193] The results show that subjects vaccinated with JACI-35.054 demonstrated an antibody response with strong recognition of the C-terminal portion of the tau antigen sequence and with antibodies binding in a similar manner to phosphorylated and non-phosphorylated tau peptides.

[0194] References

[0195] [Table 9]

[0196] Sequence Listing

[0197] [ka]

[0198] [ka]

[0199] [ka]

Claims

1. 1. A composition for use in a method of inducing antibodies against tau, preferably phosphorylated tau and at least one of enriched paired helical filaments (ePHF), in a human subject in need thereof, said composition comprising a pharmaceutically acceptable carrier and 5 μg to 200 μg of a conjugate per dose, said method comprising administering said composition to said human subject, said conjugate having a structure represented by formula (I): 【Chemical 1】 or Formula (II): 【Chemistry 2】 (In the formula, x is an integer from 0 to 10, preferably from 2 to 6, most preferably 3; n is an integer of 3 to 15, preferably 3 to 12. having the structure the carrier represents an immunogenic carrier selected from the group consisting of keyhole limpet hemocyanin (KLH), tetanus toxoid, CRM197, and outer membrane protein mixture (OMP) from N. meningitidis, or derivatives thereof; The composition, wherein the tau peptide represents a tau phosphopeptide having an amino acid sequence selected from the group consisting of SEQ ID NO:1 to SEQ ID NO:3 and SEQ ID NO:5 to SEQ ID NO:

12.

2. The carrier is CRM197; Optionally, the tau phosphopeptide has the amino acid sequence of SEQ ID NO:

2. Preferably, the conjugate comprises: 【Chemistry 3】 wherein n is an integer from 3 to 7, and VYKS(p)PVVSGDTS(p)PRHL-CONH 2 comprises the phospho-tau peptide of SEQ ID NO:

2.

2. The composition for use according to claim 1, having the structure:

3. the composition further comprises at least one adjuvant; Optionally, the at least one adjuvant comprises a TLR9 agonist. Preferably, the TLR9 agonist is a CpG oligonucleotide having a nucleotide sequence selected from the group consisting of SEQ ID NO: 14 to SEQ ID NO:

18. For example, the CpG oligonucleotide has the nucleotide sequence of SEQ ID NO:

14.

2. The composition for use according to claim 1, wherein optionally the at least one adjuvant comprises aluminum hydroxide.

4. 1. A composition for use in a method for inducing antibodies against at least one of phosphorylated tau and condensed paired helical filaments (ePHF) in a human subject in need thereof, the composition comprising a pharmaceutically acceptable carrier, a CpG oligonucleotide having the nucleotide sequence of SEQ ID NO: 14, and 5 μg to 200 μg per dose of a conjugate, the method comprising administering the composition to the subject, the conjugate comprising: 【Chemistry 4】 (wherein n is an integer of 3 to 7, and VYKS(p)PVVSGDTS(p)PRHL-CONH ２ comprises the phospho-tau peptide of SEQ ID NO: 2) A composition having the structure:

5. The composition for use according to claim 4, wherein the composition further comprises aluminum hydroxide.

6. The method comprising administering to the human subject the composition comprising 5 μg, 10 μg, 15 μg, 20 μg, 25 μg, 30 μg, 35 μg, 40 μg, 45 μg, 50 μg, 60 μg, 70 μg, 80 μg, 90 μg, 100 μg, 110 μg, 120 μg, 130 μg, 140 μg, 150 μg, 160 μg, 170 μg, 180 μg, 190 μg, or any value therebetween, of the conjugate per dose. Optionally, (a) administering to the human subject the composition comprising 15 μg of conjugate per dose; (b) administering to the human subject the composition comprising 45-60 μg, e.g., 45 μg, 50 μg, 55 μg, 60 μg, or any value therebetween, of the conjugate per dose; or (c) administering to the human subject the composition comprising 120-150 μg, e.g., 120 μg, 125 μg, 130 μg, 135 μg, 140 μg, 145 μg, 150 μg, or any value therebetween, of the conjugate per dose; A composition for use according to claim 1.

7. The composition is administered intramuscularly or subcutaneously. Optionally, (a) the antibodies comprise IgG antibodies against phosphorylated tau (p-tau), preferably with an anti-p-tau IgG titer at least 50, 60, 70, 80, 90, 100 or more times higher than that of a placebo control; (b) the antibodies comprise IgG antibodies to non-phosphorylated tau, preferably with an anti-tau IgG titer at least 50, 60, 70, 80, 90, 100 or more times higher than that of a placebo control; and / or (c) the antibodies comprise IgG antibodies to concentrated paired helical fibrils (ePHF), preferably having an anti-ePHF IgG titer that is at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more times higher than that of a placebo control; A composition for use according to claim 1.

8. The method further comprising administering to the subject a second dose of the composition comprising a pharmaceutically acceptable carrier and 5 μg to 200 μg, e.g., 15 μg or 60 μg, of the conjugate per dose, 4 to 12 weeks, e.g., 8 weeks, after the initial administration of the composition. Optionally, administration of the second dose of the composition can boost an antibody response induced by the composition, including, for example, an anti-p-tau IgG response and / or an anti-ePHF IgG response, preferably wherein the antibody response is increased by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% or more when measured at least two weeks after administration of the second dose of the composition. A composition for use according to claim 1.

9. The method further comprising administering to the subject a third dose of the composition comprising a pharmaceutically acceptable carrier and 5 μg to 200 μg, e.g., 15 μg or 60 μg, of the conjugate per dose, 20 to 28 weeks, e.g., 24 weeks, after the initial administration of the composition. Optionally, administration of the third dose of the composition can boost an antibody response induced by the composition, e.g., an antibody response comprising an anti-p-tau IgG response and / or an anti-ePHF IgG response, preferably wherein the antibody response is increased by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% or more when measured at least two weeks after administration of the third dose of the composition. Preferably, the method further comprises administering to the subject a fourth dose of the composition comprising a pharmaceutically acceptable carrier and 5 μg to 200 μg, e.g., 15 μg or 60 μg, of the conjugate per dose, 44 to 52 weeks, e.g., 48 weeks, after the first administration of the composition. More preferably, administration of said fourth dose of said composition is capable of boosting an antibody response induced by said composition, including, for example, an anti-p-tau IgG response and / or an anti-ePHF IgG response, preferably wherein said antibody response is increased by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100% or more when measured at least two weeks after administration of said fourth dose of said composition. A composition for use according to claim 8.

10. 1. A priming vaccine and / or first booster vaccine for use in a method for inducing a sustained immune response against phosphorylated tau protein (p-tau) in a human subject in need thereof, comprising: The method comprises: i. intramuscularly administering to said subject a priming vaccine comprising an effective amount of a conjugate; and ii. 6 to 10 weeks after administration of the priming vaccine, administering to the subject a first booster vaccine comprising the effective amount of the conjugate intramuscularly. Including, the sustained immune response persists for at least about 20 weeks after administration of the priming vaccine; The conjugate has formula (I): 【Chemistry 5】 or having the structure of formula (II): 【Chemistry 6】 (In the formula, x is an integer from 0 to 10, preferably from 2 to 6, most preferably 3; n is an integer of 3 to 15, preferably 3 to 12. having the structure the carrier represents an immunogenic carrier selected from the group consisting of keyhole limpet hemocyanin (KLH), tetanus toxoid, CRM197, and outer membrane protein mixture (OMP) from Neisseria meningitidis, or derivatives thereof; Tau peptide refers to a tau phosphopeptide having an amino acid sequence selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 3 and SEQ ID NO: 5 to SEQ ID NO: 12; A priming vaccine and / or a first booster vaccine, wherein said effective amount of conjugate comprises 5 μg to 200 μg of said conjugate per dose. (a) the carrier is CRM197; (b) the tau phosphopeptide has the amino acid sequence of SEQ ID NO: 2; Optionally, the conjugate comprises: 【Chemistry 7】 (In the formula, n is an integer from 3 to 7, and VYKS(p)PVVSGDTS(p)PRHL-CONH 2 comprises the phospho-tau peptide of SEQ ID NO:

2. and / or (c) the composition further comprises at least one adjuvant; For example, the at least one adjuvant comprises aluminum hydroxide. Optionally, the at least one adjuvant comprises a TLR9 agonist. Preferably, the TLR9 agonist is a CpG oligonucleotide having a nucleotide sequence selected from the group consisting of SEQ ID NO: 14 to SEQ ID NO:

18. More preferably, the CpG oligonucleotide has the nucleotide sequence of SEQ ID NO:

14. A priming vaccine and / or first booster vaccine for use according to claim 10. (a) the effective amount of the conjugate comprises 15 μg of the conjugate per dose, or the effective amount of the conjugate comprises 60 μg of the conjugate per dose, and / or (b) the method further comprises intramuscularly administering to the subject a second booster vaccine composition comprising the effective amount of the conjugate 20 to 26 weeks after administration of the priming vaccine, wherein the sustained immune response persists for at least about 36 weeks after administration of the priming vaccine. Optionally, the second booster vaccine composition is administered 24 weeks after administration of the priming vaccine, and the sustained immune response persists for at least about 48 weeks after administration of the priming vaccine. Preferably, the method further comprises intramuscularly administering to the subject a third booster vaccine composition comprising the effective amount of the conjugate 45 to 50 weeks after administration of the priming vaccine, wherein the sustained immune response persists for at least about 67 weeks after administration of the priming vaccine. More preferably, the third booster vaccine composition is administered 48 weeks after administration of the priming vaccine, and the sustained immune response persists for at least about 74 weeks after administration of the priming vaccine. A priming vaccine and / or first booster vaccine for use according to claim 10. (a) the sustained immune response comprises an IgG response to phosphorylated tau (p-tau), preferably with an anti-p-tau IgG titer at least 50, 60, 70, 80, 90, 100 or more times higher than that of a placebo control; (b) the sustained immune response comprises an IgG response to non-phosphorylated tau, preferably with an anti-tau IgG titer at least 50, 60, 70, 80, 90, 100 or more times higher than that of a placebo control; and / or (c) the sustained immune response comprises an IgG response to concentrated paired helical fibrils (ePHF), preferably with an anti-ePHF IgG titer that is at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more times higher than that of a placebo control; A priming vaccine and / or first booster vaccine for use according to claim 10. (a) the subject is in need of clearance of tau aggregates; and / or (b) the subject is in need of treatment for a neurodegenerative disease or disorder caused by or associated with the formation of neurofibrillary lesions; Optionally, the subject is in need of treatment for Alzheimer's disease. Preferably, the subject is in need of treatment for early Alzheimer's disease or mild cognitive impairment (MCI) due to Alzheimer's disease.

14. A composition for use, or a priming vaccine and / or a first booster vaccine for use, according to any one of claims 1 to 13.

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