Methods and uses of BAG3 for the treatment of cardiac amyloidosis

Abstract

Heart failure with preserved ejection fraction (HFpEF) affects approximately 3 million people in the United States. TIFF2025531194000005.tif85128

Description

[Technical Field]

[0001] Related Applications This application claims priority to U.S. Provisional Patent Application No. 63 / 376,014, filed September 16, 2022. The entire contents of the foregoing application are incorporated herein by reference, including all text, tables, sequence listings, and figures. [Background technology]

[0002] Introduction Heart failure with preserved ejection fraction (HFpEF) affects approximately 3 million people in the United States. Approximately 15 percent of these affected individuals have increased cardiac amyloid. Furthermore, approximately half of cardiac amyloidosis cases are of nonfamilial origin or etiology. Summary of the Invention

[0003] overview It has now been discovered that reduced levels or activity of BCL2-associated anthanogene 3 (BAG3) result in a significant increase in beta-amyloid levels in cells and tissues, such as those in the heart, liver, skeletal muscle, and cells or tissues of the central nervous system (CNS). Thus, in certain embodiments, a method for increasing the level or activity of BAG3 in a subject in need thereof comprises administering to the patient a therapeutically effective amount of an agent, wherein the agent modulates the expression or amount of a BAG3 molecule, its protein, or peptide in a target cell or tissue. In certain embodiments, a method for treating a subject with a cardiac disease, such as HFpEF, wherein the subject has at least one BAG3 nucleotide variant (NV) in-frame insertion compared to a control Bcl2-associated anthanogene 3 (BAG3) nucleic acid sequence, comprises administering to the subject a therapeutically effective amount of an agent, wherein the agent modulates the expression or amount of a BAG3 molecule, its protein, or peptide in a target cell or tissue. In certain embodiments, a method of treating a subject having a neurodegenerative disease associated with increased levels of APP or beta-amyloid, wherein the subject has at least one BAG3 nucleotide variant (NV) in-frame insertion compared to a control Bcl2-associated anthanogene 3 (BAG3) nucleic acid sequence, comprises administering to the subject a therapeutically effective amount of an agent, thereby reducing the level of APP or beta-amyloid in the subject, wherein the agent modulates the expression or amount of a BAG3 molecule, its protein, or peptide in a target cell or tissue.In certain embodiments, a method of treating a subject having a neurodegenerative disease associated with increased levels of APP or β-amyloid, wherein the subject has at least one BAG3 nucleotide variant (NV) in-frame insertion compared to a control Bcl2-associated anthanogene 3 (BAG3) nucleic acid sequence, comprises administering to the subject a therapeutically effective amount of an agent, thereby reducing the level of APP or β-amyloid in the subject and treating the disease, wherein the agent modulates the expression or amount of a BAG3 molecule, its protein, or peptide in a target cell or tissue.

[0004] In certain embodiments, a method of treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent, thereby treating the amyloidosis, wherein the agent modulates the expression or amount of a nucleic acid encoding BCL2-associated athanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide. In certain embodiments, the nucleic acid encoding BAG3 comprises an expression vector that expresses a BAG3 protein or an active fragment thereof.

[0005] In certain embodiments, a method for treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent, thereby treating amyloidosis, wherein the agent modulates the expression or amount of a nucleic acid encoding BCL2-associated asanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide, wherein the nucleic acid encoding BAG3 comprises an expression vector expressing the BAG3 protein or an active fragment thereof, and the expression vector further comprises a promoter, wherein the promoter comprises an inducible promoter, a constitutive promoter, a bicistronic promoter, or a tissue-specific promoter. In certain embodiments, the expression vector comprises a viral vector, a cardiotropic vector, a plasmid, or a yeast vector. In certain embodiments, the expression vector is a pseudotyped viral vector.

[0006] In certain embodiments, a method of treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent, thereby treating the amyloidosis, wherein the agent modulates the expression or amount of a nucleic acid encoding BCL2-associated aganocyte gene 3 (BAG3), a BAG3 protein, or a BAG3 peptide, and the cardiotropic vector comprises an adenovirus vector, an adeno-associated virus vector (AAV), a coxsackievirus vector, a cytomegalovirus vector, an Epstein-Barr virus vector, a parvovirus vector, or a hepatitis virus vector.

[0007] In certain embodiments, the AAV vector comprises a capsid protein having 90% or more sequence identity to any of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7m AAV8, AAV9, AAV10, AAV11, or AAV12.

[0008] In certain embodiments, a method for treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent that modulates the expression or amount of a nucleic acid encoding BCL2-associated athanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide, thereby treating amyloidosis, wherein the patient has or is at risk of having heart failure with preserved ejection fraction (HFpEF). In certain embodiments, the patient has or is at risk of having cardiac amyloid deposits or increased cardiac amyloid compared to age-, sex-, and / or race-matched controls.

[0009] In certain embodiments, a method of treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent, thereby treating the amyloidosis, wherein the agent modulates the expression or amount of a nucleic acid encoding BCL2-associated athanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide, and the patient expresses lower than normal levels of BAG3 in cardiac tissue.

[0010] In certain embodiments, a method of treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent that modulates the expression or amount of a nucleic acid encoding BCL2-associated athanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide, thereby treating the amyloidosis, including cardiac amyloidosis or Alzheimer's disease. In certain embodiments, the patient does not have a mutation or defect in the transthyretin (ATTAR) gene.

[0011] In certain embodiments, a method for treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent, thereby treating amyloidosis, wherein the agent modulates the expression or amount of a nucleic acid encoding BCL2-associated athanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide, wherein the nucleic acid encoding BAG3 comprises an expression vector that expresses the BAG3 protein or an active fragment thereof, and the expression vector further comprises a promoter, which optionally comprises an inducible promoter, a constitutive promoter, a bicistronic promoter, or a tissue-specific promoter. In certain embodiments, the inducible promoter confers expression in cardiac tissue or the CNS.

[0012] In certain embodiments, a method of treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent, thereby treating the amyloidosis, wherein the agent modulates the expression or amount of a nucleic acid encoding BCL2-associated athanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide, wherein the nucleic acid encoding BAG3 comprises an expression vector that expresses the BAG3 protein or an active fragment thereof, and wherein the expression vector further comprises AAV inverted terminal repeats (ITRs).

[0013] In certain embodiments, a method of treating a patient suffering from or at risk of developing amyloidosis comprises administering to the patient a therapeutically effective amount of an agent, thereby treating the amyloidosis, wherein the agent modulates the expression or amount of a nucleic acid encoding BCL2-associated athanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide, wherein the nucleic acid encoding BAG3 comprises an expression vector that expresses the BAG3 protein or an active fragment thereof, and wherein the expression vector further comprises a polyadenylation sequence and / or a stop codon. [Brief explanation of the drawings]

[0014] [Figure 1] Figures 1A-1B show confocal images of cardiac cells from a BAG3 haploinsufficient (BAG3+ / -) mouse experiencing heart failure with preserved ejection fraction (HFpEF). As indicated, cells were stained to visualize either APP or β-amyloid. [Figure 2] Figures 2A-2B provide quantification of the amount of β-amyloid in cardiac cells from either BAG3+ / − or wild-type mice, both of which experienced HFpEF. DETAILED DESCRIPTION OF THE INVENTION

[0015] Detailed Description 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. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, certain materials and methods are described herein. In describing and claiming the present invention, the following terminology is used:

[0016] The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. By way of example, "an element" means one element or more than one element. Thus, for example, reference to "a cell" includes multiple cells of the same type. Furthermore, to the extent the terms "including," "includes," "having," "has," "having," or variations thereof are used in either the detailed description and / or the appended claims, such terms are intended to be inclusive in the same manner as the term "comprising."

[0017] As used herein, the terms "comprising," "comprise," or "comprised," and variations thereof, in reference to defined or described elements of an item, composition, apparatus, method, process, system, etc., are meant to be inclusive or open-ended, indicating that additional elements are permitted, thereby indicating that the defined or described item, composition, apparatus, method, process, system, etc. includes those specified elements (or their equivalents, as appropriate), and that other elements can be included and still fall within the scope / definition of the defined item, composition, apparatus, method, process, system, etc.

[0018] When used herein to refer to measurable values ​​such as amounts and time durations, "about" means to include ±20%, ±10%, ±5%, ±1%, or ±0.1% variation from the specified value, when such variation is appropriate for carrying out the method of the present disclosure. Alternatively, particularly in relation to biological systems or processes, the term can mean within 5 times, and also within 2 times, the order of magnitude of the value. When a specific value is described in this application and the appended claims, unless otherwise stated, the term "about" should be assumed to mean within the acceptable error range for the specific value.

[0019] As used herein, "BAG3," "BAG3 molecule," "BCL2-associated asanogene 3 (BAG3) gene," and "BCL2-associated asanogene 3 (BAG3) molecule" include all family members, variants, cDNA sequences, alleles, fragments, species, coding and non-coding sequences, sense and antisense polynucleotide strands, and the like (HGNC (939) Entrez Gene (9531) Ensembl (ENSG00000151929) OMIM (603883) UniProtKB (O95817)). Similarly, "BAG3," "BAG3 molecule," and "BCL2-associated asanogene 3 (BAG3) molecule" also refer to BAG3 polypeptides or fragments, proteins, variants, derivatives, and the like thereof. Thus, the term "molecule" encompasses both nucleic acid and amino acid sequences of BAG3.

[0020] Bcl-2-associated anthanogene-3 (BAG3), also known as BCL2-associated anthanogene 3; MFM6; Bcl-2-binding protein Bis; CAIR-1; docking protein CAIR-1; BAG family molecular chaperone regulatory factor 3; BAG-3; BCL2-associated anthanogene 3, or BIS, is a cytoprotective polypeptide that competes with Hip-1 for binding to HSP 70. The NCBI reference amino acid sequence of BAG3 can be found in Genbank under accession number NP_004272.2; publication GI:14043024. The amino acid sequence of Genbank accession number NP_004272.2; publication GI:14043024 is referred to herein as SEQ ID NO: 1. The NCBI reference nucleic acid sequence of BAG3 can be found in Genbank under accession number NM_004281.3 GI:62530382. The nucleic acid sequence of Genbank Accession No. NM_004281.3 GI:62530382 is referred to as SEQ ID NO: 2. Other BAG3 amino acid sequences include, for example, but are not limited to, 095817.3 GI:12643665 (SEQ ID NO: 3); EAW49383.1 GI:119569768 (SEQ ID NO: 4); EAW49382.1 GI:119569767 (SEQ ID NO: 5); and CAE55998.1 GI:38502170 (SEQ ID NO: 6). The BAG3 polypeptide of the present invention can be a variant of the polypeptides described herein, so long as it retains functionality.

[0021] As used herein, the term "biological sample" includes solid samples and bodily fluid samples. Biological samples used in the present invention can include cells, protein or membrane extracts of cells, blood, or biological fluids such as ascites or cerebral fluid (e.g., cerebrospinal fluid). Examples of solid biological samples include, but are not limited to, samples taken from tissues of the central nervous system, bone, breast, kidney, cervix, endometrium, head / neck, gallbladder, parotid gland, prostate, pituitary gland, muscle, esophagus, stomach, small intestine, colon, liver, spleen, pancreas, thyroid, heart, lung, bladder, fat, lymph nodes, uterus, ovaries, adrenal glands, testes, tonsils, thymus, and skin, or samples taken from tumors. Examples of "bodily fluid samples" include, but are not limited to, blood, serum, semen, prostatic fluid, seminal plasma, urine, feces, saliva, sputum, mucus, bone marrow, lymph, and tears.

[0022] As used herein, "heart disease" refers to any type of heart disease, including heart failure, myocardial disease, cardiomyopathy, hypertrophic cardiomyopathy, dilated cardiomyopathy, atherosclerosis, coronary artery disease, non-ischemic heart disease, ischemic heart disease, myocarditis, viral infection, wound, hypertensive heart disease, valvular disease, congenital heart disease, myocardial infarction, congestive heart failure, arrhythmia, diseases that result in cardiac remodeling, etc. Heart disease can be caused by any reason, such as damage to cardiac tissue, such as reduced contractility (e.g., as may be demonstrated by a reduced ejection fraction). Cardiac damage or disorders characterized by insufficient cardiac function include any impairment or absence of normal cardiac function, or the presence of abnormal cardiac function. Abnormal cardiac function can be the result of disease, injury, and / or aging. As used herein, "abnormal cardiac function" includes morphological and / or functional abnormalities of cardiomyocytes, populations of cardiomyocytes, or the heart itself. Non-limiting examples of morphological and functional abnormalities include physical deterioration and / or death of cardiomyocytes, abnormal growth patterns of cardiomyocytes, abnormalities in the physical connections between cardiomyocytes, under- or overproduction of substances by cardiomyocytes, inability of cardiomyocytes to produce substances that they normally produce, and transmission of electrical impulses in abnormal patterns or at abnormal times. Abnormalities at a more gross level include movement abnormalities, reduced ejection fraction, changes as seen by echocardiography (e.g., dilation), changes in EKG, changes in exercise tolerance, reduced capillary perfusion, and changes as seen by angiography. Abnormal cardiac function is seen in many disorders, including, for example, non-ischemic or ischemic heart disease, e.g., angina pectoris, myocardial infarction, chronic ischemic heart disease, hypertensive heart disease, pulmonary heart disease (cor pulmonale), valvular heart disease, e.g., rheumatic fever, mitral valve prolapse, calcification of the mitral valve annulus, carcinoid heart disease, infective endocarditis, congenital heart disease, myocardial disease, e.g., myocarditis, dilated cardiomyopathy, hypertensive cardiomyopathy, cardiac disorders leading to congestive heart failure, and cardiac tumors, e.g., primary sarcomas and secondary tumors.Cardiac injuries also include wounds, such as knife wounds; biological injuries (eg, viruses; autoimmune diseases) or chemical injuries (eg, chemotherapy, drugs); surgery; and transplants.

[0023] As used herein, "amyloidosis" refers to the abnormal accumulation, increase, or buildup of amyloid protein, such as beta-amyloid, in one or more cells, tissues, and / or organs and interferes with their normal function.

[0024] As used herein, the phrase "diagnosing" refers to classifying a disease or condition, determining the severity of a disease, monitoring the progression of a disease, and predicting the outcome and / or likelihood of recovery from a disease. The term "detecting" can optionally encompass any of the above. Diagnosis of a disease according to the present invention can be performed by determining the level of the polynucleotide or polypeptide of the present invention in a biological sample obtained from a subject, wherein the determined level can be correlated with a predisposition to the disease or the presence or absence of the disease. It should be noted that a "biological sample obtained from a subject" can optionally include a sample that has not been physically removed from the subject.

[0025] As used herein, the term "diagnostic" means identifying the presence or nature of a pathological condition. Diagnostic methods vary in their sensitivity and specificity. The "sensitivity" of a diagnostic assay is the percentage of diseased individuals who test positive (percent of "true positives"). Diseased individuals not detected by the assay are "false negatives." Subjects who are not diseased and test negative in the assay are called "true negatives." The "specificity" of a diagnostic assay is 1 minus the false positive rate, where the "false positive" rate is defined as the proportion of those without the disease who test positive. While a particular diagnostic method may not provide a definitive diagnosis of a condition, it suffices if the method provides a positive indication that aids in diagnosis.

[0026] As used herein, "effective amount" means an amount that provides a therapeutic or prophylactic benefit.

[0027] "Encoding" refers to the inherent property of a specified nucleotide sequence in a polynucleotide, e.g., a gene, cDNA, or mRNA, to serve as a template for the synthesis of other polymers and macromolecules in biological processes, having either a defined nucleotide sequence (i.e., rRNA, tRNA, and mRNA) or a defined amino acid sequence and the biological properties attributed thereto. Thus, a gene encodes a protein when the protein is produced in a cell or other biological system by transcription and translation of the mRNA corresponding to that gene. Both the coding strand, whose nucleotide sequence is identical to the mRNA sequence and is usually provided in a sequence listing, and the non-coding strand used as a template for transcription of a gene or cDNA can be said to encode the protein or other product of that gene or cDNA.

[0028] The term "expression" as used herein is defined as the transcription and / or translation of a particular nucleotide sequence driven by its promoter.

[0029] An "expression vector" refers to a vector containing a recombinant polynucleotide comprising an expression control sequence operably linked to a nucleotide sequence to be expressed. An expression vector contains sufficient cis-acting elements for expression; other elements for expression can be supplied by the host cell or in an in vitro expression system. Expression vectors include all those known in the art, such as cosmids, plasmids (e.g., naked or contained in liposomes), and viruses (e.g., lentiviruses, retroviruses, adenoviruses, and adeno-associated viruses) that incorporate a recombinant polynucleotide.

[0030] "Inhibitory nucleic acid" refers to a double-stranded RNA, siRNA, shRNA, or antisense RNA, or a portion thereof, or a mimetic thereof, that, when administered to a mammalian cell, results in a reduction (e.g., 10%, 25%, 50%, 75%, or even 90-100%) in expression of a target gene. Typically, a nucleic acid inhibitor comprises at least a portion of a target nucleic acid molecule or its ortholog, or comprises at least a portion of the complementary strand of a target nucleic acid molecule. For example, an inhibitory nucleic acid molecule comprises at least a portion of any or all of the nucleic acids delineated herein.

[0031] "Isolated" means altered or removed from its natural context. For example, a nucleic acid or peptide naturally occurring in a living animal is not "isolated," but the same nucleic acid or peptide partially or completely separated from the coexisting materials of its natural context is "isolated." An isolated nucleic acid or protein can exist in a substantially purified form, or it can exist in a non-native environment, such as, for example, a host cell.

[0032] An "isolated nucleic acid" refers to a segment or fragment of nucleic acid that has been separated from sequences that flank it in its naturally occurring context; i.e., a DNA fragment that has been removed from sequences that normally flank the DNA fragment, i.e., the sequences that flank the DNA fragment in the genome in which it naturally occurs. The term also applies to nucleic acids that have been substantially purified from other components that naturally accompany the nucleic acid, i.e., the RNA or DNA, or proteins that naturally accompany it in the cell. Thus, the term includes, for example, recombinant DNA that has been incorporated into a vector, an autonomously replicating plasmid or virus, or into the genomic DNA of a prokaryote or eukaryote, or that exists as a separate molecule independent of other sequences (i.e., as a cDNA or genomic fragment or cDNA fragment generated by PCR or restriction enzyme digestion). The term also includes recombinant DNA that is part of a hybrid gene encoding additional polypeptide sequences, complementary DNA (cDNA), and linear or circular oligomers or polymers of natural and / or modified monomers or linkages, including deoxyribonucleosides, ribonucleosides, substituted and alpha-anomeric forms thereof, peptide nucleic acids (PNAs), locked nucleic acids (LNAs), phosphorothioates, and methylphosphonates, and the like.

[0033] Nucleic acid sequences can be "chimeric," i.e., composed of different regions. In the context of the present invention, a "chimeric" compound is an oligonucleotide containing two or more chemical regions, such as DNA regions, RNA regions, PNA regions, etc. Each chemical region is made up of at least one monomer unit, i.e., nucleotide. These sequences typically contain at least one region in which the sequence has been modified to exhibit one or more desired properties.

[0034] As used herein, the term "kit" refers to any delivery system for delivering materials. In the context of a reaction assay, such a delivery system includes a system that allows the storage, transport, or delivery of reaction reagents (e.g., oligonucleotides, enzymes, etc. in appropriate containers) and / or supporting materials (e.g., buffers, written instructions for conducting the assay, etc.) from one location to another. For example, a kit includes one or more enclosed containers (e.g., boxes) containing relevant reaction reagents and / or supporting materials. As used herein, the term "fragmented kit" refers to a delivery system that includes two or more separate containers, each containing a subportion of the total components of the kit. The containers can be delivered to the intended recipient together or separately. For example, a first container can contain an enzyme for use in an assay, while a second container contains an oligonucleotide. The term "fragmented kit" is intended to encompass, but is not limited to, a kit containing analyte-specific reagents (ASRs) regulated under Section 520(e) of the Federal Food, Drug, and Cosmetic Act. Indeed, any delivery system comprising two or more separate containers, each containing a subportion of all the components of the kit, is encompassed by the term "fragmented kit." In contrast, a "combined kit" refers to a delivery system containing all the components of a reaction assay in a single container (e.g., in a single box housing each of the desired components). The term "kit" encompasses both fragmented and composite kits.

[0035] The term "target nucleic acid" sequence refers to a nucleic acid (often derived from a biological sample) to which an oligonucleotide is designed to specifically hybridize. The presence or absence of the target nucleic acid is detected, or the amount of the target nucleic acid is quantified. The target nucleic acid has a sequence complementary to the nucleic acid sequence of a corresponding oligonucleotide directed to the target. The term target nucleic acid can refer to a specified subsequence of a larger nucleic acid, or to the entire sequence (e.g., a gene or mRNA) to which the oligonucleotide is directed. Differences in usage will be clear from the context.

[0036] In the context of the present invention, the following abbreviations are used for commonly occurring nucleobases: "A" refers to adenosine, "C" refers to cytosine, "G" refers to guanosine, "T" refers to thymidine, and "U" refers to uridine.

[0037] Unless otherwise specified, a "nucleotide sequence encoding" an amino acid sequence includes all nucleotide sequences that are degenerate versions of each other and that encode the same amino acid sequence. The phrase nucleotide sequence encoding a protein or RNA can also include introns, to the extent that a nucleotide sequence encoding a protein may, in some versions, contain introns.

[0038] "Parenteral" administration of the immunogenic compositions includes, for example, subcutaneous (sc), intravenous (iv), intramuscular (im), or intrasternal injection or infusion techniques.

[0039] As used herein, the terms "patient" or "individual" or "subject" are used interchangeably and refer to a mammalian subject being treated, with a human patient being one example. In some cases, the methods of the present invention find use in laboratory animals, in veterinary applications, and in the development of animal models of disease, including, but not limited to, rodents, including mice, rats, and hamsters, and primates.

[0040] The term "percent sequence identity" or "having sequence identity" refers to the degree of identity between a given query sequence and a subject sequence.

[0041] The term "pharmaceutically acceptable" (or "pharmacologically acceptable") refers to molecular entities and compositions that do not produce adverse, allergic, or other untoward reactions when administered to an animal or human, as appropriate. The term "pharmaceutically acceptable carrier," as used herein, includes any and all solvents, dispersion media, coatings, antibacterial agents, isotonic and absorption delaying agents, buffers, excipients, binders, lubricants, gels, surfactants, and the like, that can be used as a vehicle for a pharmaceutically acceptable substance.

[0042] The term "polynucleotide" refers to a chain of nucleotides, also known as a "nucleic acid" or a "nucleic acid sequence," and includes, but is not limited to, all nucleic acid sequences obtained by any means available in the art, i.e., both naturally occurring and synthetic nucleic acids, complementary DNA (cDNA), and linear or circular oligomers or polymers of natural and / or modified monomers or linkages, such as deoxyribonucleosides, ribonucleosides, their substituted and alpha-anomeric forms, peptide nucleic acids (PNAs), locked nucleic acids (LNAs), phosphorothioates, and methylphosphonates. Nucleic acid sequences can be "chimeric," i.e., composed of different regions. In the context of the present invention, a "chimeric" compound is an oligonucleotide containing two or more chemical regions, e.g., DNA regions, RNA regions, PNA regions, etc. Each chemical region is made of at least one monomer unit, i.e., nucleotides. These sequences typically contain at least one region in which the sequence has been modified to exhibit one or more desired properties.

[0043] The terms "peptide," "polypeptide," and "protein" are used interchangeably and refer to compounds composed of amino acid residues covalently linked by peptide bonds. A protein or peptide must contain at least two amino acids, and there is no limit to the maximum number of amino acids that can comprise a protein or peptide sequence. A polypeptide includes any peptide or protein containing two or more amino acids linked to each other by peptide bonds. As used herein, the term refers to both short chains, commonly referred to in the art as peptides, oligopeptides, and oligomers, and longer chains, commonly referred to in the art as proteins, of which there are many types. "Polypeptide" includes, for example, biologically active fragments, substantially homologous polypeptides, oligopeptides, homodimers, heterodimers, polypeptide variants, modified polypeptides, derivatives, analogs, and fusion proteins, among others. A polypeptide includes natural peptides, recombinant peptides, synthetic peptides, or combinations thereof.

[0044] The terms "transfected" or "transformed" or "transduced" refer to the process by which exogenous nucleic acid is transferred or introduced into a host cell. A "transfected" or "transformed" or "transduced" cell is one that has been transfected, transformed, or transduced with exogenous nucleic acid. Transfected / transformed / transduced cells include the primary subject cell and its progeny.

[0045] "Treating" a disease, as the term is used herein, means reducing the frequency or severity of at least one sign or symptom of the disease or disorder experienced by a subject. Treating a disease or disorder includes eradicating the virus.

[0046] "Treatment" is an intervention performed with the intent to prevent the onset of a disorder or to alter the pathology or symptoms of a disorder. Thus, "treatment" refers to both therapeutic treatment and prophylactic or preventative measures. "Treatment" may also be specified as palliative care. Those in need of treatment include those who already have a disability as well as those in whom disability is to be prevented. Thus, "treating" or "treatment" of a condition, disorder, or state includes: (1) eradicating the virus; (2) preventing or delaying the appearance of clinical symptoms of the condition, disorder, or state that develops in humans or other mammals that may be susceptible to or predisposed to the condition, disorder, or state, but that have not yet experienced or exhibited clinical or subclinical symptoms of the condition, disorder, or state; (3) inhibiting the condition, disorder, or state, i.e., arresting, reducing, or delaying the onset of the disease or its recurrence (in the case of maintenance treatment), or at least one clinical or subclinical symptom thereof; or (4) palliating the disease, i.e., causing regression of the condition, disorder, or state, or at least one clinical or subclinical symptom thereof. The benefit to the treated individual is either statistically significant or at least perceptible to the patient or physician.

[0047] The term "variant," when used in the context of a polynucleotide sequence, can encompass a polynucleotide sequence related to a wild-type gene. This definition can also include, for example, "allelic," "splice," "species," or "polymorphic" variants. Splice variants can have significant identity to a reference molecule but generally have a greater or lesser number of polynucleotides due to alternative splicing of exons during mRNA processing. The corresponding polypeptide may possess additional functional domains or may lack domains. Species variants are polynucleotide sequences that vary from species to species. Particularly useful in the present invention are variants of wild-type gene products. Variants can result from at least one mutation in the nucleic acid sequence and can result in an altered mRNA or a polypeptide that may or may not have altered structure or function. Any given natural or recombinant gene can have none, one, or many allelic forms. Common mutational changes that give rise to variants are generally ascribed to naturally occurring deletions, additions, or substitutions of nucleotides. Each of these types of changes can occur alone, or in combination with the others, one or more times in a given sequence.

[0048] A "vector" is a composition that contains an isolated nucleic acid and can be used to deliver the isolated nucleic acid into a cell. Examples of vectors include, but are not limited to, linear polynucleotides, polynucleotides associated with ionic or amphiphilic compounds, plasmids, and viruses. Thus, the term "vector" includes autonomously replicating plasmids or viruses. The term is also intended to include non-plasmid and non-viral compounds that facilitate the transfer of nucleic acids into cells, such as polylysine compounds and liposomes. Examples of viral vectors include, but are not limited to, adenovirus vectors, adeno-associated virus vectors, and retrovirus vectors.

[0049] Ranges: Throughout this disclosure, various aspects of the invention may be expressed in a range format. The description in range format should be understood merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the invention. Accordingly, the description of a range should be considered to have specifically disclosed all possible subranges, as well as each individual numerical value within that range. For example, the description of a range such as 1 to 6 should be considered to have specifically disclosed each individual numerical value within that range, e.g., 1, 2, 2.7, 3, 4, 5, 5.3, and 6, as well as subranges such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, etc. This applies regardless of the breadth of the range.

[0050] Where any nucleic acid or amino acid sequence is specifically referred to by a SwissProt. or GENBANK accession number, the sequence is incorporated herein by reference. Information associated with the accession number, such as identification of the signal peptide, extracellular domain, transmembrane domain, promoter sequence, and translation start, is also incorporated herein by reference in its entirety. [Example]

[0051] Example 1: Amyloid is increased in the hearts of BAG3+ / - mice experiencing heart failure with preserved ejection fraction (HFpEF) Murine cardiac cells from BAG3 haploinsufficient (BAG3+ / −) mice were stained to visualize either APP or β-amyloid, as depicted in Figure 1. The results indicate that β-amyloid is increased in cardiac cells from BAG3+ / − mice.

[0052] Figures 2A and 2B provide the amount of β-amyloid normalized to the amount of tubulin in BAG3+ / - mice compared to wild-type mice.

[0053] The results indicated that β-amyloid accumulated in cardiac cells of HFpEF mice to levels significantly exceeding those observed in wild-type mice.

[0054] Sequence Listing TIFF2025531194000002.tif245170TIFF2025531194000003.tif250164TIFF2025531194000004.tif211164

Claims

1. 1. A method of treating a patient suffering from or at risk of developing amyloidosis, comprising the steps of: Administering a therapeutically effective amount of an agent to the patient, thereby treating amyloidosis, wherein the agent modulates the expression or amount of a nucleic acid encoding BCL2-associated athanogene 3 (BAG3), a BAG3 protein, or a BAG3 peptide.

2. The method of claim 1, wherein the nucleic acid encoding BAG3 comprises an expression vector that expresses a BAG3 protein or an active fragment thereof.

3. 3. The method of claim 2, wherein the expression vector further comprises a promoter, wherein the promoter comprises an inducible promoter, a constitutive promoter, a bicistronic promoter, or a tissue-specific promoter.

4. 4. The method of claim 2 or 3, wherein the expression vector comprises a viral vector, a cardiotropic vector, a plasmid, or a yeast vector.

5. 5. The method of any one of claims 1 to 4, wherein the cardiotropic vector comprises an adenovirus vector, an adeno-associated virus vector (AAV), a coxsackievirus vector, a cytomegalovirus vector, an Epstein-Barr virus vector, a parvovirus vector, or a hepatitis virus vector.

6. The method of claim 5, wherein the AAV vector comprises a capsid protein having 90% or more sequence identity to any of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7m AAV8, AAV9, AAV10, AAV11, or AAV12.

7. The method of claim 2 or 3, wherein the expression vector is a pseudotyped viral vector.

8. 8. The method of any one of claims 1 to 7, wherein the patient has or is at risk of having heart failure with preserved ejection fraction (HFpEF).

9. The method of any one of claims 1 to 8, wherein the patient has or is at risk of having cardiac amyloid deposits or increased cardiac amyloid compared to age-, sex-, and / or race-matched controls.

10. The method of any one of claims 1 to 9, wherein the patient expresses lower than normal levels of BAB3 in cardiac tissue.

11. 11. The method of any one of claims 1 to 10, wherein the amyloidosis comprises cardiac amyloidosis or Alzheimer's disease.

12. 12. The method of claim 11, wherein the patient does not have a mutation or defect in the transthyretin (ATTAR) gene.

13. 13. The method of any one of claims 2 to 12, wherein the expression vector further comprises a promoter, optionally comprising an inducible promoter, a constitutive promoter, a bicistronic promoter, or a tissue-specific promoter.

14. The method of claim 13 , wherein the inducible promoter confers expression in cardiac tissue or the CNS.

15. The method of any one of claims 2 to 14, wherein the expression vector further comprises AAV inverted terminal repeats (ITRs).

16. The method of any one of claims 2 to 14, wherein the expression vector further comprises a polyadenylation sequence and / or a stop codon.

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