Treatment methods for cognitive impairment

Abstract

This disclosure relates to the treatment of cognitive impairment, such as age-related cognitive impairment. In certain embodiments, this disclosure describes a method for assaying a sample obtained from a subject having or suspected having cognitive impairment for one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1. Cognitive impairment is caused by neurodegenerative diseases such as Alzheimer's disease. The method further includes identifying subjects who are likely or unlikely to respond positively to plasmapheresis. In further embodiments, this disclosure describes a method for treating cognitive impairment in a subject by plasmapheresis, wherein the subject is identified, based on specific protein expression data, as likely or unlikely to respond positively to plasmapheresis. Plasmapheresis may be full and / or low-volume plasmapheresis. Kits suitable for carrying out the methods disclosed herein are also provided.

Description

TECHNICAL FIELD

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS Pursuant to 35 U.S.C. § 119(e), this application claims priority to the filing date of U.S. Provisional Patent Application No. 63 / 539,280 filed on September 19, 2023, the disclosure of which is incorporated herein by reference in its entirety.

[0002] REFERENCE TO ELECTRONIC SEQUENCE LISTING The contents of the electronic sequence listing (ALKA-034WO_SEQ_LIST.xml; size: 6,935 bytes; creation date: September 5, 2024) are incorporated herein by reference in their entirety.

[0003] I. INTRODUCTION Field The present disclosure relates to methods of treating a subject with cognitive impairment.

[0004] Background The following is provided solely as background information and is not admitted to be prior art to the present invention.

[0005] Aging is a significant risk factor for several human diseases, including cognitive impairment, cancer, arthritis, vision loss, osteoporosis, diabetes, cardiovascular disease, and stroke. Age-related neuronal degeneration is a major cause of cognitive impairment. Therefore, aging remains the single most dominant risk factor for dementia-related neurodegenerative diseases such as Alzheimer's disease (AD) (Bishop, NA et al., Neural mechanisms of aging and cognitive decline. Nature 464(7288), 529-535 (2010); Heeden, T. et al., Insights into the aging mind: a view from cognitive neuroscience. Nat. Rev. Neurosci. 5(2), 87-96 (2004); Mattson, MP, et al., Ageing and neuronal vulnerability. Nat. Rev. Neurosci. 7(4), 278-294 (2006)).

[0006] Aging affects all tissues and functions of the body, including the central nervous system, and neurodegeneration and declines in function, such as cognitive or motor skills, can have a serious impact on quality of life. Treatments for cognitive decline, motor impairment, and neurodegenerative disorders have had limited success in preventing and restoring impairment. Furthermore, while certain treatments, such as plasma-based therapies or plasma exchange therapy, have shown promise (Boada et al., Alzheimers Dement.; 16:1412-25, 2020), these treatments are not always successful. In other words, certain patients show positive results with such treatments for cognitive impairment, while others do not. This leads to the loss of valuable time and resources as the disease progresses.

[0007] Therefore, it is desirable to identify patients who are likely to respond positively to or not to treatment for cognitive impairment before such treatment is administered. [Overview of the Initiative]

[0008] II. Overview In certain embodiments, the present disclosure provides methods for treating cognitive impairment. Such treatments may include plasmapheresis. In some cases, subjects are identified as likely to respond positively or not to plasmapheresis. The determination of whether a subject is likely to respond positively or not may be based on the results of assays of a sample of the subject for one or more proteins that indicate a high responsiveness of the subject to the treatment. Certain such proteins may be one or more of delta-like canonical notch ligand 1 (DLL1), vascular non-inflammatory molecule 2 (VNN2), guanine nucleotide exchange factor (VAV3), and sulfatase modifier 1 (SUMF1). In certain embodiments, the proteins are assayed using aptamer-based multiplex proteomics assays such as the SomaScan® assay.

[0009] Accordingly, in certain embodiments, the present disclosure provides a method for assaying a sample for one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1; the subject is someone who has or is suspected of having cognitive impairment.

[0010] Cognitive impairment can be caused by neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, and vascular dementia.

[0011] Further aspects of this disclosure provide a kit comprising reagents for assaying one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1 in a sample.

[0012] Built-in by reference. All publications and patent applications referenced herein are incorporated herein by reference to the same extent as each individual publication or patent application is specifically and individually incorporated herein by reference. [Brief explanation of the drawing]

[0013] IV. Brief Description of the Drawings [Figure 1] A schematic diagram of an example of plasma exchange therapy is shown. In the example shown in Figure 1, Alzheimer's disease management with albumin replacement (AMBAR) treatment included a 6-week intensive period of conventional therapeutic PE (TPE, also called FPE - total plasma exchange) processing 1 plasma volume [≒2500-3000 mL], followed by 5% albumin replacement (1 TPE / week) and a subsequent 12-month maintenance period of low-volume PE (LVPE: removing approximately 1 / 3 of the plasma volume [≒690-880 mL]) and 20% albumin replacement or IVIG (1 LVPE / month), applied to all three active arms. In one arm, 20 grams of Albumin 20% was used for LVPE replacement, while in the other two arms, Albumin 20% replacement (20 or 40 grams) was alternated with Flebogamma DIF 5% (10 or 20 grams). IVIG (intravenous immunoglobulin): Albutein and Flebogamma DIF are commercially available products. [Figure 2A] Proteomics analysis of samples. A. A brief explanation of statistical analysis for identifying protein biomarkers. [Figure 2B] Proteomics analysis of the sample. B. Flowchart of the process for identifying one or more proteins that can indicate the subject's responsiveness to treatment for cognitive impairment. [Figure 3A] Specific examples of predicted clinical changes in subjects stratified based on the protein DLL1 are shown. A. Relationship between baseline plasma DLL1 levels and improvement in Clinical Dementia Rating-Sum of Boxes (CDR.Sb). Higher baseline levels of DLL1 are associated with improvement in CDR.Sb (measured as a change from baseline) (Spearman correlation coefficient (rho) = -0.552, area under the curve (AUC) = 0.770 for better than baseline, AUC = 0.791 for not worse than baseline). [Figure 3B]Specific examples of predicted clinical changes in subjects stratified based on protein DLL1 are shown. B. Receiver operating characteristics (ROC) of DLL1 to predict whether the CDR.Sb bl score will be better than at end of study (EOS). [Figure 3C] This shows specific examples of predicted clinical changes in subjects stratified based on protein DLL1. C. Estimated changes in the stratified DLL1 group after plasma exchange treatment. [Figure 4A] Specific examples of predicted clinical changes in subjects stratified based on protein VNN2 are shown. A. Relationship between baseline plasma VNN2 levels and improvement in CDR.Sb. Higher baseline levels of VNN2 are associated with improvement in CDR.Sb (rho=-0.476, AUC=0.761 for better than baseline, AUC=0.726 for not worse than baseline). [Figure 4B] This shows specific examples of predicted clinical changes in stratified subjects based on protein VNN2. ROC of VNN2 for predicting whether CDR.Sb is better than bl score in B.EOS. [Figure 4C] This shows specific examples of predicted clinical changes in subjects stratified based on protein VNN2. C. Estimated changes in CDR.Sb in stratified VNN2 groups after plasma exchange. [Figure 5A] Specific examples of predicted clinical changes in subjects stratified based on the protein VAV3 are shown. A. Relationship between baseline plasma VAV3 levels and improvement in CDR.Sb. Higher baseline levels of VAV3 are associated with improvement in CDR.Sb (rho=-0.494, AUC=0.788 for better than baseline, AUC=0.724 for not worse than baseline). [Figure 5B] This shows specific examples of predicted clinical changes in stratified subjects based on the protein VAV3. ROC of VAV3 for predicting whether CDR.Sb is better than bl score in B.EOS. [Figure 5C]This shows specific examples of predicted clinical changes in subjects stratified based on the protein VAV3. C. Estimated changes in CDR.Sb in the stratified VAV3 group after plasma exchange. [Figure 6A] Specific examples of predicted clinical changes in subjects stratified based on the protein SUMF1 are shown. A. Relationship between baseline plasma SUMF1 levels and improvement in CDR.Sb. Higher baseline levels of SUMF1 are associated with improved CDR.Sb (rho=-0.489, AUC=0.696 for better than baseline, AUC=0.756 for not worse than baseline). [Figure 6B] This shows specific examples of predicted clinical changes in stratified subjects based on the protein SUMF1. ROC of SUMF1 for predicting bl score or higher in CDR.Sb in B.EOS. [Figure 6C] This shows specific examples of predicted clinical changes in subjects stratified based on the protein SUMF1. C. Estimated changes in CDR.Sb in the stratified SUNF1 group after plasma exchange. [Modes for carrying out the invention]

[0014] V. Detailed explanation A. Introduction The present invention relates to the treatment of cognitive disorders, for example, aging-related cognitive disorders. In certain aspects, the present disclosure provides a method of assaying a sample obtained from a subject having or suspected of having a cognitive disorder for one or more proteins selected from DLL1, VNN2, VAV3 and SUMF1. In certain embodiments, the cognitive disorder is caused by a neurodegenerative disease such as AD. The method further comprises identifying subjects who are more or less likely to respond positively to plasma exchange therapy. In a still further aspect, the present disclosure describes a method of treating a cognitive disorder in a subject by plasma exchange therapy, wherein the subject is identified as being more or less likely to respond positively to plasma exchange therapy based on specific protein expression data. The plasma exchange therapy may be full and / or low-volume plasma exchange. In some cases, plasma exchange comprises albumin replacement. Kits suitable for carrying out the methods of the present disclosure are also provided.

[0015] Before describing the present invention in detail, it is to be understood that this disclosure is not limited to the particular methods or compositions described, and as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, as the scope of the present disclosure is to be limited solely by the appended claims.

[0016] The publications discussed herein are provided solely for their disclosure prior to the filing date of the present application. Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication presented may differ from the actual publication dates, which may need to be independently confirmed.

[0017] Where a range of values ​​is provided, unless explicitly indicated in the context, each intermediate value between the upper and lower limits of that range, up to one-tenth of the lower limit, is also specifically disclosed. Each smaller range between any stated value or intermediate value within the stated range and any other stated value or intermediate value within that stated range is included in the present invention. The upper and lower limits of these smaller ranges may be included in or excluded from the range independently, and each range that includes either limit, does not include either limit, or includes both limits is also included in the present invention, subject to any specifically excluded limits within the stated range. Where a stated range includes one or both limits, the range excluding one or both of the limits that they include is also included in the present invention.

[0018] It should be noted that claims may be drafted to exclude optional elements. Therefore, this statement is intended to serve as a prior art for using exclusive terms such as “exclusively” or “only” in relation to the enumeration of elements in the claims or the use of “negative” limitations.

[0019] As will be apparent to those skilled in the art upon reading this disclosure, each of the individual embodiments described and illustrated herein has separate components and features that can be readily separated from or combined with features of any of several other embodiments without departing from the scope or spirit of the invention. Any method of description may be carried out in the order of the described events or in any other logically possible order.

[0020] B. Definition Unless otherwise defined, all scientific and technical terms used herein have the same meaning as those generally understood by those skilled in the art to which the invention pertains. Any methods and materials similar to or equivalent to those described herein may be used in the practice or testing of the invention, but some potential and preferred methods and materials are described below. All publications referenced herein are incorporated herein by reference to disclose and describe the relevant methods and / or materials from which the publications are cited. This disclosure is understood to supersede any disclosure in the incorporated publications to the extent that a conflict exists.

[0021] It should be noted that, as used herein and in the appended claims, the singular forms "a," "an," and "the" refer to multiple objects unless the context clearly indicates otherwise. Therefore, for example, a reference to "sample" includes multiple such samples.

[0022] In describing the methods of the present invention, the terms “host,” “subject,” “individual,” and “patient” are used interchangeably and refer to any mammal that requires such treatment by the disclosed methods. Such mammals include, for example, humans, sheep, cattle, horses, pigs, dogs, cats, non-human primates, mice, and rats. In certain embodiments, the subject is a non-human mammal. In some embodiments, the subject is livestock. In other embodiments, the subject is a pet. In some embodiments, the subject is a mammal. In certain cases, the subject is a human. Other subjects may include domestic pets (e.g., dogs and cats), livestock (e.g., cattle, pigs, goats, horses, etc.), rodents (e.g., mice, guinea pigs, and rats, such as in animal models of diseases), and non-human primates (e.g., chimpanzees and monkeys). Thus, the subjects of the present invention include, but are not limited to, mammals, such as humans and other primates, such as chimpanzees and other apes and monkey species, and in certain embodiments, the subject is a human. The term "subject" also includes any person or organism of any age, weight or other physical characteristics, and the subject may be an adult, child, infant or newborn.

[0023] As used herein, “treatment” means the reduction or elimination of cognitive impairment. Treatment may be administered therapeutically, i.e., after the onset of the disease. Effects may include improvement of cognitive abilities in subjects with cognitive impairment. Thus, as used herein, the term “treatment” encompasses any treatment of cognitive impairment in mammals, including (a) inhibiting the disease, i.e., halting its progression; or (b) alleviating the disease, i.e., causing disease regression. Treatment may result in a variety of different physical manifestations, such as regulation of gene expression, rejuvenation of tissues or organs, etc. Therapies may be administered during or after the onset of cognitive impairment. Treatment of ongoing cognitive impairment that stabilizes or reduces the patient’s undesirable clinical symptoms are of interest. Such treatments may be administered before the complete loss of function in the affected tissue. Therapies of subjects may be administered during the symptomatic phase of the disease, and in some cases after the symptomatic phase.

[0024] "Cognitive impairment" is a impairment of an individual's cognitive abilities compared to a healthy individual, for example, a healthy individual matched for the same age, or to an earlier point in time, for example, two weeks ago, one month ago, two months ago, three months ago, six months ago, one year ago, two years ago, five years ago, or ten years ago, or earlier.

[0025] "Cognitive ability" or "cognition" means mental processes including attention and concentration, learning complex tasks and concepts, memory (acquiring, retaining, and retrieving new information in the short and / or long term), information processing (processing information gathered through the five senses), visuospatial functioning (visual perception, depth perception, use of mental images, copying drawings, constructing objects or shapes), language production and comprehension, language fluency (word discovery), problem solving, decision-making, and executive functioning (planning and prioritizing). "Cognitive decline" means a progressive decline in one or more of these abilities, such as a decline in memory, language, thinking, or judgment.

[0026] In some embodiments, cognitive impairment is “age-related cognitive impairment.” “Age-related cognitive impairment” typically means impairment of cognitive abilities associated with aging, and includes, for example, cognitive impairment associated with the natural aging process, such as mild cognitive impairment (MCI); and cognitive impairment associated with age-related impairment, i.e., impairments that are seen more frequently with the progression of old age, such as cognitive impairment associated with neurodegenerative conditions such as AD, Parkinson's disease, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, and vascular dementia.

[0027] In some embodiments, the cognitive impairment in question is caused by neuroinflammation, for example, age-related increases in neuroinflammation in an individual. "Neuroinflammation" refers to the biochemical and cellular response of the nervous system to injury, infection, or neurodegenerative disease. Such a response is aimed at reducing the triggers by engaging the central nervous system immune system to defend against potential harm. Neurodegeneration occurs in the central nervous system and is characterized by loss of neuronal structure and function. Neuroinflammatory diseases or neuroinflammation-related conditions or diseases include, but are not limited to, neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and multiple sclerosis.

[0028] C. Assay Method Certain aspects of this disclosure provide a method for analyzing a sample obtained from a subject, and this method is: This involves assaying a sample for one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1; The subjects are those who have or are suspected of having a cognitive impairment.

[0029] The DLL1 protein is found in several organisms, including humans. In humans, one example of the DLL1 protein is listed in the Uniprot database with ID NO.: O00548. The sequence of the human DLL1 protein is shown below: (Sequence ID 1).

[0030] Accordingly, in certain embodiments, the methods disclosed herein include assaying samples from subjects with or suspected of having cognitive impairment for a protein having the sequence of SEQ ID NO: 1 or a protein having at least 70%, for example, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to the sequence in the case of SEQ ID NO: 1. For example, those skilled in the art can readily identify homologs of the human DLL1 protein having the sequence of SEQ ID NO: 1 in an organism of interest, and then assay such DLL1 proteins in samples obtained from such organisms. Such embodiments are within the scope of this disclosure.

[0031] The VNN2 protein is also found in several organisms, including humans. In humans, one example of the VNN2 protein is listed in the Uniprot database with ID NO.: O95498. The sequence of the human VNN2 protein is shown below: MVTSSFPISVAVFALITLQVGTQDSFIAAVYEHAVILPNKTETPVSQEDALNLMNENIDILETAIKQAAEQGARIIVTPEDALYGWKFTRETVFPYLEDIPDPQVNWIPCQDPHRFGHTPVQARLSCLAKDN SIYVLANLGDKKPCNSRDSTCPPNGYFQYNTNVVYNTEGKLVARYHKYHLYSEPQFNVPEKPELVTFNTAFGRFGIFTCFDIFFYDPGVTLVKDFHVDTILFPTAWMNVLPLLTAIEFHSAWAMGMGVNLLV ANTHHVSLNMTGSGIYAPNGPKVYHYDMKTELGKLLLSEVDSHPLSSLAYPTAVNWNAYATTIKPFPVQKNTFRGFISRDGFNFTELFENAGNLTVCQKELCCHLSYRMLQKEENEVYVLGAFTGLHGRRRREYWQVCTLLKCKTTNLTTCGRPVETASTRFEMFSLSGTFGTEYVFPEVLLTEIHLSPGKFEVLKDGRLVNKNGSSGPILTVSLFGRWYTKDSLYSSCGTSNSAITYLLIFILLMIIALQNIVML(Sequence ID 2).

[0032] Accordingly, in certain embodiments, the methods disclosed herein include assaying samples from subjects with or suspected of having cognitive impairment for a protein having the sequence of SEQ ID NO: 2 or a protein having at least 70%, for example, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to the sequence in the case of SEQ ID NO: 2. For example, a person skilled in the art can readily identify homologs of the human VNN2 protein having the sequence of SEQ ID NO: 2 in an organism of interest, and then assay such VNN2 proteins in samples obtained from such organisms. Such embodiments are within the scope of this disclosure.

[0033] Similarly, the VAV3 protein is also found in several organisms, including humans. In humans, one example of the VAV3 protein is listed in the Uniprot database with ID NO.:Q9UKW4. The sequence of the human VAV3 protein is shown below: (Sequence ID 3).

[0034] Accordingly, in certain embodiments, the methods disclosed herein include assaying samples from subjects with or suspected of having cognitive impairment for a protein having the sequence of SEQ ID NO: 3 or a protein having at least 70%, for example, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to the sequence in the case of SEQ ID NO: 3. For example, a person skilled in the art can readily identify homologs of the human VAV3 protein having the sequence of SEQ ID NO: 3 in an organism of interest, and then assay such VAV3 proteins in samples obtained from such organisms. Such embodiments are within the scope of this disclosure.

[0035] Similarly, the SUMF1 protein is also found in several organisms, including humans. In humans, one example of the SUMF1 protein is listed in the Uniprot database with ID NO.: P01042. The sequence of the human SUMF1 protein is shown below: MAAPALGLVCGRCPELGLVLLLLLLSLLCGAAGSQEAGTGAGAGSLAGSCGCGTPQRPGAHGSSAAAHRYSREANAPGPVPGERQLAHSKMVPIPAGVFTMGTDDPQIKQDGEAPARRVTIDAFYMDAYEVSNTEFEKFVNSTGYLTEAEKFGDSFVFEGMLSEQVKTNIQQAVAAAPWWLPVKGANWRHPEGPDSTILHRPDHPVLHVSWNDAVAYCTWAGKRLPTEAEWEYSCRGGLHNRLFPWGNKLQPKGQHYANIWQGEFPVTNTGEDGFQGTAPVDAFPPNGYGLYNIVGNAWEWTSDWWTVHHSVEETLNPKGPPSGKDRVKKGGSYMCHRSYCYRYRCAARSQNTPDSSASNLGFRCAADRLPTMD(Sequence ID 4).

[0036] Accordingly, in certain embodiments, the methods disclosed herein include assaying samples from subjects with or suspected of having cognitive impairment for a protein having the sequence of SEQ ID NO: 4 or a protein having at least 70%, for example, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% sequence identity to the sequence in the case of SEQ ID NO: 4. For example, a person skilled in the art can readily identify homologs of the human SUMF1 protein having the sequence of SEQ ID NO: 4 in an organism of interest, and then assay such SUMF1 proteins in samples obtained from such organisms. Such embodiments are within the scope of this disclosure.

[0037] The protein(s) being assayed are also referred to herein as “target protein(s).”

[0038] In some cases, the methods disclosed herein include qualitatively assaying a sample for one or more target proteins.

[0039] Certain non-limiting examples of qualitative assays of target proteins include determining the sequence of a target protein(s) and / or post-translational modifications of a target protein(s).

[0040] Any suitable method may be used for such a qualitative assay of one or more target proteins in a sample. Samples may be obtained from subjects who have or are suspected of having cognitive impairment. Samples may be any convenient biological samples. "Biological sample" means, but is not limited to, both a naturally occurring organism or a subset of its tissues, as well as homogenates, lysates, or extracts prepared from a subset of an organism or its tissues, including, for example, plasma, serum, cerebrospinal fluid, lymph, skin, respiratory, gastrointestinal, cardiovascular and urogenital tract sections, tears, saliva, milk, blood cells, tumors, and organs. A biological sample may be any type of living tissue, including both healthy tissue and diseased tissue (e.g., cancerous, malignant, necrotic, etc.). In certain embodiments, the biological sample is a liquid sample such as blood or its derivatives, e.g., plasma, tears, urine, semen, and in some cases, the sample is a blood sample including whole blood, such as blood obtained from venous or fingertip puncture (the blood may or may not be combined with any reagents such as preservatives and anticoagulants before the assay). Examples of additional biological samples are well known to those skilled in the art, and such embodiments are within the scope of the present invention. Methods for obtaining such samples from subjects are also well known in the art, and such embodiments are within the scope of this disclosure.

[0041] In some cases, the levels of proteins described herein are analyzed in the sample of interest using the SomaScan® assay. Specific details of the SomaScan® assay are described in reference to Gold et al. (2010), PLoS One; 5(12):e15004, which is incorporated herein by reference in its entirety. Briefly, the SomaScan® assay uses aptamers called SOMAmers that specifically bind to target proteins. The assay involves binding SOMAmers to the target protein in the sample and isolating the SOMAmers, thereby generating a complex mixture of SOMAmers containing SOMAmers in proportion to the proteins present in the sample. The resulting mixture of SOMAmers is quantified using DNA microarray technology to generate a readout of relative fluorescence units (RFUs). Thus, the SomaScan® assay provides levels of various proteins in the sample as “relative fluorescence units” (RFUs) or derivatives of RFUs such as log2RFUs.

[0042] Further non-limiting examples of methods that may be used to determine the sequence of a target protein include Edman degradation using mass spectrometry or a protein sequencer.

[0043] Non-limiting examples of methods that may be used to determine post-translational modifications of target proteins include mass spectrometry and antibody-binding assays based on post-translational modifications, such as Western blotting or immunoassays.

[0044] In some cases, the methods disclosed herein involve quantitatively assaying a sample for one or more target proteins. Specific non-limiting examples of quantitatively assaying one or more target proteins include immunoassays, mass spectrometry, and protein detection array analysis.

[0045] An immunoassay typically involves contacting a sample with a binder that specifically binds to a target protein, and then detecting the binding between the binder and the target protein. Such detection may involve detecting a label conjugated to the binder, or a label conjugated to a second binder that specifically binds to the binder and / or the binder-target protein complex. The binder is typically an antibody or an antigen-binding fragment of an antibody. The binder can also be an aptamer or a peptide bond member.

[0046] Non-exclusive examples of immunoassays include Western blot analysis, enzyme-linked immunosorbent assays (ELISA), radioimmunoassays (RIA), lateral flow immunoassays, particle-based immunoassays, and quantum dot-based immunoassays. Details of methods for performing specific exemplary immunoassays are described in the review article by Rizzo (2022), Chemosensors, 10(8), 326, which is incorporated herein by reference in its entirety.

[0047] Mass spectrometry for detecting target proteins typically involves digesting a protein-containing sample into peptides, ionizing the peptides, and analyzing the ionized peptides to determine the identity of the digested protein. Due to the unique sequence of the protein, specific digestion of the target protein produces unique digested and ionized peptides, which are identified by mass spectrometry. The presence and quantity of such unique ionized peptides facilitate the identification of the target protein.

[0048] Non-limiting examples of mass spectrometry methods for assaying one or more target proteins include quadrupole mass spectrometry, time-of-flight mass spectrometry, magnetic sector mass spectrometry, electrostatic sector mass spectrometry, quadrupole ion trap mass spectrometry, and ion cyclotron resonance spectrometry. Details of methods for performing specific exemplary mass spectrometry-based assays are described in the review article Ma (2022), Molecules, 27, 6466, which is incorporated herein by reference in its entirety.

[0049] Protein detection array analysis involves detecting the binding of a target protein in a sample to a binder, typically an antibody localized at a predetermined location on a support. The sample is brought into contact with such an array, and the binding of the target protein to the corresponding binder, such as an antibody, at a localized spot on a solid support is visualized, for example, using detectable labeling. For example, the captured target protein can be detected and / or quantified by a labeled secondary antibody, such as a fluorescent dye-labeled secondary antibody.

[0050] Instead of localized binders on a solid support, lysate microarrays involve immobilizing the sample lysate onto a support, such as a nitrocellulose-coated glass slide. The immobilized proteins are then detected using a detectable labeled conjugate solution-phase binder, such as a fluorescently labeled solution-phase-specific antibody. Differentially labeled binders, such as various fluorescently labeled antibodies against different target proteins, enable multiple detection of proteins and thus allow simultaneous assay of two or more proteins.

[0051] Non-limiting examples of protein detection array analysis include analytical protein microarrays, functional protein microarrays, and reversed-phase protein microarrays. Those skilled in the art can determine which protein microarrays are suitable for use in the methods disclosed herein. Details of methods for performing specific exemplary protein detection array analysis are described in the review article by Neagu et al. (2019), World Acad. Sci. J., 1:113-124, which is incorporated herein by reference in its entirety.

[0052] In some cases, this method involves assaying the sample for one of DLL1, VNN2, VAV3, and SUMF1.

[0053] In some cases, this method involves assaying a sample for any two of DLL1, VNN2, VAV3, and SUMF1, for example, the following combinations: DLL1 and VNN2; DLL1 and VAV3; DLL1 and SUMF1; VNN2 and VAV3; VNN2 and SUMF1; and VAV3 and SUMF1.

[0054] In further embodiments, the method includes assaying a sample for any three of DLL1, VNN2, VAV3, and SUMF1, for example, the following combinations: DLL1, VNN2, and VAV3; DLL1, VAV3, and SUMF1; DLL1, VNN2, and SUMF1; and VNN2, VAV3, and SUMF1.

[0055] In certain embodiments, the method includes assaying the sample for all four components: DLL1, VNN2, VAV3, and SUMF1.

[0056] In some cases, this method involves assaying a sample for only one of DLL1, VNN2, VAV3, and SUMF1, and in some cases, not assaying additional proteins except for a specific control protein.

[0057] In some cases, this method involves assaying a sample for only two of DLL1, VNN2, VAV3, and SUMF1, and not assaying additional proteins except for specific control proteins. For example, the following protein combinations may be assayed: DLL1 and VNN2; DLL1 and VAV3; DLL1 and SUMF1; VNN2 and VAV3; VNN2 and SUMF1; and VAV3 and SUMF1, with one or more control proteins also being assayed.

[0058] In further embodiments, the method includes assaying a sample for only three of DLL1, VNN2, VAV3, and SUMF1, and not assaying other proteins except for a specific control protein. For example, the following protein combinations may be assayed: DLL1, VNN2, and VAV3; DLL1, VAV3, and SUMF1; DLL1, VNN2, and SUMF1; and VNN2, VAV3, and SUMF1, with one or more control proteins also being assayed.

[0059] In certain embodiments, the method involves assaying a sample for only the following four proteins: DLL1, VNN2, VAV3, and SUMF1, and in some cases not assaying additional proteins except for a specific control protein.

[0060] As used herein, “control protein” refers to a protein that does not distinguish between subjects likely to respond positively to plasmapheresis for the treatment of cognitive impairment and subjects not likely to respond positively to plasmapheresis for the treatment of cognitive impairment. In other words, the level of the control protein is the same in subjects regardless of their responsiveness to plasmapheresis for the treatment of cognitive impairment. Those skilled in the art will know or be able to easily determine such proteins.

[0061] Methods for assaying one or more proteins disclosed herein may further include comparing the assay results to a threshold, reference value, or reference sample.

[0062] A threshold or reference value may be predetermined, for example, based on the concentration of one or more assayed proteins in individuals known to have responded to therapy for cognitive impairment. For example, retrospective assays of one or more proteins may be performed in a group of subjects who have responded to therapy for cognitive impairment. These retrospective assays can be used to determine a threshold or reference value that actually indicates the likelihood that a subject will respond to therapy for cognitive impairment. In certain embodiments, the therapy is, for example, plasmapheresis, as described in detail below.

[0063] Alternatively, such thresholds or reference values ​​may be predetermined based, for example, on the concentrations of one or more assayed proteins in individuals known not to have responded to therapy for cognitive impairment. For example, retrospective assays of one or more proteins may be performed in a group of subjects who did not respond to therapy for cognitive impairment. These retrospective assays may be used to determine thresholds or reference values ​​that actually indicate the likelihood that a subject will respond to therapy for cognitive impairment. In certain embodiments, the therapy is plasmapheresis, as described in detail below.

[0064] The reference sample may contain one or more target proteins at concentrations known to show a high response to plasma exchange therapy for treating cognitive impairment.

[0065] Samples for assaying one or more target proteins in the methods disclosed herein may be obtained from subjects with cognitive impairment. Alternatively, samples for assaying one or more target proteins in the methods disclosed herein may be obtained from subjects suspected of having cognitive impairment.

[0066] As used herein, the term "individual with cognitive impairment" refers to an individual exhibiting symptoms of cognitive impairment.

[0067] As used herein, the term “individual suspected of having cognitive impairment” refers to a subject exhibiting symptoms of other conditions associated with cognitive impairment. For example, as will be detailed later, a subject may exhibit certain signs of a neurodegenerative disease such as AD or Parkinson’s disease, but have not yet developed cognitive impairment. Such subjects are included in the term “subject suspected of having cognitive impairment.”

[0068] In certain embodiments, cognitive impairment is caused by neurodegenerative diseases. Non-exclusive examples of neurodegenerative diseases that can cause cognitive impairment include Alzheimer's disease (AD), Parkinson's disease, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, and vascular dementia. These diseases are described below, and additional information regarding these diseases is well known in the art.

[0069] In certain embodiments, the neurodegenerative disease is AD, i.e., the subject has cognitive impairment caused by AD.

[0070] In certain embodiments, the sample is obtained from a subject who is a candidate for plasma exchange therapy to treat cognitive impairment.

[0071] Any suitable sample from the subject can be assayed according to the methods disclosed herein. In certain embodiments, the sample is a blood sample, serum sample, plasma sample, or cerebrospinal fluid sample.

[0072] In some cases, the sample may be aqueous humor, vitreous fluid, bile, chyle, endolymph, perilymph, lymph, mucus (including nasal discharge and sputum), pericardial fluid, ascites, pleural fluid, pus, eye discharge, saliva, sputum, or synovial fluid. Examples of additional biological samples are well known to those skilled in the art, and such embodiments are within the scope of the present invention. Methods for obtaining such samples from subjects are also well known in the art, and such embodiments are within the scope of this disclosure.

[0073] D. Diagnostic Methods Certain aspects of this disclosure further provide assaying one or more proteins in a sample obtained from a subject having or suspected of having cognitive impairment, and determining, based on the assay results, whether or not the subject is likely to respond positively to a therapy for treating cognitive impairment in the subject. In some embodiments, the therapy is plasma exchange therapy. Details of plasma exchange therapies that can be administered in accordance with this disclosure are discussed in detail below and are applicable to the diagnostic methods described herein.

[0074] In some embodiments, subjects are identified as likely or unlikely to respond positively to plasmapheresis for cognitive impairment based on the results of assays of the subject sample for one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1. For example, a subject may be identified as likely or unlikely to respond positively to plasmapheresis for cognitive impairment based on the results of assays of the subject sample for DLL1. Alternatively, a subject may be identified as likely or unlikely to respond positively to plasmapheresis for cognitive impairment based on the results of assays of the subject sample for VNN2. Furthermore, a subject may be identified as likely or unlikely to respond positively to plasmapheresis for cognitive impairment based on the results of assays of the subject sample for VAV3. Furthermore, a subject may be identified as likely or unlikely to respond positively to plasmapheresis for cognitive impairment based on the results of assays of the subject sample for SUMF1.

[0075] In further embodiments, subjects can be identified as likely or unlikely to respond positively to plasmapheresis for treating cognitive impairment based on the results of assays of the subject's sample for two or more proteins selected from DLL1, VNN2, VAV3, and SUMF1. Such combinations include DLL1 and VNN2; DLL1 and VAV3; DLL1 and SUMF1; VNN2 and VAV3; VNN2 and SUMF1; and VAV3 and SUMF1.

[0076] Furthermore, subjects can be identified as likely or unlikely to respond positively to plasmapheresis for treating cognitive impairment based on the results of assays of their samples for three or more proteins selected from DLL1, VNN2, VAV3, and SUMF1. Such combinations include DLL1, VNN2, and VAV3; DLL1, VAV3, and SUMF1; DLL1, VNN2, and SUMF1; and VNN2, VAV3, and SUMF1.

[0077] Furthermore, subjects can be identified as likely or unlikely to respond positively to plasmapheresis for treating cognitive impairment based on the assay results of their samples for all proteins DLL1, VNN2, VAV3, and SUMF1.

[0078] In certain embodiments, determining whether a subject is likely to respond positively to a therapy for treating cognitive impairment in the subject based on the assay results includes comparing the assay results to a threshold, i.e., a reference value.

[0079] As described above, such thresholds or reference values ​​may be predetermined, for example, based on the concentrations of one or more assayed proteins in individuals known to have responded to therapy for cognitive impairment. For example, retrospective assays of one or more proteins may be performed in a group of subjects that have responded to therapy for cognitive impairment. These retrospective assays may be used to determine the thresholds or reference values ​​that actually indicate the likelihood that a subject will respond to therapy for cognitive impairment. In certain embodiments, the therapy is plasmapheresis, as described in detail below.

[0080] Furthermore, as described above, such thresholds or reference values ​​may be predetermined based, for example, on the concentrations of one or more assayed proteins in individuals known not to have responded to therapy for cognitive impairment. For example, retrospective assays of one or more proteins may be performed in a group of subjects who did not respond to therapy for cognitive impairment. These retrospective assays may be used to determine thresholds or reference values ​​that actually indicate the likelihood that a subject will respond to therapy for cognitive impairment. In certain embodiments, the therapy is plasma exchange therapy, as described in detail below.

[0081] The reference sample may contain one or more target proteins at concentrations known to show a high response to plasma exchange therapy for treating cognitive impairment.

[0082] In some embodiments, subjects are identified as likely or unlikely to respond positively to plasmapheresis to treat cognitive impairment based on increased levels of one or more assayed proteins in a sample obtained from the subject compared to a threshold or reference value or reference sample.

[0083] As described above, in some cases, the levels of the proteins described herein are analyzed in patient samples using the SomaScan® assay. Tables 2 and 3 below provide specific thresholds for proteins quantified using the SomaScan® assay that indicate a high response to therapy for cognitive impairment in the target population.

[0084] Therefore, log2RFU values ​​determined by the SomaScan® assay for DLL protein in a sample from a subject, such as at least 9–13, e.g., at least 9, at least 10, at least 11, at least 12, at least 13, or at least 11.2, indicate that the subject is likely to respond positively to therapies such as plasmapheresis to treat cognitive impairment. In some cases, plasmapheresis is as described in Figure 1 or in the experimental section below.

[0085] Similarly, log2RFU values ​​determined by the SomaScan® assay for VNN2 protein in a sample from a subject, such as at least 8–12, e.g., at least 8, at least 9, at least 10, at least 11, at least 12, or at least 10.6, indicate that the subject is likely to respond positively to therapies such as plasmapheresis to treat cognitive impairment. In some cases, plasmapheresis is as described in Figure 1 or in the experimental section below.

[0086] Furthermore, log2 RFU values ​​determined by the SomaScan® assay for VAV3 protein in a sample from a subject, which are at least 8–12, for example, at least 8, at least 9, at least 10, at least 11, at least 12, or at least 10.4, indicate that the subject is likely to respond positively to therapies such as plasmapheresis to treat cognitive impairment. In some cases, plasmapheresis is as described in Figure 1 or in the experimental section below.

[0087] Furthermore, log2RFU values ​​determined by the SomaScan® assay for SUMF1 protein in a sample from a subject, which are at least 10–14, for example, at least 10, at least 11, at least 12, at least 13, at least 14, or at least 12.1, indicate that the subject is likely to respond positively to therapies such as plasmapheresis to treat cognitive impairment. In some cases, plasmapheresis is as described in Figure 1 or in the experimental section below.

[0088] As also stated above, the protein levels of the proteins described herein can be determined using techniques other than the SomaScan® assay. Based on the SomaScan® assay results and associated log2RFU values ​​of the proteins described herein, those skilled in the art can determine the actual protein concentrations that indicate whether a patient is likely to respond to or not to respond positively to therapies such as plasmapheresis for treating cognitive impairment. Certain such techniques are described above in the discussion of suitable methods that may be used for such qualitative assays of one or more target proteins in a sample, and such embodiments are within the scope of this disclosure.

[0089] When specific methods other than the SomaScan® assay are used, those skilled in the art can determine appropriate thresholds or reference values ​​or reference samples that can be used to compare the results of the assay to determine elevated levels of one or more proteins. For example, such thresholds or reference values ​​may indicate the highest levels of one or more proteins that may show high or low responsiveness to plasma exchange therapy for cognitive impairment. Thus, levels of one or more proteins above such thresholds or reference values ​​indicate that the subject is responsive to plasma therapy for treating cognitive impairment.

[0090] In some cases, the results of an assay for a single protein can be compared to a threshold or reference value for that corresponding protein. As those skilled in the art will recognize, the control sample or reference value for comparison depends on the type of sample, e.g., blood sample or cerebrospinal fluid sample.

[0091] In certain other cases, the results of assays for two, three, or four proteins can be compared to thresholds or reference values ​​for corresponding combinations of two, three, or four proteins. When comparing results for two or more proteins, specific statistical analyses can be performed on the results for individual proteins to arrive at combination results for two or more proteins, which can then be compared to thresholds or reference values.

[0092] For example, results for a combination of two proteins may indicate a statistically increased level of the two proteins when the combination is considered to exist even if one of the proteins shows no change or even a decrease in level. Similarly, results for a combination of three proteins may indicate a statistically increased level of the three proteins when the combination is considered to exist even if one or two proteins show no change or even a decrease in level. Furthermore, results for a combination of four proteins may indicate a statistically increased level of the four proteins when the combination is considered to exist even if one, two, or three proteins show no change or even a decrease in level. Statistical analyses for obtaining combined values ​​for the levels of two or more proteins are well known in the art. For example, in some cases, protein ratios and multivariate linear and nonlinear modeling can be used to compare protein expression data.

[0093] statistical analysis The following are specific terms used in the statistical analysis of data obtained by analyzing bodily fluid samples from subjects. These statistical analyses can be used to identify, based on the subject's biomarker levels, whether or not the subject is likely to respond positively to treatment for cognitive impairment.

[0094] First, the benefit of treatment was confirmed by comparing the change in clinical outcomes between the treatment group and the placebo group. Next, candidate proteomics biomarkers were identified by a specific assay (here, the SomaScan assay, using a significant Spearman correlation coefficient between baseline protein levels, expressed as log2 (RFU), and the change in cognitive outcomes at the end of the study). The predictive power of these candidate proteomics biomarkers indicating whether subjects have a better cognitive outcome after treatment (treatment benefit) or not (no treatment benefit) was assessed by receiver operating characteristics (ROC), measured by the area under the curve (AUC) of the ROC. ROC is a plot of sensitivity to specificity for multiple thresholds for a biomarker. ROC is used to show the predictive power of continuous biomarkers for binary outcomes (e.g., having treatment benefit vs. not having treatment benefit).

[0095] AUC represents the probability of correctly ordering patients, regardless of whether they receive a treatment benefit, based on sequential biomarkers. Typically, AUC > 70% is considered acceptable, and AUC > 80% is considered to have excellent predictive power.

[0096] Next, high biomarker (here, baseline protein level) and low biomarker levels are defined by an optimal decision threshold (here, the best cutoff point or reference value of baseline protein level) based on Youden's index that maximizes accuracy (the overall percentage of correct classifications shown in Table 1 below). The best binary biomarkers were ultimately identified based on their predictive power, measured by accuracy, specificity, and sensitivity (their calculations are described below and shown in Table 1).

[0097] The A1, B1, A0, and B0 used in the following paragraphs are as shown in Table 1 below. [Table 1]

[0098] Accuracy represents the percentage of correctly classified cases, i.e., (number of cases that truly benefited from treatment + number of cases that did not truly benefit from treatment) / total number of predictions. ((A1 + B0) / N, where N = total number of predictions.)

[0099] Specificity represents the true negative rate, i.e., the percentage of subjects who truly did not benefit from treatment out of the total number of subjects predicted not to benefit from treatment (B0 / (B1+B0)).

[0100] Sensitivity represents the true positive rate, i.e., the percentage of subjects who actually benefit from treatment out of the total number of subjects predicted to benefit from treatment (A1 / (A1+A0)).

[0101] These identified biomarkers were then validated in silico, including in other clinical outcomes, randomly re-selected samples, and trials in other visits or / or similar clinical trials.

[0102] According to the methods described herein, at least 60%, e.g., at least 70%, at least 80%, or at least 90% of subjects identified as likely to respond positively to treatment for cognitive impairment actually respond positively to the treatment. In some cases, plasmapheresis is performed as described in Figure 1 or in the experimental section below.

[0103] Conversely, according to the methods described herein, at least 60%, e.g., at least 70%, at least 80%, or at least 90% of patients identified as unlikely to respond positively to treatment for cognitive impairment actually do not respond positively to the treatment. In some cases, plasmapheresis is as described in Figure 1 or in the experimental section below.

[0104] In some cases, patients identified as unlikely to respond positively to treatments such as plasmapheresis for cognitive impairment are not subjected to such treatments.

[0105] In some cases, the baseline biomarker levels indicating whether a subject is likely or unlikely to respond positively to plasma therapy for cognitive impairment are as shown in Table 2 below: [Table 2] [Table 3]

[0106] In some embodiments, the method may include providing a report indicating whether a subject is likely or unlikely to respond to plasma therapy for treating cognitive impairment. In some embodiments, this step may include calculating a score based on the results of assays of one or more proteins, the score may be a number such as probability, likelihood, or a score out of 10, which correlates with responsiveness. In these embodiments, the method may include inputting the amount of each protein into one or more algorithms or calculations, performing the algorithms or calculations, and receiving a score based on the calculations. In these embodiments, other measures from the subject, such as whether the subject is male or female, the subject's age, and the current degree of cognitive impairment, may be taken into consideration in the algorithms or calculations.

[0107] In some embodiments, the method may include, for example, creating a report in electronic format and forwarding the report to a physician or other healthcare professional to help identify an appropriate action process, such as identifying a subject as suitable for plasmapheresis to treat cognitive impairment. The report may be used in conjunction with other metrics to determine an appropriate action process.

[0108] In some cases, a report can be forwarded to a “remote location,” where “remote location” means a location other than where the report is generated. For example, a remote location could be another location within the same city (e.g., an office, laboratory, etc.), another location in a different city, another location in a different state, or another location in a different country. Thus, if one item is indicated as “away” from another item, the two items could be in the same room but separated, or in at least different rooms or different buildings, and could be at least one mile, ten miles, or at least 100 miles apart. To “communicate” information means to transmit data representing that information as electrical signals over an appropriate communication channel (e.g., a private or public network). To “transfer” an item means any means of moving that item from one location to another, whether by physical transport or by any other means (where possible), and, at least in the case of data, includes physically transporting the medium carrying the data or communicating the data. Examples of communication media include wireless or infrared transmission channels, as well as network connections to other computers or network devices, and the Internet, or email transmissions and information recorded on websites. In certain embodiments, the report may be analyzed by an MD or other qualified medical professional, and the report based on the results of the image analysis may be forwarded to the subject from which the sample was obtained.

[0109] In computer-related embodiments, the system may include a computer comprising a processor, a storage component (i.e., memory), a display component, and other components typically found in a general-purpose computer. The storage component stores processor-accessible information, including instructions that can be executed by the processor and data that can be retrieved, manipulated, or stored by the processor.

[0110] The memory component contains instructions for determining whether a subject is likely to respond to plasma exchange therapy to treat cognitive impairment, using the results of assays for one or more of the proteins described above. A computer processor is coupled to the memory component and is configured to execute the instructions stored in the memory component, receive patient data, and analyze the patient data according to one or more algorithms or calculations. A display component may display information regarding the patient's responsiveness to plasma therapy to treat cognitive impairment.

[0111] The memory component may be any type capable of storing information accessible by the processor, such as a hard drive, memory card, ROM, RAM, DVD, CD-ROM, USB flash drive, writable, and read-only memory. The processor may be any well-known processor, such as an Intel Corporation processor. Alternatively, the processor may be a dedicated controller, such as an ASIC.

[0112] Instructions may be any set of instructions, executed directly by the processor (such as machine code) or indirectly (such as a script). In this regard, the terms “instruction,” “step,” and “program” may be used interchangeably herein. Instructions may be stored in object code form for direct processing by the processor, or in any other computer language, including a script or set of independent source code modules that are interpreted on demand or pre-compiled.

[0113] The data may be retrieved, stored, or modified by the processor according to instructions. For example, the systems disclosed herein are not limited by any particular data structure, but the data may be stored in computer registers, relational databases as tables having multiple different fields and records, XML documents, or flat files. The data may also be formatted in any computer-readable format, such as binary values, ASCII, or Unicode, but are not limited. Furthermore, the data may include any information sufficient to identify the relevant information, such as numbers, descriptive text, proprietary codes, pointers, references to data stored in other memory (including other network locations), or information used by functions to compute the relevant data.

[0114] E. Treatment Method Certain aspects of this disclosure further provide determining whether a subject is likely to respond positively to treatment for cognitive impairment and treating the subject for cognitive impairment by administering therapy to the subject.

[0115] Accordingly, certain aspects of the present disclosure provide a method for treating a subject for cognitive impairment, comprising administering plasmapheresis to the subject, wherein the subject is identified as likely to respond positively to plasmapheresis for treating cognitive impairment or not.

[0116] In some cases, subjects are identified as likely or unlikely to respond positively to plasmapheresis to treat cognitive impairment based on the results of assays of the subject's sample for one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1.

[0117] Details on identifying whether a subject is likely to respond positively to plasmapheresis for cognitive impairment based on the results of assays of the subject's sample for one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1 are described above under “Diagnostic Methods.”

[0118] In short, subjects may be identified as likely or unlikely to respond positively to plasmapheresis to treat cognitive impairment based on the results of assays of their samples for one, two, three, or four proteins from DLL1, VNN2, VAV3, and SUMF1.

[0119] Subjects may be identified as likely or unlikely to respond positively to plasmapheresis for treating cognitive impairment based on increased levels of one, two, three, or four assayed proteins in samples obtained from the subject compared to reference values ​​or a reference sample.

[0120] "Plasma exchange therapy" refers to replacing all or part of a patient's plasma with a plasma replacement solution. Typically, during plasma exchange therapy, blood is gradually removed from the patient, blood components such as blood cells and platelets are separated from the plasma, the separated blood components are mixed with the plasma replacement solution, and the resulting mixture is returned to the patient.

[0121] Plasma exchange therapy may include replacing substantially all of the target plasma with a plasma replacement solution, which is referred to herein as “total plasma exchange.” In total plasma exchange, substantially all of the target plasma, such as at least 80%, at least 85%, at least 90%, at least 95%, or at least 99%, is replaced with a plasma exchange solution. In some cases of total plasma exchange, the plasma replacement solution is an albumin solution having 3% to 10% w / v albumin, e.g., 3%, 4%, 5%, 6%, 7%, 8%, 9%, or 10% w / v albumin solution. In one embodiment, the plasma replacement solution for total plasma exchange is a 5% w / v albumin solution.

[0122] Plasma exchange therapy may also include replacing a portion of the target plasma with a plasma replacement solution, which is referred to herein as “low-volume plasma exchange.” In low-volume plasma exchange, 10% to 50% of the target plasma, e.g., 15% to 45%, 20% to 40%, 25% to 35%, or about 30%, is replaced with a plasma exchange solution. In some cases of low-volume plasma exchange, the plasma replacement solution is an albumin solution having 15% to 25% w / v albumin, e.g., 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, or 25% w / v albumin.

[0123] In certain embodiments, plasma exchange therapy includes a process of complete plasma exchange. In some cases, the process of complete plasma exchange includes one complete plasma exchange per week over a period of one to eight weeks, for example, one, two, three, four, five, six, seven, or eight weeks. In certain such embodiments, the plasma replacement solution contains 5% w / v albumin.

[0124] In further embodiments, plasma exchange therapy includes a process of low-volume plasma exchange with a second albumin solution following a process of complete plasma exchange. The process of low-volume plasma exchange may be administered once a month over a period of at least one month to at least ten months, for example, 12 to 16 months, for example, 12, 13, 14, 15, or 16 months. In certain such embodiments, the plasma replacement solution contains 20% w / v albumin.

[0125] In certain embodiments, plasma therapy includes a therapy as shown in Figure 1. Such therapy includes weekly total plasma exchange for 6 weeks using a plasma replacement solution containing 5% w / v albumin. Following such a course of total plasma exchange, plasma exchange therapy includes monthly low-volume plasma exchange therapy for 10 to 16 months, for example, 10, 11, 12, 13, 14, 15, or 16 months. Low-volume plasma exchange therapy includes replacing 15% to 25%, for example 20%, of the target plasma with a plasma replacement solution containing 20 gm to 40 gm of albumin, for example 20 g, 30 g, or 40 g of albumin. In some cases, low-volume plasma exchange therapy involves replacing 15% to 25%, for example 20%, of the target plasma with a plasma replacement solution containing 20g to 40gm³ of albumin, for example 20g, 30g, or 40g of albumin and 10g to 20g of immunoglobulin, for example 10g, 15g, or 20g of immunoglobulin.

[0126] In some cases, plasmapheresis involves weekly total plasmapheresis over six weeks using a plasma replacement solution containing 5% w / v albumin. For example, during total plasmapheresis, 2500–3000 mL of the patient's plasma is replaced with the same volume of 5% albumin solution (e.g., 5% albutein). Following total plasmapheresis, plasmapheresis involves monthly low-volume plasmapheresis over 10–16 months, for example, 10, 11, 12, 13, 14, 15, or 16 months. During low-volume plasmapheresis, 600–900 mL (e.g., 650–880 mL) of the patient's plasma is replaced with 100 mL or 200 mL of 20% albumin (e.g., 20% albutein). In some cases, low-volume plasmapheresis involves using 20-40g, 20g, 30g, and 40g of albumin as a substitution, for example, in a 20% albumin solution (e.g., albutein 20%). In some cases, albumin substitution is alternated with immunoglobulin substitution, for example, 10-30g, for example, 10g, 20g, or 30g of immunoglobulin as a 5% immunoglobulin solution (e.g., Flebogamma DIF 5%).

[0127] plasma replacement solution In certain cases, any of the plasma replacement solutions listed below may be used for plasma exchange therapy.

[0128] There are two common categories of albumin plasma preparations ("APPs"): plasma protein fractions ("PPFs") and human albumin solutions ("HASs"). PPFs are derived from processes that have a higher yield than HASs, but have a lower minimum albumin purity than HASs (>83% for PPFs and >95% for HASs) (Generation of Human Albumin Solution: Continuously Occurring Colloids, P. Matejtschuk et al., British J. of Anaesthesia 85(6):887-95, at 888(2000)). In some cases, PPFs have an albumin purity of 83%–95% or alternatively 83%–96%. Albumin purity can be determined by electrophoresis or other quantitative assays such as mass spectrometry.

[0129] Those skilled in the art will recognize that several commercial sources of PPF exist or have existed ("Commercial PPF Preparations"). These include Plasma-Plex® PPF (Armour Pharmaceutical Co., Tarrytown, NY), Plasmanate® PPF (Grifols, Clayton, NC), Plasmatein® (Alpha Therapeutics, Los Angeles, CA), and Protenate® PPF (Baxter Labs, Inc., Deerfield, IL).

[0130] Those skilled in the art will also recognize that several commercial sources of HAS ("Commercially Available HAS Preparations") exist or have existed. These include Albuminar® (CSL Behring), AlbuRx® (CSL Behring), Albutein® (Grifols, Clayton, NC), Buminate® (Baxatla, Inc., Bannockburn, IL), Flexbumin® (Baxatla, Inc., Bannockburn, IL), and Plasbumin® (Grifols, Clayton, NC).

[0131] Plasma protein fraction (human) (PPF) According to the U.S. Food and Drug Administration ("FDA"), "Plasma Protein Fraction (Human)," or PPF, is the formal name of a product defined as "a sterile solution of proteins consisting of albumin and globulin derived from human plasma" (21 CFR 640.90, Code of Federal Regulations "CFR" incorporated herein by reference). The source material for PPF is plasma recovered from whole blood prepared as specified in 21 CFR 640.1–640.5 (incorporated herein by reference), or source plasma prepared as specified in 21 CFR 640.60–640.76 (incorporated herein by reference).

[0132] PPF is tested to determine whether it meets the following criteria according to 21 CFR 640.92 (incorporated herein by reference): (a) The final product shall be a 5.0+ / -0.30% protein solution; and (b) The total protein in the final product shall consist of at least 83% albumin and 17% or less globulin. Gamma globulin shall constitute 1% or less of the total protein. The protein composition shall be determined by a method approved by each manufacturer by the Director of the Center for Biologics Evaluation and Research, Food and Drug Administration.

[0133] As used herein, “Plasma Protein Fraction” or “PPF” refers to a sterile solution of proteins composed of albumin and globulin derived from human plasma, which, as determined by electrophoresis, contains at least 83% albumin, 17% or less globulin (including α1, α2, β, and γ globulins) and other plasma proteins, and 1% or less gamma globulin. (Hink, JH, Jr., et al., Preparation and Properties of a Heat-Treated Human Plasma Protein Fraction, VOX SANGUINIS 2(174)(1957)). PPF may also refer to a solid form having a similar composition when suspended in a solvent. The total globulin fraction can be determined by subtracting albumin from the total protein. (Busher, J., Serum Albumin and Globulin, CLINICAL METHODS: THE HISTORY, PHYSICAL, AND LABORATORY EXAMINATIONS, Chapter 10, Walker HK, Hall WD, Hurst JD, eds. (1990)).

[0134] Albumin (human) (HAS) According to the FDA, “Human Albumin” (also known herein as “HAS”) is the formal name of the product defined as “Sterile Solution of Albumin Derived from Human Plasma” (21 CFR 640.80, Code of Federal Regulations “CFR” incorporated herein by reference). The source material for Human Albumin is plasma recovered from whole blood prepared as specified in 21 CFR 640.1–640.5 (incorporated herein by reference), or source plasma prepared as specified in 21 CFR 640.60–640.76 (incorporated herein by reference). Other requirements for Human Albumin are listed in 21 CFR 640.80–640.84 (incorporated herein by reference).

[0135] Test human albumin to determine if it meets the following criteria according to 21 CFR 640.82. (a) Protein concentration. The final product shall conform to one of the following concentrations: 4.0+ / -0.25%; 5.0+ / -0.30%; 20.0+ / -1.2%; and 25.0+ / -1.5% protein solution. (b) Protein composition. At least 96% of the total protein in the final product shall be albumin, determined by a method approved by each manufacturer by the Director of the Center for Biologics Evaluation and Research, Food and Drug Administration.

[0136] As used herein, “Albumin (Human)” or “HAS” refers to a sterile solution of proteins composed of albumin and globulin derived from human plasma, containing at least 95% albumin and 5% or less globulin (including α1, α2, β, and γ globulins) and other plasma proteins. HAS may also refer to a solid form having a similar composition when suspended in a solvent. The total globulin fraction can be determined by subtracting albumin from the total protein.

[0137] As can be recognized by those skilled in the art, the PPF and HAS fractions can also be lyophilized or in other solid forms. Such preparations can be used, for example, to make tablets, powders, granules, or capsules, with appropriate additives. The solid forms can be incorporated into injectable preparations by dissolving, suspending, or emulsifying them in aqueous solutions, and, if necessary, using conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifiers, stabilizers, and preservatives.

[0138] F. Indications The methods and compounds are used to treat cognitive impairments, such as cognitive impairments caused by neuroinflammation. In some cases, the cognitive impairment is age-related cognitive impairment, such as a decline in an individual's cognitive abilities due to age.

[0139] Non-limited examples of such cognitive impairment include age-related dementia, immunological symptoms, and physical or functional decline. Individuals who have or are suspected of having cognitive impairment and who would benefit from the treatments disclosed herein include individuals who are about 50 years of age or older, e.g., 60 years of age or older, 70 years of age or older, 80 years of age or older, 90 years of age or older and 100 years of age or older, i.e., about 50 to 100 years of age, e.g., 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 or about 100 years of age and who have cognitive impairment related to the natural aging process, e.g., mild cognitive impairment (MCI); and individuals who are about 50 years of age or older, e.g., 60 years of age or older, 70 years of age or older, 80 years of age or older, 90 years of age or older, usually up to 100 years of age, i.e., about 50 to 90 years of age, e.g., 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 or about 100 years of age and who have not yet begun to show symptoms of cognitive impairment. Examples of cognitive impairment / conditions that may be caused by natural aging include the following:

[0140] 1. Mild Cognitive Impairment (MCI) Mild cognitive impairment is a mild disturbance of cognition that manifests as problems with memory or other mental functions, such as planning, following instructions, or making decisions, where overall mental function and daily activities are not impaired but worsen over time. Thus, significant neuronal death does not typically occur, but neurons in the aging brain are vulnerable to sublethal age-related changes in synaptic structure, synaptic integrity, and molecular processing, all of which impair cognitive function. Individuals with or suspected of having age-related cognitive impairment who would benefit from treatment with the methods disclosed herein.

[0141] The subjects envisioned herein include individuals of any age with cognitive impairment resulting from age-related disorders; and individuals of any age who have been diagnosed with age-related disorders typically accompanied by cognitive impairment. Examples of such age-related disorders include:

[0142] 2. Alzheimer's disease Alzheimer's disease is a progressive, irreversible loss of cognitive function associated with an excessive number of senile plaques in the cerebral cortex and subcortical gray matter, which also include neurofibrillary tangles composed of β-amyloid and tau proteins. The most common form develops in people over 60 years of age, and its incidence increases with age. It accounts for over 65% of dementia in older adults.

[0143] The cause of Alzheimer's disease (AD) is unknown. Approximately 15–20% of cases are familial. The remaining so-called sporadic cases have several genetic determinants. While the disease exhibits an autosomal dominant inheritance pattern in most early-onset cases and some late-onset cases, penetrance in later development varies. Environmental factors are a focus of active research.

[0144] During the course of the disease, synapses, and ultimately neurons, are lost within the cerebral cortex, hippocampus, and subcortical structures (including selective cell loss in the nucleus basalis of Meynert), locus coeruleus, and dorsal raphe nucleus (nucleus raphae dorsalis). Brain glucose use and perfusion are reduced in several areas of the brain (parietal lobe and temporal cortex in early disease, prefrontal cortex in later disease). Senile plaques (composed of neurites, astrocytes, and glial cells surrounding an amyloid core) and neurofibrillary tangles (composed of pairs of helical filaments) play a role in the pathogenesis of AD. Senile plaques and neurofibrillary tangles occur with normal aging, but are far more common in people with AD.

[0145] 3. Parkinson's disease Parkinson's disease (PD) is an idiopathic, slowly progressive, degenerative central nervous system disorder characterized by slow, reduced movement (bradykinesia), muscle rigidity, resting tremor (dystonia), muscle freezing, and postural instability. Originally considered primarily a movement disorder, PD is now recognized as also causing depression and emotional changes. PD can also affect cognition, behavior, sleep, autonomic nervous system function, and sensory function. The most common cognitive impairments include difficulties with attention and concentration, working memory, executive function, language production, and visuospatial function. A characteristic feature of PD is that symptoms associated with reduced motor function usually precede symptoms associated with cognitive impairment, which is helpful in diagnosing the disease.

[0146] In primary Parkinson's disease, pigment neurons in the substantia nigra, locus coeruleus, and other brainstem dopaminergic cell groups degenerate. The cause is unknown. The loss of substantia nigra neurons projecting to the caudate nucleus and putamen leads to depletion of the neurotransmitter dopamine in these areas. Onset generally occurs after age 40, with an increased incidence in older adults.

[0147] Parkinson's disease is newly diagnosed in approximately 60,000 Americans each year, and currently affects about 1 million Americans. While PD itself is not fatal, its complications are the 14th leading cause of death in the United States. Currently, PD is incurable, and treatment is generally prescribed to control symptoms, with surgery prescribed for more severe cases.

[0148] Treatment options for Parkinson's disease (PD) include the administration of medications to help manage movement disorders. These options increase or replace dopamine, a neurotransmitter that PD patients have low brain concentrations of. Such drug therapies include carbidopa / levodopa (which produces more dopamine in the brain); apomorphine, pramipexolole, ropinirole, and rotingotine (dopamine agonists); selegiline and rasagiline (MAO-B inhibitors that prevent the breakdown of dopamine); entacapone and tolcapone (catechol-O-methyltransferase [COMT] inhibitors that make more levodopa available in the brain); benztropine and trihexyphenidyl (anticholinergics); and amantadine (which controls tremors and rigidity). Exercise / physical therapy is also commonly prescribed to help maintain physical and mental function.

[0149] Current treatment options address the symptoms of Parkinson's disease (PD) but are not curative and fail to prevent disease progression. Furthermore, current drug therapies tend to lose efficacy in late-stage PD. Levodopa, the most commonly prescribed drug, generally causes adverse effects within 5–10 years of initiating drug therapy. These adverse effects can be severe and may result in motor variability and unpredictable fluctuations in motor control between doses, as well as seizures / convulsions (dyskinesia), which are difficult to manage and can be as detrimental as the symptoms of PD itself. Therefore, there is still a need for new therapies with novel mechanisms of action that can be administered alone or in combination with current PD drug therapies.

[0150] 4. Parkinsonism Secondary parkinsonism (also known as atypical parkinson's disease or parkinson's disease+) results from the loss or interference of dopamine action in the basal ganglia due to other idiopathic degenerative diseases, drugs, or exogenous toxins. The most common cause of secondary parkinsonism is the ingestion of antipsychotic drugs or reserpine, which cause parkinsonism by blocking dopamine receptors. Less common causes include carbon monoxide or manganese poisoning, hydrocephalus, structural lesions (tumors or infarcts affecting the midbrain or basal ganglia), subdural hematoma, and degenerative disorders, including nigrostriatal degeneration. Certain disorders, such as progressive supranuclear palsy (PSP), multiple system atrophy (MSA), corticobasal degeneration (CBD), and Lewy body dementia (DLB), may present with parkinsonist symptoms before the basic symptoms necessary for a specific diagnosis are present, and may therefore be labeled as “parkinsonism.”

[0151] 5. Frontotemporal dementia Frontotemporal dementia (FTD) is a condition resulting from the progressive deterioration of the frontal lobe of the brain. Over time, the degeneration can progress to the temporal lobe. FTD is the second most common type of dementia after Alzheimer's disease (AD), accounting for 20% of pre-senile dementia cases. The symptoms are classified into three groups based on the function of the frontal and temporal lobes affected.

[0152] Symptomatic behavioral variants of FTD (bvFTD) include apathy and loss of spontaneity on the one hand, and disinhibition on the other; progressive non-fluent aphasia (PNFA) where speech fluency is impaired due to dysarthria, phonological and / or syntactic errors, but word comprehension is preserved; and semantic dementia (SD) where the patient remains fluent in normal phonological and syntactic but has increasing difficulties with naming and word comprehension. Other cognitive symptoms common to all FTD patients include impairments in executive function and concentration. Other cognitive abilities, including perception, spatial skills, memory, and ideomotor abilities, are typically intact. FTD can be diagnosed by the observation of clear frontal and / or anterior-temporal lobe atrophy on structural MRI scans.

[0153] Several forms of FTD exist, all of which can be treated or prevented using targeted methods and compositions. For example, one form of frontotemporal dementia is semantic dementia (SD). SD is characterized by a loss of semantic memory in both the linguistic and non-linguistic domains. Patients with SD often complain of difficulty finding words. Clinical signs include fluent aphasia, anomalism, difficulty understanding the meaning of words, and relevance visual agnosia (inability to match semantically relevant images or objects). As the disease progresses, behavioral and personality changes are often similar to those seen in frontotemporal dementia, although cases have been described as “pure” semantic dementia with little to no late-stage behavioral symptoms. Structural MRI imaging shows a characteristic pattern of temporal lobe atrophy (primarily on the left side), with greater atrophy in the inferior temporal lobe than in the superior temporal lobe, and greater atrophy in the anterior temporal lobe than in the posterior temporal lobe.

[0154] Another example, a form of frontotemporal dementia, is Pick's disease (PiD, or PcD). A distinctive feature of this disease is the accumulation of tau protein in neurons, which accumulates in silver-stained spherical aggregates known as "Pick bodies." Symptoms include loss of speech (aphasia) and dementia. Patients with orbitofrontal dysfunction may become aggressive and socially inappropriate. They may engage in kleptomania or exhibit compulsive or repetitive stereotyped behaviors. Patients with dorsomedial or dorsolateral frontal dysfunction may exhibit a lack of concern, apathy, or reduced spontaneity. Patients may exhibit a lack of self-monitoring, abnormal self-awareness, and an inability to understand meaning. Patients with gray matter loss in the bilateral posterolateral orbitofrontal cortex and the right anterior insula may exhibit changes in eating behavior, such as a pathological sweet tooth. Patients with more localized gray matter loss in the anterolateral orbitofrontal cortex may develop hypereating. While some symptoms may be initially alleviated, the disease progresses, and patients often die within 2 to 10 years.

[0155] 6. Huntington's disease Huntington's disease (HD) is a hereditary, progressive neurodegenerative disorder characterized by the onset of emotional, behavioral, and psychiatric abnormalities; loss of intellectual or cognitive function; and movement disorders. Classical signs of HD include the onset of chorea (involuntary, rapid, irregular, or spasmodic movements that may affect the face, arms, legs, or trunk), as well as cognitive decline, including a progressive loss of thought processing and acquired intellectual abilities; impairment of memory, abstract thinking, and judgment; disorientation (inappropriate perception of time, place, or people); increased agitation; and personality changes (disintegration). While symptoms typically become apparent between the ages of 30 and 40, the age of onset varies, ranging from infancy to late adulthood (e.g., 70s or 80s).

[0156] HD is transmitted within families as an autosomal dominant trait. This disorder results from an abnormally long sequence or "repetition" of the coded instructions in a gene (4p16.3) on chromosome 4. The progressive loss of nervous system function associated with HD is due to the loss of neurons in specific areas of the brain, including the basal ganglia and cerebral cortex.

[0157] 7. Amyotrophic lateral sclerosis (ALS) Amyotrophic lateral sclerosis (ALS) is a rapidly progressing, always fatal neurological disease that attacks motor neurons. Signs of muscle weakness and atrophy, as well as anterior horn cell dysfunction, are most frequently seen in the hands initially and less frequently in the feet. The site of onset is random, and progression is asymmetrical. Muscle spasms are common and may precede muscle weakness. Rarely, patients survive for 30 years; 50% die within 3 years of onset, 20% survive for 5 years, and 10% survive for 10 years.

[0158] Diagnostic features include middle- or late-adult onset and progressive generalized motor impairment without sensory disturbances. Nerve conduction velocity remains normal until later stages of the disease. Recent studies have also demonstrated the presentation of cognitive impairment, particularly a decline in immediate verbal memory, visual memory, language, and executive function.

[0159] A decrease in cell body region, synapse number, and total synaptic length has been reported even in normally appearing neurons in ALS patients. It has been suggested that continuous synaptic loss can lead to dysfunction once the plasticity of the active region reaches its limit. Promoting the formation of new synapses or preventing synaptic loss may preserve neuronal function in these patients.

[0160] 8. Multiple sclerosis Multiple sclerosis (MS) is characterized by a variety of symptoms and signs of central nervous system (CNS) dysfunction, with periods of remission and relapsing exacerbations. The most common presenting symptoms are unilateral paresthesia of one or more limbs, trunk, or face; weakness or clumsiness of the legs or hands; or visual impairment, such as partial blindness and pain in one eye (retrobulbar optic neuritis), decreased visual acuity, or scotoma. Common cognitive impairments include difficulties with memory (acquisition, retention, and retrieval of new information), attention and concentration (especially splitting of attention), information processing, executive function, visuospatial function, and verbal fluency. Common early symptoms include ophthalmoplegia resulting in diplopia, transient weakness of one or more limbs, slight rigidity or abnormal fatigability of the limbs, mild gait disturbance, difficulty controlling bladder function, dizziness, and mild emotional disturbances; all indicating scattered CNS involvement and often occurring months or years before the disease is recognized. Excessive fever can worsen symptoms and signs.

[0161] The course of the disease is highly diverse and unpredictable, and in most patients it is relapsing-remitting. Initially, remissions of several months or years may isolate episodes, especially if the disease begins with retrobulbar optic neuritis. However, some patients experience frequent seizures and rapid incapacitation; in some cases, the course can be rapidly progressive.

[0162] 9. Glaucoma Glaucoma is a common neurodegenerative disease that affects retinal ganglion cells (RGCs). Evidence supports the presence of compartmentalized degenerative programs in synapses and dendrites, including RGCs. Recent evidence also shows a correlation between cognitive impairment and glaucoma in older adults (Yochim BP, et al. Prevalence of cognitive impairment, depression, and anxiety symptoms among older adults with glaucoma. J Glaucoma. 2012;21(4):250-254).

[0163] 10. Myotonic dystrophy Myotonic dystrophy (DM) is an autosomal dominant multiple system disorder characterized by dystrophic muscle weakness and myotonia. The molecular defect is an extended trinucleotide (CTG) repeat in the 3' untranslated region of the myotonin protein kinase gene on chromosome 19q. Symptoms can occur at any age, and the range of clinical severity is wide. Myotonia is prominent in the muscles of the hands, and ptosis is common even in mild cases. In severe cases, marked peripheral muscle weakness occurs, often accompanied by cataracts, premature alopecia, axe-shaped facies, cardiac arrhythmias, testicular atrophy, and endocrine abnormalities (e.g., diabetes mellitus). Intellectual disability is common in severe congenital forms, but age-related declines in frontal and temporal cognitive function, particularly language and executive function, are observed in milder adult forms of the disorder. Severely affected individuals die by their early 50s.

[0164] 11. Dementia Dementia represents a class of disorders characterized by symptoms that significantly impair thinking and social abilities, often severe enough to interfere with daily functioning. In addition to dementia observed in the later stages of age-related disorders as described above, other forms of dementia include vascular dementia and Lewy body dementia, which will be discussed later.

[0165] In vascular dementia, also known as "multiple-infarct dementia," cognitive impairment is caused by problems with blood supply to the brain, typically resulting from a series of minor strokes, or occasionally from a single major stroke preceding or following other smaller strokes. Vascular lesions can be the result of diffuse cerebrovascular disease, such as small vessel disease, or focal lesions, or both. Patients with vascular dementia present with acute or subacute cognitive impairment following an acute cerebrovascular event, followed by progressive cognitive decline. The cognitive impairment is similar to that observed in Alzheimer's disease, including impairments in language, memory, complex visual processing, or executive function, but the associated brain changes are not due to the pathophysiology of AD, but rather to chronic reduced blood flow to the brain, ultimately leading to dementia. Single-photon emission tomography (SPECT) and positron emission tomography (PET) neuroimaging, along with assessments including mental state testing, can be used to confirm the diagnosis of multiple-infarct dementia.

[0166] Lewy body dementia (DLB, also known by various other names including Lewy body dementia, diffuse Lewy body disease, cortical Lewy body disease, and Lewy-type senile dementia) is a type of dementia anatomically characterized by the presence of Lewy bodies (clumps of alpha-synuclein and ubiquitin proteins) in neurons detectable in postmortem brain histology. Its main feature is a decline in cognitive function, particularly executive function. Arousal levels and short-term memory may increase or decrease.

[0167] Persistent or recurrent visual hallucinations with vivid, detailed images are often an early diagnostic symptom. DLB is often confused with Alzheimer's disease and / or vascular dementia in its early stages, but while Alzheimer's disease usually develops very gradually, DLB often has a rapid or acute onset. DLB symptoms also include motor symptoms similar to those of Parkinson's disease. DLB is distinguished from dementia that occurs occasionally in Parkinson's disease by the time frame in which dementia symptoms appear compared to Parkinsonian symptoms. Parkinson's disease with dementia (POD) would be diagnosed when the onset of dementia occurs more than one year after the onset of Parkinson's disease. DLB is diagnosed when dementia symptoms begin simultaneously with or within one year of Parkinsonian symptoms.

[0168] 12. CADASIL Autosomal dominant arterial disease (CADASIL) with subcortical infarction and leukoencephalopathy is a hereditary disorder associated with mutations in the NOTCH 3 gene (Locatelli M, et al., Front. Pharmacol. 11:321 (2020)). It typically occurs in middle-aged adults with symptoms including cognitive impairment leading to dementia and physical disability (ibid.). Other symptoms include mood disorders, migraines with aura, and recurrent strokes. Because the mode of disease development (etiology) remains uncertain, effective treatments have not yet been established (ibid.). CADASIL is the most common hereditary subcortical type of vascular dementia (Kalimo H, et al., Future Neurology, 3(6) (2008)).

[0169] CADASIL is characterized by four main common symptoms: migraine with aura, recurrent ischemic stroke, psychiatric disorder, and cognitive decline. The first is usually the onset symptom and occurs in 20–40% of affected individuals. The second symptom occurs in 60–85% of symptomatic individuals. Thirdly, psychiatric disorder occurs in 25–30% of patients in the form of moderate / major depression, bipolar disorder, panic disorder, schizophrenia, and apathy. Cognitive impairment occurs in 60% of patients, becomes clinically detectable between the ages of 35 and 50, and gradually worsens with age. (Ibid.) In younger patients, attention, memory, and executive function impairments are dominant. (Buffon F, et al., J Neurol Neurosurg Psychiatry 77(2):175-80 (2006)). Visuospatial abilities and reasoning decline with age, mainly after the age of 60. Dementia is present in 25% of patients, and 75% of those are over 60 years old. However, the number of ischemic attacks is not associated with dementia. (Ibid.)

[0170] CADASIL is a progressive and fatal disease. To date, no disease-modifying treatments have been developed (Locatelli et al., previously cited). Symptomatic treatment is the only option for clinicians, based on routine clinical care such as acetazolamide or sodium valproate for migraines; daily aspirin to reduce the likelihood of heart attack or stroke; and supportive care for cognitive loss. Notably, no drug has yet demonstrated a clear benefit for cognitive loss associated with CADASIL (ibid.). Interventions that have been studied but failed include donepezil (used to improve cognitive impairment in Alzheimer's), galantamine (an acetylcholinesterase inhibitor used to treat cognitive impairment in Alzheimer's), and L-dopa (used in Alzheimer's and Parkinson's diseases).

[0171] 13. Progressive supranuclear palsy Progressive supranuclear palsy (PSP) is a brain disorder that causes serious and progressive problems with gait and balance control, along with problems with complex eye movements and thinking. One of the typical signs of this disorder is the inability to properly direct the eyes, which results from lesions in the brain regions that control eye movements. Some individuals describe this effect as blurred vision. Affected individuals often exhibit mood and behavioral changes, including depression and apathy, as well as progressive mild dementia. The long name of the disorder indicates that the disease starts slowly, continues to worsen (progressive), and causes debilitation (paralysis) by damaging specific parts of the brain (supranuclear) above pea-sized structures called nuclei that control eye movements. PSP was first described as a distinct disorder in 1964 when three scientists published a paper differentiating the symptoms from Parkinson's disease. It is sometimes called Steele-Richardson-Olszewski syndrome, combining the names of the scientists who defined the disorder. While PSP gradually worsens, no one dies from PSP itself.

[0172] 14. Ataxia People with ataxia have problems with coordination because parts of the nervous system that control movement and balance are affected. Ataxia can affect the movement of fingers, hands, arms, legs, body, speech, and eyes. The term ataxia is often used to describe a symptom of coordination disorder that may be associated with infection, injury, other disease, or degenerative changes in the central nervous system. Ataxia is also used to refer to a group of specific degenerative disorders of the nervous system called hereditary and sporadic ataxia, which are the main focus of the National Ataxia Foundation.

[0173] 15. Multiple system atrophy Multiple system atrophy (MSA) is a degenerative neurological disorder. MSA is associated with the degeneration of nerve cells in specific areas of the brain. This cellular degeneration causes problems with movement, balance, and other autonomic nervous system functions of the body, such as bladder control or blood pressure regulation.

[0174] The cause of MSA is unknown, and no specific risk factors have been identified. Approximately 55% of cases occur in men, with a typical age of onset in the late 50s to early 60s. MSA often presents with some of the same symptoms as Parkinson's disease. However, MSA patients generally have a minimal response, if any, to dopamine therapy used for Parkinson's disease.

[0175] 16. Frailty Frailty syndrome ("frailty") is a geriatric syndrome characterized by functional and physical decline, including reduced motor function, muscle weakness, physical retardation, decreased endurance, low physical activity, malnutrition, and involuntary weight loss. Such declines often accompany and result from diseases such as cognitive impairment and cancer. However, frailty can occur even without disease. Individuals suffering from frailty have a higher risk of poor prognosis from fractures, accidental falls, handicaps, comorbidities, and premature death. (C. Bugues, et al. Effect of a Prebiotic Formulation on Frailty Syndrome: A Randomized, Double-Blind Clinical Trial, Int. J. Mol. Sci. 2016, 17, 932). Furthermore, individuals suffering from frailty have a higher incidence of medical expenses. (Ibid.)

[0176] Common symptoms of frailty can be determined by certain types of tests. For example, unintentional weight loss is defined as a loss of at least 10 pounds; or more than 5% of the previous year's body weight; muscle weakness can be determined by a reduction in grip strength by the bottom 20% at baseline (adjusted for sex and BMI); physical stagnation can be based on the time required to walk a distance of 15 feet; decreased endurance can be determined by the individual's self-reported fatigue; and low physical activity can be measured using a standardized questionnaire. (Z. Palace et al., The Frailty Syndrome, Today's Geriatric Medicine 7(1), at 18(2014)).

[0177] 17. Neuromyelitis Optica Spectrum Disorder Neuromyelitis optica spectrum disorder (NMOSD), also known as Devic's disease, is a rare inflammatory disorder of the central nervous system. It is characterized by optic neuritis (inflammation of the optic nerve) and myelitis (inflammation of the spinal cord). Typically, patients experience recurrent episodes of inflammation separated by periods of remission. The disease is thought to be caused often by autoantibodies targeting myelin oligodendrocyte glycoprotein (MOG-IgG) or aquaporin 4 (AQP4-IgG), resulting in demyelination and axonal damage in the optic nerve and spinal cord.

[0178] 18. Postoperative cognitive impairment Postoperative cognitive decline occurs after anesthesia and surgical procedures. It is common in patients over 60 years of age and is diagnosed by preoperative and postoperative cognitive testing. Patients typically present with memory impairment, delirium, and impaired ability to perform intellectual tasks.

[0179] 19. Chronic traumatic encephalopathy Chronic traumatic encephalopathy (CTE) is a neurodegenerative brain disorder most commonly seen in athletes, veterans, or others with a history of repeated head injuries. It is one of many tauopathies characterized by an excess of tau protein in the brain of patients, resulting in neuronal loss. Symptoms include memory loss, mood or personality changes, confusion, impaired judgment, impulse control, aggression, and depression.

[0180] 20. Traumatic brain injury Traumatic brain injury (TBI) is caused by a severe blow to the head or body. It can also be caused by an object penetrating brain tissue during the injury. This results in bleeding, tissue rupture, as well as physical damage and cell death to brain cells. Physical symptoms vary but may include loss of consciousness, headache, nausea, extreme fatigue, speech difficulties, sleep disturbances, dizziness, blurred vision, sensitivity to light or sound, memory loss, and difficulty concentrating.

[0181] In some embodiments, the methods and compositions in question are used to slow the progression of age-related cognitive impairment. In other words, cognitive impairment in an individual will decline more slowly after treatment with the disclosed methods than before or in the absence of treatment with the disclosed methods. In some such cases, the treatment method in question includes measuring the progression of cognitive impairment after treatment and determining that the progression of decline has decreased. In some such cases, the determination is made by comparison with a reference (e.g., the rate of decline in the individual before treatment, which is determined, for example, by measuring cognitive ability at two or more time points prior to administration of the blood product in question).

[0182] The methods and compositions can also be used to stabilize the cognitive abilities of individuals, for example, individuals with age-related cognitive impairment or suspected of having age-related cognitive decline. For example, an individual may exhibit some age-related cognitive impairment, and the progression of cognitive impairment observed before treatment by the disclosed methods will be halted after treatment by the disclosed methods.

[0183] As another example, an individual may be at risk of developing age-related cognitive decline (for example, the individual may be 50 years of age or older, or may have been diagnosed with age-related disorder), and the individual's cognitive abilities may not have changed substantially after treatment by the disclosed method compared to before treatment by the disclosed method; that is, cognitive decline cannot be detected.

[0184] The methods and compositions are also used to reduce cognitive impairment in individuals with age-related disorders. In other words, affected cognitive abilities are improved in individuals after treatment with the methods. For example, cognitive abilities in an individual increase by, for example, more than 2 times, more than 5 times, more than 10 times, more than 15 times, more than 20 times, more than 30 times, or more than 40 times (including more than 50 times, more than 60 times, more than 70 times, more than 80 times, more than 90 times, or more than 100 times) after treatment with the methods compared to cognitive abilities observed in the individual before treatment with the methods.

[0185] In some cases, treatment with the methods and compositions restores the cognitive abilities of individuals with age-related cognitive impairment to levels they were, for example, at around 40 years of age or younger. In other words, cognitive or motor impairments are resolved.

[0186] G. Monitoring methods for improvement In some cases, among various methods for monitoring disease progression and improvement of cognitive impairment, the following types of assessments are used alone or in combination with subjects having cognitive impairment. The following types of methods are presented as examples and are not limited to those listed. Any convenient method for monitoring the disease may be used in the implementation of the present invention as needed. These methods are also intended by the methods of the present invention.

[0187] i. General cognition Certain embodiments of the methods of the present invention further include methods for monitoring the effects of pharmacotherapy or treatment on a subject for treating cognitive impairment, such as age-related cognitive impairment. Certain such methods include comparing cognitive function before and after treatment. Methods for assessing cognitive function are well known in the art. For example, but not limited to, methods may include assessments of cognitive function based on medical history, family history, physical and neurological examinations by a clinician specializing in cognitive function, laboratory tests, and neuropsychological assessments. Additional embodiments contemplated by the present invention include assessments of mental state, such as the Glasgow Coma Scale (EMV); the Ambidextrous Test Score (AMTS) or Mini-Mental State Examination (MMSE) (Folstein et al., J. Psychiatr. Res 1975;12:1289-198); a global assessment of higher-order functions; and assessments of consciousness, such as those using estimation of intracranial pressure by fundus examination. In one embodiment, monitoring the effect on cognitive impairment, such as age-related cognitive impairment, involves examining improvements of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 points using the Alzheimer's Disease Assessment Scale-Cognitive Subscale (ADAS-COG).

[0188] In one embodiment, cognitive function may be assessed using tests of the peripheral nervous system, including any one of the following: sense of smell, visual field and visual acuity, eye movements and pupils (sympathetic and parasympathetic), facial sensory function, strength of facial and shoulder girdle muscles, hearing, taste, pharyngeal movements and reflexes, tongue movements (which can be tested individually; for example, visual acuity can be tested by the Snellen chart; the reflex hammer is used to test reflexes including the masseter, biceps and triceps tendons, patellar tendon, Achilles tendon reflex and plantar flexion (i.e., Babinski sign)); muscle strength, often on an MRC scale of 1-5; signs of muscle tone and stiffness.

[0189] Certain embodiments of the methods of this disclosure further include monitoring the effect of a cognitive impairment treatment on motor function before and after treatment. Certain such methods include evaluating motor function in a subject having PD and being treated for cognitive impairment according to the methods disclosed herein. Methods for evaluating motor function are well known in the art. For example, but not limited to, methods may include evaluation of motor function based on medical history, family history, physical and neurological examinations by a clinician specializing in neurodegeneration and motor disorders, laboratory tests, and neurodegenerative assessments. Additional embodiments contemplated by this disclosure include the use of evaluation scales discussed below.

[0190] Several assessment scales are used to evaluate the progression of Parkinson's Disease (PD). The most widely used scales include the Unified Parkinson's Disease Rating Scale (UPDRS, introduced in 1987) (J. Rehabil Res. Dev., 2012 49(8):1269-76) and the Hoehn and Yahr scale (Neurology, 1967 17(5):427-42). Further scales include the Movement Disorder Society's (MDS) updated UPDRS scale (MDS-UPDRS) and the Schwab and England Activities of Daily Living (ADL) Scale.

[0191] The UPDRS scale assesses 31 items that contribute to the following three subscales: (1) mental, behavioral, and mood; (2) activities of daily living; and (3) motor skills. The Hoehn and Yahr scale classifies PD into five stages with discrete substages: 0 - no signs of disease; 1 - symptoms on only one side; 1.5 - symptoms affecting not only one side but also the cervical and spinal areas; 2 - bilateral symptoms without balance impairment; 2.5 - mild bilateral symptoms that resolve on a "pull" test; 3 - balance impairment with mild to moderate disease; 4 - severe handicap but the ability to walk or stand without assistance; and 5 - wheelchair required or bedridden without assistance. The Schwab and England scale classifies PD into several percentages (from 100% - completely independent to 10% - fully dependent).

[0192] General motor function can be assessed using widely used scales, including the General Motor Function Scale (GMF). This test includes three components: dependence, pain, and anxiety. (Aberg AC, et al. (2003) Disabil. Rehabil. 2003 May 6;25(9):462-72.). Motor function can also be assessed using home monitoring or wearable sensors. For example, accelerometers can be used to detect gait (movement speed, variability, leg stiffness); gyroscopes can detect posture (trunk tilt); accelerometers can detect leg movement; accelerometers and gyroscopes can detect hand movement; accelerometers can detect tremor (amplitude, frequency, duration, asymmetry); accelerometers can detect falls; accelerometers can detect gait freeze; accelerometers, gyroscopes, and inertial sensors can detect dyskinesia; accelerometers and gyroscopes can detect bradykinesia (duration and frequency); and microphones can be used to detect aphasia (pitch). (Pastorino M, et al., Journal of Physics: Conference Series 450 (2013) 012055).

[0193] Certain embodiments of the methods disclosed herein further include monitoring the effect of treatment for cognitive impairment on the progression or improvement of neurodegeneration. Certain such methods include evaluating the progression or improvement of neurodegeneration in a subject having a neurodegenerative disease, e.g., MS, HD, ALS, glaucoma, PSP, and who has been treated for cognitive impairment according to the methods disclosed herein.

[0194] Methods for monitoring the progression or improvement of neurodegeneration are well known to those skilled in the art. Monitoring can be performed, but not limited to, through techniques such as cerebrospinal fluid (CSF) monitoring; magnetic resonance imaging (MRI) for detecting the development of lesions and demyelinating plaques; evoked potential testing; and gait monitoring.

[0195] CSF analysis can be performed, for example, by lumbar puncture, to obtain pressure, appearance, and CSF content. Normal values ​​are typically within the following ranges: pressure (70-180 mmH2O); appearance is colorless and clear; total protein (15-60 mg / 100 mL); IgG is 3-12% of total protein; glucose is 50-80 mg / 100 mL; cell count is 0-5 white blood cells, with no red blood cells; chloride (110-125 mEq / L). Abnormal results may indicate the presence or progression of MS.

[0196] MRI is another technique that may be used to monitor disease progression and improvement. Typical criteria for monitoring MS using MRI include the appearance of abnormal white matter patches in the cerebral hemispheres and paraventricular regions, and lesions present in the cervical or thoracic regions of the cerebellum and / or brainstem, as well as the spinal cord.

[0197] Evoked potentials can be used to monitor the progression and improvement of MS in subjects. Evoked potentials measure the delay of electrical impulses such as visual evoked responses (VER), brainstem auditory evoked responses (BAER), and somatosensory evoked responses (SSER). Abnormal responses can help indicate a decrease in the conduction velocity of the central sensory pathway.

[0198] Gait monitoring can also be used to monitor disease progression and improvement in MS patients. MS often involves motor impairment and abnormal gait, partly due to fatigue. Monitoring can be performed, for example, using a mobile monitoring device worn by the patient. (Moon, Y., et al., Monitoring gait in multiple sclerosis with novel wearable motion sensors, PLOS One, 12(2):e0171346(2017)).

[0199] This disclosure also aims to treat or improve neurogenesis in subjects where neurogenesis is reduced or impaired, which may be evident, for example, by a reduction in cognitive or motor function, or in association with neuroinflammation.

[0200] One embodiment of the present invention also intends to determine the level of neurogenesis before, during, and / or after therapies according to the present disclosure, such as plasmapheresis. Non-invasive techniques for evaluating neurogenesis have been reported (Tamura Y. et al., J. Neurosci. (2016) 36(31):8123-31). Positron emission tomography (PET) used with the tracer [18F]FLT, in combination with the BBB transporter inhibitor probenecid, allows for the accumulation of the tracer in neurogenic regions of the brain. Such imaging allows for the evaluation of neurogenesis in patients undergoing treatment for neurodegenerative diseases.

[0201] H. Exercise Exercise can be characterized by aerobic or anaerobic activity and may involve high and moderate thermal activity. Exercise may involve strength training (e.g., weight training or isometric exercise). Exercise may also include, for example, running, cycling, walking, dancing, marching, swimming, yoga, tai chi, balance exercises, leg flexion, jump rope, surfing, rowing, arm or leg rotations or flexion, gardening, cleaning, active games such as bowling, aerobics, Pilates, and martial arts.

[0202] An exercise regimen may include performing a single exercise at a specific frequency, or a combination of exercises at a specific frequency. The frequency may be once, twice, three, four, five, six, or seven times per week. The frequency may vary from week to week. The exercise regimen may be at the same intensity and / or frequency as the subject performed prior to administration of the composition of the present invention. The exercise regimen may also be at a higher intensity and / or frequency compared to the level the subject performed prior to administration of the therapy of this disclosure. The exercise regimen may be suggested or prescribed by a health or fitness professional, or the exercise regimen may be initiated by the subject themselves.

[0203] I. Kits, reagents, and devices Certain aspects of this disclosure provide a kit comprising reagents for measuring the amount of one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1 in a sample.

[0204] As described above, assays of one or more target proteins can be carried out by any suitable method, such as a qualitative or quantitative method. Therefore, the kits disclosed herein may be designed for qualitative or quantitative assays of one or more target proteins.

[0205] Accordingly, in certain cases, the kits disclosed herein provide reagents and / or devices for determining the sequence of a target protein. Certain such reagents and / or devices can be used when performing qualitative target protein analysis using mass spectrometry. Such reagents and / or devices include reagents for dissolving proteins in a sample, such as enzymes for digesting proteins, such as trypsin; various solvents such as water, methanol, acetonitrile, chloroform, tetrahydrofuran, and dichloromethane; and various buffering reagents for preparing buffers used in the analysis.

[0206] Certain other reagents and / or devices can be used when performing qualitative target protein analysis using Edman degradation with a protein sequencer. Such reagents include phenyl isothiocyanates. Other reagents used for Edman degradation sequencing are well known in the art, and kits containing such reagents are within the scope of this disclosure.

[0207] In some cases, the kits disclosed herein are designed to quantitatively assay a sample for one or more proteins. Certain non-limiting examples include kits for quantitatively assaying one or more target proteins via immunoassays, mass spectrometry, and protein detection array analysis.

[0208] As described above, non-limiting examples of immunoassays include Western blot analysis, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), lateral flow immunoassay, particle-based immunoassay, and quantum dot-based immunoassay. Accordingly, certain embodiments of this disclosure provide kits for performing one or more of these assays on one or more target proteins disclosed herein.

[0209] In one embodiment, the kit includes a lateral flow immunoassay device. Such a device enables the detection and / or quantification of one or more target proteins from DLL1, VNN2, VAV3, and SUMF1. The lateral flow immunoassay device may also include an internal control indicating a reference value or threshold.

[0210] In some cases, lateral flow immunoassay devices allow for the detection and / or quantification of only one, two, three, or four proteins from DLL1, VNN2, VAV3, and SUMF1. In addition to one or more target proteins, lateral flow immunoassay devices may allow for the detection and / or quantification of one or more control proteins.

[0211] Certain kits may enable the detection of one or more target proteins via a protein detection array. A particular such protein detection array may contain antibodies that specifically bind to one or more proteins from specific conjugates, e.g., DLL1, VNN2, VAV3, and SUMF1. The antibodies localize to specific spots on a solid support. The kit may further contain reagents that facilitate visualization of the specific binding between the target protein in the sample and the conjugate on the array. Such reagents may include labeled secondary antibodies, e.g., fluorescently labeled secondary antibodies.

[0212] In certain embodiments, the kit enables the detection of one or more target proteins by lysate microarray analysis. Such a kit allows for the preparation of a sample lysate and the immobilization of the lysate on a support, such as a nitrocellulose-coated glass slide. The target proteins are then detected using a labeling solution-phase specific conjugate for one or more target proteins, such as a fluorescently labeled antibody that specifically binds to one or more target proteins. Thus, the kit includes a labeling conjugate for one or more target proteins, in particular a fluorescently labeled antibody that specifically binds to one or more target proteins. In some cases, antibodies against different target proteins are differentially labeled, thereby enabling multiple detection and / or quantification of two or more target proteins.

[0213] In some cases, the kit is designed to allow a sample to be assayed for one of DLL1, VNN2, VAV3, and SUMF1. For example, a particular kit allows a sample to be assayed for DLL1. A particular other kit allows a sample to be assayed for VNN2. Further embodiments describe a kit that allows a sample to be assayed for VAV3. Yet another kit allows a sample to be assayed for SUMF1.

[0214] In some cases, the kit is designed to allow a sample to be assayed for any two of DLL1, VNN2, VAV3, and SUMF1, for example, the following combinations: DLL1 and VNN2; DLL1 and VAV3; DLL1 and SUMF1; VNN2 and VAV3; VNN2 and SUMF1; and VAV3 and SUMF1.

[0215] In further embodiments, the kit is designed to allow a sample to be assayed for any three of DLL1, VNN2, VAV3, and SUMF1, for example, in the following combinations: DLL1, VNN2, and VAV3; DLL1, VAV3, and SUMF1; DLL1, VNN2, and SUMF1; and VNN2, VAV3, and SUMF1.

[0216] In certain embodiments, the kit is designed to allow the sample to be assayed for all four DLL1, VNN2, VAV3, and SUMF1.

[0217] In some cases, the kit is designed to allow a sample to be assayed for only one of DLL1, VNN2, VAV3, and SUMF1, and to avoid assaying additional proteins except for a specific control protein.

[0218] In some cases, the kit is designed to allow assaying of a sample for only two of DLL1, VNN2, VAV3, and SUMF1, and to exclude additional proteins from assaying, except for specific control proteins. For example, the following protein combinations may be assayed: DLL1 and VNN2; DLL1 and VAV3; DLL1 and SUMF1; VNN2 and VAV3; VNN2 and SUMF1; and VAV3 and SUMF1, with one or more control proteins also being assayed.

[0219] In further embodiments, the kit is designed to allow assaying of a sample for only three of DLL1, VNN2, VAV3, and SUMF1, and to exclude other proteins from assaying except for specific control proteins. For example, the following protein combinations may be assayed: DLL1, VNN2, and VAV3; DLL1, VAV3, and SUMF1; DLL1, VNN2, and SUMF1; and VNN2, VAV3, and SUMF1, with one or more control proteins also being assayed.

[0220] In certain embodiments, the kit is designed to allow assaying of a sample for only the following four proteins: DLL1, VNN2, VAV3, and SUMF1, and to avoid assaying additional proteins except for a specific control protein.

[0221] In some cases, each of the one or more specific binding members in the kit described herein independently comprises an antibody or its antigen-binding fragment, aptamer, or peptide-binding member. The one or more specific binding members are labeled with a detectable portion, such as an optically detectable portion.

[0222] In some cases, the kits disclosed herein are intended for use in the methods disclosed herein, namely, for use in identifying whether a subject is likely to respond positively to plasmapheresis for treating cognitive impairment in that subject.

[0223] In addition to the components described above, the kit may further include instructions for carrying out the method. These instructions may be present in the kit in various forms, and one or more of these forms may be present in the kit. One possible form of these instructions is printed information on a suitable medium or substrate, e.g., one or more sheets of paper on which the information is printed, the kit's packaging, or accompanying documents. Yet another means may be a computer-readable medium on which the information is recorded, e.g., a diskette, CD, or portable flash drive. Yet another possible means may be a website address that can be used via the internet to access the information at a remote site. In certain cases, the kit may also provide information about a smartphone app, which then provides the relevant information. Any other convenient means of conveying the information may be present in the kit. [Examples]

[0224] VI. Experimental Examples A. Example 1 - Identification of proteins that predict responsiveness to plasma exchange therapy for treating cognitive impairment. Example 1 describes an exemplary method for identifying protein biomarkers to predict the response of subjects to plasma exchange (PE) therapy for treating cognitive impairment. Firstly, the benefit of treatment was confirmed by group comparisons of altered clinical outcomes between the treatment group and the placebo group based on appropriate statistical tests such as t-tests, ANOVA, MMRM, linear mixed models, CHIsq tests, Fisher's exact test, and logistic regression. Such analyses were also performed in subgroups where sufficient samples were available.

[0225] In this example, the Clinical Dementia Rating Box Sum (CDR.sb) was used for clinical outcomes. It is a comprehensive assessment of AD patients and was a secondary endpoint in the AMBAR clinical trial. Six categories were assessed: memory, orientation, judgment and problem-solving, social life, home and hobbies, and personal care. A higher CDR.sb indicates more severe dementia.

[0226] Following PE treatment, a significant improvement of 1.1 points in EOS (End of Life Scale) was observed in the treatment group (p=0.002). A similar improvement (1.5 points) (p=0.01) was observed in moderate AD, and although both the PE and control groups worsened, PE treatment reduced the worsening. Mild AD PE treatment showed improvement, while the placebo group showed cognitive deterioration. (Boada et al. (2020), Alzheimer's & Dementia, 16:10, pp.1412-1425.)

[0227] In a subset of patients, baseline predictive biomarkers were identified in relation to the responsiveness to PE-Alb treatment for clinical improvement based on CDR.sb. Spearman correlation analysis was used to identify baseline plasma proteins associated with CDR.sb changes at EOS. Such analyses demonstrated the intensity of proteomic biomarkers associated with continuous clinical changes.

[0228] We reported the Spearman correlation coefficient or Spearman's rho and the associated p-values. A positive rho indicated a positive relationship, and a negative rho indicated a negative relationship.

[0229] An absolute value of rho greater than 0.3 (abs(rho)) was considered a true correlation, rho values ​​between 0.3 and 0.5 were considered low correlation, rho values ​​between 0.5 and 0.7 were considered moderate correlation, rho values ​​between 0.7 and 0.9 were considered high correlation, and rho values ​​greater than 0.9 were considered very high correlation. (See Mukaka (2012), Malawi Med J., 24(3):69-71.)

[0230] Based on concepts from diagnostic trials, altered CDR.sb at EOS was converted into two binary variables compared to baseline: better or worse than baseline, and not worse than clinically meaningful change (a 1-point increase in CDR.sb). ROCs were generated, AUCs were calculated, and the diagnostic / predictive power of candidate proteomics biomarkers was compared.

[0231] Next, high or low baseline protein levels were defined by an optimal decision threshold (reference value) based on Youden's index. Then, binary biomarkers (higher or lower than the reference value) were evaluated based on their predictive power, measured by precision, sensitivity, and specificity. Finally, the estimated mean change in CDR.sb in each new group (high biomarker group and low biomarker group) was calculated based on a linear mixed model adjusted for age, sex, and baseline CDR.sb score.

[0232] These identified candidate biomarkers were validated by in silico validation, including trials in other clinical outcomes (ADAS_COg and ADCS_ADL from the AMBAR study), in randomly re-selected samples (1001 resampled datasets, 80% of the original sample size).

[0233] Based on the above results, a final list of candidate biomarkers was prepared using the flowchart shown in Figure 2B.

[0234] Selected single plasma protein biomarkers are provided in Table 4, and additional predictive information is provided in Tables 5 and 6. [Table 4] [Table 5] [Table 6]

[0235] The data in Tables 5 and 6 demonstrate that the DLL1 level can be used as an exclusion criterion when looking for better outcomes than baseline after treatment, given its specificity of 92.5%. The DLL1 level has a balanced sensitivity and specificity, as well as a precision of 78.4%, so it can be used as a predictive biomarker when looking for deterioration of less than 1 point after treatment.

[0236] Table 7 shows in silico validation of random sampling and other clinical outcomes, AUC, to predict improvement from baseline or at least not deterioration beyond CMC. [Table 7] 1. What are the clinically significant changes in ADAS-Cog? (2012) J. Neurol Neurosurg Psychiatry;83(2):171-3.

[0237] Figures 3A to 3C describe the predicted clinical changes in stratified subjects based on the candidate predictive biomarker baseline DLL1.

[0238] Figures 4A to 4C describe the predicted clinical changes in stratified subjects based on the candidate predictive biomarker baseline VNN2.

[0239] Figures 5A to 5C describe the predicted clinical changes in stratified subjects based on the candidate predictive biomarker baseline VAV3.

[0240] Figures 6A to 6C describe the predicted clinical changes in stratified subjects based on the candidate predictive biomarker baseline SUMF1.

[0241] Notwithstanding the attached claims, this disclosure is also defined by the following clauses:

[0242] 1. A method for analyzing samples obtained from a subject, The assay involves assaying a sample for one or more proteins selected from delta-like canonical notch ligand 1 (DLL1), vascular non-inflammatory molecule 2 (VNN2), guanine nucleotide exchange factor (VAV3), and sulfatase modifier 1 (SUMF1); A method in which the subject has or is suspected of having a cognitive impairment.

[0243] 2. The method according to Clause 1, comprising assaying a sample for DLL1.

[0244] 3. The method according to Clause 1, comprising assaying a sample for VNN2.

[0245] 4. The method according to Clause 1, comprising assaying a sample for VAV3.

[0246] 5. The method according to Clause 1, comprising assaying a sample for SUMF1.

[0247] 6. The method according to any one of clauses 1 to 5, comprising qualitatively assaying a sample for one or more proteins.

[0248] 7. The method according to any one of the clauses 1 to 5, comprising quantitatively assaying a sample for one or more proteins.

[0249] 8. The method described in any one of the clauses 1 to 7, wherein the subject has a cognitive impairment.

[0250] 9. The method described in any one of the paragraphs 1 to 7, for a subject suspected of having a cognitive impairment.

[0251] 10. The method described in any one of the preceding clauses, in which cognitive impairment is caused by a neurodegenerative disease.

[0252] 11. The method according to Clause 10, wherein the neurodegenerative disease is Alzheimer's disease (AD), Parkinson's disease, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, or vascular dementia.

[0253] 12. The method according to Clause 11, wherein the neurodegenerative disease is AD.

[0254] 13. The method described in any one of the preceding clauses, wherein the subject is a candidate for plasma exchange therapy to treat cognitive impairment.

[0255] 14. The method according to any one of the preceding clauses, comprising assaying one or more proteins in an immunoassay, mass spectrometry, or protein detection array analysis.

[0256] 15. The method according to any one of clauses 1 to 13, comprising assaying one or more proteins in an aptamer-based multiplex proteomic assay.

[0257] 16. The method according to any one of the preceding clauses, comprising comparing an assay result to a threshold value or a reference value.

[0258] 17. The method according to any one of the preceding clauses, wherein the sample is a blood sample, a serum sample, a plasma sample, or a cerebrospinal fluid sample.

[0259] 18. Determining, based on protein expression data, whether a subject is likely to respond positively to plasma exchange therapy for treating cognitive impairment in the subject, The method according to any one of the preceding clauses.

[0260] 19. The method according to clause 18, further comprising treating the subject for cognitive impairment by administering plasma exchange therapy to the subject if the subject is determined to be likely to respond positively to plasma exchange therapy.

[0261] 20. A method for treating a subject for cognitive impairment, the method comprising administering plasma exchange therapy to the subject, wherein the subject is identified as being likely to respond positively to plasma exchange therapy for treating cognitive impairment.

[0262] 21. The method according to clause 20, wherein the subject is identified as being likely to respond positively to plasma exchange therapy for treating cognitive impairment based on results of assaying a sample from the subject for one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1.

[0263] 22. The method according to clause 20 or 21, wherein the subject is identified as likely to respond positively to plasmapheresis for treating cognitive impairment based on the results of assays of the subject's sample for two or more proteins selected from DLL1, VNN2, VAV3, and SUMF1.

[0264] 23. The method according to any one of clauses 20-22, wherein the subject is identified as likely to respond positively to plasmapheresis for the treatment of cognitive impairment based on the results of assays of the subject's sample for three or more proteins selected from DLL1, VNN2, VAV3, and SUMF1.

[0265] 24. The method according to any one of the clauses 20-23, wherein the subject is identified as likely to respond positively to plasmapheresis for the treatment of cognitive impairment based on the results of assays of the subject's sample for all of the proteins DLL1, VNN2, VAV3 and SUMF1.

[0266] 25. The method according to Clause 20 or 24, wherein the subject is identified, based on the results of an assay of the subject's sample for DLL1, as likely to respond positively to plasmapheresis for treating cognitive impairment.

[0267] 26. The method according to Clause 20 or 21, wherein the subject is identified as likely to respond positively to plasmapheresis for treating cognitive impairment based on the results of an assay of the subject's sample for VNN2.

[0268] 27. The method according to Clause 20 or 21, wherein the subject is identified as likely to respond positively to plasmapheresis for treating cognitive impairment based on the results of an assay of the subject's sample for VAV3.

[0269] 28. The method according to Clause 20 or 21, wherein the subject is identified as likely to respond positively to plasmapheresis for treating cognitive impairment based on the results of an assay of the subject's sample for SUMF1.

[0270] 29. The method according to any one of the clauses 20-28, wherein the subject is identified as likely to respond positively to plasmapheresis for treating cognitive impairment based on increased levels of one or more assayed proteins in a sample obtained from the subject compared to a threshold, reference value, or reference sample.

[0271] 30. Plasma exchange therapy is the method described in any one of the clauses 20 to 29, including the process of complete plasma exchange.

[0272] 31. The method according to Clause 30, wherein total plasma exchange comprises replacing substantially all of the plasma in question with a first albumin solution.

[0273] 32. The method according to clause 30 or 31, wherein the process of total plasma exchange includes one total plasma exchange per week for 5 to 8 weeks.

[0274] 33. The method according to any one of the clauses 30 to 32, wherein the process of total plasma exchange includes total plasma exchange once a week for six weeks.

[0275] 34. The method according to any one of the clauses 31 to 33, wherein the first albumin solution contains 5% albumin.

[0276] 35. The method according to any one of the clauses 30 to 34, wherein plasma exchange therapy includes a step of low-volume plasma exchange with a second albumin solution after a step of total plasma exchange.

[0277] 36. The method according to Clause 35, wherein low-volume plasma exchange comprises exchanging 20% ​​to 40% of the target plasma with a second albumin solution.

[0278] 37. The method according to clause 35 or 36, wherein the course of low-volume plasma exchange comprises low-volume plasma exchange once a month for a period of at least 10 months.

[0279] 38. The method according to any one of clauses 35 to 37, wherein the course of low-volume plasma exchange comprises low-volume plasma exchange once a month for a period of 12 to 16 months.

[0280] 39. The method according to any one of clauses 35 to 38, wherein the second albumin solution comprises 20% albumin.

[0281] 40. A kit comprising a reagent for measuring the amount of one or more proteins selected from DLL1, VNN2, VAV3 and SUMF1 in a sample.

[0282] 41. The kit according to clause 40, comprising a reagent for measuring the amount of DLL1 in a sample.

[0283] 42. The kit according to clause 40, comprising a reagent for measuring the amount of VNN2 in a sample.

[0284] 43. The kit according to clause 40, comprising a reagent for measuring the amount of VAV3 in a sample.

[0285] 44. The kit according to clause 40, comprising a reagent for measuring the amount of SUMF1 in a sample.

[0286] 45. The kit according to any one of clauses 40 to 44, wherein the reagent comprises one or more specific binding members that specifically bind to the corresponding one or more proteins.

[0287] 46. The kit according to clause 45, wherein each of the one or more specific binding members is independently an antibody or antigen-binding fragment thereof, an aptamer, or a peptide binding member.

[0288] 47. The kit according to clause 45 or 46, wherein the one or more specific binding members are labeled with a detectable moiety.

[0289] 48. The kit described in Clause 47, wherein the detectable portion is an optically detectable portion.

[0290] 49. A kit as described in any one of clauses 40-48, suitable for performing immunoassays, Western blot analysis, mass spectrometry, or protein detection array analysis.

[0291] 50. A kit suitable for performing an aptamer-based multiple proteomics assay, as described in any one of clauses 40-48.

[0292] 51. A kit as described in any one of the clauses 40-50, for use in identifying whether a subject is likely to respond positively to plasmapheresis for the treatment of cognitive impairment in that subject.

[0293] In at least some of the embodiments described above, one or more elements used in the embodiments may be interchangeably used in other embodiments, provided that such substitution is not technically impossible. Those skilled in the art will understand that various other omissions, additions, and modifications can be made to the above methods and structures without departing from the scope of the claimed subject matter. All such modifications and changes are intended to fall within the scope of the subject matter, as defined by the appended claims.

[0294] In general, the terms used herein, and in particular in the appended claims (e.g., the body of the appended claims), will be understood by those skilled in the art to be generally intended as “open” terms (for example, “including” should be interpreted as “including but not limited to,” “having” as “having at least,” and “includes” as “includes but not limited to,” and so on). Where a certain number of introduced claims are intended, such intention will be explicitly stated in the claims, and where there is no such statement, such intention will not be present, will be understood by those skilled in the art to be further understood. For example, to aid understanding, the following appended claims may contain the use of the introductory phrases “at least one” and “one or more” to introduce the claims. However, the use of such phrases should not be interpreted as meaning that the introduction of a claim description by the indefinite article "a" or "an" limits any particular claim containing such introduced claim description to only one embodiment containing such description, even if the same claim contains the introductory phrase "one or more" or "at least one" and an indefinite article such as "a" or "an" (for example, "a" and / or "an" should be interpreted as meaning "at least one" or "one or more"); the same applies to the use of definite articles used to introduce claim descriptions.Furthermore, even if a specific number of claims introduced is explicitly stated, a person skilled in the art will recognize that such a statement should be interpreted as meaning at least the number stated (for example, the bare statement “two statements” without other modifiers means at least two statements, or two or more statements). Furthermore, when a convention similar to “at least one of A, B, and C, etc.” is used, it is generally intended to be a construction that a person skilled in the art will understand as such (for example, “a system having at least one of A, B, and C” includes, but is not limited to, a system having only A, a system having only B, a system having only C, a system having both A and B, a system having both A and C, a system having both B and C, and / or a system having both A, B, and C, etc.). Where conventions similar to “at least one of A, B, or C” are used, they are generally intended to be constructed in a way that a person skilled in the art would understand (for example, “a system having at least one of A, B, or C” includes, but is not limited to, a system having only A, a system having only B, a system having only C, a system having both A and B, a system having both A and C, a system having both B and C, and / or a system having both A, B, and C). A person skilled in the art will further understand that substantially any disjunctive word and / or phrase presenting two or more alternative terms should be understood as construing the possibility of including one of the terms, either of the terms, or both of the terms, whether in the specification, claims, or drawings. For example, the phrase “A or B” is understood to include the possibilities of “A” or “B” or “A and B”.

[0295] Furthermore, where any feature or aspect of the present disclosure is described in relation to the Markush group, a person skilled in the art will recognize that the present disclosure also describes any individual member or subgroup of a member of the Markush group.

[0296] As will be understood by those skilled in the art, for any and all purposes, for providing, for example, written explanations, all scopes disclosed herein also encompass any and all possible subscopes and combinations thereof. Any enumerated scope can be readily recognized as sufficiently explainable and enabling the division of the same scope into at least equal halves, thirds, quarters, fifths, tenths, etc. As a non-limiting example, each scope described herein can readily be broken down into a lower third, a middle third, an upper third, etc. Also, as will be understood by those skilled in the art, all language such as “up to,” “at least,” “greater than,” and “less than” includes the enumerated numbers and refers to scopes that can be broken down into subscopes as described above. Finally, as will be understood by those skilled in the art, a scope includes each individual member. Thus, for example, a group having 1 to 3 articles refers to a group having 1, 2, or 3 articles. Similarly, a group having 1 to 5 articles refers to a group having 1, 2, 3, 4, or 5 articles, and so on.

[0297] While the aforementioned inventions are described in some detail as examples and illustrations for clarity, it will be readily apparent to those skilled in the art that certain changes and modifications can be made in light of the teachings of the present invention without departing from the spirit or scope of the appended claims.

[0298] Therefore, the above merely illustrates the principles of the present invention. Those skilled in the art will understand that various configurations embodying the principles of the present invention and falling within its spirit and scope can be devised, although not expressly described or illustrated herein. Furthermore, all examples and conditional statements enumerated herein are intended primarily to help the reader understand the principles of the present invention and the concepts to which the inventors have contributed to advancing the art, and should be construed as not being limited to such specifically enumerated examples and conditions. Moreover, all descriptions herein enumerating the principles, aspects, and embodiments of the present invention, as well as specific examples thereof, are intended to encompass both structural and functional equivalents. In addition, such equivalents are intended to include both currently known equivalents and equivalents to be developed in the future, i.e., any developed elements that perform the same function regardless of their structure. Furthermore, nothing disclosed herein is intended to be made available to the public, whether such disclosure is expressly described in the claims or not.

[0299] Accordingly, the scope of the present invention is not intended to be limited to the exemplary embodiments illustrated and described herein. Rather, the scope and spirit of the present invention are embodied in the appended claims. In the claims, 35 U.SC § 112(f) or 35 U.SC § 112(6) are explicitly defined as being invoked for limitation in a claim only if the exact phrase “means for” or the exact phrase “step for” is placed at the beginning of such limitation in the claim. If such exact phrase is not used in limitation of the claim, 35 U.SC § 112(f) or 35 U.SC § 112(6) is not invoked.

Claims

1. A method for analyzing samples obtained from a subject, The assay includes assaying the sample for one or more proteins selected from delta-like canonical notch ligand 1 (DLL1), vascular non-inflammatory molecule 2 (VNN2), guanine nucleotide exchange factor (VAV3), and sulfatase modifier 1 (SUMF1); A method in which the subject has or is suspected of having a cognitive impairment.

2. The method according to claim 2, comprising qualitatively assaying the sample for one or more proteins.

3. The method according to claim 1 or 2, comprising quantitatively assaying the sample for one or more proteins.

4. The method according to any one of claims 1 to 3, wherein the subject has the cognitive impairment or is suspected of having the cognitive impairment.

5. The method according to any one of claims 1 to 4, wherein the cognitive impairment is caused by a neurodegenerative disease.

6. The method according to any one of claims 1 to 5, wherein the subject is a candidate for plasma exchange therapy to treat the cognitive impairment.

7. The method according to any one of claims 1 to 6, comprising assaying one or more proteins in an immunoassay, mass spectrometry, protein detection array analysis, or aptamer-based multiplexed proteomics assay.

8. The method according to any one of claims 1 to 7, comprising comparing the results of the assay with a threshold or reference value.

9. The method according to any one of claims 1 to 8, wherein the sample is a blood sample, serum sample, plasma sample, or cerebrospinal fluid sample.

10. The process includes determining, based on the protein expression data, whether the subject is likely to respond positively to plasma exchange therapy to treat cognitive impairment in the subject. The method according to any one of claims 1 to 9.

11. The method according to claim 10, further comprising treating the subject for cognitive impairment by administering the plasma exchange therapy to the subject if it is determined that the subject is likely to respond positively to the plasma exchange therapy.

12. A method for treating cognitive impairment in a subject, The method comprises administering plasmapheresis to the subject, wherein the subject is identified as likely to respond positively to the plasmapheresis for treating the cognitive impairment.

13. The method according to claim 12, wherein the subject is identified as likely to respond positively to plasma exchange therapy for treating the cognitive impairment, based on the results of assays of the subject sample for one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1.

14. The method according to claim 12 or 13, wherein the subject is identified as likely to respond positively to plasma exchange therapy for treating the cognitive impairment, based on the results of assays of the subject sample for two or more proteins selected from DLL1, VNN2, VAV3, and SUMF1.

15. A kit containing reagents for measuring the amount of one or more proteins selected from DLL1, VNN2, VAV3, and SUMF1 in a sample.

Images