Gene related to multiple system atrophy, gene product thereof, and test and diagnostic method

Abstract

To provide a method and a kit related to the diagnosis, examination or the like of multiple system atrophy (MSA).SOLUTION: The present invention provides a method for examining whether or not a subject is likely to suffer from, has suffered from, or is likely to suffer from MSA by examining the number of AGG repeats in intron 1 of BIN1 gene; a kit for carrying out such a method; a use of a BIN1 gene containing an AGG repeat in intron 1; and MSA model cells, MSA model animals, and the like.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to the diagnosis and testing of multiple system atrophy. [Background technology]

[0002] Multiple system atrophy (MSA) is a neurodegenerative disease that develops in adulthood. It is classified into type P (MSA-P), which exhibits autonomic dysfunction and Parkinsonian symptoms, and type C (MSA-C), which exhibits cerebellar symptoms. There is no effective treatment, and the prognosis is poor, with patients becoming wheelchair-bound approximately five years after onset, becoming bedridden within eight years, and the disease duration reportedly reaching nine years. Therefore, elucidation of the cause and development of a treatment are urgently needed. The number of patients in Japan is estimated to be approximately 12,000, which is comparable to the approximately 10,000 patients of amyotrophic lateral sclerosis (ALS), another incurable disease. However, research into MSA has been significantly less advanced than that for ALS.

[0003] A pathological hallmark of MSA is the accumulation of insoluble α-synuclein in oligodendroglia, known as glial cytoplasmic inclusions (GCIs), which lead to progressive cell degeneration and loss. While α-synuclein accumulates in neurons as Lewy bodies in Parkinson's disease, in MSA it accumulates as GCIs in glial cells. Interestingly, recent cryo-electron microscopy findings have revealed that the structure of the accumulated α-synuclein in each type of disease is distinct. However, the mechanism of GCI formation remains largely unknown. While familial cases of Parkinson's disease account for approximately 5-10% of cases, MSA is extremely rare, occurring in approximately 1% or less. This, in part, has led to limited progress in genetic research into MSA's etiology, mechanisms, diagnosis, and treatment. Furthermore, genome-wide association studies (GWAS) of MSA have not revealed any significant associations at the genome level (see Non-Patent Documents 1 and 2). [Prior art documents] [Non-patent literature]

[0004] [Non-Patent Document 1] Sailer, A. et al. A genome-wide association study in multiple system atrophy. Neurology 87, 1591-1598 (2016). [Non-patent document 2] Hopfner, F. et al. Common Variants Near ZIC1 and ZIC4 in Autopsy-Confirmed Multiple System Atrophy. Mov Disord 37, 2110-2121 (2022). Summary of the Invention [Problem to be solved by the invention]

[0005] There was a need to find a way to diagnose and test for MSA. [Means for solving the problem]

[0006] The present inventors have conducted extensive research to solve the above problems and have discovered that AGG repeats are expanded in intron 1 of the BIN1 (bridging integrator 1) gene in MSA patients, thereby completing the present invention.

[0007] Thus, the present invention provides: (1) A method for testing whether a subject may have, have had, or may be suffering from MSA by determining the number of AGG repeats in intron 1 of the BIN1 gene in a sample from the subject, and comparing the number of repeats, if greater than the number of repeats in individuals not suffering from MSA, with the criteria that the subject may have, may have had, or may be suffering from MSA. (2) The method according to (1), wherein the criterion is that if the number of repeats is 80 or more, the subject may have, may have had, or may be suffering from MSA. (3) The method according to (1) or (2), wherein the sample is blood, cerebrospinal fluid, brain, or postmortem brain. (4) A method for diagnosing whether a subject may have, have had, or may be suffering from MSA, comprising determining the number of AGG repeats in intron 1 of the BIN1 gene in a sample derived from the subject, and diagnosing that the subject may have, have had, or may be suffering from MSA if the number of repeats is greater than the number of repeats in an individual not suffering from MSA. (5) The method according to (4), wherein if the number of repeats is 80 or more, the subject is diagnosed as possibly having, having had, or being likely to have MSA. (6) The method according to (4) or (5), wherein the sample is blood, cerebrospinal fluid, brain, or postmortem brain. (7) Use of the number of AGG repeats in intron 1 of the BIN1 gene in a sample from a subject as a marker for use in diagnosing whether the subject may have, have had, or may be suffering from MSA. (8) The use according to (7), wherein the subject is diagnosed as possibly having, having had, or being likely to have MSA if the number of repeats is greater than the number of repeats in an individual not having MSA, or if the number of repeats is 80 or more. (9) The use according to (7) or (8), wherein the sample is blood, cerebrospinal fluid, brain, or postmortem brain. (10) A kit for diagnosing whether a subject may have, have had, or may be suffering from MSA, comprising the means necessary for determining the number of AGG repeats in intron 1 of the BIN1 gene in a sample derived from the subject. (11) The kit according to (10), which diagnoses that a subject may have, have had, or may be suffering from MSA if the number of repeats is greater than the number of repeats in an individual not suffering from MSA, or if the number of repeats is 80 or more. (12) The kit according to (10) or (11), wherein the sample is blood, cerebrospinal fluid, brain, or postmortem brain. (13) A BIN1 gene containing AGG repeats in intron 1, the number of repeats being greater than the number of repeats in individuals not affected by MSA. (14) The gene according to (13), having 80 or more repeats. (15) MSA model cells expressing the BIN1 gene containing an AGG repeat in intron 1. (16) MSA model animals expressing the BIN1 gene containing an AGG repeat in intron 1. [Effects of the Invention]

[0008] According to the present invention, it is possible to determine whether a subject is likely to have, have had, or will have MSA. [Brief explanation of the drawings]

[0009] [Figure 1] Figure 1 shows the pedigree of an MSA family. The arrow indicates the index patient. Numbers represent the number of AGG repeats in BIN1, and bold indicates expanded alleles. [Figure 2] Figure 2 shows the brain MRI images of individual II-2. The left image is a T2-weighted axial slice, and the right image is a T1-weighted sagittal slice. [Figure 3] Figure 3 is a bar graph showing the relationship between the number of repeats and the number of reads in intron 1 of the BIN1 gene in the peripheral blood sample of individual II-2. Red indicates the number of repeats on the + strand, and blue indicates the number of repeats on the - strand. [Figure 4]Figure 4 shows the distribution of AGG repeat numbers in intron 1 of the BIN1 gene in autopsy brains of normal controls (left) and MSA patients (right). [Figure 5] Figure 5 shows the results of staining GCIs in the brain of an MSA patient with BIN1 antibody. The arrow indicates the GCI. [Figure 6] FIG. 6 shows Western blotting using BIN1 antibody on sarkosyl-insoluble fractions of brain tissue from MSA patients with and without repeat expansions, and controls. [Figure 7] Figure 7 shows fluorescence in situ hybridization-immunofluorescence (FISH-IF) images of brain tissue from an MSA patient. The upper left panel shows an image of the AGG repeat-containing BIN1 gene (mRNA) in brain tissue (red). The upper right panel shows immunofluorescent staining of phosphorylated α-synuclein (green). The lower left panel shows nuclear staining (blue). The lower right panel is a merged image of these three images. The scale bar in the lower right panel is 10 μm and applies to all panels. [Figure 8] The left panel of Figure 8 shows a fluorescence in situ hybridization (FISH) image of oligodendrocytes differentiated from iPS cells derived from an MSA patient. Red indicates the fluorescence of the BIN1 gene (mRNA) containing AGG repeats. Blue indicates the fluorescence of nuclear staining. The right panel of Figure 8 shows an image of nuclear staining (blue) in control cells. DETAILED DESCRIPTION OF THE INVENTION

[0010] The present invention provides, in one aspect, A method for testing whether a subject may have, may have had, or may be suffering from MSA by determining the number of AGG repeats in intron 1 of the BIN1 gene in a sample from the subject, and comparing the number of repeats to a criterion that the subject may have, may have had, or may be suffering from MSA if the number of repeats is greater than the number of repeats in individuals not suffering from MSA. to provide.

[0011] As used herein, "possibly having MSA" means that the possibility of having MSA is 10% or more, preferably 30% or more, more preferably 50% or more, and even more preferably 70% or more. "possibly having had MSA" means that the possibility of having MSA is 10% or more, preferably 30% or more, more preferably 50% or more, and even more preferably 70% or more. "possibly having MSA" means that the possibility of having MSA is 10% or more, preferably 30% or more, more preferably 50% or more, and even more preferably 70% or more.

[0012] In another aspect, the present invention provides a method for producing a pharmaceutical composition comprising: A method for diagnosing whether a subject may have, have had, or is likely to have MSA, comprising determining the number of AGG repeats in intron 1 of the BIN1 gene in a sample from the subject, and diagnosing that the subject may have, have had, or is likely to have MSA if the number of repeats is greater than the number of repeats in an individual not suffering from MSA. to provide.

[0013] In yet another aspect, the present invention provides a method for producing a pharmaceutical composition comprising: Use of the number of AGG repeats in intron 1 of the BIN1 gene in a sample from a subject as a marker for use in diagnosing whether the subject may have, has had or may have, or may have MSA. to provide.

[0014] The subject is a mammal, preferably a primate, more preferably a human.

[0015] The sample may be any sample obtained from a subject, as long as it contains the BIN1 gene. The sample may be, for example, a body fluid such as blood, brain, or cerebrospinal fluid. The blood may be peripheral blood. The sample may be obtained from a living subject or a deceased subject. An example of a sample obtained from a deceased subject is postmortem brain. The method of the present invention may be used to make a definitive diagnosis of postmortem brain.

[0016] The BIN1 protein, a product of the BIN1 gene, is known to be essential for the formation of membrane structures in skeletal muscle cells and is also known to be a genetic risk factor for Alzheimer's disease. However, the relationship between the BIN1 gene and MSA was unknown. The present inventors have now discovered for the first time that AGG repeats are expanded in intron 1 of the BIN1 gene in MSA patients. Based on this finding, testing, examination, diagnosis (including definitive diagnosis), etc. of MSA can be performed. Furthermore, since the present invention has discovered AGG repeat expansion in intron 1 of the BIN1 gene in sporadic MSA patients, the present invention is useful for the diagnosis of sporadic MSA.

[0017] As used herein, "AGG repeat" refers to a base sequence containing or consisting of multiple base sequences A(G)n [where n is a natural number of 1 or greater]. Usually, n is 1-10, typically 1-5. An "AGG repeat" may be, for example, 5'-AGGAGGAGAGGGAGGAGGAGGAGGGGAGGAGG-3'. As used herein, "number of AGG repeats" refers to the number of A(G)n in the AGG repeat. In the above example, the number of AGG repeats is 10. As used herein, "AGG repeat expansion" refers to a state in which the number of AGG repeats is increased.

[0018] The position of the AGG repeat in intron 1 is not particularly limited, and may be, for example, 1000 to 2000 base pairs from the 5' end of intron 1. Intron 1 may have two or more AGG repeats.

[0019] A subject can be considered to have, have had, or be likely to have MSA if the number of AGG repeats in intron 1 of the BIN1 gene of the subject is greater than the number of AGG repeats in intron 1 of the BIN1 gene of an individual not suffering from MSA. Based on this criterion, it may be tested whether the subject may have, have had, or be likely to have MSA.

[0020] If the number of AGG repeats in intron 1 of the BIN1 gene in a sample from a subject is greater than the number of AGG repeats in intron 1 of the BIN1 gene in an individual not suffering from MSA, it may be diagnosed that the subject may have, have had, or may be suffering from MSA.

[0021] The number of AGG repeats in intron 1 of the BIN1 gene in a sample from a subject may be used as a marker to diagnose that the subject may have, have had, or may be suffering from MSA if the number is greater than the number of AGG repeats in intron 1 of the BIN1 gene in an individual who does not have MSA.

[0022] The number of AGG repeats in intron 1 of the BIN1 gene of an individual not affected by MSA may be the average number of repeats in a population of individuals not affected by MSA. To improve diagnostic accuracy, it is preferable that the population contains a large number of individuals. For example, the number of individuals may be tens, hundreds, thousands, or tens of thousands. Identification of an individual as not affected by, or possibly affected by, MSA may be based on the presence or absence of major symptoms such as cerebellar symptoms, parkinsonism, autonomic neuropathy, complex tract symptoms, and cognitive function, as well as imaging findings.

[0023] A subject may be considered to have, have had, or may be susceptible to MSA if the number of AGG repeats in intron 1 of the BIN1 gene of the subject is equal to or greater than a certain number. The certain number may be 50 or greater. The certain number may be, for example, 50, 60, 70, 80, 90, 100, or any number between these numbers.

[0024] Methods for determining AGG repeat expansion or the number of AGG repeats in the method of the present invention are known, including, for example, a method using a long-read sequencer, a method in which the repeats are amplified by PCR and then estimated from the product length, a method in which the repeats are detected by repeat-primed PCR, and a method in which the repeats are detected by Southern blotting.

[0025] The AGG repeat expansion or the number of AGG repeats on the + strand of the BIN1 gene may be examined, or the AGG repeat expansion or the number of AGG repeats on the - strand of the BIN1 gene may be examined.

[0026] The method of the present invention may be carried out using an imaging diagnostic probe. The use of an imaging diagnostic probe can contribute to the rapid implementation of the method of the present invention. The imaging diagnostic probe used in the method of the present invention may be a substance that specifically binds to BIN1.

[0027] In yet another aspect, the present invention provides a method for producing a pharmaceutical composition comprising: Provided is a kit for diagnosing whether a subject may have, have had, or may be suffering from MSA, which kit includes the means necessary to determine the number of AGG repeats in intron 1 of the BIN1 gene in a sample derived from the subject.

[0028] The above kit can be used to determine the number of AGG repeats in intron 1 of the BIN1 gene in a sample from a subject, and if the number of repeats is greater than the number of repeats in an individual who does not have MSA, or if the number of repeats is 50 or more, for example 80 or more, it can be diagnosed that the subject may have, have had, or may be suffering from MSA.

[0029] The means included in the kit may be, for example, a means for determining the number of AGG repeats in intron 1 of the BIN1 gene using a long-read sequencer, and may be a method of PCR amplification followed by estimation from the length of the product, a method of detection by repeat-primed PCR, a method of detection by Southern blotting, etc. Usually, the kit comes with an instruction manual.

[0030] In yet another aspect, the present invention provides a method for producing a pharmaceutical composition comprising: Provided is a BIN1 gene containing an AGG repeat expansion in intron 1, the number of repeats of which is greater than the number of repeats in individuals not affected by MSA.

[0031] The BIN1 gene was first identified by the present inventors. Preferably, the gene is isolated.

[0032] Based on the number of AGG repeats in intron 1 of the BIN1 gene, it is possible to test, investigate, determine, diagnose, etc., whether a subject is likely to have, have had, or will have MSA. For this reason, the BIN1 gene is useful.

[0033] If the number of AGG repeats is greater than the number of repeats in individuals who do not have MSA, or if the number of repeats is 50 or more, for example 80 or more, it can be considered that the subject may have, have had, or may be susceptible to MSA.

[0034] The methods, uses, kits and / or genes of the present invention may be used in conjunction with known methods for diagnosing MSA.

[0035] In a further aspect, the present invention provides an MSA model cell. The MSA model cell of the present invention expresses a gene associated with the onset of MSA. A specific example of the MSA model cell of the present invention is an MSA model cell that expresses the BIN1 gene, which contains an AGG repeat in intron 1.

[0036] The MSA model cells of the present invention can be produced using known methods. For example, iPS cells can be obtained by introducing a reprogramming gene into somatic cells derived from an MSA patient according to known methods, and the obtained iPS cells can be differentiated into central nervous system cells such as oligodendrocytes, thereby obtaining the MSA model cells of the present invention. Alternatively, for example, iPS cells can be obtained by introducing a reprogramming gene into somatic cells derived from a healthy subject according to known methods, and a gene associated with MSA onset can be introduced into the iPS cells using genome editing technology, and the gene-introduced iPS cells can be differentiated into central nervous system cells such as oligodendrocytes, thereby obtaining the MSA model cells of the present invention.

[0037] In a further aspect, the present invention provides an MSA animal model. The MSA animal model of the present invention expresses a gene associated with the onset of MSA and preferably develops MSA. A specific example of the MSA animal model of the present invention is an MSA animal model that expresses the BIN1 gene containing an AGG repeat in intron 1. The species of the MSA animal model of the present invention is not particularly limited, and includes mammals (excluding humans) such as mice, rats, dogs, cats, and monkeys.

[0038] The MSA animal model of the present invention can be produced using known methods, for example, by introducing a gene associated with MSA onset into animal ES cells or fertilized eggs using gene modification techniques such as homologous recombination or genome editing.

[0039] Unless otherwise specified, terms used in this specification are to be interpreted as they are commonly understood in the fields of medicine, pharmacology, biochemistry, chemistry, etc. Numerical values ​​used in this specification are to be interpreted as having a range of ±20%, preferably ±10%, and more preferably ±5% of the numerical value.

[0040] The present invention will be explained in more detail and specifically below by showing examples, but the examples should not be construed as limiting the scope of the present invention. [Example]

[0041] Example 1 - Genetic analysis for familial MSA We analyzed a patient with MSA whose siblings were affected (individual II-2 in Figure 1). The patient's younger brother (individual II-3 in Figure 1) died of MSA at the age of 67. Individual II-2 first experienced unsteady walking at age 75, and his motor function gradually deteriorated. At age 76, he began to have difficulty moving his upper limbs and speaking, and was diagnosed with MSA-cerebellar type (MSA-C) by a neurologist. Furthermore, Individual II-2 experienced dizziness and vocalization during sleep. MRI scans of Individual II-2 showed hot cross bun signs and atrophy of the cerebral cortex and cerebellum, confirming his diagnosis of MSA (Figure 2). Similar MRI images were obtained for Individual II-3, confirming his diagnosis of MSA.

[0042] First, we obtained peripheral blood DNA from individual II-2 and performed exome sequencing to confirm the absence of rare mutations in SNCA, LRRK2, and COQ2, which have been reported to be associated with MSA5. To evaluate repeat expansions, we performed whole-genome sequencing using a long-read sequencer (PromethION). Whole-genome sequencing was performed using a long-read sequencer as follows: 1.5 μg of DNA sample from peripheral blood was used with a ligation sequencing kit (SQK-LSK-110, Oxford Nanopore Technologies) for library preparation according to the manufacturer's protocol. Sequencing was performed using a PromethION with an R9.4.1 flow cell (Oxford Nanopore Technologies). Data analysis was performed as previously described. Briefly, mapping was performed using LAST (version 983, https: / / github.com / mcfrith / last-genome-alignments), and repeat length was assessed using tandem genotypes (https: / / github.com / mcfrith / tandem-genotypes).

[0043] Analysis using Tandem Genotype software confirmed an AGG repeat expansion in intron 1 of BIN1 (chr2:127,105,468-127,105,503 [hg38]). The number of AGG repeats ranged from 200 to 800 (Figure 3). Using long-read sequencing data, we constructed a consensus sequence for this repeat, which revealed that the typical repeat number was 257, with irregular variations in the repeat unit sequence. An example of the observed repeat pattern was (AGG) 68 (AGGG)1(AGG) 70 (AG)1(AGG) 102 (AG)1(AGG)3(AGGG)1(AGG)7(AGGGG)1(AGG)2.

[0044] Repeat-primed PCR fragment analysis was used to assess repeat expansion in intron 1 of BIN1 in additional family members (individuals III-3 and III-4). Individual III-3, like individual II-2, displayed a sawtooth pattern in the assay, suggesting repeat expansion (data not shown). Cas9-mediated enrichment sequencing revealed that III-3 had 159 AGG repeats. Although individual III-3 had no neurological symptoms at age 55, continued observation was warranted to monitor for MSA.

[0045] Example 2 - Screening for repeat expansions on MSAs Next, we screened for AGG repeat expansion in intron 1 of BIN1 using repeat-primed PCR in autopsy brains of control and MSA patients, and determined the repeat number using amplicon length PCR or Cas9-mediated enrichment sequencing. As a result, we found AGG repeat expansion in 10 of 68 MSA cases, approximately 15%. We observed a statistically significant increase in the frequency of repeat numbers greater than 80 in the MSA group compared to the control group (Figure 4, p=0.002). This result demonstrates the high accuracy of our method.

[0046] Example 3 - Association of BIN1 protein with MSA pathology We performed immunohistochemical analysis to evaluate the localization of BIN1. BIN1-positive cells were found in the cerebellar white matter lesions of MSA patients. High-magnification images revealed that the BIN1-positive cells were not neurons but glial cytoplasmic inclusions (GCIs). GCIs were stained with a BIN1 antibody (Figure 5). Some GCIs were stained with a phosphorylated α-synuclein antibody, some with both phosphorylated α-synuclein and BIN1 antibodies, and some with BIN1 antibody alone. Western blotting using a BIN1 antibody on sarkosyl-insoluble fractions of brain tissue from MSA patients with and without repeat expansions, and controls, revealed BIN1 bands in MSA patient samples, regardless of whether the repeat expansion was present, but not in control samples (Figure 6). These results demonstrate that the BIN1 protein is involved in the pathogenesis of MSA genetically, neuropathologically, and biochemically.

[0047] Example 4 - Detection of AGG repeat-containing BIN1 gene expression in brain tissue of MSA patients Brain tissue sections were obtained from MSA patients and subjected to the following procedures: (1) detection of the BIN1 gene containing an AGG repeat using a FISH probe, (2) immunofluorescence staining for phosphorylated α-synuclein, and (3) nuclear staining. In (1), a FISH probe targeting the AGG repeat was ordered and synthesized by IDT. In (2), the primary antibody was Anti-Phosphorylated α-Synuclein Monoclonal Antibody (cat# pSyn#64, Wako Chemical, Tokyo, Japan), and the secondary antibody was Goat anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor 488® (Thermo Fisher Scientific, cat# A11029). In (3), DAPI (4',6-diamidino-2-phenylindole) was used for staining. As shown in Figure 7, the image of the BIN1 gene (mRNA) containing AGG repeats in brain tissue (upper left panel, red), the immunofluorescent staining image of phosphorylated α-synuclein (upper right panel, green), and the nuclear staining image (lower left panel, blue) were superimposed (lower right panel). These results confirmed the expression of the BIN1 gene containing AGG repeats in brain tissue.

[0048] Example 5 - Preparation of MSA model cells According to a known method, iPS cells were obtained by introducing reprogramming factors into lymphocytes derived from MSA patients. These iPS cells were then cultured after introducing transcription factors for oligodendrocyte differentiation to generate MSA model cells. As a control, iPS cells derived from lymphocytes derived from healthy individuals were used. Gene expression in the cell samples was analyzed by FISH and gene staining. FISH and gene staining were performed as in Example 4. As shown in the left panel of Figure 8, the fluorescence (red) of the BIN1 gene (mRNA) containing AGG repeats overlapped with the fluorescence (blue) of the nuclear stain. In the control cells, only the fluorescence (blue) of the nuclear stain was observed (right panel of Figure 8). These results confirmed that oligodendrocytes expressing the BIN1 gene containing AGG repeats were obtained. [Industrial Applicability]

[0049] The present invention can be used in the fields of diagnostic agents and test reagents, research into brain diseases, and the like.

Claims

1. A method for testing whether a subject may have, have had, or may be suffering from MSA by determining the number of AGG repeats in intron 1 of the BIN1 gene in a sample from the subject, and comparing the number of repeats to a criterion that the subject may have, have had, or may be suffering from MSA if the number of repeats is greater than the number of repeats in individuals who do not have MSA.

2. 2. The method of claim 1, wherein the criterion is that if the number of repeats is 80 or more, the subject may have, may have had, or may be suffering from MSA.

3. 3. The method of claim 1 or 2, wherein the sample is blood, cerebrospinal fluid, brain, or postmortem brain.

4. A method for diagnosing whether a subject may have, have had, or may be suffering from MSA, comprising determining the number of AGG repeats in intron 1 of the BIN1 gene in a sample from the subject, and diagnosing that the subject may have, have had, or may be suffering from MSA if the number of repeats is greater than the number of repeats in an individual not suffering from MSA.

5. The method of claim 4, wherein if the number of repeats is 80 or more, the subject is diagnosed as possibly having, having had, or being likely to have MSA.

6. The method of claim 4 or 5, wherein the sample is blood, cerebrospinal fluid, brain, or postmortem brain.

7. Use of the number of AGG repeats in intron 1 of the BIN1 gene in a sample from a subject as a marker for use in diagnosing whether the subject may have, have had, or may be suffering from MSA.

8. The use of claim 7, wherein a subject is diagnosed as having, having had, or being likely to have MSA if the number of repeats is greater than the number of repeats in an individual not having MSA, or if the number of repeats is 80 or more.

9. 9. The use according to claim 7 or 8, wherein the sample is blood, cerebrospinal fluid, brain, or post-mortem brain.

10. A kit for diagnosing whether a subject may have, have had, or may be suffering from MSA, comprising the necessary means for determining the number of AGG repeats in intron 1 of the BIN1 gene in a sample derived from the subject.

11. The kit of claim 10, which diagnoses that a subject may have, have had, or may be suffering from MSA if the number of repeats is greater than the number of repeats in an individual not suffering from MSA, or if the number of repeats is 80 or more.

12. 12. The kit of claim 10 or 11, wherein the sample is blood, cerebrospinal fluid, brain, or postmortem brain.

13. A BIN1 gene containing an AGG repeat in intron 1, the number of repeats being greater than the number of repeats in individuals not affected with MSA.

14. The gene according to claim 13, wherein the number of repeats is 80 or more.

15. MSA model cells expressing the BIN1 gene containing an AGG repeat in intron 1.

16. MSA model animal expressing the BIN1 gene containing an AGG repeat in intron 1.

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