Synaptojanin 2 (SYNJ2) Variants And Uses Thereof

Abstract

The present disclosure provides methods of treating patients having hearing loss, methods of identifying subjects having an increased risk of developing hearing loss, methods of detecting human Synaptojanin-2 (SYNJ2) variant nucleic acid molecules and variant polypeptides, and SYNJ2 variant nucleic acid molecules and variant polypeptides.

Claims

1. A method of identifying a human subject having an increased risk for developing hearing loss, wherein the method comprises: determining or having determined the presence or absence of an Synaptojanin-2 (SYNJ2) rs2256014 predicted loss-of-function variant nucleic acid molecule in a biological sample obtained from the subject; wherein: when the human subject is SYNJ2 reference, then the human subject does not have an increased risk for developing hearing loss; and when the human subject is heterozygous or homozygous for SYNJ2 rs2256014, then the human subject has an increased risk for developing hearing loss.

2-6. (canceled)

7. The method according to claim 1, wherein the determining step is carried out in vitro.

8. The method according to claim 1, wherein the determining step comprises sequencing at least a portion of the nucleotide sequence of the SYNJ2 genomic nucleic acid molecule in the biological sample, wherein the sequenced portion comprises: a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof; wherein when the sequenced portion of the SYNJ2 genomic nucleic acid molecule in the biological sample comprises a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3, then the SYNJ2 genomic nucleic acid molecule in the biological sample is a SYNJ2 predicted loss-of-function variant genomic nucleic acid molecule.

9-10. (canceled)

11. The method according to claim 1, wherein the determining step comprises: a) contacting the biological sample with a primer hybridizing to a portion of the nucleotide sequence of the SYNJ2 genomic nucleic acid molecule that is proximate to a position corresponding to position 99,219 according to SEQ ID NO:3; b) extending the primer at least through the position of the nucleotide sequence of the SYNJ2 genomic nucleic acid molecule corresponding to position 99,219 according to SEQ ID NO:3; and c) determining whether the extension product of the primer comprises a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3.

12-13. (canceled)

14. The method according to claim 8, wherein the determining step comprises sequencing the entire nucleic acid molecule.

15. The method according to claim 1, wherein the determining step comprises: a) amplifying at least a portion of the genomic nucleic acid molecule that encodes the human SYNJ2 polypeptide, wherein the portion comprises a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof; b) labeling the amplified nucleic acid molecule with a detectable label; c) contacting the labeled nucleic acid molecule with a support comprising an alteration-specific probe, wherein the alteration-specific probe comprises a nucleotide sequence which hybridizes under stringent conditions to the nucleic acid sequence of the amplified nucleic acid molecule comprising a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof; and d) detecting the detectable label.

16-18. (canceled)

19. The method according to claim 15, wherein the detecting step comprises: contacting the nucleic acid molecule in the biological sample with an alteration-specific probe comprising a detectable label, wherein the alteration-specific probe comprises a nucleotide sequence which hybridizes under stringent conditions to the nucleotide sequence of the amplified nucleic acid molecule comprising a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof; and detecting the detectable label.

20-21. (canceled)

22. The method according to claim 1, wherein when the human subject is heterozygous or homozygous for SYNJ2 rs2256014, the human subject is further administered a therapeutic agent that treats or inhibits the hearing loss.

23. A method of treating a patient with a therapeutic agent that treats or inhibits hearing loss, wherein the patient is suffering from hearing loss, the method comprising the steps of: determining whether the patient has a Synaptojanin-2 (SYNJ2) rs2256014 predicted loss-of-function variant nucleic acid molecule by: obtaining or having obtained a biological sample from the patient; and performing or having performed a genotyping assay on the biological sample to determine if the patient has a genotype comprising SYNJ2 rs2256014; and when the patient is SYNJ2 reference, then administering or continuing to administer to the patient the therapeutic agent that treats or inhibits hearing loss in a standard dosage amount; and when the patient is heterozygous or homozygous for SYNJ2 rs2256014, then administering or continuing to administer to the patient the therapeutic agent that treats or inhibits hearing loss in an amount that is the same as or greater than a standard dosage amount; wherein the presence of a genotype having SYNJ2 rs2256014 indicates the patient has an increased risk of developing hearing loss.

24-28. (canceled)

29. The method according to claim 23, wherein the genotyping assay comprises sequencing at least a portion of the nucleotide sequence of the SYNJ2 genomic nucleic acid molecule in the biological sample, wherein the sequenced portion comprises a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof; wherein when the sequenced portion of the SYNJ2 genomic nucleic acid molecule in the biological sample comprises a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3; then the SYNJ2 genomic nucleic acid molecule in the biological sample is a SYNJ2 predicted loss-of-function variant genomic nucleic acid molecule.

30-31. (canceled)

32. The method according to claim 23, wherein the genotyping assay comprises: a) contacting the biological sample with a primer hybridizing to a portion of the nucleotide sequence of the SYNJ2 genomic nucleic acid molecule that is proximate to a position corresponding to position 99,219 according to SEQ ID NO:3; b) extending the primer at least through the position of the nucleotide sequence of the SYNJ2 genomic nucleic acid molecule corresponding to position 99,219 according to SEQ ID NO:3; and c) determining whether the extension product of the primer comprises a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3.

33-34. (canceled)

35. The method according to claim 29, wherein the detecting step comprises sequencing the entire nucleic acid molecule.

36. The method according to claim 23, wherein the genotyping assay comprises: a) amplifying at least a portion of the nucleic acid molecule that encodes the human SYNJ2 polypeptide, wherein the portion comprises a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof; b) labeling the amplified nucleic acid molecule with a detectable label; c) contacting the labeled nucleic acid molecule with a support comprising an alteration-specific probe, wherein the alteration-specific probe comprises a nucleotide sequence which hybridizes under stringent conditions to the nucleic acid sequence of the amplified nucleic acid molecule comprising a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof; and d) detecting the detectable label.

37-39. (canceled)

40. The method according to claim 36, wherein the genotyping assay comprises: contacting the genomic nucleic acid molecule in the biological sample with an alteration-specific probe comprising a detectable label, wherein the alteration-specific probe comprises a nucleotide sequence which hybridizes under stringent conditions to the nucleotide sequence of the amplified nucleic acid molecule comprising a thymine at a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof; and detecting the detectable label.

41-42. (canceled)

43. The method according to claim 23, wherein the nucleic acid molecule is present within a cell obtained from the human subject.

44-65. (canceled)

66. An isolated alteration-specific probe or alteration-specific primer comprising at least about 15 nucleotides, wherein the alteration-specific probe or alteration-specific primer comprises a nucleotide sequence which is complementary to a portion pf human Synaptojanin-2 (SYNJ2) rs2256014 variant nucleic acid molecule, wherein the portion comprises a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof.

67-72. (canceled)

73. The alteration-specific probe or alteration-specific primer according to claim 66, wherein the alteration-specific probe or alteration-specific primer comprises DNA.

74. The alteration-specific probe or alteration-specific primer according to claim 66, wherein the alteration-specific probe or alteration-specific primer comprises RNA.

75. The alteration-specific probe or alteration-specific primer according to claim 66, wherein the alteration-specific probe or alteration-specific primer comprises a label.

76. The alteration-specific probe or alteration-specific primer according to claim 75, wherein the label is a fluorescent label, a radiolabel, or biotin.

77. A support comprising a substrate to which an alteration-specific probe or alteration-specific primer according to claim 66 is attached.

78. The support according to claim 77, wherein the support is a microarray.

79. A molecular complex comprising an alteration-specific primer or an alteration-specific probe hybridized to a SYNJ2 rs2256014 genomic nucleic acid molecule, wherein the alteration-specific primer or the alteration-specific probe is hybridized to thymine at a position corresponding to position 99,219 according to SEQ ID NO:3, or the complement thereof.

80-86. (canceled)

87. The molecular complex according to claim 79, wherein the alteration-specific probe or alteration-specific primer comprises a label.

88. The molecular complex according to claim 87, wherein the label is a fluorescent label, a radiolabel, or biotin.

89. The molecular complex according to claim 79, further comprising a non-human polymerase.

90. (canceled)

Description

EXAMPLES

[0151] The following examples are put forth so as to provide those of ordinary skill in the art with a complete disclosure and description of how the compounds, compositions, articles, devices and/or methods claimed herein are made and evaluated, and are intended to be purely exemplary and are not intended to limit the scope of what the inventors regard as their invention. Efforts have been made to ensure accuracy with respect to numbers (e.g., amounts, temperature, etc.), but some errors and deviations should be accounted for. Unless indicated otherwise, parts are parts by weight, temperature is in ° C. or is at ambient temperature, and pressure is at or near atmospheric.

Example 1

Genomic Data Analyses

[0152] Genotype and exome sequence analysis of the UK Biobank 500 k (genotype) and 150 k (exome sequence) dataset identified that two variants, including a rare missense variant, in SYNJ2 significantly associate with increased risk for hearing loss (see, Table 1 and Table 2), and also with increased mean speech reception threshold (SRT) (see, Table 4), wherein speech reception threshold is defined as the minimum intensity (dB) at which a person can recognize 50% of spoken words (individuals with hearing problems will have an increased mean SRT). Further, association analyses of one of the associated variants in SYNJ2 (p.Thr656Met) with hearing loss in the Geisinger 90 k cohort showed a trend towards increased risk of hearing loss in variant carriers, consistent with results from UK Biobank, although this result was not statistically significant (see, Table 4).

TABLE-US-00001 TABLE 1 Association of variants in SYNJ2 with increased risk for self-reported hearing loss in UKB (500 k imputed/genotyped) dataset Name repEffect hgvsc hgvsp OR_95_CI Pval 6:158071628:C:T missense c.1967C > T p.Thr656Met 1.3873 1.37E−12 (1.2672, 1.5188) 6:158081105:G:T splice_region c.2568-4G > T   NA 1.0345 1.87E−08 (1.0223, 1.0468) Name repEffect hgvsc Hgvsp AAF Ncase_RR_RA_AA Nctrl_RR_RA_AA 6:158071628:C:T missense c.1967C > T p.Thr656Met 0.0048 100,633 99,857|770|6 250,910 249,326|1,583|1 6:158081105:G:T splice_region c.2568-4G > T   NA 0.5137 101,158 25,1941 23,362|50,363|27,433 60,151|126,059|65,731

TABLE-US-00002 TABLE 2 Association of SYNJ2 Thr656Met variant with self-reported hearing loss in UKB 150 k exome dataset cDNA; OR Case_N Control_N Position Amino acid Cohort (LCI, UCI) Pvalue AAF RR|RA|AA RR|RA|AA 6:158071628:C:T c.1967C > T; UKB 150 1.343 3.8E−08 0.0053 35,860 106,734 p.Thr656Met exome (1.209, 1.492) 35,396|457|6 105,685|1,039|1

TABLE-US-00003 TABLE 3 Association of SYNJ2 Thr656Met with an increase in mean speech reception threshold (SRT) in UKB 500 k imputed/genotyped and 150 k exome datasets Effect in standard deviation units Sample N Position Variant Cohort (LCI, UCI) Pvalue AAF RR|RA|AA 6:158071628:C:T c.1967C > T; UKB 0.144 3.59E−07 0.0048 145,877 p.Thr656Met 500K (0.088, 0.199) 144,916|958|3 Imputed UKB 0.15 1.3E−04 0.0053 63,117 150K (0.072, 0.22) 62,449|657|3 exome

TABLE-US-00004 TABLE 4 Association of SYNJ2 Thr656Met with ICD10 codes for hearing loss in GHS90k exome meta analysis dataset Cases Controls Phenotype Cohort Variant OR (CI) Pvalue AAF N RR|RA|AA N RR|RA|AA ICD10 3D: GHS 90 k c.1967C > T; 1.236 0.0060 3,673 75,437 Conductive and Meta p.Thr656Met (0.926, 1.648) 1.50E−01 3,621|52|0 74,546|889|2 sensorineural hearing loss ICD10 3D: GHS 90 k c.1967C > T; 1.080 5.48E−01 0.0060 5,478 72,729 Other and Meta p.Thr656Met (0.840, 1.390) 5,410|67|1 71,866|861|2 unspecified hearing loss

Example 2

Detection

[0153] The presence of a certain genetic variant in a subject can indicate that the subject has an increased risk of having or developing hearing loss. A sample, such as a blood sample, can be obtained from a subject. Nucleic acids can be isolated from the sample using common nucleic acid extraction kits. After isolating the nucleic acid from the sample obtained from the subject, the nucleic acid is sequenced to determine if there is a genetic variant present. The sequence of the nucleic acid can be compared to a control sequence (wild type sequence). Finding a difference between the nucleic acid obtained from the sample obtained from the subject and the control sequence indicates the presence of a genetic variant. These steps can be performed as described in the examples above and throughout the present disclosure. The presence of one or more genetic variants is indicative of the subject's increased risk for having or developing early-onset inflammatory bowel disease.

[0154] Various modifications of the described subject matter, in addition to those described herein, will be apparent to those skilled in the art from the foregoing description. Such modifications are also intended to fall within the scope of the appended claims. Each reference (including, but not limited to, journal articles, U.S. and non-U.S. patents, patent application publications, international patent application publications, gene bank accession numbers, and the like) cited in the present application is incorporated herein by reference in its entirety and for all purposes.