Genetic Alterations on Chromosome 16 and Methods of Use Thereof for the Diagnosis and Treatment of Type 1 Diabetes
Compositions and methods for the detection and treatment of T1D are provided.
1 . A method for detecting the presence or absence of at least one genetic alteration in a target nucleic acid isolated from a patient for assessing susceptibility for developing type 1 diabetes (T1D), said method comprising:
a) providing a target nucleic acid from a patient sample, said target nucleic acid having a predetermined sequence in the normal population;
b) assessing said target nucleic acid for the presence of a single nucleotide polymorphism which is indicative of an increased or decreased susceptibility of developing T1D.
2 . The method as claimed in claim 1 , wherein said genetic alteration is selected from the group consisting of inversion, deletion, duplication, and insertion of at least one nucleotide in said sequence.
3 . The method of claim 1 , wherein said target nucleic acid is assessed for genetic alterations via a method selected from the group consisting of size analysis, hybridization of allele specific probes, allele-specific primer extension, oligomer ligation, DNA sequencing, single-stranded conformation polymorphism, and quantitative PCR.
4 . The method as claimed in claim 1 , wherein said genetic alteration is a single nucleotide polymorphism (SNP) at the 16p13 region of chromosome 16, said SNP is selected from the group consisting of an A at rs2903692, a C at rs725613, or a G at rs17673553.
5 . The method as claimed in claim 1 , wherein said genetic alteration is a single nucleotide polymorphism at the KIAA0350 locus region of chromosome 16, said SNP being associated with reduced risk of developing T1D, and selected from the group consisting of an A at rs2903692, a C at rs725613, or a G at rs17673553.
6 . The method as claimed in claim 1 , wherein said genetic alteration is a single nucleotide polymorphism at the KIAA0350 locus region of chromosome 16, said SNP being associated with increased risk of developing T1D, and being an A at rs7200786.
7 . The method as claimed in claim 1 , wherein said genetic alteration comprises at least one of the single nucleotide polymorphisms set forth in Tables 1-3.
8 . The method as claimed in claim 1 , wherein said SNP is found in SEQ ID NO: 583.
9 . A method for determining the presence or absence of at least one specific nucleotide in a target nucleic acid for the diagnosis of T1D as claimed in claim 1 , said method comprising the steps of:
a) providing a detectable amount of a target nucleic acid polymer isolated from a chromosomal region known to be associated with diabetes,
b) hybridizing said detectable amount of the nucleic acid polymer with one or more oligonucleotide primers, wherein each primer has a nucleotide sequence that is complementary to a sequence in the target nucleic acid polymer,
c) exposing the hybridized nucleic acid polymer to a polymerization agent in a mixture containing at least one deoxynucleotide, said deoxynucleotide comprising a detectable label,
d) analyzing the polymerization mixture of step (c) for the presence or absence of the primer extension product containing the labeled deoxynucleotide, whereby the identity of the specific nucleotide at the defined site is determined; and
e) assessing said target nucleic acid for the presence of a genetic alteration at said at least one single nucleotide loci, the presence of the polymorphism being associated with a reduced risk of developing T1D.
10 . A kit for practicing the method of claim 9 .
11 . A nucleic acid comprising at least one SNP identified in Tables 1-3 of SEQ ID No: 583.
12 . A microarray comprising the at least one nucleic acid of claim 11 , said nucleic acid optionally being between 10 and 50 nucleotides in length.
13 . A method of management for T1D comprising:
a) identifying a patient who is at an increased risk of developing T1D by detecting at least one of the polymorphisms in Tables 1-3;
b) administering to said patient a therapeutic agent in a therapeutically effective amount to manage T1D.
14 . The method of claim 13 , wherein said therapeutic agent can modulate signaling mediated via the KIAA0350 gene product.
15 . The method of claim 14 , wherein said therapeutic agent can modulate the activity of the protein encoded by SEQ ID NO: 583.
16 . The method according to claim 13 , wherein said therapeutic agent is selected from the group consisting of a small molecule, an antibody, a protein, an oligonucleotide, or an siRNA molecule.
17 . The method according to claim 16 , wherein at least one siRNA molecule is set forth in Table 4.
18 . The method of claim 16 , wherein said therapeutic agent is delivered to an inflammatory cell.
19 . The method of claim 16 , wherein said therapeutic agent modulates natural killer cell activity.
20 . The method of claim 16 , wherein said therapeutic agent modulates signaling in an insulin-producing beta cell.
21 . A method for detecting a SNP which confers protection against developing T1D as claimed in claim 1 , said method comprising: detecting the presence of a single nucleotide polymorphism on chromosome 16p13 in a target nucleic acid from a patient, wherein said SNP is an A at rs2903692, a C at rs725613, or a G at rs17673553 of SEQ ID NO: 583 and is associated with a decreased risk of developing T1D.
22 . The method of claim 21 , wherein said target nucleic acid is amplified prior to detection.
23 . The method of claim 21 , wherein the target nucleic acid is DNA.
24 . A single nucleotide polymorphism associated with a decreased risk of developing T1D as claimed in claim 1 , selected from the group consisting of an A at rs2903692m a C at rs725613, a G at rs17673553 or a SNP in Table IC.
25 . A method for identifying agents which modulate autoimmune beta cell destruction leading to T1D comprising:
a) providing cells expressing a SNP selected from the group consisting of an A at rs2903692m a C at rs725613, or a G at rs17673553;
b) providing cells which express the cognate sequence lacking the SNPs of step a);
c) contacting the cells of steps a) and b) with an agent; and
d) determining whether said agent alters beta cell destruction of the cells of step a) relative to those of step b), thereby identifying agents which modulate autoimmune beta cell destruction.
26 . An siRNA composition comprising at least one nucleotide sequence selected from the group consisting of Table 4 in a pharmaceutically acceptable carrier for delivery to a patient.
27 . A method of inhibiting the expression of CLEC16A in a patient comprising administering to said patient at least one siRNA molecule as claimed in claim 26 that directs cleavage of a target CLEC16A mRNA sequence present in said patient.
28 . The method of claim 27 , wherein said siRNA is introduced directly into said patient.
29 . The method of claim 28 , wherein the siRNA is combined with other conventional agents for treating T1D.
30 . A method for detecting a SNP which confers protection against developing T1D as claimed in claim 1 , said method comprising: detecting the presence of a single nucleotide polymorphism on chromosome 16p13 in a target nucleic acid from a patient, wherein said SNP is an A at rs7200786 of SEQ ID NO: 583 and is associated with a increased risk of developing T1D.
31 . A method of treating T1D in a patient as claimed in claim 13 , comprising administering into said patient an effective amount of a composition that modulates the function, activity, or expression of a gene, transcript, or variant protein identified in Tables 1-3.
32 - 33 . (canceled)
34 . A method for identifying a combination of siRNA molecules as claimed in claim 26 , effective for down-regulating the expression of CLEC16A, comprising:
a) contacting a cell with an effective amount of at least two siRNA molecules provided in Table 4;
b) assessing the effect of said siRNAs on down regulation of CLEC16A expression relative to untreated cells.
35 . The method of claim 34 , wherein said cells are selected from the group consisting of INS cells, PC12 cells, MIN6 cells, pancreatic beta islet cells and alpha TC6 cells.
36 . The method of claim 35 wherein modulatory effects of said siRNAs on a parameter selected from the group consisting of insulin secretion, glucagon secretion and glucosamine induced beta cell apoptosis is determined.