Detection of organ rejection
The present invention features methods and compositions for the non-invasive detection of organ rejection using a microRNA score.
1. A method for identifying a subject as having, or having a propensity to develop, organ rejection, the method comprising:
(i) transplanting an allogeneic organ into a subject in need thereof,
(ii) measuring the expression of at least 20 miRNAs selected from the group consisting of:
mmu-miR-134,
(SEQ ID NO: 1)
UGUGACUGGUUGACCAGAGGGG,
mmu-miR-144,
(SEQ ID NO: 2)
UACAGUAUAGAUGAUGUACUAG,
mmu-miR-466,
(SEQ ID NO: 3)
AUACAUACACGCACACAUAAGAC,
mmu-miR-7,
(SEQ ID NO: 4)
UGGAAGACUAGUGAUUUUGUUG,
mmu-miR-346,
(SEQ ID NO: 5)
UGUCUGCCCGAGUGCCUGCCUCU,
mmu-miR-468,
(SEQ ID NO: 6)
UAUGACUGAUGUGCGUGUGUCU,
mmu-miR-188,
(SEQ ID NO: 7)
CAUCCCUUGCAUGGUGGAGGGU,
mmu-miR-298,
(SEQ ID NO: 8)
GGCAGAGGAGGGCUGUUCUUCC,
mmu-miR-467,
(SEQ ID NO: 9)
AUAUACAUACACACACCUACAC,
mmu-miR-292-5p,
(SEQ ID NO: 10)
ACUCAAACUGGGGGCUCUUUUG,
mmu-miR-186,
(SEQ ID NO: 11)
CAAAGAAUUCUCCUUUUGGGCUU,
mmu-miR-486,
(SEQ ID NO: 12)
UCCUGUACUGAGCUGCCCCGAG,
mmu-miR-451,
(SEQ ID NO: 13)
AAACCGUUACCAUUACUGAGUU,
mmu-miR-18,
(SEQ ID NO: 14)
UAAGGUGCAUCUAGUGCAGAUA,
mmu-miR-25,
(SEQ ID NO: 15)
CAUUGCACUUGUCUCGGUCUGA,
mmu-miR-223
(SEQ ID NO: 16)
UGUCAGUUUGUCAAAUACCCC,
mmu-miR-320,
(SEQ ID NO: 17)
AAAAGCUGGGUUGAGAGGGCGAA,
mmu-miR-148b,
(SEQ ID NO: 18)
UCAGUGCAUCACAGAACUUUGU,
mmu-miR-21,
(SEQ ID NO: 19)
UAGCUUAUCAGACUGAUGUUGA,
mmu-miR-124a,
(SEQ ID NO: 20)
UAAGGCACGCGGUGAAUGCC,
mmu-miR-19a,
(SEQ ID NO: 21)
UGUGCAAAUCUAUGCAAAACUGA,
mmu-miR-301
(SEQ ID NO: 22)
CAGUGCAAUAGUAUUGUCAAAGC,
mmu-miR-31,
(SEQ ID NO: 23)
AGGCAAGAUGCUGGCAUAGCUG,
mmu-miR-20,
(SEQ ID NO: 24)
UAAAGUGCUUAUAGUGCAGGUAG,
mmu-miR-29c,
(SEQ ID NO: 25)
UAGCACCAUUUGAAAUCGGU,
mmu-miR-148a,
(SEQ ID NO: 26)
UCAGUGCACUACAGAACUUUGU,
mmu-miR-17-5p,
(SEQ ID NO: 27)
CAAAGUGCUUACAGUGCAGGUAGU,
mmu-miR-185,
(SEQ ID NO: 28)
UGGAGAGAAAGGCAGUUC,
mmu-miR-106a,
(SEQ ID NO: 29)
CAAAGUGCUAACAGUGCAGGUA,
mmu-miR-106b,
(SEQ ID NO: 30)
UAAAGUGCUGACAGUGCAGAU,
mmu-miR-375,
(SEQ ID NO: 31)
UUUGUUCGUUCGGCUCGCGUGA,
mmu-miR-216,
(SEQ ID NO: 32)
UAAUCUCAGCUGGCAACUGUG,
mmu-miR-217,
(SEQ ID NO: 33)
UACUGCAUCAGGAACUGACUGGAU,
mmu-miR-200a,
(SEQ ID NO: 34)
UAACACUGUCUGGUAACGAUGU,
mmu-miR-200b,
(SEQ ID NO: 35)
UAAUACUGCCUGGUAAUGAUGAC,
mmu-miR-429,
(SEQ ID NO: 36)
UAAUACUGUCUGGUAAUGCCGU,
mmu-miR-200c,
(SEQ ID NO: 37)
UAAUACUGCCGGGUAAUGAUGG,
mmu-miR-141
(SEQ ID NO: 38)
UAACACUGUCUGGUAAAGAUGG,
mmu-miR-148a,
(SEQ ID NO: 39)
UCAGUGCACUACAGAACUUUGU,
mmu-miR-152,
(SEQ ID NO: 40)
UCAGUGCAUGACAGAACUUGGG,
mmu-miR-182,
(SEQ ID NO: 41)
UUUGGCAAUGGUAGAACUCACA,
mmu-miR-99a,
(SEQ ID NO: 42)
ACCCGUAGAUCCGAUCUUGU,
mmu-miR-130a,
(SEQ ID NO: 43)
CAGUGC AAUGUU AAAAGGGC AU,
mmu-miR-127,
(SEQ ID NO: 44)
UCGGAUCCGUCUGAGCUUGGC,
mmu-miR-100,
(SEQ ID NO: 45)
AACCCGUAGAUCCGAACUUGUG,
mmu-miR-199a,
(SEQ ED NO: 46)
CCCAGUGUUCAGACU ACCUGUUC,
mmu-miR-199b,
(SEQ ED NO: 47)
CCCAGUGUUU AGACU ACCUGUUC,
mmu-miR-125a,
(SEQ ED NO: 48)
UCCCUGAGACCCUUUAACCUGUG,
mmu-miR-22,
(SEQ ID NO: 49)
AAGCUGCCAGUUGAAGAACUGU,
mmu-miR-434-3p,
(SEQ ID NO: 50)
UUUGAACCAUCACUCGACUCC,
mmu-miR-34a,
(SEQ ID NO: 51)
UGGCAGUGUCUUAGCUGGUUGUU,
mmu-miR-181c,
(SEQ ID NO: 52)
AACAUUCAACCUGUCGGUGAGU,
mmu-miR-139,
(SEQ ID NO: 53)
UCUACAGUGCACGUGUCU,
mmu-miR-130a,
(SEQ ID NO: 54)
CAGUGCAAUGUUAAAAGGGCAU,
mmu-miR-322,
(SEQ ID NO: 55)
CAGCAGCAAUUCAUGUUUUGGA,
mmu-miR-181a,
(SEQ ID NO: 56)
AACAUUCAACGCUGUCGGUGAGU,
mmu-miR-125a,
(SEQ ID NO: 57)
UCCCUGAGACCCUUUAACCUGUG,
mmu-miR-200b,
(SEQ ID NO: 58)
UAAUACUGCCUGGUAAUGAUGAC,
mmu-miR-145,
(SEQ ID NO: 59)
GUCCAGUUUUCCCAGGAAUCCCUU,
mmu-miR-127,
(SEQ ID NO: 60)
UCGGAUCCGUCUGAGCUUGGC,
mmu-miR-199a,
(SEQ ID NO: 61)
CCCAGUGUUCAGACUACCUGUUC,
mmu-miR-425,
(SEQ ID NO: 62)
AUCGGGAAUGUCGUGUCCGCC,
mmu-miR-99b,
(SEQ ID NO: 63)
CACCCGU AGAACCGACCUUGCG,
mmu-let-7e,
(SEQ ID NO: 64)
UGAGGUAGGAGGUUGUAUAGU,
mmu-miR-195,
(SEQ ID NO: 65)
UAGCAGCACAGAAAUAUUGGC,
mmu-miR-152,
(SEQ ID NO: 66)
UCAGUGCAUGACAGAACUUGGG,
mmu-miR-125b,
(SEQ ID NO: 67)
UCCCUGAGACCCUAACUUGUGA,
mmu-miR-187,
(SEQ ID NO: 68)
UCGUGUCUUGUGUUGCAGCCGG,
mmu-miR-324-3p,
(SEQ ID NO: 69)
CCACUGCCCCAGGUGCUGCUGG,
mmu-miR-150,
(SEQ ID NO: 70)
UCUCCCAACCCUUGUACCAGUG,
mmu-miR-28,
(SEQ ID NO: 71)
AAGGAGCUCACAGUCUAUUGAG,
and
mmu-miR-143
(SEQ ID NO: 72)
UGAGAUGAAGCACUGUAGCUCA,
in a biological sample from the subject, and
(ii) calculating altered expression of the at least 20 miRNAs relative to a reference, thereby identifying a subject as having, or having a propensity to develop, organ rejection.
2. The method of claim 1 , wherein at least one miRNA from the at least 20 miRNAs is selected from the group consisting of: miR-182, miR-467, miR-434-3p, miR-181a, miR-19a, and miR-375, wherein an increase in the level of miR-467 or miR-19a or wherein a decrease in the level of miR-375, miR-182, miR-434-3p, or miR-181a indicates an increased likelihood of organ rejection.
3. The method of claim 1 , wherein the transplanted organ is selected from the group consisting of: heart, kidney, liver, lung, and pancreas.
4. The method of claim 3 , wherein the transplanted organ is kidney.
5. The method of claim 1 , wherein expression of at least one miRNA from the at least 20 miRNAs is increased relative to the reference.
6. The method of claim 1 , wherein expression of at least one miRNA from the at least 20 miRNAs is decreased relative to the reference.
7. The method of claim 1 , wherein the amount of expression is determined using a microarray.
8. The method of claim 7 , wherein the microarray comprises any member from the group consisting of a: chip, plate, bead, and membrane.
9. The method of claim 1 , wherein the biological sample is selected from the group consisting of: blood cells, biopsy specimens, urine cells/urine sediment, and cells found in sputum.
10. A method for hybridizing one or more microRNAs in a sample, the method comprising:
(i) amplifying at least one miRNA obtained from a biological sample and selected from the group consisting of:
mmu-miR-134,
(SEQ ID NO: 1)
UGUGACUGGUUGACCAGAGGGG,
mmu-miR-144,
(SEQ ID NO: 2)
UACAGUAUAGAUGAUGUACUAG,
mmu-miR-466,
(SEQ ID NO: 3)
AUACAUACACGCACACAUAAGAC,
mmu-miR-7,
(SEQ ID NO: 4)
UGGAAGACUAGUGAUUUUGUUG,
mmu-miR-346,
(SEQ ID NO: 5)
UGUCUGCCCGAGUGCCUGCCUCU,
mmu-miR-468,
(SEQ ID NO: 6)
UAUGACUGAUGUGCGUGUGUCU,
mmu-miR-188,
(SEQ ID NO: 7)
CAUCCCUUGCAUGGUGGAGGGU,
mmu-miR-298,
(SEQ ID NO: 8)
GGCAGAGGAGGGCUGUUCUUCC,
mmu-miR-467,
(SEQ ID NO: 9)
AUAUACAUACACACACCUACAC,
mmu-miR-292-5p,
(SEQ ID NO: 10)
ACUCAAACUGGGGGCUCUUUUG,
mmu-miR-186,
(SEQ ID NO: 11)
CAAAGAAUUCUCCUUUUGGGCUU,
mmu-miR-486,
(SEQ ID NO: 12)
UCCUGUACUGAGCUGCCCCGAG,
mmu-miR-451,
(SEQ ID NO: 13)
AAACCGUUACCAUUACUGAGUU,
mmu-miR-18,
(SEQ ID NO: 14)
UAAGGUGCAUCUAGUGCAGAUA,
mmu-miR-25,
(SEQ ID NO: 15)
CAUUGCACUUGUCUCGGUCUGA,
mmu-miR-223
(SEQ ID NO: 16)
UGUCAGUUUGUCAAAUACCCC,
mmu-miR-320,
(SEQ ID NO: 17)
AAAAGCUGGGUUGAGAGGGCGAA,
mmu-miR-148b,
(SEQ ID NO: 18)
UCAGUGCAUCACAGAACUUUGU,
mmu-miR-21,
(SEQ ID NO: 19)
UAGCUUAUCAGACUGAUGUUGA,
mmu-miR-124a,
(SEQ ID NO: 20)
UAAGGCACGCGGUGAAUGCC,
mmu-miR-19a,
(SEQ ID NO: 21)
UGUGCAAAUCUAUGCAAAACUGA,
mmu-miR-301
(SEQ ID NO: 22)
CAGUGCAAUAGUAUUGUCAAAGC,
mmu-miR-31,
(SEQ ID NO: 23)
AGGCAAGAUGCUGGCAUAGCUG,
mmu-miR-20,
(SEQ ID NO: 24)
UAAAGUGCUUAUAGUGCAGGUAG,
mmu-miR-29c,
(SEQ ID NO: 25)
UAGCACCAUUUGAAAUCGGU,
mmu-miR-148a,
(SEQ ID NO: 26)
UCAGUGCACUACAGAACUUUGU,
mmu-miR-17-5p,
(SEQ ID NO: 27)
CAAAGUGCUUACAGUGCAGGUAGU,
mmu-miR-185,
(SEQ ID NO: 28)
UGGAGAGAAAGGCAGUUC,
mmu-miR-106a,
(SEQ ID NO: 29)
CAAAGUGCUAACAGUGCAGGUA,
mmu-miR-106b,
(SEQ ID NO: 30)
UAAAGUGCUGACAGUGCAGAU,
mmu-miR-375,
(SEQ ID NO: 31)
UUUGUUCGUUCGGCUCGCGUGA,
mmu-miR-216,
(SEQ ID NO: 32)
UAAUCUCAGCUGGCAACUGUG,
mmu-miR-217,
(SEQ ID NO: 33)
UACUGCAUCAGGAACUGACUGGAU,
mmu-miR-200a,
(SEQ ID NO: 34)
UAACACUGUCUGGUAACGAUGU,
mmu-miR-200b,
(SEQ ID NO: 35)
UAAUACUGCCUGGUAAUGAUGAC,
mmu-miR-429,
(SEQ ID NO: 36)
UAAUACUGUCUGGUAAUGCCGU,
mmu-miR-200c,
(SEQ ID NO: 37)
UAAUACUGCCGGGUAAUGAUGG,
mmu-miR-141
(SEQ ID NO: 38)
UAACACUGUCUGGUAAAGAUGG,
mmu-miR-148a,
(SEQ ID NO: 39)
UCAGUGCACUACAGAACUUUGU,
mmu-miR-152,
(SEQ ID NO: 40)
UCAGUGCAUGACAGAACUUGGG,
mmu-miR-182,
(SEQ ID NO: 41)
UUUGGCAAUGGUAGAACUCACA,
mmu-miR-99a,
(SEQ ID NO: 42)
ACCCGUAGAUCCGAUCUUGU,
mmu-miR-130a,
(SEQ ID NO: 43)
CAGUGC AAUGUU AAAAGGGC AU,
mmu-miR-127,
(SEQ ID NO: 44)
UCGGAUCCGUCUGAGCUUGGC,
mmu-miR-100,
(SEQ ID NO: 45)
AACCCGUAGAUCCGAACUUGUG,
mmu-miR-199a,
(SEQ ED NO: 46)
CCCAGUGUUCAGACU ACCUGUUC,
mmu-miR-199b,
(SEQ ED NO: 47)
CCCAGUGUUU AGACU ACCUGUUC,
mmu-miR-125a,
(SEQ ED NO: 48)
UCCCUGAGACCCUUUAACCUGUG,
mmu-miR-22,
(SEQ ID NO: 49)
AAGCUGCCAGUUGAAGAACUGU,
mmu-miR-434-3p,
(SEQ ID NO: 50)
UUUGAACCAUCACUCGACUCC,
mmu-miR-34a,
(SEQ ID NO: 51)
UGGCAGUGUCUUAGCUGGUUGUU,
mmu-miR-181c,
(SEQ ID NO: 52)
AACAUUCAACCUGUCGGUGAGU,
mmu-miR-139,
(SEQ ID NO: 53)
UCUACAGUGCACGUGUCU,
mmu-miR-130a,
(SEQ ID NO: 54)
CAGUGCAAUGUUAAAAGGGCAU,
mmu-miR-322,
(SEQ ID NO: 55)
CAGCAGCAAUUCAUGUUUUGGA,
mmu-miR-181a,
(SEQ ID NO: 56)
AACAUUCAACGCUGUCGGUGAGU,
mmu-miR-125a,
(SEQ ID NO: 57)
UCCCUGAGACCCUUUAACCUGUG,
mmu-miR-200b,
(SEQ ID NO: 58)
UAAUACUGCCUGGUAAUGAUGAC,
mmu-miR-145,
(SEQ ID NO: 59)
GUCCAGUUUUCCCAGGAAUCCCUU,
mmu-miR-127,
(SEQ ID NO: 60)
UCGGAUCCGUCUGAGCUUGGC,
mmu-miR-199a,
(SEQ ID NO: 61)
CCCAGUGUUCAGACUACCUGUUC,
mmu-miR-425,
(SEQ ID NO: 62)
AUCGGGAAUGUCGUGUCCGCC,
mmu-miR-99b,
(SEQ ID NO: 63)
CACCCGU AGAACCGACCUUGCG,
mmu-let-7e,
(SEQ ID NO: 64)
UGAGGUAGGAGGUUGUAUAGU,
mmu-miR-195,
(SEQ ID NO: 65)
UAGCAGCACAGAAAUAUUGGC,
mmu-miR-152,
(SEQ ID NO: 66)
UCAGUGCAUGACAGAACUUGGG,
mmu-miR-125b,
(SEQ ID NO: 67)
UCCCUGAGACCCUAACUUGUGA,
mmu-miR-187,
(SEQ ID NO: 68)
UCGUGUCUUGUGUUGCAGCCGG,
mmu-miR-324-3p,
(SEQ ID NO: 69)
CCACUGCCCCAGGUGCUGCUGG,
mmu-miR-150,
(SEQ ID NO: 70)
UCUCCCAACCCUUGUACCAGUG,
mmu-miR-28,
(SEQ ID NO: 71)
AAGGAGCUCACAGUCUAUUGAG,
and
mmu-miR-143
(SEQ ID NO: 72)
UGAGAUGAAGCACUGUAGCUCA,
in the presence of one or more fluorescent nucleotides to generate one or more labeled cDNAs, wherein the biological sample is obtained from a subject having a transplanted organ,
(ii) contacting the one or more labeled cDNAs with one or more complementary oligonucleotides bound to a microarray, and
(ii) detecting conformational changes in the cDNA by detecting the presence of at least one hybridization product, wherein the at least one hybridization product is detected using a signal from the fluorescent nucleotides.
11. The method of claim 10 , wherein at least 20 miRNAs are amplified to produce at least 20 labeled miRNAs.
12. The method of claim 1 , wherein the at least 20 miRNAs consist essentially of miR-375, miR-216, miR-217, miR-200a, miR-200b, miR-429, miR-200c, miR-141, miR-134, miR-148a, miR-144, miR-466, miR-7, miR-346, miR-468, miR-188, miR-152, miR-298, miR-182, and miR-99a.