GENE EXPRESSION SIGNATURE FOR CLASSIFICATION OF KIDNEY TUMORS
The present invention provides a method for classification of kidney tumors through the analysis of the expression patterns of specific microRNAs and nucleic acid molecules relating thereto. Classification according to a microRNA expression framework allows optimization of treatment, and determination of specific therapy.
1 . A method for the detection of a specific subtype of kidney tumor, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining an expression profile in said sample of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-75, or a sequence having at least about 80% identity thereto; and
(c) comparing said expression profile to a reference value;
whereby an altered expression levels of the nucleic acid sequence allows the detection of the specific subtype of kidney tumor in said sample.
2 . The method of claim 1 , wherein said specific subtype is selected from the group consisting of oncocytoma, clear cell RCC (conventional), papillary (chromaphil) RCC and chromophobe RCC.
3 . The method of claim 1 , wherein said altered expression level is a change in a score based on a combination of expression levels of said nucleic acid sequences.
4 . A method for distinguishing between benign and malignant kidney tumor, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining in said sample an expression profile of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-27, 47, 49, a fragment thereof or a sequence having at least 80% identity thereto; and
(c) comparing said expression profile to a reference value;
whereby a relative abundance of said nucleic acid sequences allows the detection of said kidney tumor.
5 . The method of claim 4 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 12-15, 18, 19, 25-27 is indicative of the presence of benign kidney tumor.
6 . The method of claim 4 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-11, 16, 17, 20-24, 47, 49 is indicative of the presence of malignant kidney tumor.
7 . A method for distinguishing between chromophobe RCC and oncocytoma, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining in said sample an expression profile of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 14-15, 28-35, a fragment thereof or a sequence having at least 80% identity thereto; and
(c) comparing said expression profile to a reference value; whereby a relative abundance of said nucleic acid sequences allows the detection of said oncocytoma or chromophobe RCC.
8 . The method of claim 7 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 14-15, 28-31, is indicative of the presence of oncocytoma.
9 . The method of claim 7 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 32-35 is indicative of the presence of chromophobe RCC.
10 . A method for distinguishing between clear cell RCC and oncocytoma, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining in said sample an expression profile of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-3, 6, 7, 11, 13, 16, 17, 19-27, 30, 31, 36-40, 47-49, a fragment thereof or a sequence having at least 80% identity thereto; and
(c) comparing said expression profile to a reference value;
whereby a relative abundance of said nucleic acid sequences allows the detection of said oncocytoma or clear cell RCC.
11 . The method of claim 10 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NO: 13, 19, 25-27, 30, 31, 36-40, 48 is indicative of the presence of oncocytoma.
12 . The method of claim 10 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-3, 6, 7, 11, 16, 17, 20-24, 47, 49 is indicative of the presence of clear cell RCC.
13 . A method for distinguishing between chromophobe RCC and clear cell RCC, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining in said sample an expression profile of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-3, 6, 7, 22, 23, 32-37, 43, 44, a fragment thereof or a sequence having at least 80% identity thereto; and
(c) comparing said expression profile to a reference value;
whereby a relative abundance of said nucleic acid sequences allows the detection of said RCC.
14 . The method of claim 13 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-3, 6, 7, 22, 23 is indicative of the presence of clear cell RCC.
15 . The method of claim 13 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 32-37, 43, 44 is indicative of the presence of chromophobe RCC.
16 . A method for distinguishing between clear cell RCC and chromaphil RCC, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining in said sample an expression profile of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 4, 5, 8, 9, 30, 31, 41, 42, 50, 51, a fragment thereof or a sequence having at least 80% identity thereto; and
(c) comparing said expression profile to a reference value;
whereby a relative abundance of said nucleic acid sequences allows the detection of said RCC.
17 . The method of claim 16 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 41, 42 is indicative of the presence of clear cell RCC.
18 . The method of claim 16 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 4, 5, 8, 9, 30, 31, 50, 51, is indicative of the presence of papillary (chromaphil) RCC.
19 . A method for distinguishing between chromophobe RCC and papillary (chromaphil) RCC, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining in said sample an expression profile of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-5, 8-11, 13, 24-27, 30-35, 43, 44, 47, 49, 52, 53, a fragment thereof or a sequence having at least 80% identity thereto; and
(c) comparing said expression profile to a reference value;
whereby a relative abundance of said nucleic acid sequences allows the detection of said RCC.
20 . The method of claim 19 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 13, 25-27, 32-35, 43, 44, 52, 53, is indicative of the presence of chromophobe RCC.
21 . The method of claim 19 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 1-5, 8-11, 24, 30, 31, 47, 49 is indicative of the presence of papillary RCC.
22 . A method for distinguishing between papillary RCC and oncocytoma, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining in said sample an expression profile of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 4, 5, 8-13, 25-27, 41-42, 45-47, a fragment thereof or a sequence having at least 80% identity thereto; and
(c) comparing said expression profile to a reference value;
whereby a relative abundance of said nucleic acid sequences allows the detection of said papillary RCC or oncocytoma.
23 . The method of claim 22 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 4, 5, 8-11, 45-47 is indicative of the presence of papillary RCC.
24 . The method of claim 22 , wherein a relative abundance of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 12, 13, 25-27, 41-42 is indicative of the presence of oncocytoma.
25 . A method for distinguishing between chromophobe RCC, clear cell RCC, papillary RCC and oncocytoma, the method comprising:
(a) obtaining a biological sample from a subject;
(b) determining in said sample an expression profile of nucleic acid sequences selected from the group consisting of SEQ ID NOS: 4, 5, 13-15, 20, 21, 25, 34, 35, 41, 42; a fragment thereof and a sequence having at least 80% identity thereto; and
(c) comparing said expression profile to a reference value;
whereby a relative abundance of said nucleic acid sequences allows the detection of said RCC.
26 . The method of claim 1 , wherein said biological sample is selected from the group consisting of bodily fluid, a cell line and a tissue sample.
27 . The method of claim 26 , wherein said tissue is a fresh, frozen, fixed, wax-embedded or formalin fixed paraffin-embedded (FFPE) tissue.
28 . The method of claim 26 , wherein said tissue sample is a kidney tumor sample.
29 . The method of claim 1 , wherein the expression levels are determined by a method selected from the group consisting of nucleic acid hybridization, nucleic acid amplification, and a combination thereof.
30 . The method of claim 29 , wherein the nucleic acid hybridization is performed using a solid-phase nucleic acid biochip array.
31 . The method of claim 29 , wherein the nucleic acid hybridization is performed using in situ hybridization.
32 . The method of claim 31 , wherein the in situ hybridization method comprises hybridization with a probe.
33 . The method of claim 32 , wherein the probe comprises a nucleic acid sequence that is complementary to a sequence selected from the group consisting of SEQ ID NOS: 1-75 and sequences at least about 80% identical thereto.
34 . The method of claim 29 , wherein the nucleic acid amplification method is real-time PCR(RT-PCR).
35 . The method of claim 34 , wherein the RT-PCR method comprises forward and reverse primers.
36 . The method of claim 35 , wherein the forward primer comprises a sequence selected from the group consisting of SEQ 11) NOS: 82-87, a fragment thereof and sequences at least about 80% identical thereto.
37 . The method of claim 35 , wherein the reverse primer comprises SEQ ID NO: 90, a fragment thereof and sequences at least about 80% identical thereto.
38 . The method of claim 35 , wherein the RT-PCR method further comprises hybridization with a probe.
39 . The method of claim 38 , wherein the probe comprises a nucleic acid sequence that is complementary to a sequence selected from the group consisting of SEQ ID NOS: 1-75, a fragment thereof and sequences at least about 80% identical thereto.
40 . The method of claim 39 , wherein the probe comprises a nucleic acid sequence selected from the group consisting of SEQ ID NOS: 76-81, a fragment thereof and sequences at least about 80% identical thereto.
41 . A kit for renal tumor classification, said kit comprises a probe comprising a nucleic acid sequence that is complementary to a sequence selected from the group consisting of SEQ ID NOS: 1-75, a fragment thereof and sequences having at least about 80% identity thereto.
42 . The kit of claim 41 , wherein the probe comprising a nucleic acid sequence that is complementary to a sequence selected from the group consisting of
SEQ ID NOS: 76-81, a fragment thereof and sequences at least about 80% identical thereto.
43 . The kit of claim 41 , further comprises forward and reverse primers.
44 . The kit of claim 43 , wherein the forward primer comprising a sequence selected from the group consisting of SEQ ID NOS: 82-87, a fragment thereof and sequences having at least about 80% identity thereto.
45 . The kit of claim 43 , wherein the reverse primer comprises SEQ ID NO: 90, a fragment thereof and sequences having at least about 80% identity thereto.
46 . The kit of claim 41 , wherein the kit comprises reagents for performing in situ hybridization analysis.