Thyroid tumors identified

Abstract

The invention relates to methods and kits for detecting thyroid cancer by detecting differences in the expression of genes that are differentially expressed in thyroid cancer cells.

1 . A method of detecting whether thyroid cancer cells are present in a test tissue or cell sample which comprises
 observing test levels of RNA or protein expression in the test tissue or cell sample for one or more differentially expressed genes;   comparing the test levels of expression to one or more standard or control levels of expression, to ascertain whether higher or lower levels of expression of any of the differentially genes is present in the test tissue or cell sample; and   thereby detecting whether thyroid cancer cells are present in the test tissue or cell sample;   wherein the one or more differentially expressed genes are selected from the group consisting of ANK2, ARHGAP6, C11orf17, CAPN3, CDH16, ChGn, CITED 1, CITED 2, CKB, COL9A3, CSRP2, DAPK2, DIO1, DPP4, DTX4, DUSP4, EFEMP1, ELMO1, FGFR2, FLRT1, FMOD, GALNT7, GATM, HGD, HMGA2, IGFBP6, KIT, LRP4, MATN2, MET, MYH10, PFAAP5, PGF, PIPS-E, PKNOX2, PRKACB, PROS1, PSD3, QPCT, RAB27A, RAB27A, RXRG, SDC4, SERPINA1, SLC25A15, SLC4A4, SLIT1, SPTAN1, TFCP2L1, TIAM1, TIMP1, TNS3, TSPAN12, UPP1, and a combination thereof.   
     
     
         2 . The method of  claim 1 , wherein the one or more differentially expressed genes are selected from the group consisting of DIO1, DTX4, GALNT7, HMGA2, IGFBP6, MET, PROS1, SDC4, SERPINA1, SLC4A4, TIAM1, TIMP1, UPP1 and a combination thereof. 
     
     
         3 . The method of  claim 1 , wherein the one or more differentially expressed genes are selected from the group consisting of ANK2, ARHGAP6, CDH16, CITED 1, CITED 2, COL9A3, ChGn, DUSP4, EFEMP1, ELMO1, FGFR2, FLRT1, FMOD, GATM, KIT, LRP4, MATN2, SLIT1, SPTAN1, TFCP2L1, PIPS-E, PSD3, TNS3, TSPAN12, TIAM1 and a combination thereof. 
     
     
         4 . The method of  claim 1 , wherein the one or more differentially expressed genes are selected from the group consisting of C11orf17, CAPN3, CAPN3, CKB, CSRP2, DAPK2, DPP4, HGD, MYH10, PFAAP5, PGF, PKNOX2, PRKACB, QPCT, RAB27A, RXRG, and SLC25A15 and a combination thereof. 
     
     
         5 . The method of  claim 1 , wherein the test levels of RNA expressed are detected by microarray analysis or by nucleic acid amplification. 
     
     
         6 . The method of  claim 5 , wherein the test levels of RNA expressed are detected by microarray analysis comprising use of one or more probes on the microarray that can hybridize to one or more of the differentially expressed genes, or an RNA or DNA copy of the one or more differentially expressed genes. 
     
     
         7 . The method of  claim 6 , wherein the one or more probes hybridize to any of SEQ ID NO:119-172. 
     
     
         8 . The method of  claim 6 , wherein the one or more probes hybridize to one or more of the differentially expressed genes, or an RNA or DNA copy of the one or more differentially expressed genes under moderate to highly stringent hybridization conditions. 
     
     
         9 . The method of  claim 8 , wherein the hybridization conditions are highly stringent hybridization conditions. 
     
     
         10 . The method of  claim 5 , wherein the nucleic acid amplification comprises reverse transcription polymerase chain reaction, real time polymerase chain reaction, and/or quantitative polymerase chain reaction. 
     
     
         11 . The method of  claim 5 , wherein the test levels of RNA expressed are detected by nucleic acid amplification using one or more primers that hybridize to one or more of the differentially expressed genes, or an RNA or DNA copy of the one or more differentially expressed genes under moderate to highly stringent hybridization conditions. 
     
     
         12 . The method of  claim 11 , wherein the hybridization conditions are highly stringent hybridization conditions. 
     
     
         13 . The method of  claim 11 , wherein the one or more primers hybridize to any of SEQ ID NO:119-172. 
     
     
         14 . The method of  claim 11 , wherein the one or primers are selected from the group consisting of SEQ ID NO:3-118. 
     
     
         15 . The method of  claim 1 , wherein the one or more standard or control levels of expression comprise:
 an expression level observed for a malignant thyroid cancer cell or tissue;   an expression level observed for a benign thyroid cell or tissue;   an expression level observed for a follicular adenoma with nuclear atypia;   an expression level observed for a borderline thyroid cell or tissue;   an expression level observed for a normal non-cancerous thyroid cell or tissue; or   an expression level observed for a constitutively expressed gene.   
     
     
         16 . The method of  claim 1 , wherein the method can distinguish between benign, malignant and borderline thyroid cells or tissues. 
     
     
         17 . The method of  claim 1 , wherein the method can distinguish between benign thyroid cells or tissues, malignant thyroid cells or tissues, and follicular adenomas with nuclear atypia (FANA). 
     
     
         18 . The method of  claim 1 , wherein the test tissue or cell sample is obtained from a patient with thyroid cancer or suspected of having thyroid cancer. 
     
     
         19 . A kit comprising:
 (a) at least one set of oligonucleotide primers, wherein a first primer in the set contains a sequence complementary to a region in one strand of a nucleic acid sequence template and primes the synthesis of a first extension product, and a second primer contains a sequence complementary to a region in said first extension product and primes the synthesis of a nucleic acid strand complementary to said first extension product, and wherein the template is a differentially expressed gene, or an RNA or DNA copy of the differentially expressed gene; and   (b) instructions for using the at least one set of oligonucleotide primers;   wherein differentially expressed gene is selected from the group consisting of ANK2, ARHGAP6, C11orf17, CAPN3, CDH16, ChGn, CITED 1, CITED 2, CKB, COL9A3, CSRP2, DAPK2, DIOL, DPP4, DPP4, DTX4, DUSP4, EFEMP1, ELMO1, FGFR2, FLRT1, FMOD, GALNT7, GATM, HGD, HMGA2, IGFBP6, KIT, LRP4, MATN2, MET, MYH10, PFAAP5, PGF, PIPS-E, PKNOX2, PRKACB, PROS1, PSD3, PSD3, QPCT, RAB27A, RAB27A, RXRG, SDC4, SERPINA1, SERPINA1, SLC25A15, SLC4A4, SLIT1, SPTAN1, TFCP2L1, TIAM1, TIMP1, TNS3, TSPAN12, UPP1, and a combination thereof.   
     
     
         20 . The kit of  claim 19 , wherein the differentially expressed gene is selected from the group consisting of DIO1, DTX4, GALNT7, HMGA2, IGFBP6, MET, PROS1, SDC4, SERPINA1, SLC4A4, TIAM1, TIMP1, UPP1 and a combination thereof. 
     
     
         21 . The kit of  claim 19 , wherein the differentially expressed gene is selected from the group consisting of ANK2, ARHGAP6, CDH16, CITED 1, CITED 2, COL9A3, ChGn, DUSP4, EFEMP1, ELMO1, FGFR2, FLRT1, FMOD, GATM, KIT, LRP4, MATN2, SLIT1, SPTAN1, TFCP2L1, PIPS-E, PSD3, TNS3, TSPAN12, TIAM1 and a combination thereof. 
     
     
         22 . The kit of  claim 19 , wherein the differentially expressed gene is selected from the group consisting of C11orf17, CAPN3, CAPN3, CKB, CSRP2, DAPK2, DPP4, HGD, MYH10, PFAAP5, PGF, PKNOX2, PRKACB, QPCT, RAB27A, RXRG, and SLC25A15 and a combination thereof. 
     
     
         23 . The kit of  claim 19 , wherein the first primer and/or the second primer comprise a label. 
     
     
         24 . The kit of  claim 19 , comprising more than one set of primers. 
     
     
         25 . The kit of  claim 19 , wherein the first primer and/or the second primer hybridize to any of SEQ ID NO:119-172. 
     
     
         26 . The kit of  claim 19 , wherein the first primer and/or the second primer are selected from the group consisting of SEQ ID NO:3-118. 
     
     
         27 . The kit of  claim 19 , further comprising a container of nucleotides for use as subunits in the synthesis of and amplified product. 
     
     
         28 . The kit of  claim 27 , wherein one or more nucleotides further comprises a label. 
     
     
         29 . The kit of  claim 27 , wherein the nucleotides are deoxyribonucleotides. 
     
     
         30 . The kit of  claim 19 , wherein the instructions describe a method for amplifying an mRNA, cRNA or cDNA corresponding to the differentially expressed gene(s). 
     
     
         31 . The kit of  claim 19 , wherein the first primer and/or the second primer hybridize to an mRNA, cRNA or cDNA corresponding to the differentially expressed gene under moderate to highly stringent hybridization conditions. 
     
     
         32 . The kit of  claim 31 , wherein the hybridization conditions are highly stringent hybridization conditions. 
     
     
         33 . A kit comprising (a) a microarray with covalently attached probes that can hybridize to a differentially expressed gene selected from the group consisting of ANK2, ARHGAP6, C11orf17, CAPN3, CDH16, ChGn, CITED 1, CITED 2, CKB, COL9A3, CSRP2, DAPK2, DIO1, DPP4, DPP4, DTX4, DUSP4, EFEMP1, ELMO1, FGFR2, FLRT1, FMOD, GALNT7, GATM, HGD, HMGA2, IGFBP6, KIT, LRP4, MATN2, MET, MYH10, PFAAP5, PGF, PIPS-E, PKNOX2, PRKACB, PROS1, PSD3, PSD3, QPCT, RAB27A, RAB27A, RXRG, SDC4, SERPINA1, SERPINA1, SLC25A15, SLC4A4, SLIT1, SPTAN1, TFCP2L1, TIAM1, TIMP1, TNS3, TSPAN12, UPP1, and a combination thereof; and (b) instructions for using the microarray. 
     
     
         34 . The kit of  claim 33 , wherein the differentially expressed gene is selected from the group consisting of DIOL, DTX4, GALNT7, HMGA2, IGFBP6, MET, PROS1, SDC4, SERPINA1, SLC4A4, TIAM1, TIMP1, UPP1 and a combination thereof. 
     
     
         35 . The kit of  claim 33 , wherein the differentially expressed gene is selected from the group consisting of ANK2, ARHGAP6, CDH16, CITED 1, CITED 2, COL9A3, ChGn, DUSP4, EFEMP1, ELMO1, FGFR2, FLRT1, FMOD, GATM, KIT, LRP4, MATN2, SLIT1, SPTAN1, TFCP2L1, PIP3-E, PSD3, TNS3, TSPAN12, TIAM1 and a combination thereof. 
     
     
         36 . The kit of  claim 33 , wherein the differentially expressed gene is selected from the group consisting of C11orf17, CAPN3, CAPN3, CKB, CSRP2, DAPK2, DPP4, HGD, MYH10, PFAAP5, PGF, PKNOX2, PRKACB, QPCT, RAB27A, RXRG, and SLC25A15 and a combination thereof. 
     
     
         37 . The kit of  claim 33 , wherein the probes hybridize to an mRNA, cRNA or cDNA corresponding to the differentially expressed gene under moderate to highly stringent hybridization conditions. 
     
     
         38 . The kit of  claim 33 , wherein the hybridization conditions are highly stringent hybridization conditions. 
     
     
         39 . The kit of  claim 33 , wherein the probes hybridize to any of SEQ ID NO:119-172. 
     
     
         40 . The kit of  claim 33 , further comprising one or more standard or control probes. 
     
     
         41 . The kit of  claim 33 , where the one or more standard or control probes comprise a probe for a constitutively expressed gene. 
     
     
         42 . A method of detecting a mutation in a human BRAF gene comprising: (a) obtaining a test sample of genomic DNA from a human; (b) amplifying a segment of BRAF DNA from the genomic DNA using primers with SEQ ID NO: 1 and SEQ ID NO:2; and (c) detecting whether the mutation exists in the segment amplified; wherein the mutation consists of a glutamate substituted for valine at codon 600. 
     
     
         43 . The method of  claim 42 , further comprising detecting whether the human has thyroid cancer by observing test levels of RNA or protein expression in the test tissue or cell sample for any differentially expressed gene, and
 comparing the test levels of expression to one or more standard or control levels of expression, to ascertain whether higher or lower levels of expression of any of the genes is present in the test tissue or cell sample, and   thereby detecting whether thyroid cancer cells are present in the test tissue or cell sample;   wherein the differentially expressed gene is selected from the group consisting of ANK2, ARHGAP6, C11orf17, CAPN3, CDH16, ChGn, CITED 1, CITED 2, CKB, COL9A3, CSRP2, DAPK2, DIO1, DPP4, DPP4, DTX4, DUSP4, EFEMP1, ELMO1, FGFR2, FLRT1, FMOD, GALNT7, GATM, HGD, HMGA2, IGFBP6, KIT, LRP4, MATN2, MET, MYH10, PFAAP5, PGF, PIPS-E, PKNOX2, PRKACB, PROS1, PSD3, PSD3, QPCT, RAB27A, RAB27A, RXRG, SDC4, SERPINA1, SERPINA1, SLC25A15, SLC4A4, SLIT1, SPTAN1, TFCP2L1, TIAM1, TIMP1, TNS3, TSPAN12, UPP1, and a combination thereof.