Method, array and use for determining the presence of pancreatic cancer

Abstract

The present invention relates to a method for determining the presence of pancreatic cancer in an individual comprising or consisting of the steps of: (a) providing a sample to be tested from the individual, and (b) determining a biomarker signature of the test sample by measuring the expression in the test sample of one or more biomarkers selected from the group defined in Table III, wherein the expression in the test sample of one or more biomarkers selected from the group defined in Table III is indicative of the individual having pancreatic cancer. The invention also comprises arrays and kits of parts for use in the method of the invention.

1 . A method for determining the presence of pancreatic cancer in an individual comprising or consisting of the steps of:
 a) providing a sample to be tested from the individual;   b) determining a biomarker signature of the test sample by measuring the expression in the test sample of one or more biomarkers selected from the group defined in Table III;   wherein the expression in the test sample of one or more biomarkers selected from the group defined in Table III is indicative of the individual having pancreatic cancer; and   wherein step (b) comprises or consists of measuring the expression of one or more of the biomarkers listed in Table IV(A) and/or Table IV(B).   
     
     
         2 . The method according to  claim 1  further comprising or consisting of the steps of:
 c) providing a control sample from an individual not afflicted with pancreatic cancer; 
 d) determining a biomarker signature of the control sample by measuring the expression in the control sample of the one or more biomarkers measured in step (b); 
 wherein the presence of pancreatic cancer is identified in the event that the expression in the test sample of the one or more biomarkers measured in step (b) is different from the expression in the control sample of the one or more biomarkers measured in step (d). 
 
     
     
         3 . The method according to  claim 1  or  2  further comprising or consisting of the steps of:
 e) providing a control sample from an individual afflicted with pancreatic cancer; 
 f) determining a biomarker signature of the control sample by measuring the expression in the control sample of the one or more biomarkers measured in step (b); 
 wherein the presence of pancreatic cancer is identified in the event that the expression in the test sample of the one or more biomarkers measured in step (b) corresponds to the expression in the control sample of the one or more biomarkers measured in step (f). 
 
     
     
         4 . The method according to  claim 1 ,  2  or  3 , wherein step (b) comprises or consists of measuring the expression of one or more of the biomarkers listed in Table IV(A), for example, at least 2 of the biomarkers listed in Table IV(A). 
     
     
         5 . The method according to any one of the preceding claims, wherein step (b) comprises or consists of measuring the expression of interleukin-7 (IL-7) and/or integrin alpha-10, for example, measuring the expression of interleukin-7, measuring the expression of integrin alpha-10, or measuring the expression of interleukin-7 and integrin alpha-10. 
     
     
         6 . The method according to any one of the preceding claims, wherein step (b) comprises or consists of measuring the expression of each the biomarkers listed in Table IV(A). 
     
     
         7 . The method according to any one of the preceding claims, wherein step (b) comprises or consists of measuring the expression of 1 or more of the biomarkers listed in Table IV(B), for example at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 of the biomarkers listed in Table IV(B). 
     
     
         8 . The method according to any one of the preceding claims, wherein step (b) comprises or consists of measuring the expression of all of the biomarkers listed in Table IV(B). 
     
     
         9 . The method according to any one of the preceding claims wherein step (b) comprises or consists of measuring the expression of 1 or more biomarkers from the biomarkers listed in Table IV(C), for example at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40 or 41 of the biomarkers listed in Table IV(C). 
     
     
         10 . The method according to any one of the preceding claims, wherein step (b) comprises or consists of measuring the expression of all of the biomarkers listed in Table IV(C). 
     
     
         11 . The method according to any one of the preceding claims wherein step (b) comprises or consists of measuring the expression in the test sample of all of the biomarkers defined in Table IV. 
     
     
         12 . The method according to any one of the preceding claims, wherein the method is for differentiating between pancreatic cancer (PaC) and any other disease state or states. 
     
     
         13 . The method according to  claim 12 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(A), for example at least 2, 3, 4, 5, 6, 7, 8, 9 or 10 of the biomarkers listed in Table V(A). 
     
     
         14 . The method according to  claim 12  or  13 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(B), for example at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 of the biomarkers listed in Table V(B). 
     
     
         15 . The method according to  claim 12 ,  13  or  14 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(C), for example at least 2, 3, 4 or 5 of the biomarkers listed in Table V(C). 
     
     
         16 . The method according to any one of  claims 12  to  15 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(D), for example at least 2 or 3 of the biomarkers listed in Table V(D). 
     
     
         17 . The method according to any one of  claims 12  to  16 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(F), for example at least 2, 3, 4, 5 or 6 of the biomarkers listed in Table V(F). 
     
     
         18 . The method according to any one of  claims 12  to  17 , wherein step (b) comprises or consists of measuring the expression in the test sample of all of the biomarkers listed in Table V(A), Table V(B), Table V(C), Table V(D) and/or Table V(F). 
     
     
         19 . The method according to any one of  claims 12  to  18 , wherein the other disease state or states is chronic pancreatitis (ChP), acute inflammatory pancreatitis (AIP) and/or normal, for example, the other disease state or states may be chronic pancreatitis alone; acute inflammatory pancreatitis alone; chronic pancreatitis and acute inflammatory pancreatitis; chronic pancreatitis and normal; acute inflammatory pancreatitis and normal; or, chronic pancreatitis, acute inflammatory pancreatitis and normal. 
     
     
         20 . The method according to any one of  claims 1  to  11 , wherein the method is for differentiating between pancreatic cancer and chronic pancreatitis (ChP). 
     
     
         21 . The method according to  claim 20 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(A), for example at least 2, 3, 4, 5, 6, 7, 8, 9 or 10 of the biomarkers listed in Table V(A). 
     
     
         22 . The method according to  claim 20  or  21 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(C), for example at least 2, 3, 4, or 5 of the biomarkers listed in Table V(C). 
     
     
         23 . The method according to  claim 20 ,  21 , or  22 , wherein step (b) comprises or consists of measuring the expression in the test sample of all of the biomarkers listed in Table V(A) and/or Table V(C). 
     
     
         24 . The method according to any one of  claims 1  to  11 , wherein the method is for differentiating between pancreatic cancer and acute inflammatory pancreatitis (AIP). 
     
     
         25 . The method according to  claim 24 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(A), for example at least 2, 3, 4, 5, 6, 7, 8, 9 or 10 of the biomarkers listed in Table V(A). 
     
     
         26 . The method according to  claim 24  or  25 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(B), for example at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 of the biomarkers listed in Table V(B). 
     
     
         27 . The method according to  claim 24 ,  25  or  26 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(C), for example at least 2, 3, 4 or 5 of the biomarkers listed in Table V(C). 
     
     
         28 . The method according to any one of  claims 24  to  27 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(E). 
     
     
         29 . The method according to any one of  claims 24  to  28 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(F), for example at least 2, 3, 4, 5 or 6 of the biomarkers listed in Table V(F). 
     
     
         30 . The method according to any one of  claims 24  to  29 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(H), for example at least 2 or 3 of the biomarkers listed in Table V(H). 
     
     
         31 . The method according to any one of  claims 24  to  30 , wherein step (b) comprises or consists of or consists of measuring the expression in the test sample of all of the biomarkers listed in Table V(A), Table V(B), Table V(C), Table V(E), Table V(F) and/or Table IV(H). 
     
     
         32 . The method according to any one of  claims 1  to  11 , wherein the method is for differentiating between pancreatic cancer and normal individuals (N). 
     
     
         33 . The method according to  claim 32 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(A), for example at least 2, 3, 4, 5, 6, 7, 8, 9 or 10 of the biomarkers listed in Table V(A). 
     
     
         34 . The method according to  claim 32  or  33 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(B), for example at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24 of the biomarkers listed in Table V(B). 
     
     
         35 . The method according to  claim 32 ,  33  or  34 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(D), for example at least 2 or 3 of the biomarkers listed in Table V(D). 
     
     
         36 . The method according to any one of  claims 32  to  35 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(E). 
     
     
         37 . The method according to any one of  claims 32  to  36 , wherein step (b) comprises or consists of measuring the expression in the test sample of 1 or more biomarkers from the biomarkers listed in Table V(G), for example at least 2 or 3 of the biomarkers listed in Table V(G). 
     
     
         38 . The method according to any one of  claims 32  to  37 , wherein step (b) comprises or consists of measuring the expression in the test sample of all of the biomarkers listed in Table V(A), Table V(B), Table V(D), Table V(E) and/or Table IV(G). 
     
     
         39 . The method according to any one of  claims 2  to  38 , wherein the individual not afflicted with pancreatic cancer is not afflicted with pancreatic cancer (PaC), chronic pancreatitis (ChP) or acute inflammatory pancreatitis (AIP). 
     
     
         40 . The method according to  claim 39 , wherein the individual not afflicted with pancreatic cancer is not afflicted with any pancreatic disease or condition. 
     
     
         41 . The method according to  claim 39  or  40 , wherein the individual not afflicted with pancreatic cancer is not afflicted with any disease or condition. 
     
     
         42 . The method according to  claim 39 ,  40  or  41 , wherein the individual not afflicted with pancreatic cancer is a healthy individual. 
     
     
         43 . The method according to any one of  claims 2  to  38 , wherein the individual not afflicted with pancreatic cancer is afflicted with chronic pancreatitis. 
     
     
         44 . The method according to any one of  claims 2  to  38 , wherein the individual not afflicted with pancreatic cancer is afflicted with acute inflammatory pancreatitis. 
     
     
         45 . The method according to any one of the preceding claims wherein the pancreatic cancer is selected from the group consisting of adenocarcinoma, adenosquamous carcinoma, signet ring cell carcinoma, hepatoid carcinoma, colloid carcinoma, undifferentiated carcinoma, and undifferentiated carcinomas with osteoclast-like giant cells. 
     
     
         46 . The method according to any one of the preceding claims wherein the pancreatic cancer is an adenocarcinoma. 
     
     
         47 . The method according to any one of the preceding claims wherein step (b), (d) and/or step (f) is performed using a first binding agent capable of binding to the one or more biomarkers. 
     
     
         48 . The method according to  claim 47  wherein the first binding agent comprises or consists of an antibody or an antigen-binding fragment thereof. 
     
     
         49 . The method according to  claim 48  wherein the antibody or antigen-binding fragment thereof is a recombinant antibody or antigen-binding fragment thereof. 
     
     
         50 . The method according to  claim 48  or  49  wherein the antibody or antigen-binding fragment thereof is selected from the group consisting of: scFv; Fab; a binding domain of an immunoglobulin molecule. 
     
     
         51 . The method according to any one of  claims 48  to  50  wherein the first binding agent is immobilised on a surface. 
     
     
         52 . The method according to any one of  claims 1  to  25  wherein the one or more biomarkers in the test sample are labelled with a detectable moiety. 
     
     
         53 . The method according to any one of  claims 2  to  25  wherein the one or more biomarkers in the control sample(s) are labelled with a detectable moiety. 
     
     
         54 . The method according to  claim 52  or  53  wherein the detectable moiety is selected from the group consisting of: a fluorescent moiety; a luminescent moiety; a chemiluminescent moiety; a radioactive moiety; an enzymatic moiety. 
     
     
         55 . The method according to  claim 52  or  53  wherein the detectable moiety is biotin. 
     
     
         56 . The method according to any one of  claims 47  to  55  wherein step (b), (d) and/or step (f) is performed using an assay comprising a second binding agent capable of binding to the one or more biomarkers, the second binding agent comprising a detectable moiety. 
     
     
         57 . The method according to any one of  claim 56  wherein the second binding agent comprises or consists of an antibody or an antigen-binding fragment thereof. 
     
     
         58 . The method according to  claim 57  wherein the antibody or antigen-binding fragment thereof is a recombinant antibody or antigen-binding fragment thereof. 
     
     
         59 . The method according to  claim 57  or  58  wherein the antibody or antigen-binding fragment thereof is selected from the group consisting of: scFv; Fab; a binding domain of an immunoglobulin molecule. 
     
     
         60 . The method according to any one of  claims 56  to  59  wherein the detectable moiety is selected from the group consisting of: a fluorescent moiety; a luminescent moiety; a chemiluminescent moiety; a radioactive moiety; an enzymatic moiety. 
     
     
         61 . The method according to  claim 60  wherein the detectable moiety is fluorescent moiety (for example an Alexa Fluor dye, e.g. Alexa647). 
     
     
         62 . The method according to any one of the preceding claims wherein the method comprises or consists of an ELISA (Enzyme Linked Immunosorbent Assay). 
     
     
         63 . The method according to any one of the preceding claims wherein step (b), (d) and/or step (f) is performed using an array. 
     
     
         64 . The method according to  claim 63  wherein the array is a bead-based array. 
     
     
         65 . The method according to  claim 63  wherein the array is a surface-based array. 
     
     
         66 . The method according to any one of  claims 63  to  65  wherein the array is selected from the group consisting of: macroarray; microarray; nanoarray. 
     
     
         67 . The method according to any one of the preceding claims wherein the method comprises:
 (v) labelling biomarkers present in the sample with biotin;   (vi) contacting the biotin-labelled proteins with an array comprising a plurality of scFv immobilised at discrete locations on its surface, the scFv having specificity for one or more of the proteins in Table III;   (vii) contacting the immobilised scFv with a streptavidin conjugate comprising a fluorescent dye; and   (viii) detecting the presence of the dye at discrete locations on the array surface wherein the expression of the dye on the array surface is indicative of the expression of a biomarker from Table III in the sample.   
     
     
         68 . The method according to any one of claims wherein, step (b), (d) and/or (f) comprises measuring the expression of a nucleic acid molecule encoding the one or more biomarkers. 
     
     
         69 . The method according to  claim 68 , wherein the nucleic acid molecule is a cDNA molecule or an mRNA molecule. 
     
     
         70 . The method according to  claim 68 , wherein the nucleic acid molecule is an mRNA molecule. 
     
     
         71 . The method according to  claim 68 ,  69  or  70 , wherein measuring the expression of the one or more biomarker(s) in step (b), (d) and/or (f) is performed using a method selected from the group consisting of Southern hybridisation, Northern hybridisation, polymerase chain reaction (PCR), reverse transcriptase PCR (RT-PCR), quantitative real-time PCR (qRT-PCR), nanoarray, microarray, macroarray, autoradiography and in situ hybridisation. 
     
     
         72 . The method according to any one of  claims 68 - 71 , wherein measuring the expression of the one or more biomarker(s) in step (b) is determined using a DNA microarray. 
     
     
         73 . The method according to any one of  claims 68  to  72 , wherein measuring the expression of the one or more biomarker(s) in step (b), (d) and/or (f) is performed using one or more binding moieties, each individually capable of binding selectively to a nucleic acid molecule encoding one of the biomarkers identified in Table III. 
     
     
         74 . The method according to  claim 73 , wherein the one or more binding moieties each comprise or consist of a nucleic acid molecule. 
     
     
         75 . The method according to  claim 74  wherein, the one or more binding moieties each comprise or consist of DNA, RNA, PNA, LNA, GNA, TNA or PMO. 
     
     
         76 . The method according to  claim 74  or  75 , wherein the one or more binding moieties each comprise or consist of DNA. 
     
     
         77 . The method according to any one of  claims 74 - 76  wherein the one or more binding moieties are 5 to 100 nucleotides in length. 
     
     
         78 . The method according to any one of  claims 74 - 76  wherein the one or more nucleic acid molecules are 15 to 35 nucleotides in length. 
     
     
         79 . The method according to any one of  claims 74 - 78  wherein the binding moiety comprises a detectable moiety. 
     
     
         80 . The method according to  claim 79  wherein the detectable moiety is selected from the group consisting of: a fluorescent moiety; a luminescent moiety; a chemiluminescent moiety; a radioactive moiety (for example, a radioactive atom); or an enzymatic moiety. 
     
     
         81 . The method according to  claim 80  wherein the detectable moiety comprises or consists of a radioactive atom. 
     
     
         82 . The method according to  claim 81  wherein the radioactive atom is selected from the group consisting of technetium-99m, iodine-123, iodine-125, iodine-131, indium-111, fluorine-19, carbon-13, nitrogen-15, oxygen-17, phosphorus-32, sulphur-35, deuterium, tritium, rhenium-186, rhenium-188 and yttrium-90. 
     
     
         83 . The method according to  claim 80  wherein the detectable moiety of the binding moiety is a fluorescent moiety. 
     
     
         84 . The method according to any one of the preceding claims wherein, the sample provided in step (b), (d) and/or (f) is selected from the group consisting of unfractionated blood, plasma, serum, tissue fluid, pancreatic tissue, pancreatic juice, bile and urine. 
     
     
         85 . The method according to  claim 84 , wherein the sample provided in step (b), (d) and/or (f) is selected from the group consisting of unfractionated blood, plasma and serum. 
     
     
         86 . The method according to  claim 84  or  85 , wherein the sample provided in step (b), (d) and/or (f) is plasma. 
     
     
         87 . An array for determining the presence of pancreatic cancer in an individual comprising one or more binding agent as defined in any one of  claims 47  to  61 . 
     
     
         88 . An array according to  claim 87  wherein the one or more binding agents is capable of binding to all of the proteins defined in Table III. 
     
     
         89 . Use of one or more biomarkers selected from the group defined in Table III as a diagnostic marker for determining the presence of pancreatic cancer in an individual. 
     
     
         90 . The use according to  claim 89  wherein all of the proteins defined in Table III are used as a diagnostic marker for determining the presence of pancreatic cancer in an individual. 
     
     
         91 . A kit for determining the presence of pancreatic cancer comprising:
 C) one or more first binding agent as defined in any one of  claims 47  to  55  or an array according to  claims 63  to  66  or  claim 87  or  88 ;   D) instructions for performing the method as defined in any one of  claims 1  to  67  or  68  to  86 .   
     
     
         92 . A kit according to  claim 31  further comprising a second binding agent as defined in any one of  claims 56  to  61 . 
     
     
         93 . A method or use for determining the presence of pancreatic cancer in an individual substantially as described herein. 
     
     
         94 . An array or kit for determining the presence of pancreatic cancer in an individual substantially as described herein.