Differential Identification of Pancreatic Cysts

Abstract

More than 2% of adults harbor a pancreatic cyst, a subset of which progress to invasive lesions with lethal consequences. To assess the genomic landscapes of neoplastic cysts of the pancreas, we determined the exomic sequences of DNA from the neoplastic epithelium of eight surgically resected cysts of each of the major neoplastic cyst types: serous cystadenomas (SCAs), intraductal papillary mucinous neoplasms (IPMNs), mucinous cystic neoplasms (MCNs) and solid pseudo-papillary neoplasms (SPNs). SPNs are low-grade malignancies, and IPMNs and MCNs, but not SCAs, have the capacity to progress to cancer. We found that SCAs, IPMNs, MCNs, and SPNs contained 10=4.6, 27=12, 16=7.6, and 2.9=2.1 somatic mutations per tumor, respectively. Among the mutations identified, E3 ubiquitin ligase components were of particular note. Four of the eight SCAs contained mutations of VHL, a key component of the VHL ubiquitin ligase complex that has previously been associated both with renal cell carcinomas, SCAs, and other neoplasms. Six of the eight IPMNs and three of the eight MCNs harbored mutations of RNF43, a gene coding for a protein with intrinsic E3 ubiquitin ligase activity that has not previously been found to be genetically altered in any human cancer. The preponderance of inactivating mutations in RNF43 unequivocally establish it as a suppressor of both IPMNs and MCNs. SPNs contained remarkably few genetic alterations, but always contained mutations of CTNNB1, previously demonstrated to inhibit degradation of the encoded protein (β-catenin) by E3 ubiquitin ligases. These results highlight the essential role of ubiquitin ligases in these neoplasms and have important implications for the diagnosis and treatment of patients with cystic tumors.

1 - 8 . (canceled) 
     
     
         9 . A kit comprising a nucleic acid primer complementary to RNF43, wherein the nucleic acid primer comprises an adaptor, wherein the adaptor comprises a priming site for nucleic acid sequencing or amplification, wherein the priming site is not complementary to RNF43. 
     
     
         10 . The kit of  claim 9  further comprising a nucleic acid primer complementary to VHL, GNAS, and CTNNBI. 
     
     
         11 . The kit of  claim 9  further comprising a nucleic acid primer complementary to KRAS. 
     
     
         12 . The kit of  claim 10  further comprising a nucleic acid primer complementary to KRAS. 
     
     
         13 . A kit comprising a nucleic acid probe complementary to RNF43, wherein the nucleic acid probe comprises a chemical modification. 
     
     
         14 . The kit of  claim 13  further comprising a nucleic acid probe complementary to VHL, GNAS, and CTNNBI. 
     
     
         15 . The kit of  claim 13  further comprising a nucleic acid probe complementary to KRAS. 
     
     
         16 . The kit of  claim 14  further comprising a nucleic acid probe complementary to KRAS. 
     
     
         17 . A kit comprising a set of oligonucleotides probes which are complementary to at least one gene from each of the following groups:
 a. VHL;   b. GNAS;   c. RNF43, KRAS; and   d. CTNNBI   wherein the 5′ end of each of the oligonucleotide probes is modified.   
     
     
         18 . The kit of  claim 17  wherein the 5′ end of each of the oligonucleotide probes is modified with a fluorescein label. 
     
     
         19 . The kit of  claim 17  wherein the set comprises at least two probes complementary to at least one gene from each of the groups, wherein the probes are differentially labeled. 
     
     
         20 . The kit of  claim 19  wherein the at least two probes are complementary respectively to a mutant and a wild-type allele of the at least one gene. 
     
     
         21 . The kit of  claim 17  which comprises an oligonucleotide probe complementary to RNF43. 
     
     
         22 . A kit comprising a set of oligonucleotide primers which are complementary to at least one gene from each of the following groups:
 a. VHL;   b. GNAS;   c. RNF43, KRAS; and   d. CTNNBI   wherein each primer is ligated to an adaptor nucleic acid molecule for attachment to a flow cell.   
     
     
         23 . The kit of  claim 22  wherein the set comprises at least two primers complementary to at least one gene from each of the groups. 
     
     
         24 . The kit of  claim 23  wherein the at least two primers are complementary respectively to a mutant and a wild-type allele of the at least one gene. 
     
     
         25 . The kit of  claim 23  which comprises an oligonucleotide primer complementary to RNF43