Germline and somatic mutations of the STK11/LKB1 Peutz-Jeghers gene in pancreatic and biliary cancers

Abstract

Peutz-Jeghers syndrome (PJS) is an autosomal-dominant disorder characterized by hamartomatous polyps in the gastrointestinal tract and by pigmented macules of the lips, buccal mucosa, and digits. Less appreciated is the fact that PJS also predisposes patients to an increased risk of gastrointestinal cancer, and pancreatic cancer has been reported in many PJS patients. It was recently shown that germline mutations of the STK11/LKB1 gene are responsible for PJS. We investigated the role of STK11/LKB1 in the development of pancreatic and biliary cancer in patients with and without the PJS. In a PJS patient having a germline splice site mutation in the STK11/LKB1 gene, sequencing analysis of an intestinal polyp and pancreatic cancer from this patient revealed loss of the wild-type allele of the STK11/LKB1 gene in the cancer. Inactivation of STK11/LKB1, by homozygous deletions or somatic sequence mutations coupled with loss of heterozygosity, was also demonstrated in 4-6% of 127 sporadic pancreatic and biliary adenocarcinomas. Our results demonstrate that germline and somatic genetic alterations of the STK11/LKB1 gene may play a causal role in carcinogenesis and that the same gene contributes to the development of both sporadic and familial forms of cancer.

Figure 1.

Histological findings of patient PJS1. A: The Peutz-Jeghers polyp of the duodenum (original magnification, ×50). B: Poorly differentiated adenocarcinoma of the pancreas (original magnification, ×250). Hematoxylin & eosin staining.

Figure 2.

Germline mutation of STK11/LKB1 in patient PJS1 and loss of the wild-type allele of STK11/LKB1 in the pancreatic cancer of the same patient. A: The germline mutation of STK11/LKB1 in the family of patient PJS1 was confirmed (lanes 2) ( Ref. 15 ). CFPAC1 (lanes 1) and two pancreatic cancer xenografts (lanes 3 and 4) exhibited normal sequences. The samples were sequenced with a forward primer, STK11-E3-SEQ-F2. The arrow indicates the site of the nucleotide insertion mutation. B: DNA from normal tissue of a non-Peutz-Jeghers patient (lane 1), cancer of patient PJS1 (lane 2), and polyps of patient PJS1 (lane 3) were PCR-amplified and sequenced with a reverse primer, STK11-E3-SEQ-R. The cancer, but not the polyp, of patient PJS1 showed loss of the remaining allele of STK11/LKB1.

Figure 3.

Duplex PCR analysis of homozygous deletions in pancreatic and biliary cancers. Detection of homozygous deletions in the genomic DNAs of pancreatic cancer xenograft PX30 and biliary cancer xenograft PX115 by duplex PCR using pairs of internal control primers (INTB4-B or MKK4-E) and STK11/LKB1-specific primers. The entire coding region of STK11/LKB1 was deleted in PX30. Only exon 1 of STK11/LKB1 was deleted in PX115.

Figure 4.

Somatic mutations in the exonic sequences of STK11/LKB1. A: PX68 (lanes 2) and PX68-1A (lanes 3) are two parallel xenografts of the same pancreatic cancer, and both exhibited the same somatic nucleotide substitution that created a stop codon (a non-sense mutation). The mutation was not detected in the normal tissue of the patient (PN68, lanes 1 and 4). Lanes 5 belong to another pancreatic tumor xenograft, which did not harbor a somatic mutation in STK11/LKB1. The arrow points to the site of the mutation. B: Xenograft tumor PX104 (lanes 1) and its corresponding primary cancer, PC104 (lanes 3), harbored the same nucleotide deletion, which was not detected in the normal tissue of the patient (PN104, lanes 2) or in another pancreatic xenograft (lanes 4). The arrow indicates the site of the mutation.