Mark M. Entius

Germline and Somatic Mutations of the STK11/LKB1 Peutz-Jeghers Gene in Pancreatic and Biliary Cancers

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.

Peutz-Jeghers syndrome: 78-year follow-up of the original family.

BACKGROUND The association between heredity, gastrointestinal polyposis, and mucocutaneous pigmentation in Peutz-Jeghers syndrome (PJS) was first recognised in 1921 by Peutz in a Dutch family. This original family has now been followed-up for more than 78 years. We did mutation analysis in this family to test whether the recently identified LKB1 gene is indeed the PJS gene in this family. METHODS The original family was retraced and the natural history of PJS was studied in six generations of this kindred by interview, physical examination, chart view, and histological review of tissue specimens. DNA-mutation analysis was done in all available descendants. FINDINGS Clinical features in this family included gastrointestinal polyposis, mucocutaneous pigmentation, nasal polyposis, and rectal extrusion of polyps. Survival of affected family members was reduced by intestinal obstruction and by the development of malignant disease. A novel germline mutation in the LKB1 gene was found to cosegregate with the disease phenotype in the original family. The mutant LKB1 allele carried a T insertion at codon 66 in exon 1 resulting in frameshift and stop at codon 162 in exon 4. INTERPRETATION The morbidity and mortality in this family suggest that PJS is not a benign disease. An inactivating germline mutation in the LKB1 gene is involved in the PJS phenotype in the original and oldest kindred known to be affected by PJS.

NOVEL MUTATIONS IN THE LKB1/STK11 GENE IN DUTCH PEUTZ-JEGHERS FAMILIES

The Peutz‐Jeghers syndrome (PJS) is a rare hereditary disorder in which gastrointestinal hamartomatous polyposis, mucocutaneous pigmentation, and a predisposition for developing cancer are transmitted in an autosomal dominant fashion. The recently identified LKB1/STK11 gene located at chromosome 19p13.3 is mutated in a number of PJS pedigrees. We performed mutation analysis in 19, predominantly Dutch, PJS families. In 12 of these families, we identified LKB1/STK11 mutations, none of which has been described before. These 12 novel LKB1/STK11 mutations consist of one nonsense mutation, three frameshift deletions, three frameshift insertions, two acceptor splice site mutations, and three missense mutations. In addition, we detected four polymorphisms in LKB1/STK11. In the remaining seven PJS families, we found no apparent abnormalities of the LKB1/STK11 gene, which could reflect the existence of locus heterogeneity in PJS. None of the mutations occurred in more than one family, and a number were demonstrated to have arisen de novo. The diverse array of mutations found, the apparent high mutation rate, as well as the existence of a possible second PJS locus, renders diagnostic or predictive genetic testing in individual patients difficult, although future identification of additional mutations or even gene(s) will help in increasing the yield of direct mutation analysis. Hum Mutat 13:476–481, 1999.

Molecular genetic alterations in hamartomatous polyps and carcinomas of patients with Peutz-Jeghers syndrome

Aim—To investigate whether mutations in the STK11/LKB1 gene and genes implicated in the colorectal adenoma–carcinoma sequence are involved in Peutz-Jeghers syndrome (PJS) related tumorigenesis. Methods—Thirty nine polyps and five carcinomas from 17 patients (from 13 families) with PJS were analysed for loss of heterozygosity (LOH) at 19p13.3 (STK11/LKB1 gene locus), 5q21 (APC gene locus), 18q21–22 (Smad4 and Smad2 gene locus), and 17p13 (p53 gene locus), and evaluated for immunohistochemical staining of p53. In addition, mutational analysis of K-ras codon 12, APC, and p53 and immunohistochemistry for Smad4 expression were performed on all carcinomas. Results—LOH at 19p was seen in 15 of the 39 polyps and in all carcinomas (n = 5). Interestingly, six of the seven polyps from patients with cancer had LOH, compared with nine of the 31 polyps from the remaining patients (p = 0.01). In one polyp from a patient without a germline STK11/LKB1 mutation, no LOH at 19p or at three alternative PJS candidate loci (19q, 6p, and 6q) was found. No LOH at 5q was observed. However, mutational analysis revealed an APC mutation in four of the five carcinomas. LOH at 17p was not seen in polyps or carcinomas; immunohistochemistry showed expression of p53 in one carcinoma and focal expression in three polyps. At subsequent sequence analysis, no p53 mutation was found. One carcinoma had an activating K-ras codon 12 mutation and another carcinoma showed 18q LOH; however, no loss of Smad4 expression was seen. Conclusions—These results provide further evidence that STK11/LKB1 acts as a tumour suppressor gene, and may be involved in the early stages of PJS tumorigenesis. Further research is needed to see whether LOH in PJS polyps could be used as a biomarker to predict cancer. Differences in molecular genetic alterations noted between the adenoma–carcinoma sequence and PJS related tumours suggest the presence of a distinct pathway of carcinogenesis.

Peutz-Jeghers polyps, dysplasia, and K-ras codon 12 mutations

Background—Peutz-Jeghers syndrome (PJS) is a rare, autosomal dominant, polyposis syndrome, associated with an increased risk of gastrointestinal and extragastrointestinal malignancy. Occasionally dysplasia occurs in PJS polyps. Aims—In colorectal carcinomas, mutations in codon 12 of the K-ras oncogene are common and are found at similar frequency in precursor adenomas. Therefore, K-rascodon 12 point mutations in PJS polyps were evaluated. Materials and methods—Fifty two PJS polyps, including four with dysplasia, collected from 19 patients with PJS, were analysed for mutations in the K-ras codon 12 by a mutant enriched polymerase chain reaction procedure, followed by allele specific oligodeoxynucleotide hybridisation. Results—A K-ras codon 12 mutation was identified in one colonic polyp with dysplasia. The mutation was found in the non-neoplastic epithelial cells and not in the dysplastic component of the polyp. Conclusions—K-ras codon 12 point mutations are very rare in PJS polyps, by contrast with colorectal adenomas. The findings support previous evidence that there seems to be no intrinsic relation between K-ras codon 12 mutation and dysplasia.