Germline DICER1 Mutations Research Articles

DICER1 Syndrome: A New Cancer Syndrome

Recently, germline mutations of DICER1 have been identified in patients with rare neoplasms suggesting the existence of a newly discovered cancer prone syndrome. Initially, DICER1 mutations were identified in patients with familial pleuropulmonary blastoma. Subsequently, additional manifestations of the syndrome have been identified including cystic nephroma, medulloepithelioma, Sertoli-Leydig cell tumor and others. The DICER1 gene encodes an enzyme that is involved in the biogenesis of microRNAs. The entire tumor spectrum and the respective tumor risks are unknown. We are in the process of launching a natural history study aimed at identifying more information on this new cancer syndrome.

Abstract 3803: Genome-wide approach to identify gene targets of pleuropulmonary blastoma.

Abstract Pleuropulmonary blastoma (PPB) is an extremely rare and highly aggressive pediatric pulmonary malignancy that is often part of inherited cancer syndrome. Recently, germline mutations of DICER1, a gene encoding an endoribonuclease type III critical to the generation of small noncoding regulatory RNAs were reported in the majority of familial PPB. However, little is known about other genetic changes than DICER1 mutations that are responsible for the development and/or progression of PPB. Therefore, to assess the genetic alterations underlying the pathogenesis of PPB, genome-wide analyses using whole-exome sequencing and 250K SNP array were performed in total of ten cases with non-familial PPB. Whole-exome sequencing was performed in two cases which primary tumor-relapse tumor pairs were available. Although most mutations were commonly detected in both the primary and relapse tumors, several alterations were private to either of both samples, suggesting that clonal evolution derives genetic divergence in relapsed/metastatic tumors of PPB. In accordance with this, SNP array analysis also revealed several shared or private copy change variations, the most common finding was gain of the long arm of chromosome 8 in 7 cases, which was previously reported common in PPB. Whole-exome sequencing disclosed that DICER1 mutation, major cause of familial PPB, was recurrently found in the primary and relapse tumors of one case, but not detected in the another case. In the validation cohort of 8 cases of non-familial PPB, all of them had DICER1 mutations, and 6 cases had compound heterozygous mutations of DICER1. Notably, homozygous DICER1 mutation was observed in one case, and subsequent SNP array analysis revealed uniparental disomy of 14q lesion harboring DICER1 in this case. Somatic origins of missense changes were confirmed in 2 of 10 cases, for which DNA was obtained from peripheral blood. Meanwhile, whole-exome sequencing identified 13 candidate genes including the cadherin-related gene and a negative regulator of cell cycle in one case without DICER1 mutation. These gene mutations shared in primary and relapse samples, suggests that these are driver mutations rather than passenger mutations or non-functional SNPs. In conclusion, DICER1 mutation might be a driver mutation not only in familial PPB, but also in non-familial PPB. While DICER1 mutation was reported only germline heterozygous mutation in familial PPB, we notably found compound heterozygous or homozygous DICER1 mutations in non-familial PPB. Furthermore, there were different clones between primary and relapse samples, suggests that clonal evolution contributes to relapse tumor progression. Citation Format: Masafumi Seki, Riki Nisimura, Yusuke Okuno, Kenichi Yoshida, Yuichi Shiraishi, Masashi Sanada, Keisuke Kato, Katsuyoshi Koh, Ryoji Hanada, Yasuhide Hayashi, Satoru Miyano, Seishi Ogawa, Junko Takita. Genome-wide approach to identify gene targets of pleuropulmonary blastoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3803. doi:10.1158/1538-7445.AM2013-3803

Germ-line DICER1 mutations do not make a major contribution to the etiology of familial testicular germ cell tumours

BackgroundThe RNase III enzyme DICER1 plays a central role in maturation of microRNAs. Identification of neoplasia-associated germ-line and somatic mutations in DICER1 indicates that mis-expression of miRNAs in cancer may result from defects in their processing. As part of a recent study of DICER1 RNase III domains in 96 testicular germ cell tumors, a single RNase IIIb domain mutation was identified in a seminoma. To further explore the importance of DICER1 mutations in the etiology of testicular germ cell tumors (TGCT), we studied germ-line DNA samples from 43 probands diagnosed with familial TGCT.FindingsWe carried out High Resolution Melting Curve Analysis of DICER1 exons 2–12, 14–19, 21 and 24–27. All questionable melt curves were subjected to confirmatory Sanger sequencing.Sanger sequencing was used for exons 13, 20, 22 and 23. Intron-exon boundaries were included in all analyses. We identified 12 previously reported single nucleotide polymorphisms and two novel single nucleotide variants. No likely deleterious variants were identified; notably no mutations that were predicted to truncate the protein were identified.ConclusionsTaken together with previous studies, the findings reported here suggest a very limited role for either germ-line or somatic DICER1 mutations in the etiology of TGCT.

Exploring the endocrine manifestations of DICER1 mutations

The discovery of each new cancer susceptibility gene answers one set of questions but poses many more. In this article, we outline a recent example: a new cancer syndrome caused by germline mutations in DICER1, responsible for microRNA processing. In particular, we discuss the endocrine manifestations of mutations in this crucial gene.

Germline DICER1 mutation and associated loss of heterozygosity in a pineoblastoma

Pineoblastomas occur in RB-1 mutation carriers,1 but are rarely seen in association with other hereditary syndromes.2 Germline DICER1 mutations are associated with embryonal tumours such as pleuropulmonary blastoma (PPB),3 ,4...

Embryonal rhabdomyosarcoma of the uterine cervix: a report of 14 cases and a discussion of its unusual clinicopathological associations

Embryonal rhabdomyosarcoma of the uterine cervix is an uncommon presentation of the most common soft-tissue sarcoma in the first decades of life. Unlike embryonal rhabdomyosarcoma in other anatomic sites, in which 70–80% of cases present before 9 years of age, the average age in our series of 14 cervical cases was 12.4 years (median, 13 years), with an age range of 9 months to 32 years at diagnosis. Of the 14 cases, 12 presented as a polyp at the cervical os; two patients had an infiltrative mass in the cervix without a botryoid polyp. The polyps measured 1.5–5 cm and all had the histopathological pattern of the sarcoma botryoides variant of embryonal rhabdomyosarcoma, with condensations of primitive and differentiated rhabdomyoblasts beneath the surface epithelium and around endocervical glands. Nodules of benign-appearing cartilage were present in the stroma of six cases (43%). One of the embyronal rhabdomyosarcomas from the youngest patient, 9 months old, also had a distinctive microscopic focus of immature tubular profiles in a primitive stroma; these tubules expressed epithelial and neuroendocrine markers. Two patients had a pleuropulmonary blastoma, one diagnosed 9 years before the embryonal rhabdomyosarcoma of the cervix and the other recognized synchronously. This latter 9-year old had a DICER1 germline mutation. One patient presented with hirsutism and had a Sertoli–Leydig cell tumor, an incidentally detected cervical embryonal rhabdomyosarcoma, and nodular hyperplasia of the thyroid. Although a pleuropulmonary blastoma was not documented in the latter patient, ovarian sex-cord stromal tumors and nodular hyperplasia of the thyroid are manifestations of the pleuropulmonary blastoma family tumor and dysplasia syndrome (OMIM 601200). Embryonal rhabdomyosarcoma of the cervix must be distinguished from other rare entities, including adenosarcoma, malignant mixed Mullerian tumor and low-grade stromal sarcoma, as the former has a better prognosis; 12 of our 14 patients remain disease-free following conservative surgery and chemotherapy. Our study suggests that cervical embryonal rhabdomyosarcoma may be another pathological manifestation in the spectrum of extrapulmonary pathology in the setting of pleuropulmonary blastoma.

Recurrent Somatic DICER1 Mutations in Nonepithelial Ovarian Cancers

BackgroundGermline truncating mutations in DICER1, an endoribonuclease in the RNase III family that is essential for processing microRNAs, have been observed in families with the pleuropulmonary blastoma–family tumor and dysplasia syndrome. Mutation carriers are at risk for nonepithelial ovarian tumors, notably sex cord–stromal tumors.MethodsWe sequenced the whole transcriptomes or exomes of 14 nonepithelial ovarian tumors and noted closely clustered mutations in the region of DICER1 encoding the RNase IIIb domain of DICER1 in four samples. We then sequenced this region of DICER1 in additional ovarian tumors and in certain other tumors and queried the effect of the mutations on the enzymatic activity of DICER1 using in vitro RNA cleavage assays.ResultsDICER1 mutations in the RNase IIIb domain were found in 30 of 102 nonepithelial ovarian tumors (29%), predominantly in Sertoli–Leydig cell tumors (26 of 43, or 60%), including 4 tumors with additional germline DICER1 mutations. These mutations were restricted to codons encoding metal-binding sites within the RNase IIIb catalytic centers, which are critical for microRNA interaction and cleavage, and were somatic in all 16 samples in which germline DNA was available for testing. We also detected mutations in 1 of 14 nonseminomatous testicular germ-cell tumors, in 2 of 5 embryonal rhabdomyosarcomas, and in 1 of 266 epithelial ovarian and endometrial carcinomas. The mutant DICER1 proteins had reduced RNase IIIb activity but retained RNase IIIa activity.ConclusionsSomatic missense mutations affecting the RNase IIIb domain of DICER1 are common in nonepithelial ovarian tumors. These mutations do not obliterate DICER1 function but alter it in specific cell types, a novel mechanism through which perturbation of microRNA processing may be oncogenic. (Funded by the Terry Fox Research Institute and others.)

DICER1 mutations in embryonal rhabdomyosarcomas from children with and without familial PPB-tumor predisposition syndrome.

Embryonal rhabdomyosarcoma (ERMS) is the most common childhood sarcoma and is a component of the familial pleuropulmonary blastoma (PPB)-predisposition syndrome. Using the PPB model, we hypothesized that DICER1 mutations would be found in familial and sporadic forms of ERMS. Blood samples from four children with familial PPB and ERMS, and 52 sporadic ERMS tumors were tested for DICER1 mutations. Germline DICER1 mutations were found in all four patients with familial PPB and 2 of 52 (3.8%) sporadic ERMS had somatic mutations. Our findings confirm the pathogenetic relationship between ERMS and PPB suggesting that ERMS may result from abnormal miRNA regulation.

Extending the phenotypes associated withDICER1mutations

DICER1 is crucial for embryogenesis and early development. Forty different heterozygous germline DICER1 mutations have been reported worldwide in 42 probands that developed as children or young adults, pleuropulmonary blastoma (PPB), cystic nephroma (CN), ovarian sex cord-stromal tumors (especially Sertoli-Leydig cell tumor [SLCT]), and/or multinodular goiter (MNG). We report DICER1 mutations in seven additional families that manifested uterine cervix embryonal rhabdomyosarcoma (cERMS, four cases) and primitive neuroectodermal tumor (cPNET, one case), Wilms tumor (WT, three cases), pulmonary sequestration (PS, one case), and juvenile intestinal polyp (one case). One carrier developed (age 25 years) a pleomorphic sarcoma of the thigh; another carrier had transposition of great arteries (TGA). These observations show that cERMS, cPNET, WT, PS, and juvenile polyps fall within the spectrum of DICER1-related diseases. DICER1 appears to be the first gene implicated in the etiology of cERMS, cPNET, and PS. Young adulthood sarcomas and perhaps congenital malformations such as TGA may also be associated.

Ovarian sex cord-stromal tumors, pleuropulmonary blastoma and DICER1 mutations: A report from the International Pleuropulmonary Blastoma Registry

Objective Pleuropulmonary blastoma (PPB) is a childhood cancer arising from pleuropulmonary mesenchyme. This neoplasm is a sentinel disease in a familial tumor syndrome recently found to be associated with germline mutations in DICER1. Observations of ovarian sex cord-stromal tumors (OSCST) in PPB kindreds led to further study. We sought to characterize ovarian tumors seen in probands and families with PPB and PPB-related conditions and define germline DICER1 status. Methods Patient and family records of pathology-reviewed PPB cases enrolled in the International PPB Registry (IPPBR) were searched for ovarian tumors. Ovarian tumor pathology specimens were obtained and centrally reviewed. Germline DNA from patients with ovarian tumors was tested for DICER1 mutations. Three additional OSCST patients registered in the IPPBR were also tested for mutations in DICER1. Results Among 296 kindreds including 325 children with PPB, we observed three children with both PPB and Sertoli–Leydig cell tumors (SLCT)/Sertoli cell tumors. Among family members of PPB patients, we identified six OSCST (three SLCT, one Sertoli cell tumor, one juvenile granulosa cell tumor, one gynandroblastoma). Age at ovarian tumor diagnosis was youngest in PPB probands and younger in family members than in OSCST in general. Germline DICER1 mutations were identified in four of six patients with OSCST from PPB kindreds and in two of three children with OSCST and no personal or family history of PPB. Conclusions Primary ovarian neoplasms, particularly OSCST, are a manifestation of the familial PPB syndrome and may be the initial clinical presentation of DICER1 mutations within a family.

<emph type="ital">DICER1</emph> Mutations in Familial Multinodular Goiter With and Without Ovarian Sertoli-Leydig Cell Tumors

Nontoxic multinodular goiter (MNG) is frequently observed in the general population, but little is known about the underlying genetic susceptibility to this disease. Familial cases of MNG have been reported, and published reports describe 5 families that also contain at least 1 individual with a Sertoli-Leydig cell tumor of the ovary (SLCT). Germline mutations in DICER1, a gene that codes for an RNase III endoribonuclease, have been identified in families affected by pleuropulmonary blastoma (PPB), some of whom include cases of MNG and gonadal tumors such as SLCTs. To determine whether familial MNG with or without SLCT in the absence of PPB was associated with mutations in DICER1. From September 2009 to September 2010, we screened 53 individuals from 2 MNG and 3 MNG/SLCT families at McGill University for mutations in DICER1. We investigated blood lymphocytes and MNG and SLCT tissue from family members for loss of the wild-type DICER1 allele (loss of heterozygosity), DICER1 expression, and microRNA (miRNA) dysregulation. Detection of germline DICER1 gene mutations in familial MNG with and without SLCT. We identified and characterized germline DICER1 mutations in 37 individuals from 5 families. Two mutations were predicted to be protein truncating, 2 resulted in in-frame deletions, and 1 was a missense mutation. Molecular analysis of the 3 SLCTs showed no loss of heterozygosity of DICER1, and immunohistochemical analysis in 2 samples showed strong expression of DICER1 in Sertoli cells but weak staining of Leydig cells. miRNA profiling of RNA from lymphoblastoid cell lines from both affected and unaffected members of the familial MNG cases revealed miRNA perturbations in DICER1 mutation carriers. DICER1 mutations are associated with both familial MNG and MNG with SLCT, independent of PPB. These germline DICER1 mutations are associated with dysregulation of miRNA expression patterns.

A Genetic Defect in Exportin-5 Traps Precursor MicroRNAs in the Nucleus of Cancer Cells

The global impairment of mature microRNAs (miRNAs) is emerging as a common feature of human tumors. One interesting scenario is that defects in the nuclear export of precursor miRNAs (pre-miRNAs) might occur in transformed cells. Exportin 5 (XPO5) mediates pre-miRNA nuclear export and herein we demonstrate the presence of XPO5-inactivating mutations in a subset of human tumors with microsatellite instability. The XPO5 genetic defect traps pre-miRNAs in the nucleus, reduces miRNA processing, and diminishes miRNA-target inhibition. The XPO5 mutant form lacks a C-terminal region that contributes to the formation of the pre-miRNA/XPO5/Ran-GTP ternary complex and pre-miRNAs accumulate in the nucleus. Most importantly, the restoration of XPO5 functions reverses the impaired export of pre-miRNAs and has tumor-suppressor features.