High Frequency of Germline TP53 Mutations in a Prospective Adult-Onset Sarcoma Cohort

et al. (2013) High Frequency of Germline TP53 Mutations in a Prospective Adult-Onset Sarcoma Cohort. PLoS ONE 8(7): e69026. doi:10.1371/journal.pone.0069026 High Frequency of Germline TP53 Mutations in a Prospective Adult-Onset Sarcoma Cohort Gillian Mitchell 0 Mandy L. Ballinger 0 Stephen Wong 0 Chelsee Hewitt 0 Paul James 0 Mary- 0 Anne Young 0 Arcadi Cipponi 0 Tiffany Pang 0 David L. Goode 0 Alex Dobrovic 0 David M. Thomas 0 on behalf of the International Sarcoma Kindred Study 0 Mathias Toft, Oslo University Hospital, Norway 0 1 Department of Cancer Medicine, Peter MacCallum Cancer Centre , Melbourne, Victoria , Australia , 2 Research Division, Peter MacCallum Cancer Centre , Melbourne, Victoria , Australia , 3 Department of Pathology, Peter MacCallum Cancer Centre , Melbourne, Victoria , Australia , 4 Sir Peter MacCallum Department of Oncology, University of Melbourne , Melbourne, Victoria , Australia , 5 Department of Pathology, University of Melbourne , Melbourne, Victoria , Australia Sarcomas are a key feature of Li-Fraumeni and related syndromes (LFS/LFL), associated with germline TP53 mutations. Current penetrance estimates for TP53 mutations are subject to significant ascertainment bias. The International Sarcoma Kindred Study is a clinic-based, prospective cohort of adult-onset sarcoma cases, without regard to family history. The entire cohort was screened for mutations in TP53 using high-resolution melting analysis and Sanger sequencing, and multiplexligation-dependent probe amplification and targeted massively parallel sequencing for copy number changes. Pathogenic TP53 mutations were detected in blood DNA of 20/559 sarcoma probands (3.6%); 17 were germline and 3 appeared to be somatically acquired. Of the germline carriers, one appeared to be mosaic, detectable in the tumor and blood, but not epithelial tissues. Germline mutation carriers were more likely to have multiple cancers (47% vs 15% for non-carriers, P = 3.061023), and earlier cancer onset (33 vs 48 years, P = 1.1961023). The median survival of mutation carriers following first cancer diagnosis was not significantly different from non-carriers. Only 10/17 (59%) pedigrees met classical or Chompret criteria for LFS. In summary, germline TP53 mutations are not rare in adult patients with sarcoma, with implications for screening, surveillance, treatment and genetic counselling of carriers and family members. - Funding: This work was supported by a Johanna Sewell Research Grant, the Rainbows for Kate Foundation, the Victorian Cancer Agency, and the National Health and Medical Research Council (Project Grant 1004017). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. . These authors contributed equally to this work. " These authors also contributed equally to this work. Germline TP53 mutations result in the classical Li-Fraumeni or Li-Fraumeni-like syndromes (LFS/LFL) [1], rare inherited syndromes with a lifetime cancer penetrance up to 73% for males and ,100% for females[27]. Historically there has been little enthusiasm in the medical community for germline TP53 testing in LFS/LFL. Reasons include the perceptions of rarity, a lack of proven risk management strategies, and the potential for psychological harm by identifying people with an unmodifiable, extreme cancer risk [8]. However, developments in breast and whole body MRI screening [9,10], pre-implantation genetic diagnosis for family planning, and the use of genetic information to guide cancer therapy, may influence decision-making for TP53 genetic testing. Mutation frequencies and penetrance estimates are largely derived from pedigree-ascertained pediatric, cohorts[46,8,11], fraught with ascertainment biases. Moreover, studies of LFSassociated cancers[1216] suggest many germline TP53 mutation carriers have little family history, or will be increasingly identified through genomic screens of cancer populations unselected for family history. Accurate risk counselling to the carriers identified in these ways will require study of the impact of TP53 mutations outside of familial settings. Sarcomas are the most common cancer type seen in LFS [17]; approximately 90% of sarcomas occur in adults [18]. To determine the incidence and clinical spectrum of germline TP53 mutations in adult-onset sarcoma populations, a systematic screen using multiplexed ligation-dependent probe amplification and Sanger sequencing was undertaken in 559 probands consecutively recruited from adult sarcoma clinicsagnostic to family historyon the Australian arm of the International Sarcoma Kindred Study (ISKS; http:// www.anzctr.org.au; http://www.australiansarcomagroup.org/ sarcomakindredstudy/index.html). Pathogenic or putatively pathogenic TP53 events occurred in the peripheral blood DNA of 20/559 probands (3.6%), comprising 18 single nucleotide mutations or indels, and 2 whole gene deletions (Table 1). Pathogenicity was assigned as described in the Materials and Methods, and in Figure S1. Most were previously s me p r s e e a 3 0 3 e R g c 0 1 0 0 0 0 5 0 9 2 1 1 0 0 4 0 0 0 1 1 h reported somatically [19], but 10 are reported here for the first time in the germline. Six variants were regarded as putatively pathogenic. The age of sarcoma onset in individuals carrying pathogenic variants was not significantly different from those carrying putatively pathogenic variants (mean6standard deviation: 38617 years versus 40619 years, compared to 48618 years for non-carriers in the ISKS cohort). Seventeen were putative germline events, with the mutant allele also detected in tumor DNA and 8 tumors also demonstrating loss of heterozygosity. The remaining 3 cases suggested somatic origin: Case 18 had clinical evidence of myelodysplasia (MDS), and neither parent carried the TP53 mutation. Both cases 19 and 20 demonstrated heterozygous whole gene deletion. While neither cases 19 or 20 had clinical evidence of MDS, both cases 18 and 19 showed widespread copy number changes in the peripheral bloodincluding the RB1 locus in case 19suggestive of somatic tumor changes rather than germline events. Only case 18 had been exposed to chemotherapy prior to blood sampling. Ten of 17 (59%) germline carriers had classical LFS or Chompret pedigrees that would have prompted genetic testing (Table 2). Case 14 showed somatic mosaicism, with 2025% of mutant alleles (estimated by both Sanger sequencing and HRM analysis) in the peripheral blood, heterozygosity in the tumor, and absent in adjacent buccal mucosa (Figure 1). Somatic mosaicism for TP53 mutations has been previously reported [20]. Of the putatively pathogenic variants, 1 occurred in a family fitting classical Li-Fraumeni criteria (case 4); two occurred in individuals fitting Chompret criteria (cases 6 & 8), and three did not demonstrate an unusual family history of cancer (cases 1 (...truncated)