Detection of BRCA1/2 mutations in breast cancer patients from Thailand and Pakistan

Transcription

1 Clin Genet 2012: 82: Printed in Singapore. All rights reserved Letter to the Editor 2012 John Wiley & Sons A/S. Published by Blackwell Publishing Ltd CLINICAL GENETICS doi: /j x Detection of BRCA1/2 mutations in breast cancer patients from Thailand and Pakistan To the Editor: Women carrying a highly penetrant mutation in the genes BRCA1 (breast cancer 1) or BRCA2 have a 60 85% lifetime risk of breast cancer and a 15 40% lifetime risk of ovarian cancer (1). In Asia, the contribution of BRCA1/2 to breast cancer and the ethnic distribution of the different type of mutations remain relatively unexplored. We performed BRCA1/2 mutation screening using high-resolution melting curve analysis followed by Sanger sequencing (2) in an unselected hospital-based series of 77 breast cancer Pakistani patients residing in the Balochistan province (age range, 30 70; median age of disease onset, 47) and 121 early-onset patients and 9 patients with family history of breast cancer originating from Thailand (age range, 17 62; median age of disease onset, 36). All tested cases were, to our knowledge, unrelated. In BRCA1, one known nonsense mutation and three novel truncating mutations were observed in one or the other population. In BRCA2, one missense mutation causing aberrant splicing of the gene (3) and three novel truncating mutations were identified (Table 1). None of these mutations were observed in healthy subjects of respective population (85 controls from Balochistan and 190 controls from Thailand, respectively). (a) We also identified two variants of uncertain clinical significance (VUS) having relatively high sequence analysis-based prior probabilities in favour of pathogenicity (4), although functional or familial data are lacking to confirm the in silico predictions. Interestingly, a known pathogenic mutation occurring at the same position as the non-conservative BRCA1 VUS p.g1788c (Fig. 1a) had been reported (p.g1788v) in a patient of European ancestry (4, 5), and another VUS at same codon had been recorded in the Breast Cancer Information Core (BIC) (p.g1788d) but insufficient data exist to assess clinical or functional implications of this sequence change. The BRCA2 VUS p.g2901d (Fig. 1b) had been observed in Finnish familial ovarian cancer cases, was absent in Finnish controls, and analysis of tumour material from one case showed loss of the allele not carrying the variant (6). However, one Thai control carried it but her mother had cancer of the cervix at the age of 44. Thus, data on this p.g2901d are somewhat conflicting. Of note, a Pakistani patient carried the already reported BRCA1 in frame deletion c.3328_3330del. Since it falls in a region where sequence variations have an extremely low prior probability in favour of pathogenicity, it is far less likely to be pathogenic than the two missense substitutions discussed above. p. G1788C (b) p. G2901D C D Fig. 1. (a) A segment from our reference BRCA1 protein multiple sequence alignment centred on glycine The residue falls in the functionally important BRCA1 C terminus (BRCT) domain, is invariant from human to sea urchin, and is flanked by other strongly conserved residues. (b) A segment from our BRCA2 protein multiple sequence alignment centred on glycine The residue falls in the functionally important DNA binding domain, is invariant from human to sea urchin, and is flanked by other strongly conserved residues. Hsap, Homo sapiens; Ptro, Pan troglodytes (chimpanzee); Ggor, Gorilla gorilla; Ppyg, orangutan; Mmul, Macaca mulatta (monkey); Mmus (mus musculus), mouse; Cfam, Canis lupus familiaris (dog); Btau, Bos taurus (cow); mdom, Monodelphis domesticus (grey, short-tailed opossum); Ggal, Gallus gallus (red jungle fowl); Xlae, Xenopus laevis (frog); Tnig, Tetraodon nigroviridis (pufferfish, green spotted); Spur, Strongylocentrotus purpuratus (purple sea urchin); Rnor, Rattus norvegicus (rat); Xtro, X. laevis (frog); Frub, Fugu rubripes (pufferfish, fugu). 594

2 Table 1. BRCA1/2 pathogenic mutations, variants of uncertain clinical significance (VUS), and neutral variants identified in the Thai and Pakistani breast cancer patients Age at Gene Exon Position in transcript Effect on protein Variation type Population (ethnic group) Case ID b diagnosis Databases entries (a) Pathogenic mutations a BRCA1 11 c.1267insat p.s423fs FS Thai Tai Novel BRCA1 11 c.3169_3172delagta p.s1057fs FS Pakistani (Baloch) PKBC17 c 50 Novel BRCA1 11 c.3748g>t p.e1250x NS Thai Thai TaiN TaiS BIC ; UMD-BRCA1 BRCA1 21 c.5308insg p.g1770fs FS Pakistani (Baloch) PKBC47 c 47 Novel BRCA2 11 c.4729ginsa p.e1577fs FS Pakistani (Pashtoon) PKBC30 d 30 Novel Novel BRCA2 11 c.5111gdelaata p.r1704fs FS Thai Thai TaiF TaiN BRCA2 13 c.7007g>a p.r2336h SP Pakistani (Punjabi) PKBC44 e 35 BIC ; UMD-BRCA2 BRCA2 16 c.7676insaaac p.s2559fs FS Thai Tai Novel Gene Exon Position in transcript Effect on protein Variation type Population Case ID b grade Databases entries (b) Variants of uncertain clinical significance a BRCA1 11 c.823g>a p.g275s MS Pakistani (Baloch) PKBC73 C0 BIC ; UMD-BRCA1 BRCA1 11 c.1033g>t p.d345y MS Thai TaiN C0 BIC BRCA1 11 c.1847c>t p.s616f MS Thai Tai-057 C0 Novel BRCA1 11 c.2258g>c p.s753t MS Thai Tai-066 C0 Novel BRCA1 11 c.3119g>a p.s1040n MS Pakistani (Baloch) PKBC24 C0 BIC ; UMD-BRCA1 BRCA1 11 c.3328_3330delaag p.k1110del IFD Pakistani (Baloch) PKBC52 n/a BIC BRCA1 12 c.4166g>a p.s1389n MS Thai Tai-056 C0 Novel BRCA1 16 c.4892g>a p.s1631n MS Thai TaiF C0 Novel BRCA1 16 c.4942a>g p.k1648e MS Pakistani (Baloch) PKBC68 C0 Novel BRCA1 22 c.5362g>t p.g1788c MS Thai TaiS C65 f Novel BRCA2 10 c.1070a>c p.e357a MS Pakistani (Baloch) Pakistani (Pashtoon) PKBC31 PKBC70 C0 Novel BRCA2 11 c.2892a>t p.k964n MS Pakistani (Pashtoon) PKBC07 C0 Novel BRCA2 11 c.2918c>t p.s973l MS Thai Tai-010 C0 Novel BRCA2 11 c.5705a>t p.d1902v MS Pakistani (Baloch) PKBC22 C0 Novel BRCA2 14 c.7397t>c p.a2466v MS Pakistani (Punjabi) PKBC36 C0 BIC; UMD-BRCA2 BRCA2 21 c.8702g>a p.g2901d a MS Thai Tai-078 C65 f BIC BRCA2 26 c.9640a>g p.k3214e MS Thai Tai-059 C0 Novel 595

3 Table 1. Continued Frequency (%) g Gene Exon Position in transcript Effect on protein Variation type rs number Pakistan Thailand grade Databases entries (c) Neutral variants (common missense substitutions, silent substitutions and intronic variants) BRCA1 3 c.114g>a p.k38k S rs n/a dbsnp BRCA1 8 c c>t n/a I rs n/a dbsnp BRCA1 9 c.571g>a p.v191i MS 1.6 C0 BIC BRCA1 9 c.591c>t p.c197c S rs n/a dbsnp BRCA1 11 c.2082c>t p.s694s S rs n/a dbsnp BRCA1 11 c.2311t>c p.l771l S rs n/a dbsnp BRCA1 11 c.2566t>c p.y856h MS 2.3 C0 BIC BRCA1 11 c.2612c>t p.p871l MS rs C0 dbsnp BRCA1 11 c.3113a>g p.e1038g MS rs C0 dbsnp BRCA1 11 c.3548a>g p.k1183r MS rs C0 dbsnp BRCA1 11 c.3822a>c p.v1274v S 1.3 n/a Novel BRCA1 13 c.4308t>c p.s1436s S rs n/a dbsnp BRCA1 14 c a>g n/a I 1.5 n/a Novel BRCA1 16 c.4837a>g p.s1613g MS rs C0 dbsnp BRCA1 16 c.4956g>a p.m1652i MS 1.6 C0 BIC BRCA1 22 c.5388a>t p.s1796s S 0.8 n/a Novel BRCA1 22 c g>a n/a I 0.8 n/a Novel BRCA2 2 c.1-26g>a n/a P rs n/a dbsnp BRCA2 10 c.865a>c p.n289h MS rs n/a dbsnp BRCA2 10 c.943t>a p.c315s MS 0.5 C0 BIC 596

4 Table 1. Continued Frequency (%) g Gene Exon Position in transcript Effect on protein Variation type rs number Pakistan Thailand grade Databases entries BRCA2 10 c.1114a>c p.n372h MS rs n/a dbsnp BRCA2 11 c.2971a>g p.n991d MS rs n/a dbsnp BRCA2 11 c.3396a>g p.k1132k S rs n/a dbsnp BRCA2 11 c.3445a>g p.m1149v MS 0.5 C0 BIC BRCA2 11 c.4258g>t p.d1420y MS 0.5 n/a BIC BRCA2 11 c.5785a>g p.i1929v MS 1.0 C0 BIC BRCA2 14 c.7242a>g p.s2414s S rs n/a dbsnp BRCA2 17 c t>c n/a I rs n/a dbsnp BRCA2 18 c.8187g>t p.k2729n MS 0.5 C0 BIC BRCA2 27 c.10234a>g p.i3412v MS rs n/a dbsnp BIC, Breast Cancer Information Core; FS, frameshift; I, intronic; IFD, in frame deletion; MS, missense; n/a, not applicable; NS, nonsense; S, silent; SNP, single nucleotide polymorphism; SP, splice site mutation. a Pathogenic mutations and VUS in both genes were not detected in control subjects originated from the same population, with the exception of the missense substitution p.g2901d which was carried by one Thai control. b All cases were, to our knowledge, unrelated. c This patient had bilateral breast cancer. d This patient had breast cancer and ovarian cancer. e This patient had breast cancer and unconfirmed diagnostic of ovarian cancer. f This corresponds to the most likely deleterious grade output of and results in a prior probability in favour of pathogenicity of BIC database, UMD-BRCA1 Locus Specific database, UMD-BRCA2 Locus Specific database, dbsnp, g Thai and Pakistani control populations were composed of 190 and 85 healthy subjects, respectively. Pakistani controls were recruited in Quetta and were representative of the major ethnic groups in the Balochistan population. They were randomly chosen among female University faculty members or students. Thai controls were randomly selected from healthy women who visited patients admitted to the Bangkok hospital for diseases other than breast or ovarian cancer or from healthy women from the Khon-Kaen University. For both Pakistani and Thai controls, blood samples were collected after confirmation that the subject and her family had no history of breast cancer or ovarian cancer. 597

5 The rate of observed BRCA1/2 mutations in the Baloch population (5.2% of the unselected cases investigated) approaches that observed in other ethnic groups in Pakistan but observed variants were different from those identified in other series (7). Larger studies would be needed to elucidate whether they represent founder mutations. Inherited BRCA1/2 mutations were found in 4.6% of the Thai genetically enriched breast cancer cases investigated. To our knowledge, this represents the largest study conducted in this country, and the mutations we report differ from the mutations identified previously in this population (8). Extensive mutation screening of high-risk breast cancer primarily targeting early-onset cases could be undertaken in Pakistan and Thailand and could be paired with proper genetic counselling to help affected women and their at-risk relatives understand the genetic risk factors of breast cancer. These studies will contribute to the assessment of the usefulness of preventive program for mutation carriers as part of the National Public Health policies. Hence, personal risk information may help in taking preventive measures and also motivate a highrisk woman to adopt breast screening that may promote early detection and improve chances of surviving for these patients. Acknowledgements We thank all patients who participated in this study. We are thankful to Nabeela Nasreen and Rashida Hussain for their efforts to enrol the Pakistani subjects, and to Abdul Majeed Cheema and Dost Muhammad Baloch for facilitating the fieldwork at Quetta, Pakistan. We thank Drs G. M. Burdy and Khursheed Ahmad Khan, CENAR-Quetta for their help in collecting the blood samples, Nathalie Forey, Sandrine McKay-Chopin and Nivonirina Robinot for their technical help during the mutation screening, and Jocelyne Michelon for DNAs preparation. The work reported in this paper was undertaken during the tenure of a postdoctoral fellowship from the International Agency for Research on Cancer (J. A.). Recruitment of the Pakistani subjects was supported by faculty funds at BUITEMS (Ahmed Farooq Bazai, Vice Chancellor, BUITEMS). J Ahmad a,b F Le Calvez-Kelm a S Daud c C Voegele a MVallée a A Ahmad d N Kakar b JD McKay a V Gaborieau e MLéoné f O Sinilnikova f,g S Sangrajrang h SV Tavtigian i F Lesueur a a Genetic Cancer Susceptibility Group, International Agency for Research on Cancer (IARC), Lyon, France, b Department of Biotechnology and Informatics, Balochistan University of Information Technology, Engineering and Management Sciences (BUITEMS), Quetta, Pakistan, c Centre for Applied Molecular Biology, University of the Punjab, Lahore, Pakistan, d Mayo Hospital, Lahore, Pakistan; e Genetic Epidemiology Group, International Agency for Research on Cancer, Lyon, France, f Unité Mixte de Génétique Constitutionnelle des Cancers Fréquents, Centre Hospitalier Universitaire de Lyon, Centre Léon Bérard, Lyon, France, g INSERM U1052, CNRS UMR5286, Université Lyon 1, Centre de Recherche en Cancérologie Lyon-Est, Lyon, France, h National Cancer Institute, Bangkok, Thailand, and i Department of Oncological Sciences, Huntsman Cancer Institute, University of Utah School of Medicine, University of Utah, Salt Lake City, UT 84112, USA References 1. Palma MD, Domchek SM, Stopfer J et al. The relative contribution of point mutations and genomic rearrangements in BRCA1 and BRCA2 in high-risk breast cancer families. Cancer Res 2008: 68 (17): Nguyen-Dumont T, Calvez-Kelm FL, Forey N et al. Description and validation of high-throughput simultaneous genotyping and mutation scanning by high-resolution melting curve analysis. Hum Mutat 2009: 30 (6): Farrugia DJ, Agarwal MK, Pankratz VS et al. Functional assays for classification of BRCA2 variants of uncertain significance. Cancer Res 2008: 68 (9): Tavtigian SV, Byrnes GB, Goldgar DE, Thomas A. Classification of rare missense substitutions, using risk surfaces, with genetic- and molecular-epidemiology applications. Hum Mutat 2008: 29 (11): Easton DF, Deffenbaugh AM, Pruss D et al. A systematic genetic assessment of 1,433 sequence variants of unknown clinical significance in the BRCA1 and BRCA2 breast cancer-predisposition genes. Am J Hum Genet 2007: 81 (5): Roth S, Kristo P, Auranen A et al. A missense mutation in the BRCA2 gene in three siblings with ovarian cancer. Br J Cancer 1998: 77 (8): Farooq A, Naveed AK, Azeem Z, Ahmad T. Breast and ovarian cancer risk due to prevalence of BRCA1 and BRCA2 variants in Pakistani population: a Pakistani database report. J Oncol 2011: Patmasiriwat P, Bhothisuwan K, Sinilnikova OM et al. Analysis of breast cancer susceptibility genes BRCA1 and BRCA2 in Thai familial and isolated early-onset breast and ovarian cancer. Hum Mutat 2002: 20 (3): 230. Correspondence: Fabienne Lesueur Genetic Cancer Susceptibility group International Agency for Research on Cancer Lyon France Tel.: Fax: lesueurf@iarc.fr 598