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Genetics and Molecular Biology

Print version ISSN 1415-4757On-line version ISSN 1678-4685

Genet. Mol. Biol. vol.43 no.2 Ribeirão Preto  2020  Epub May 20, 2020 

Human and Medical Genetics

Haplotypic characterization of BRCA1 c.5266dupC, the prevailing mutation in Brazilian hereditary breast/ovarian cancer

Renan Gomes1

Barbara Luisa Soares1

Paula Silva Felicio2

Rodrigo Michelli2 

Cristina B. O. Netto3 

Barbara Alemar4  5

Patrícia Ashton-Prolla4  5

Edenir Inêz Palmero2  6

Miguel Ângelo Martins Moreira1

1Instituto Nacional de Câncer, Programa de Genética, Rio de Janeiro, RJ, Brazil.

2Hospital de Cancer de Barretos, Centro de Pesquisa em Oncologia Molecular, Barretos, SP, Brazil.

3Hospital de Clínicas de Porto Alegre, Serviço de Genética Médica, Porto Alegre, RS, Brazil.

4Hospital de Clínicas de Porto Alegre, Laboratório de Medicina Genômica, Porto Alegre, RS, Brazil.

5Universidade Federal do Rio Grande do Sul (UFRGS), Departamento de Genética, Programa de Pós-Graduação em Genética e Biologia Molecular, Porto Alegre, RS, Brazil.

6Faculdade de Ciências de Saúde de Barretos Dr. Paulo Prata (FACISB), Barretos, SP, Brazil.


Specific pathogenic mutations associated with breast cancer development can vary between ethnical groups. One example is BRCA1 c.5266dupC that was first described as a founder mutation in the Ashkenazi Jewish population, but was later also found in other populations. In Brazil, this mutation corresponds to 20% of pathogenic BRCA1 variants reported. Our objective was to investigate the haplotype component of a group of Brazilian families who inherited c.5266dupC in the BRCA1 gene and to verify the ancestry contribution from European, African, and Amerindian origins. Fourteen probands carrying c.5266dupC and 16 relatives (carriers and non-carriers) were investigated. The same haplotype was observed segregating within all the families analyzed, revealing no recombinants in a region of 0.68 Mb. Ancestry analysis demonstrated that the European component was predominant among probands. The BRCA1 c.5266dupC analysis indicates that there was a founder effect in the Brazilian population.

Keywords Founder mutation; Ashkenazi Jewish; BRCA1; BRCA1 c.5266dupC; hereditary breast cancer

The spectrum of pathogenic mutations found in genes related to cancer development can vary depending on the ethnic groups that are being studied. Specific pathogenic mutations associated to particular ethnic groups show a high frequency due to founder effects, or population bottleneck and consequent inbreeding. As a result, rare pathogenic mutations become more common within the population over time (Ferla et al., 2007). Inbreeding contributes to linkage disequilibrium in genomic regions that are segregated together for many generations with specific alleles in loci placed closest to the mutation site in that specific population (Fackenthal and Olopade, 2007).

A well-known example of founder effects is that of BRCA pathogenic mutations in the Ashkenazi Jewish population (Rubinstein, 2004). At least 2.6% (1/40) of this population carry one of the three founder pathogenic mutations described for BRCA1 (OMIM #113705) and BRCA2 (OMIM #600185): BRCA1c.66_67delAG (p.Glu23fs), BRCA1 c.5266dupC (p.Gln1756fs, former named 5382insC), and BRCA2c.5946delT (p.Ser1982fs) (Rubinstein, 2004; Antoniou et al., 2005; Ferla et al., 2007; Hamel et al., 2011; Tafe et al., 2015). Although these pathogenic mutations were first identified in Ashkenazi Jews, BRCA1 c.5266dupC, was later described in other populations. It has already been identified in many countries of Central and Eastern Europe (Burcos et al., 2013; Gorodetska et al., 2015) and also recurrently described in the Brazilian population (Lourenço et al., 2004; da Costa et al., 2008; Fernandes et al., 2016), representing 20% of the BRCA1 pathogenic variants reported in a recent survey (Palmero et al., 2018).

The comparison of haplotypes between families sharing the same mutation allows to distinguish whether high-frequency alleles derive from a single mutational event (a founder mutation), or if they have arisen independently more than once in a population (Hamel et al., 2011; Ossa and Torres, 2016). In a previous report, da Costa et al. (2008) showed that seven unrelated carriers of this mutation share the same haplotype of genetic marker alleles flanking BRCA1. However, data available in Brazil regarding the frequency of different pathogenic variants in individuals at risk of hereditary breast/ovary cancer and the availability of samples from the c.5266dupC mutation carriers limited the conclusions at that time with respect to a possible founder effect. In addition, da Costa et al. (2008) did not evaluate the ancestry of the carriers.

In this work, we present a haplotype analysis in an expanded set of Brazilian carriers of BRCA1 c.5266dupC. In addition, considering that the Brazilian population is highly admixed, with genetic backgrounds derived from Europeans, Africans and Amerindians, ancestry analysis was carried out to assess the contribution of these different backgrounds to the genetic diversity present in the carriers

Fourteen unrelated heterozygous probands harboring BRCA1 c.5266dupC and 26 relatives (carriers or non-carriers of eight families) were recruited for this study from collaborating research centers located in the cities of Rio de Janeiro (5 probands), Barretos (5 probands), and Porto Alegre (4 probands) between 2004 to 2016. All research procedures followed ethical guidelines and were approved by the local Ethics Committee (009/07). The five probands/families from Rio de Janeiro were also previously analyzed in da Costa et al. (2008).

Haplotypes were characterized based on three SNPs and four Short Tandem Repeat (STR) markers along ~581 kb of chromosome 13 encompassing the BRCA1 locus (Figure S1). SNPs were analyzed by PCR amplification and DNA sequencing, as described by da Costa et al. (2008).

Four microsatellite loci were used for genotyping: D17S855 (intragenic marker in intron 20), D17S1325 and D17S1326 (3’ markers) and D171321 (5’ marker); primers are listed in Table S1. For all microsatellite loci, PCR amplifications were performed in final volumes of 25 μL, with 2.0 mmol/L of MgCl2, 125 μmol/L of each dNTP, 20 pmol of each primer, 1x PCR buffer, 1 U of Platinum Taq DNA polymerase (Invitrogen) and 50 ng of genomic DNA. Reactions were submitted to 30 cycles of 94 °C for 15 s, 60 °C for 15 s, and 72 °C for 20 s. Forward primers were labeled with carboxyfluorescein, and PCR products were analyzed in an ABI-PRISM 3730 automatic sequencer (Applied Biosystems, Foster City, CA). Scoring of allele size was achieved using the internal size standard GeneScan –500 LIZ® (Applied Biosystems). Allele size was estimated using Peak ScannerTM software v1.0 (Applied Biosystems).

To estimate genetic ancestry, 46 ancestry-informative markers (AIMs) were selected. These markers were used to investigate the contribution of African, European, East Asian, and Native American populations to the genetic background of the probands. AIMs were genotyped in one multiplex PCR assay followed by capillary electrophoresis, as previously described (Pereira et al., 2012). Structure software (Pritchard et al., 2000) was used to estimate the ancestral components of the samples, and the results were validated by Admixture software. Genetic ancestry analysis was carried out for the 14 index cases.

Of all BRCA1 c.5266dupC carriers, 21 (63.6%) had been diagnosed with breast and/or ovarian cancer at ages ranging from 22 and 63 years old (median = 43 years) (Table 1). The age of cancer-unaffected mutation carriers (n=12) ranged from 22 to 66 years old (median = 47). For five of the 14 probands only the personal cancer history was available, for the remainder at least one relative developed breast or ovarian cancer (Table 1).

Table 1 Clinical features of BRCA1 c.5266dupC carriers. 

Carrier Relationship Gender Cancer type Tumor location Age at diagnostic
RJ-01 Proband F Ovarian Bilateral 47/47
RJ-02 Proband F Breast Bilateral 36/41
RJ-02C 3rd degree relative F Breast Bilateral 47
RJ-02F 3rd degree relative F Breast Unilateral 49
RJ-02G 4th degree relative F Healthy - -
RJ-03 Proband F Breast Bilateral 47/47
RJ-04 Proband F Breast Bilateral 33/38
RJ-05 Proband F Breast Unilateral 33
RJ-05A 1st degree relative F Healthy - -
SP-01 Proband F Breast Bilateral 36
SP-01A 2nd degree relative F Breast Bilateral 46
SP-01B 3rd degree relative F Healthy - -
SP-01C 5th degree relative F Healthy - 35
SP-02 Proband F Breast Unilateral 63
SP02-A 1st degree relative F Healthy - -
SP-03 Proband F Breast Bilateral 37
SP-03A 1st degree relative F Breast Unilateral 41
SP-03B 1st degree relative M Healthy - -
SP-04 Proband F Breast Bilateral 31
SP-04A 1st degree relative F Healthy - -
SP-05 Proband F Ovarian - 49
SP-05A 1st degree relative F Healthy - -
SP-05B 1st degree relative F Breast Unilateral 47
SP-05C 1st degree relative F Healthy - -
SP-05D 3rd degree relative M Healthy - -
SP-05E 3rd degree relative F Healthy - -
RS-01 Proband F Breast Bilateral 36/-
RS-01A 3rd degree relative F Ovarian Bilateral 52/54
RS-02 Proband F Breast Bilateral 23/44
RS-03 Proband F Breast Bilateral -
RS-04 Proband F Breast Bilateral 35/45
RS-04A 1st degree relative F Healthy - -
RS-05B 3rd degree relative F Breast Bilateral 49/64

Fourteen probands carrying BRCA1 c.5266dupC and 26 relatives (carriers and non-carriers) were haplotyped. The same haplotype associated with c.5266dupC was segregating within all the families analyzed, revealing no recombinants in a region of 0,68 Mb (Figure S2). On the other hand, this haplotype was not found in non-carrier relatives analyzed (n=7). Ancestry analyses showed that the European component was predominant among the probands, with an average of 81.15% (Figure 1).

Figure 1 Ancestry components of the 14 probands analyzed. Ancestry analysis was performed to investigate the contribution of African (AFR), European (EUR), East Asian (EAS), and Native American (AME) populations in our probands. 

The high frequency of breast cancer observed in our sample, especially bilateral cancer (n=13/21), was also reported in other studies and recently associated to its location (within the breast cancer cluster region of the gene) (Rubinstein, 2004; Hamel et al., 2011; Pritchard et al., 2012; Rebbeck et al., 2015; Tafe et al., 2015). The segregation of the same haplotype within BRCA1 c.5266dupC in all carrier relatives analyzed, reinforced the founder effect of this mutation in the Brazilian population.

Our data is in accordance with previous results showing a European component that exceeds 70% in the South and Southeast of Brazil (Kehdy et al., 2015), and support the European origin of BRCA1 c.5266dupC in Brazil. The predominant European ancestry observed in the carriers studied here is in line with the proposition of Hamel et al. (2011) for the Scandinavian origin of this mutation, followed by its dispersion through Central Europe 400-500 years ago. As was stated before (da Costa et al. , (2008)), a better explanation for the presence of this mutation in the Brazilian population is the immigration from Central Europe diring the 19th century encouraged by Brazilian officials (Pena et al., 2011), particularly to the Southeast and South regions of Brazil (Kehdy et al., 2015), where the mutation is nowadays more frequently found.

There are some limitations to the present study. Unfortunately, the sample size was small, considering that c.5266dupC corresponds to 20.2% of all Brazilian BRCA1 pathogenic variants (Palmero et al., 2018), although, our samples account for patients of three Brazilian states. Nonetheless, it is the largest published study revealing a single haplotype between carriers in Brazil. This unique haplotype validates the founder effect for the c.5266dupC insertion in Brazil, and the ancestry data reveal the contribution of Central Europe for the Brazilian genetic background. The frequency of this mutation is shown to be relevant especially among patients of the southern and utheastern Brazilian regions, where the European ancestry contribution is large. Our study also shows that c.5266dupC is associated with the appearance of bilateral breast tumors, which confirms what was previously observed by other authors (Ewald et al., 2011). Considering a scenario of limited resources, low cost screening focused on this recurrent pathogenic variant could be offered for patients and their families of European ancestry. However, this strategy is not adequate in view of the diversity of pathogenic BRCA1 and BRCA2 variants found in Brazil and the admixed ethnic origin of its individuals.


This work was supported by National Institute for Cancer Control (INCT para Controle do Câncer;, Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq, Brazil; grant numbers: 305873/2014-8 and 573806/2008-0), and Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ, Brazil; grant number: E26/170.026/2008).


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Received: March 01, 2019; Accepted: July 23, 2019

Send correspondence to Miguel Ângelo Martins Moreira. Instituto Nacional de Câncer, Programa de Genética, 20231-050, Rio de Janeiro, RJ, Brazil. E-mail:

Author Contributions

RG and BLS conceived the study, carried out micro-satellite analysis, and participated in writing the manuscript. RM, CBON, BA and PAP contributed with patient samples, family cancer history, and participated in writing the manuscript. PSF and EIP carried out the ancestry analysis and participated in writing the manuscript. MAMM conceived the study, contributed with patient samples, family cancer history, and participated in writing the final version of the manuscript. All authors read and approved the final version.

Conflict of Interest

The authors declare no conflict of interest.

Associate Editor:

Houtan Noushmehr

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