Clin Genet 2015: 87: 392–394
Printed in Singapore. All rights reserved
© 2014 John Wiley & Sons A/S.
Published by John Wiley & Sons Ltd
CLINICAL GENETICS
doi: 10.1111/cge.12451
Letter to the Editor
Identification of a recurrent BRCA1 exon 21-22
genomic rearrangement in Malay breast cancer
patients
To the Editor:
Germline mutations in BRCA1 and BRCA2 are associated with increased risk to breast and ovarian cancer
and their discovery has led to the accurate identification of high-risk individuals. The majority of mutations
identified in BRCA1 and BRCA2 are point mutations,
small deletions and insertions but large genomic rearrangements (LGRs), though rare, also contribute to the
spectrum of disease-causing mutations in BRCA1 and
BRCA2 (1–3).
The frequency of LGRs varies across different populations, from none in French–Canadians, to 1.3% in
Asians and 6.7% in Latin American/Caribbean high-risk
patients (1–6). To date, recurrent LGRs have been identified particularly in Dutch and Hispanic populations and
one recurrent BRCA2 LGR has been reported in two
unrelated Southern Chinese families (5, 7, 8). In this
study, we have analysed the overall prevalence of BRCA1
and BRCA2 genomic rearrangements in a multiethnic
cohort of Malaysian breast cancer patients, characterized
the breakpoints of the rearrangements and report a recurrent LGR found in three unrelated Malay families.
A total of 524 breast cancer patients were selected
for germline analysis on the basis of age of onset and
family history of breast and/or ovarian cancer (4).
Germline analysis was conducted using direct DNA
sequencing and multiplex ligation-dependent probe
amplification (MLPA) using MLPA BRCA1-P002C-1
and BRCA2-P045-B2 kits (MRC-Holland, Amsterdam,
Netherlands). The characterization of genomic rearrangement breakpoints was performed by long-range
polymerase chain reaction (PCR) using a series of
primers (Table S1, Supporting Information) flanking the putative rearrangement regions. Amplified
products of abnormal size were subjected to DNA
sequencing and sequence alignment to the human
genome (GRCh37/hg19) on the UCSC Genome
Browser (http://genome.ucsc.edu) (4). All genomic
rearrangements were annotated using HGVS nomenclature (www.hgvs.org) with the GenBank reference
sequence L78833 and AY436640 for BRCA1 and
BRCA2, respectively.
LGRs account for 7 of the 42 BRCA1 mutation carriers
and 2 of the 38 BRCA2 mutation carriers identified
among the 524 breast cancer patients tested (Table S3).
We mapped the breakpoints of the rearrangements and
392
characterized the mutational mechanisms by analysing
the breakpoints at nucleotide sequence level (Fig. 1).
Two genomic rearrangements in BRCA1 were identified in more than one family (Table 1). First, deletion of
exon 3 was identified in a Punjabi and an Indian family
and long-range PCR analysis showed distinct rearrangement breakpoints between these two families. Second,
deletion of exon 21-22, which removes 43 amino acids
from the C-terminal BRCT domain and is predicted to
lead to BRCT folding defects, was found in three unrelated Malay women. All three women shared the same
Alu-mediated deletion of 3,430 base pairs and a common
haplotype (Table S2). All three women were diagnosed
with early-onset breast cancer (mean age 34.6 years),
of which two were of the triple-negative subtype, but
none reported family history of breast or ovarian cancer (Fig. S1). This recurrent rearrangement accounts for
7.1% (3/42) of the BRCA1 mutations in our study, and
20% (2/10) of the BRCA1 mutations among the Malays.
Our study shows that LGRs in BRCA1 and BRCA2
constitute 17% of BRCA1 carriers and 5% of BRCA2
carriers, and 1.3% and 0.4% respectively of the highand moderate-risk breast cancer patients analysed. In
particular, we report a novel recurrent deletion of exon
21-22 in BRCA1 in Malay families. Unlike previous
studies in Asians, we report that LGRs are significant and
recommend that screening of LGRs should be conducted
together with whole gene sequencing in BRCA1 and
BRCA2 for high-risk patients to ensure comprehensive
detection of mutations.
Supporting Information
Additional supporting information may be found in the online
version of this article at the publisher’s web-site.
Acknowledgements
We thank participants and their families for taking part in this
study; Peter Choon Eng Kang, Sze Yee Phuah, Norhashimah Hassan, Kang In Nee, Katie Laird and Ling Zhi Ni for assistance with
DNA preparation and helpful discussions. This study was funded
by research grants from the Malaysian Ministry of Science, Technology and Innovation, Ministry of Higher Education University
Malaya (UM.C/HlR/MOHE/06), and Cancer Research Initiatives
Foundation.
Letter to the Editor
(a) BRCA1 exon 3 deletion (Case B)
8931
13986
5,056-bp del
Ex. 3
BRCA1 8896
CASE B
BRCA1 13952
(b) BRCA1 exon 3 deletion (Case C)
7693
13259
5,567-bp del
Ex. 3
AluSg
7671-7940
AluSx4
13218-13511
BRCA1 7654
CASE C
BRCA1 13221
(c) BRCA1 exon 21-22 deletion (Case E/F/G)
76870
80299
3,430-bp del
Ex. 21
Ex. 22
AluSg
76803-77098
AluSc
80234-80539
BRCA1 76833
CASE E/F/G
BRCA1 80263
(d) BRCA2 exon 22-24 deletion (Case I)
1,256-bp del/
5-bp ins
65067
Ex. 22
Ex. 23
66322
Ex. 24
AluY
66303-66607
<82-bp>
<1153-bp>
BRCA2 65044
CASE I
Fig. 1. Rearrangement breakpoints and sites of crossover event. (a–d) Schematic representations of breakpoints (arrows) and distribution of Alu
sequences (vertical lines) across the gene fragment. Sequence analyses comparing both normal and mutant alleles show genomic deletion (underlined)
and regions of complete homology (shaded) as possible sites of crossover event.
Table 1. BRCA1 and BRCA2 large genomic rearrangements in Malaysian breast cancer patients
Genomic
rearrangement
BRCA1
Exon 1-14 deletiona
Exon 3 deletion
Exon 13-15 deletiona
Exon 21-22 deletion
Patient
Case A
Case B
Case C
Case D
Case E
Case F
Case G
BRCA2
Exon 14-16 deletiona Case H
Exon 22-24 deletion
Case I
a BRCA1
Region of
rearrangement
Size of alteration
No. of
cases
Not characterized
L78833:g.8931_13986del
L78833:g.7693_13259del
L78833:g.45345_54744del
L78833:g.76870_80299del
–
5,056-bp deletion
5,567-bp deletion
9,400-bp deletion
3,430-bp deletion
1
1
1
1
3
Chinese
Punjabi
Indian
Indian
Malay/Chinese
Malay
Malay
AY436640:g.38656_45514del
AY436640:g.65067_
66322delinsTCAGA
6,859-bp deletion
1,256-bp deletion/
5-bp insertion
1
1
Indian
Chinese
Ancestry
Mechanism
–
Microhomology-mediated
AluSg/AluSx4
AluSx/AluSx
AluSg/AluSc
Microhomology-mediated
Non-homologous end-joining
and BRCA2 rearrangements were previously published in Kang et al. (4).
393
Letter to the Editor
H.N. Hasmada
K. Sivanandana
V. Leea
C.-H. Yipb
N.A. Mohd Taibc,d
S.-H. Teoa,c,d
a Cancer Research Initiatives Foundation, Sime Darby
Medical Centre, Subang Jaya, Malaysia
b Breast Surgery Unit, Sime Darby Medical Centre,
Subang Jaya, Malaysia
c Breast Cancer Research Unit, University Malaya Cancer
Research Institute, Faculty of Medicine, University Malaya
Medical Centre, University Malaya, Kuala Lumpur,
Malaysia
d Department of Surgery, Faculty of Medicine, University
Malaya Medical Centre, University Malaya, Kuala Lumpur,
Malaysia
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BRCA1 and BRCA2: a literature review. Genet Mol Biol 2009: 32:
437–446.
394
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Correspondence:
Dr Soo-Hwang Teo
Cancer Research Initiatives Foundation
2nd Floor, Outpatient Centre
Sime Darby Medical Centre
1 Jalan SS12/1A
47500 Subang Jaya
Selangor, Malaysia
Tel.: +603 5639 1874;
fax: +603 5639 1875;
e-mail: [email protected]