Original Article Diagnostic Genetics
Ann Lab Med 2016;36:463-468 http://dx.doi.org/10.3343/alm.2016.36.5.463
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ISSN 2234-3806 · eISSN 2234-3814
ANNALS OF LABORATORY MEDICINE
Germline TP53 Mutation and Clinical Characteristics of Korean Patients With Li-Fraumeni Syndrome
Kyoung-Jin Park, M.D.1,“,t, Hyun-Jung Choi, M.D.21, Soon-Pal Suh, M.D.2, Chang-Seok Ki, M.D.3, and Jong-Won Kim, M.D.1,3 Department of Health Sciences and Technology1, Samsung Advanced Institute for Health Sciences and Technology, Sungkyunkwan University, Seoul; Department of Laboratory Medicine2, Chonnam National University Medical School and Chonnam National University Hospital, Gwanju; Department of Laboratory Medicine & Genetics3, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea
Background: Little is known of the mutation and tumor spectrum of Korean patients with Li-Fraumeni syndrome (LFS). Owing to the rarity of LFS, few cases have been reported in Korea thus far. This study aimed to retrospectively review the mutations and clinical char- acteristics of Korean patients with LFS.
Methods: TP53 mutation was screened in 89 unrelated individuals at the Samsung Medi- cal Center in Korea, from 2004 to 2015. Six additional mutation carriers were obtained from the literature.
Results: We identified nine different mutations in 14 Korean patients (male to female ra- tio=0.3:1). Two such frameshift mutations (p.Pro98Leufs*25, p.Pro27Leufs*17) were novel. The recurrent mutations were located at codons 31 (n=2; p.Val31Ile), 175 (n=3; p.Arg175His), and 273 (n=4; p.Arg273His and p.Arg273Cys). The median age at the first tumor onset was 25 yr. Ten patients (71%) developed multiple primary tumors. A di- verse spectrum of tumors was observed, including breast (n=6), osteosarcoma (n=4), brain (n=4), leukemia (n=2), stomach (n=2), thyroid (n=2), lung (n=2), skin (n=2), bladder (n=1), nasal cavity cancer (n=1), and adrenocortical carcinoma (n=1).
Conclusions: There was considerable heterogeneity in the TP53 mutations and tumor spectrum in Korean patients with LFS. Our results suggest shared and different LFS char- acteristics between Caucasian and Korean patients. This is the first report on the mutation spectrum and clinical characteristics from the largest series of Korean LFS patients.
Key Words: Li-Fraumeni syndrome, TP53, Germline mutation
Received: February 28, 2016 Revision received: March 31, 2016 Accepted: June 2, 2016
Corresponding author: Jong-Won Kim Department of Laboratory Medicine & Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, 81 Irwon-ro, Gangnam-gu, Seoul 06351, Korea Tel: +82-2-3410-2705 Fax: +82-2-3410-2719 E-mail: kimjw@skku.edu
*Currently affiliated with the Department of Laboratory Medicine, Asan Medical Center, University of Ulsan College of Medicine, Seoul, Korea
+ Equally contributed
@ The Korean Society for Laboratory Medicine
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecom- mons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
INTRODUCTION
Li-Fraumeni syndrome (LFS, OMIM #151623) is an autosomal dominant cancer predisposition syndrome caused by a germ- line mutation in the TP53 tumor suppressor gene [1, 2]. LFS is characterized by an early onset of tumors and a lifetime risk of developing multiple primary tumors, with individuals likely to develop osteosarcoma, soft tissue sarcoma, pre-menopausal breast cancer, brain tumors, adrenocortical carcinoma (ACC), and acute leukemia [1]. These core component tumors com-
prise ~70% of LFS-related tumors. In addition, there is growing evidence that a wide spectrum of cancers, including lung can- cers or stomach cancers, is associated with LFS [3-6].
LFS diagnosis can be confirmed by a genetic mutation in TP53. According to the largest TP53 mutation database, the In- ternational Agency for Research on Cancer (IARC) database (version R18, April 2016, http://p53.iarc.fr/), over 700 germline mutations have been described to date [7]. Although most mu- tations are scattered throughout the gene, several hotspots such as codons 125, 158, 175, 196, 213, 220, 245, 248, 273, 282,
and 337 have been reported [7, 8]. Furthermore, the spectrum of LFS-related TP53 mutations can be categorized into two groups: (1) gain of function mutations (missense mutations) conferring a dominant-negative effect or promoting an onco- genic effect and (2) loss of function mutations such as non- sense and frameshift mutations [9, 10]. A previous study re- ported that missense mutations rather than nonsense or frame- shift mutations are associated with an earlier onset of cancer [8].
Although the IARC database provided essential information pertaining to the mutation distribution and cancer spectrum in LFS, the majority of data originated from Caucasian populations [7]. Currently, little is known about the mutation and tumor spectrum of Korean patients with LFS. Owing to the rarity of LFS, few case reports of Korean patients have been published [11-16]. However, the data presented in these case reports were too limited to assess the mutation spectrum in Korean pa- tients with LFS. We aimed to evaluate the mutation features and clinical characteristics, including the tumor spectrum, of Korean patients with LFS.
METHODS
1. Study subjects
The Institutional Review Board of the Samsung Medical Center in Korea approved this study. This retrospective review was ex- empt from the requirement for informed consent. TP53 muta- tion was screened in 89 unrelated individuals at the Samsung Medical Center from 2003 to 2015. Among them, eight patients were selected as mutation carriers (9%, 8/89). Six additional mutation carriers were identified from case reports published in the literature [11-16]. We collected information on the age at the first tumor onset, sex, familial cancer history, and tumor spectrum from the 14 mutation-positive cases.
2. TP53 mutation analysis
All 11 exons and their flanking intron regions of TP53 were am- plified by PCR using primer pairs designed with the Primer3 software in samples from 85 individuals. In the remaining four individuals, exons 4-9 and their flanking regions were targeted. Sanger sequencing was performed by using the BigDye Termi- nator Cycle Sequencing Ready Reaction Kit (Applied Biosys- tems, Foster City, CA, USA) and an ABI Prism 3100 Genetic An- alyzer (Applied Biosystems). The mutation status was assessed by using the Human Genome Mutation Database (HGMD, pro- fessional version 2014.01) and IARC TP53 database (version R18, April 2016, http://p53.iarc.fr/) [7]. The “A” of the ATG
translation initiation codon is described as position number 1 in TP53 (NM_000546.4). Mutation group and domain were based on previously reported structural data and the Pfam database, respectively [8, 17].
RESULTS
1. Spectrum of germline TP53 mutations
The mutations consisted of seven known mutations and two novel mutations in 14 unrelated Korean patients with 30 tu- mors. Six missense mutations (11/14, 79%), two frameshift de- letions (2/14, 14%), and one nonsense mutation (1/14, 7%) were detected (Table 1). All mutations were distributed among exons 3 to 8; the DNA-binding domain (n=10) and transactiva- tion domain (n=3) (Table 1). The most common mutation loca- tions were exons 8 (n=5), 5 (n=3), and 3 (n=3) in TP53. The recurrent mutations were located at codons 31 (n=2; p.Val- 31Ile), 175 (n=3; p.Arg175His), and 273 [n=4; p.Arg273His (n=3), and p.Arg273Cys (n=1)] in TP53 (Fig. 1). Two novel frameshift mutations (p.Pro98Leufs*25 and p.Pro27Leufs*17) were identified (Table 1, Fig. 2).
2. Clinical characteristics of Korean TP53 mutation carriers The TP53 mutation carriers included three males and 11 fe- males (male to female ratio of 0.3:1). Clinical criteria were ful- filled in 11 patients, with the exception of three patients for whom familial cancer history was not available (Table 1). The median age at the first tumor manifestation was 25 yr, regard- less of the mutation type.
A diverse spectrum of tumors was detected, including breast cancer (n=6), osteosarcoma/rhabdomyosarcoma (n=4/n=1), brain tumor (n=4), leukemia (n=2), stomach cancer (n=2), thyroid cancer (n=2), lung cancer (n=2), skin cancer (n=2), nasal cavity cancer (n=1), and ACC (n=1) (Table 1). Twelve patients (86%) had at least one of the core spectrum cancers (breast cancer, brain tumors, osteosarcoma, rhabdomyosar- coma, and ACC) (Table 1). Ten patients (71%) developed multi- ple primary tumors (range of 1-5). Therapy-related neoplasm was observed in three patients (Patients 1, 3, and 6) (Table 1). Among them, one patient (Patient 3) treated with radiation ther- apy was positive for chromosomal breakage analysis.
One novel mutation, p.Pro98Leufs*25 was observed in pa- tient 6, who was treated for ACC (at the age of 1 yr) and later developed osteosarcoma (4 yr), brain cancer (4 yr), and ther- apy-related neoplasm (10 yr) (Table 1). The other novel muta- tion, p.Pro27Leufs*17 was identified in patient 7 (Table 1). After
| Patient | Age* | Sex | Tumor type (age at diagnosis)* | Criteria | Exon | Domain | Group ** | NT alteration | Mutation type | AA alteration | HGMD | References |
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| 1 | 7 | F | Br (7), Breast (23), Der (35), Leu (35)* | Chompret | 5 | DNA binding | L2 | c.524G>A | missense | p.Arg175His | CM951224 | This study |
| 2 | NA | F | Os (NA) | NA | 5 | DNA binding | L2 | c.524G>A | missense | p.Arg175His | CM951224 | This study |
| 3 | NA | F | Os (NA), Leu*,§ | NA | 8 | DNA binding | S2-S2-H2 motif | c.818G>A | missense | p.Arg273His | CM920677 | This study |
| 4 | 45 | F | Thy (45)", Lm (47) | NA | 3 | Transactivation | NA | c.91G>A | missense | p.Val31Ile | CM074603 | This study |
| 5 | NA | F | Leu (NA), Breast (NA), Thy (NA) | Chompret | 8 | DNA biding | S2-S2-H2 motif | c.817C>T | missense | p.Arg273Cys | CM951233 | This study |
| 6 | 1 | F | ACC (1), Os, (4), Br (4), Leu (10)* | Chompret | 4 | NA | NA | c.293delC | frameshift | p.Pro98Leufs*25 | novel | This study |
| 7 | 28 | F | Breast (28,38)", Nf (33), Nasal cavity (38), Lung (43) | Chompret | 3 | Transactivation | NA | c.78delT | frameshift | p.Pro27Leufs*17 | novel | This study |
| 8 | 26 | M | St (26), Br (26) | LFL | 3 | Transactivation | NA | c.91G>A | missense | p.Val31Ile | CM074603 | This study |
| 9 | 25 | F | Breast (25), Lm (25) | LFL | 7 | DNA binding | L3 | c.742C>T | missense | p.Arg248Trp | CM900211 | [11] |
| 10 | 38 | M | St (38) | LFS | 8 | DNA binding | NA | c.859G>T | nonsense | p.Glu287* | CM044948 | [12] |
| 11 | 24 | F | Breast (25), Br (25) | LFL | 5 | DNA binding | L2 | c.524G>A | missense | p.Arg175His | CM951224 | [13] |
| 12 | 2 | M | Rhabdomyosarcoma (2), Br (4) | LFS | 8 | DNA binding | S2-S2-H2 motif | c.818G>A | missense | p.Arg273His | CM920677 | [14] |
| 13 | 47 | F | Breast (47) | LFL | 6 | DNA binding | L2 | c.566C>T | missense | p.Ala189Val | CM031755 | [15] |
| 14 | 17 | F | Os (17), Lung (17) | LFL | 8 | DNA binding | S2-S2-H2 motif | c.818G>A | missense | p.Arg273His | CM920677 | [16] |
The reference sequence of TP53 was NM_000546.4. Recurrent mutations are in bold. All mutations were heterozygous.
*Age at first tumor diagnosis; “In the order of tumor diagnosis; * Therapy-related myeloid neoplasm; $positive for chromosomal breakage test; “BRAF V600E mutation positive case; “BRCA1 and BRCA2 mutation negative and positive for hormone receptors (estrogen and progesterone) and ERBB2 (formerly HER2/ neu); ** Based on structural data from [8, 17].
Abbreviations: NT, nucleotide; AA, Amino acid; LFS, Li-Fraumeni syndrome; LFL, Li-Fraumeni syndrome like; NA, not applicable, Br; brain; Der, dermatofi- brosarcoma; Leu, leukemia; Os, Osteosarcoma; Thy, thyroid carcinoma; Lm, leiomyoma; ACC, adrenocortical carcinoma; Nf, neurofibroma; St, stomach; HGMD, Human Genome Mutation Database.
A
B
16
337
9
% of germline mutation
14
N of germline mutations
8
12
7
10
6
8
248
273
5
175
4
273
6
4
245
125
3
175
158
213
282
2
2
31
27
98
189
248
287
0
Il
L
1
A
Codon number (IARC TP53 database, R18, April 2016, n=741)
Codon number (Korean patients with Li-Fraumeni syndrome, n= 14)
the age of 28 yr, five primary tumors developed, including left breast cancer (28 yr), neurofibroma (33 yr), right breast cancer (38 yr), right nasal cavity squamous cell carcinoma (38 yr), and EGFR-mutated (p.Leu858Arg) lung adenocarcinoma (43 yr) in
patient 7 (Table 1). The bilateral breast cancers of patient 7 were positive for hormone receptors (estrogen and progester- one) and ERBB2 (formerly HER2/neu) while negative for BRCA1 and BRCA2 mutations.
A
c.293delC (p.Pro98Leufs*25)
G
C
T
C
T
T
G
T C
CT T T
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G
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T
Y
C
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K
R
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M
M
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S
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C
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G
T
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CA
G
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C
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K
R
A
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S
K
I
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1
Y
K
G
T
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T
K
K
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T
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M
R
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C
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G
A
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3
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a
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3
T
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a
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3
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A
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1
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c.78delT (p.Pro27Leufs*17)
G
( T
TG T
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1
DISCUSSION
In this study, we investigated the mutation and tumor spectrums of Korean patients with LFS. Nine different mutations, including seven known mutations and two novel mutations, were identi- fied in 14 unrelated patients. The codon and exon distribution of some mutations (especially codons 175 and 273) in Korean patients was similar to that in Caucasians. A notable exception was codon 337 (p.Arg337His), which was the previously re- ported founder mutation in Brazilian patients [18]. The mutation of p.Arg337His has been reported to be the most common mu- tation in the IARC database, but it was not identified in the Ko- rean patients analyzed in this study.
Furthermore, missense mutations located at the loop, which were demonstrated to bind to the minor groove of DNA (L2: co- dons 164 to 194, L3: codons 237 to 250) and major groove of DNA (L1: codons 115 to 135, S2-S2-H2 motif: codons 273 to
286), were associated with brain cancer and ACC, respectively [8]. Here, we noted that the mutations of two brain cancer carri- ers and four breast cancer carriers were located at loop L2 and L3. However, no association was evident between tumor spec- trum and mutation location, owing to the limited number of cases.
This study reported two novel frameshift mutations located at exons 3 and 4. The mutation locations and types were not pres- ent in the IARC database. The majority of previously reported mutations are missense mutations. In addition, previously re- ported mutations have been located mainly within exons 5-8 that encode the DNA-binding region. Despite the limited num- ber of cases, identification of these frameshift mutations pro- vides further insights into the TP53 mutation spectrum from Ko- rean patients.
The predominance of women among Korean patients was not necessarily associated with breast cancer. A previous study re- ported a significant gender difference, possibly due to the high
incidence of breast cancer among women with LFS [19]. How- ever, in this study, the predominance of women was observed for a variety of tumors, including osteosarcoma and brain tu- mors as well as breast cancer.
There was no significant difference in age at the first tumor diagnosis between mutation types. A previous study reported that missense variants appear to be associated with an earlier onset of cancer [8]. However, this study demonstrated that mis- sense mutations (e.g., p.Val31Ile) were identified in patients with a late-onset of cancer, while frameshift mutations (e.g. p.Pro98Leufs*25) were identified in patients with an early-onset of cancer. The age at tumor onset in TP53 mutation carriers could be influenced by genetic modifiers such as SNP309 (T>G), rs2279744 in MDM2 as well as PIN3 in TP53, together with environmental factors [20, 21]. Moreover, this might be possibly explained by variable expressivity.
There was a considerable heterogeneity in the relationship between TP53 mutations and tumor spectrum. Of note, breast cancer (n=6) was the most common tumor in Korean patients, followed by osteosarcoma/rhabdomyosarcoma (n=5), brain (n=4), and others. Our findings revealed that non-core tumors such as stomach cancer (n=2), lung cancer (n=2), and thyroid cancer (n=2) were recurrently present in Korean patients. A re- cent study reported that stomach cancer is more frequently de- tected in Asian patients compared with Caucasian patients [22]. This suggested differences in the tumor spectrum of LFS be- tween Caucasian and Asian patients [22].
Previous studies reported that, when BRCA mutations are not identified in patients with breast cancer diagnosed before 30 yr, the likelihood of having a TP53 mutation is estimated to be 4-8% [19, 23, 24]. The likelihood could increase with a family history of LFS-related cancers, personal history of a breast can- cer positive for hormone receptors, and/or ERBB2, or additional LFS-related cancers [24, 25]. This is in agreement with the find- ings from patient 7 with bilateral breast cancers.
Lastly, we reported patients with EGFR-mutated (p.Leu858Arg) lung adenocarcinoma (in patient 7) as well as therapy-related neoplasm. To date, little data on EGFR-mutated lung cancer in the context of LFS have been published [5, 6]. It is assumed that patient 7 had a germline TP53 mutation (p.Pro27Leufs*17) as the first genetic hit and a somatic EGFR mutation (p.Leu858Arg) as the second hit.
This study had several limitations. This study was performed by retrospective review in a single institution. The small number of patients as well as incomplete information might have had an impact on the results. In addition, we did not compare clinical
data between TP53 mutation-positive and -negative cases in the current study. Lastly, we did not consider genetic modifiers and environmental factors that influence the clinical phenotype. De- spite these limitations, it is important that these are the first data from the largest case series of Korean patients with LFS.
In summary, this study provides the mutation features and tu- mor spectrum of Korean patients with LFS. There was a consid- erable heterogeneity in TP53 mutations and tumor spectrum among Korean patients. This study suggests shared and differ- ent features of LFS between Caucasian and Korean patients. Further large studies are warranted to stratify the clinical man- agement and to establish surveillance strategies specific for Ko- rean patients with LFS.
Authors’ Disclosures of Potential Conflicts of Interest
No potential conflicts of interest relevant to this article were re- ported.
Acknowledgments
This study was supported by a grant from the Korea Health Technology R&D Project, Ministry of Health and Welfare, Re- public of Korea (A120030).
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