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Frontiers of Medicine

ISSN 2095-0217

ISSN 2095-0225(Online)

CN 11-5983/R

Postal Subscription Code 80-967

2018 Impact Factor: 1.847

Front. Med.    2018, Vol. 12 Issue (3) : 280-288
Genomic variations in the counterpart normal controls of lung squamous cell carcinomas
Dalin Zhang1, Liwei Qu1, Bo Zhou1,2, Guizhen Wang1, Guangbiao Zhou1()
1. Division of Molecular Carcinogenesis and Targeted Therapy for Cancer, State Key Laboratory of Membrane Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China
2. University of the Chinese Academy of Sciences, Beijing 100049, China
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Lung squamous cell carcinoma (LUSC) causes approximately 400 000 deaths each year worldwide. The occurrence of LUSC is attributed to exposure to cigarette smoke, which induces the development of numerous genomic abnormalities. However, few studies have investigated the genomic variations that occur only in normal tissues that have been similarly exposed to tobacco smoke as tumor tissues. In this study, we sequenced the whole genomes of three normal lung tissue samples and their paired adjacent squamous cell carcinomas. We then called genomic variations specific to the normal lung tissues through filtering the genomic sequence of the normal lung tissues against that of the paired tumors, the reference human genome, the dbSNP138 common germline variants, and the variations derived from sequencing artifacts. To expand these observations, the whole exome sequences of 478 counterpart normal controls (CNCs) and paired LUSCs of The Cancer Genome Atlas (TCGA) dataset were analyzed. Sixteen genomic variations were called in the three normal lung tissues. These variations were confirmed by Sanger capillary sequencing. A mean of 0.5661 exonic variations/Mb and 7.7887 altered genes per sample were identified in the CNC genome sequences of TCGA. In these CNCs, C:G→T:A transitions, which are the genomic signatures of tobacco carcinogen N-methyl-N-nitro-N-nitrosoguanidine, were the predominant nucleotide changes. Twenty five genes in CNCs had a variation rate that exceeded 2%, including ARSD (18.62%), MUC4 (8.79%), and RBMX (7.11%). CNC variations in CTAGE5 and USP17L7 were associated with the poor prognosis of patients with LUSC. Our results uncovered previously unreported genomic variations in CNCs, rather than LUSCs, that may be involved in the development of LUSC.

Keywords lung cancer      counterpart normal control      genomic variations     
Corresponding Authors: Guangbiao Zhou   
Just Accepted Date: 09 October 2017   Online First Date: 29 November 2017    Issue Date: 04 May 2018
 Cite this article:   
Dalin Zhang,Liwei Qu,Bo Zhou, et al. Genomic variations in the counterpart normal controls of lung squamous cell carcinomas[J]. Front. Med., 2018, 12(3): 280-288.
Patient ID Gene Chr Start End Ref Alt Function Transcriptor cDNA position Amino acid
712 C10orf95 10 104210754 104210754 G C Stopgain NM_024886 c.C234G p.Y78X
CNTNAP3 9 39287976 39287976 C T Splicing NM_033655 c.85+ 1G>A
DPPA4 3 109046864 109046864 C A NS NM_018189 c.G886T p.V296F
GLB1L 2 220102416 220102416 T C NS NM_024506 c.A1507G p.I503V
MACF1 1 39913789 39913789 C A NS NM_012090 c.C13876A p.P4626T
NCL 2 232325382 232325384 TCC NF NM_005381 c.807_809del p.E271del
NCOR2 12 124810072 124810072 G A NS NM_006312 c.C7421T p.A2474V
805 IGFN1 1 201178904 201178904 A G NS NM_001164586 c.A4883G p.E1628G
MADCAM1 19 501801 501802 AG CC NS NM_130760 c.[A800C;G801C] p.K267T
ZP3 7 76069902 76069902 G C NS NM_001110354 c.G1034C p.R345T
831 ACAP3 1 1233970 1233970 G T NS NM_030649 c.C840A p.S280R
CEL 9 135947032 135947032 C A NS NM_001807 c.C2152A p.P718T
SLAMF9 1 159923185 159923185 C A NS NM_001146172 c.G305T p.W102L
MUC4 3 195513461 195513461 G A NS NM_018406 c.C4990T p.P1664S
UBE2Q1 1 154530881 154530881 G T NS NM_017582 c.C149A p.S50Y
KDM4B 19 5032981 5032981 A T NS NM_015015 c.A80T p.D27V
Tab.1  Genomic variations in the normal lung tissues of three patients with LUSCs
Fig.1  Validation of genomic variations in normal lung tissues. Genomic variations were identified through the analyses of whole-genome sequencing data. Polymerase chain reaction and Sanger capillary sequencing were performed using the primers listed in Table S1and genomic DNA samples from three patients with LUSC. (A) NCOR2 in the normal lung and tumor samples of a patient with LUSC. Two sets of primers were used. (B) GLB1L in the normal lung and tumor samples of a patient with LUSC. Two sets of primers were used. (C) MACF1 in the normal lung and tumor samples of a patient with LUSC. Two sets of primers were used. (D) C10orf95 in the normal lung and tumor samples of a patient with LUSC. (E) DPPA4 in the normal lung and tumor samples of a patient with LUSC. (F) NCL in the normal lung and tumor samples of a patient with LUSC.
Exonic mutations/MB Mutated genes/sample Nonsynonymous mutations/sample Synonymous mutations/sample Rearrangements/sample
Frameshift Inframe
CNC 0.5661 7.7887 6.9184 4.1339 0.3661 1.0774
Tumor 7.0671 164.8159 152.5272 53.3745 12.2950 3.1088
P value <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001
Tab.2  Somatic mutations in CNCs and the tumor samples of TCGA LUSCs
Fig.2  Genomic variations in the CNCs of 478 patients with LUSC. (A) Frequency of each type of single base substitution in CNC and tumor samples of 478 patients with LUSC. (B) Proportion of nucleotide changes in the CNCs of nonsmokers, current smokers, and reformed smokers. (C) Frequency of base substitution in the CNCs of male and female patients with LUSC. (D) Altered genes in the CNCs of patients with LUSC. CNC samples are arranged from left to right in the top track.
Fig.3  CNC variations in nine representative genes. Schematic representations of proteins encoded by the genes are shown. Numbers refer to amino acid residues. Each “+” corresponds to an independent, mutated CNC sample, and mutations in a nonred “+” with the same color are found in the same patient. (A) Variations in ARSD. (B) Variations in MUC4. (C) Variations in RBMX. (D) Variations in MUC5B. (E) Variations in RP1L1. (F) Variations in CDC27. (G) Variations in MADCAM1. (H) Variations in ANKRD36. (I) Variations in KRTAP5-5.
Fig.4  CNC variations associated with poor patient prognosis. (A) Variations of CTAGE5 in CNC samples and Kaplan–Meier curve for the overall survival of the patients. (B) Variations of USP17L7 in the CNCs and overall survival of the patients.
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