Overexpression of OLC1, cigarette smoke, and human lung tumorigenesis.
BACKGROUND
Exposure to cigarette smoke is a major risk factor for lung cancer, but how it induces cancer is unclear. The overexpressed in lung cancer 1 (OLC1) gene is one of 50 candidate lung cancer genes identified by suppression subtractive hybridization as having higher expression in squamous cell carcinoma (SCC) than normal lung epithelia.


METHODS
We used immunohistochemistry (IHC) to measure OLC1 protein levels in primary lung cancer samples from 559 patients and used fluorescence in situ hybridization to measure OLC1 copy number in primary SCC samples from 23 patients. We compared OLC1 protein expression in SCC samples of 371 patients with and without a smoking history using the Pearson chi(2) test. We assayed OLC1 protein levels by immunoblotting in H1299 human lung cancer cells, immortalized human bronchial epithelial cells, and primary cultured normal human bronchial epithelial cells that were treated with cigarette smoke condensate. We assayed tumor formation in athymic mice using NIH3T3 mouse fibroblast cells transfected with OLC1 (eight mice) and analyzed apoptosis and colony formation of H1299 and H520 lung cancer cells transfected with scrambled (negative) or OLC1 small interfering RNAs (siRNAs) (s1).


RESULTS
OLC1 protein was overexpressed in 387 of 464 (83.4%) of primary lung cancers, as detected by IHC, and OLC1 was amplified in 14 of 23 (60%) of SCC samples. OLC1 protein overexpression was more common in SCC patients with a smoking history than those without (77.1% vs 45.8%, P < .001). In addition, cigarette smoke condensate increased OLC1 protein levels in H1299 cells, immortalized human bronchial epithelial cells, and primary cultured normal human bronchial epithelial cells. Overexpression of OLC1 induced tumor formation in athymic mice (control vs OLC1, 0% vs 100%). Knockdown of OLC1 increased apoptosis (mean percentage of apoptotic H1299 cells, s1 vs negative: 30.3% vs 6.4%, difference = 23.9%, 95% confidence interval [CI] = 19.1% to 28.5%, P = .002; mean percentage of apoptotic H520 cells, s1 vs negative: 21.6% vs 4.9%, difference = 16.7%, 95% CI = 10.6% to 22.8%, P = .007) and decreased colony formation (mean no. of colonies of H1299 cells transfected with siRNAs, negative vs s1: 84 vs 4, difference = 80, 95% CI = 71 to 88, P < .001; mean no. of colonies of H520 cells transfected with siRNAs, negative vs s1: 103 vs 24, difference = 79, 95% CI = 40 to 116, P = .005).


CONCLUSIONS
OLC1 is a candidate oncogene in lung cancer whose expression may be regulated by exposure to cigarette smoke.
Yuan
Jingsong
J
Department of Chemical Etiology and Carcinogenesis, Cancer Institute & Hospital, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing, People's Republic of China.
Ma
Jinfang
J
Zheng
Hongwei
H
Shi
Taiping
T
Sun
Wenyue
W
Zhang
Qiao
Q
Lin
Dongmei
D
Zhang
Kaitai
K
He
Jie
J
Mao
Yousheng
Y
Gao
Xia
X
Gao
Peng
P
Han
Naijun
N
Fu
Guobin
G
Xiao
Ting
T
Gao
Yanning
Y
Ma
Dalong
D
Cheng
Shujun
S
D000230:Adenocarcinoma / Q000209:etiology* / Q000235:genetics; D000818:Animals; D018288:Carcinoma, Small Cell / Q000209:etiology* / Q000235:genetics; D002294:Carcinoma, Squamous Cell / Q000209:etiology* / Q000235:genetics; D049109:Cell Proliferation; D003001:Cloning, Molecular; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D015151:Immunoblotting; D017404:In Situ Hybridization, Fluorescence; D008175:Lung Neoplasms / Q000209:etiology* / Q000235:genetics; D051379:Mice; D008819:Mice, Nude; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells; D014411:Neoplastic Stem Cells; D015513:Oncogene Proteins / Q000378:metabolism*; D009857:Oncogenes; D010957:Plasmids; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012307:Risk Factors; D014028:Tobacco Smoke Pollution / Q000009:adverse effects*; D015854:Up-Regulation
C533705:IST1 protein, human; D015513:Oncogene Proteins; D014028:Tobacco Smoke Pollution