Hyperphosphorylation of ribosomal protein S6 predicts unfavorable clinical survival in non-small cell lung cancer.
BACKGROUND
Ribosomal protein S6 (rpS6), a component of the 40S ribosomal subunit, is involved in multiple cellular bioactivities. However, its clinicopathological significance in non-small cell lung cancer (NSCLC) is poorly understood.


METHODS
Expressions of total rpS6 (t-rpS6) and phosphorylated rpS6 (Ser235/236, p-rpS6) were detected immunohistochemically in 316 NSCLC tissues and 82 adjacent controls, followed by statistical evaluation of the relationship between proteins expressions and patients' survivals to identify their prognostic values. Cytological experiments with overexpressing or silencing rpS6 by lentivirus in human bronchial epithelial (HBE) and NSCLC cell lines were performed to explore potential mechanisms by which rpS6 affects the clinical development of NSCLC. Additionally, specific RNA interference for Akt1, Akt2, Akt3, Akt inhibitor and subsequent cellular bioactivity tests were employed as well to investigate the upstream regulation of rpS6.


RESULTS
Positive rates of t-rpS6 and p-rpS6 were both significantly increased in NSCLC tissues, compared with controls (82.91 vs 62.20 % for t-rpS6; 52.22 vs 21.95 % for p-rpS6; both P < 0.001). However, only hyperphosphorylation of rpS6, expressed as either elevated p-rpS6 alone or the ratio of p-rpS6 to t-rpS6 (p-rpS6/t-rpS6) no less than 0.67, was greatly associated with the unfavorable survival of NSCLC patients, especially for cases at stage I (all P < 0.001). The independent adverse prognostic value of hyperphosphorylated rpS6 was confirmed by multivariate Cox regression analysis (hazard ratios for elevated p-rpS6 alone and p-rpS6/t-rpS6 no less than 0.67 were 2.403, 4.311 respectively, both P < 0.001). Overexpression or knockdown of rpS6, along with parallel alterations of p-rpS6, led to increased or decreased cells proliferations respectively, which were dependent on redistributions of cell cycles (all P < 0.05). Cells migration and invasion also changed with rpS6 interference (all P < 0.05). Furthermore, upstream overexpression or knockdown of Akt2 or Akt2 phosphorylation inhibition, rather than Akt1 or Akt3, resulted in striking hyperphosphorylation or dephosphorylation of mTOR, p70S6K and rpS6 (all P < 0.05), without any change in total proteins expressions. Further tests showed markedly accompanied variation of cells proliferation, cell cycle distribution and invasion (all P < 0.05).


CONCLUSIONS
Hyperphosphorylation of rpS6, probably regulated by the Akt2/mTOR/p70S6K signaling pathway, is closely relevant to the progression of NSCLC and it might be served as a promising therapeutic target for NSCLC treatment.
Department of Respiratory and Critical Care Medicine, West China Hospital of Sichuan University, No. 37, Guo Xue Street, Chengdu, Sichuan, 610041, China.
Bojiang
B
Chen
Inspectiong and Quarantine Technical Center of Sichuan Entry-Exit Inspection and Quarantine Bureau, Chengdu, China.
Zhi
Z
Tan
Department of Toxicological Inspection, Sichuan Center for Disease Prevention and Control, Chengdu, China.
Jun
J
Gao
Department of Outpatient, West China Hospital of Sichuan University, Chengdu, China.
Wei
W
Wu
Department of Toxicological Inspection, Sichuan Center for Disease Prevention and Control, Chengdu, China.
Lida
L
Liu
Department of Toxicological Inspection, Sichuan Center for Disease Prevention and Control, Chengdu, China.
Wei
W
Jin
Department of Pathology, West China Hospital of Sichuan University, Chengdu, China.
Yidan
Y
Cao
Department of Respiratory and Critical Care Medicine, West China Hospital of Sichuan University, No. 37, Guo Xue Street, Chengdu, Sichuan, 610041, China.
Shuang
S
Zhao
Department of Respiratory and Critical Care Medicine, West China Hospital of Sichuan University, No. 37, Guo Xue Street, Chengdu, Sichuan, 610041, China.|Department of Respiratory Medicine, Second Affiliated Hospital, Chongqing Medical University, Chongqing, China.
Wen
W
Zhang
Department of Respiratory and Critical Care Medicine, West China Hospital of Sichuan University, No. 37, Guo Xue Street, Chengdu, Sichuan, 610041, China.
Zhixin
Z
Qiu
Department of Respiratory and Critical Care Medicine, West China Hospital of Sichuan University, No. 37, Guo Xue Street, Chengdu, Sichuan, 610041, China.
Dan
D
Liu
Laboratory of Stem Cell Biology, West China Hospital of Sichuan University, Chengdu, China.
Xianming
X
Mo
Department of Respiratory and Critical Care Medicine, West China Hospital of Sichuan University, No. 37, Guo Xue Street, Chengdu, Sichuan, 610041, China. weimi003@yahoo.com.
Weimin
W
Li
D000328:Adult; D000368:Aged; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics / Q000378:metabolism* / Q000401:mortality* / Q000473:pathology; D016022:Case-Control Studies; D002453:Cell Cycle; D049109:Cell Proliferation; D005260:Female; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D006801:Humans; D053208:Kaplan-Meier Estimate; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000401:mortality* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D060787:Neoplasm Grading; D009367:Neoplasm Staging; D010766:Phosphorylation; D011379:Prognosis; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D038601:Ribosomal Protein S6 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D047368:Tumor Burden
D034741:RNA, Small Interfering; D038601:Ribosomal Protein S6; C494921:AKT2 protein, human; D051057:Proto-Oncogene Proteins c-akt