| Background and AimLung cancer is the most common malignancy in our country and the world wide. The incidence and mortality of lung cancer lead the first of all tumors. Chemotherapy is the most important strategy for tumor management. Both impactive factor and targets proteins of cancer chemotherapy are long of great concern. Gene abnormal expressing in tumors may be involved with the key points in oncogenesis and development of tumor, and have impacts on status of other factors and pathways associated with tumor. The abnormality of such gene may have important prognostic significance for tumor management and chemotherapy, and itself could become potential target for chemotherapy as well.eIF3a, the biggest subunit of eukaryotic translation initiation factor3, was found over-expressing in a variety kinds of tumors. It is known for us that eIF3a was involved in the regulation of translation and differentiation, and was closely related to the cell cycle. However, the mechanism and significance of its over-expression in lung cancer and other tumors are unclear. Our group had conducted preliminary exploration on the expression status of eIF3a in lung cancer and its molecular mechanisms as well, we found over-expression of eIF3a improved chemo-responses of lung cancer patients to platinum-based chemotherapy, and we confirmed that eIF3a inhibited the NER pathway in translational level to regulate DNA repairs in vitro. So expression of eIF3a might be indicative for a sensitive phenotype of NSCLC. Herein, the underlying hypotheses of these studies were the expression of eIF3a might predict clinical benefits due to increase of cisplatin sensitivity in NSCLC. On the other hands, there were some reports on the factor and signaling pathways associated with eIF3a, for example:p27/Kipl and ERK pathway. What is more, most studies in vitro suggested the over-expression of eIF3a promote malignant phenotype as well as resistance to chemotherapy. Therefore, the prognostic value of eIF3a is still not clear, as it seems participate in both the protection from and induction of oncogenesis.Non-small cell lung cancer (NSCLC), the main types of lung cancer, accounts for85%of all lung cancers. Chemotherapy improves survival in patients with NSCLC. We wish to confirm the correlation of eIF3a with its related factors and pathways in NSCLC tissues. We also design retrospective study on radically resected NSCLC patients, to investigate the clinical significance of eIF3a, as well as its associated factors, in predicting the prognosis of NSCLC and chemotherapy. Mimosine, a natural pyridone containing α-amino acid structure, can inhibite eIF3a expression and protein synthesis, and is commonly used in cell synchronization, reversibly arresting cells in late G1phase. We postulated eIF3a inhibitors may produce specific anti-tumor proliferative activity. What is more, eIF3a inhibitor might be lead compounds in the study on functions of eIF3a and its correlation with tumors. In our study, we plan to screeen derivative compounds of pyridones with the activities to inhibite eIF3a and anti-proliferation of tumor cells. MethodsBased on the above background, This research intends to use the following methods to achieve the purposes as followed:(1) preparate prokaryotic protein of eIF3a and its polyclonal antibodies;(2) investigate the expression level of eIF3a and related signaling pathways in NSCLC tissues by immunohistochemical methods, analysize the degree of correlation betwwen eIF3a and related factors with Spearman rank correlation coefficient method, analysize the correlation between the biomarkers and clinical parameters such as demographic data and pathologic characteristics using the chi-square test;(3) collect clinical information of complete surgical resection of NSCLC and Follow up of survival, analysize prognostic significance of eIF3a and its related factors in signaling pathways in radically resected NSCLC and postoperative chemotherapy with Kaplan-Meier method and Cox regression model;(4) modify the side chains of lead compound (pirfenidone) and synthesis new derivatives. Investigate the anti-proliferative activities of new derivatives on lung cancer cell A549, which.express high level eIF3a, and lung fibroblast cell NIH3T3, which express low level eIF3a. Validate the screening results in protein and RNA level. ResultsWe got following results in this research:(1) we obtained prokaryotic expression vector for eIF3a C-terminal and its polyclonal antibodies with high titer;(2) the subcellular expression of eIF3a was strongly correlated with status of p27/Kip1(Spearman rank coefficient correlation for cytoplasmic eIF3a and p27/Kipl=0.653, for nuclear staining=0.716). Expression of p-ERK was significantly lower in NSCLC tissues:the positive expression was localized spotty; relationship of p-ERK/ERK with eIF3a expression levels in the tissues was not yet evaluated. Expression of eIF3a was higher in well differentiated squamous cell carcinoma tissues; p27/Kip1highly expressed in well differentiated in NSCLC;(3) In537radically resected NSCLC patients, eIF3a and p27/Kipl expressed in the nucleus has a positive meaning on NSCLC survival. Survival analysis revealed favorable prognostic impact of nuclear eIF3a, p27, and the combination high nuclear staining on NSCLC (Hazards Ratio=0.360,95%CI=0.109-0.782, P=0.028). Chemotherapy prognostic analysis showed high expression of cytoplasmic eIF3a has predictive value for chemotherapy in stage II NSCLC patients. Moreover, In addition, interaction research between biomarkers and chemotherapy status disclosed cisplatin-based regimen trend to prolong DSS of stage II NSCLC patients with high eIF3a-C (P=0.036) and low p27-N (P=0.031);(4)Some1-substituted phenyl-5-phenoxymethyl-2-pyridones with similar Y-type structure synthesized had been improved to inhibite eIF3a and proliferation of lung cancer cell. The two aniline series exhibited significant activities on inhibition of eIF3a and anti-proliferation on A549but not NIH3T3. ConclussionWe arrived following conclusions in the study:(1) the expression status eIF3a and p27/Kipl were significantly related in NSCLC tissues;(2) eIF3a and p27/K.ipl expression in different states NSCLC tissues and their relationships, could help predict mid-term prognosis in patients with completely resected NSCLC, as well as predict the possible benefit of postoperative chemotherapy;(3)The derivative pyridones with similar structure of mimosine could inhibite the express of eIF3a and the proliferation of lung cancer cell significantly, while it does not restrain the growth of lung fibroblast cells. Such derivatives are potential anti-tumor lead compounds and merits intensive development in future. Innovation pointsWe prepared recombinant protein of C terminal in eIF3a and the corresponding antibody; we histopathology confirmed the close relationship eIF3a with p27/kipl and provide the latest clinical evidence for prognostic and predictive significance of eIF3a for lung cancer, we screened derivatives inhibiting eIF3a and tumor growth, provided novel ideas for therapy of lung cancer. |
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