Research On The Anti-lung Cancer Effect And Molecular Mechanism Of KRT7-AS

Tumorigenesis often accompanies with the activation of oncogenes and inactivation of tumor suppressor genes.In the past 40 years,the oncogenes have been extensively studied,and more than 400 oncogenes have been identified.However,the study on tumor suppressor genes is relatively insufficient.Tumor suppressor genes also play important roles in the occurrence and development of lung cancer,such as the tumor suppressor genes P53 and PTEN.PTEN expression is negatively correlated with the poor prognosis of lung cancer patients.In recent years,more than 8,000 long-chain non-coding RNAs(lncRNAs)have been successfully discovered,but little is known about their functions.In tumors,lncRNA can interact with various big biological molecules such as DNA,RNA,and protein,thereby affecting tumor development.However,the function of most lncRNAs in tumors is currently unknown.LncRNA provides new opportunity for us to study cancer.In this study,firstly,we use bioinformatics analysis methods to discover new tumor suppressor genes from the cancer genome atlas(TCGA),focusing on the analysis of the relationship between lncRNA expression and tumorigenesis.The data shows that among 33 human malignant tumor tissues,compared with normal tissues,lncRNA Keratin 7-Antisense RNA 1(KRT7-AS)shows significantly low expression in 17 types of malignant tumors,among them lung cancer tissues have the highest expression difference compared to the normal tissues.However,the impact of KRT7-AS on the tumorigenesis of the most malignant tumors including lung cancer has not been reported.Accordingly,we raise the scientific questions,does KRT7-AS play a role in lung cancer development?Could it act as a tumor suppressor gene to inhibit the occurrence and development of lung cancer?We hypothesize that KRT7-AS functions as a tumor suppressor gene in lung cancer to inhibit the occurrence and development of lung cancer.Next,we established the KRT7-AS either overexpressed or silenced lung cancer cell lines to explore the effect of KRT7-AS on lung cancer cell phenotypes,such as tumorigenicity,motility,drug sensitivity,and tumor growth in tumor-bearing mice,and we also studied the molecular mechanism of KRT7-AS in lung cancer cell tumorigenicity in vitro and in vivo in the three aspects as follows.We reveal that:(1)KRT7-AS is significantly and lowly expressed in clinical lung cancer tissue samples and lung cancer cell lines as well,and its expression levels are 5-fold reduction in lung cancer tissues as compared to the normal tissues.(2)The transcription factor RXRa could bind to the KRT7-AS gene core promoter region"GGTCA" to activate the transcription of KRT7-AS.(3)In vitro studies show that the KRT7-AS-overexpressed lung cancer cells have a 3.2-fold reduction in clonal numbers,indicating that KRT7-AS overexpression significantly inhibits the tumorigenicity of lung cancer cells.In vivo tumor bearing mice,KRT7-AS overexpression significantly suppresses tumor growth,the tumor inhibition rate is 53%.Together,these data suggest that KRT7-AS is tumor suppressor gene.(4)The bioinformatics analysis of DNA microarray chip results and cell signal pathways,and combined with the results from experiments below,indicate that PTEN protein is the main target molecule of KRT7-AS in the regulation of tumorigenicity.(5)Based on the discovery of KRT7-AS target protein PTEN,and PTEN is involved in the regulation of DNA repair and chemotherapy drug sensitization,we test the effect of KRT7-AS on the Cisplatin(clinical chemotherapy drug)sensitivity in lung cancer cells,the results show that KRT7-AS significantly enhances the sensitivity of lung cancer cells to cisplatin.(6)The molecular mechanistic study shows that KRT7-AS increases the levels of Cleaved-Caspase 3,?-H2AX,and other apoptosis-related proteins in a cisplatin-dependent manner.(7)Further mechanistic study shows that KRT7-AS regulates the ubiquitination-proteasome degradation of PTEN protein,and reveals that the binding of PTEN protein to KRT7-AS fragment 3 in lung cancer cells.In summary,KRT7-AS acts as a tumor suppressor to reduce the tumorigenicity of lung cancer cells in vitro and to inhibit tumor growth in vivo tumor bearing mice mainly through elevation of PTEN protein levels and an increase in the sensitivity of lung cancer cells to cisplatin.We firstly uncover that KRT7-AS is a new tumor suppressor gene in lung cancer.Our study provides new ideas for anti-lung cancer study,offer a new biomarker for the diagnosis of lung cancer and a novel target for development of therapeutics against lung cancer.Additionally,our findings also provide new strategies for sensitizing the lung cancer cells to the anticancer drug cisplatin and to reduce the toxicity and side effects of cisplatin.Therefore,this study has theoretical significance and potential application value.