The role of NAD(P)H:quinone oxidoreductase 1 in non-small cell lung cancer tumorigenesis and lung cancer stem cell maintenance

摘要

Lung cancer is the leading cause of cancer related deaths worldwide. The five-year survival rates for those patients suffering from non-small cell lung cancer (NSCLC), continues to be an abysmal 15%. One of the major reasons for the poor survival rate amongst NSCLC patients is the lack of early detection and subsequent late stage initial diagnosis. Tumors discovered at later stages are often refractory toward chemotherapy and radiation regimens. One theory as to why tumors become resistant to therapy relies heavily on the cells that make up the cancer stem cell (CSC) niche. This small percentage of cells within the heterogeneous tumor has been reported to be responsible for drug resistance, tumor recurrence, and metastasis. In general, CSCs have been isolated using a number of different markers, including cluster differentiation markers, somatic stem cell markers, as well as a number of functional markers such as the side population and aldehyde dehydrogenase (ALDH) activity. While some cancer types, such as breast and hematologic cancers, have been significantly investigated to identify and define their CSC population, lung cancer researchers have only recently begun to identify CSC markers in lung tumors. In addition to the CSC population, malignant cells can also alter their expression of a number of cytoprotective genes that promote tumorigenesis. NAD(P)H:quinone oxidoreductase 1 (NQO1) is a detoxifying enzyme that has been demonstrated to be highly overexpressed in a number of different malignancies. This overexpression has been utilized as a drug target, as the enzyme is expressed at low levels in normal tissue. To this point, there has been success in using NQO1 as a drug target, however little research has been conducted on understanding why NQO1 is overexpressed in these malignancies. The work presented here investigated the role of NQO1 in tumorigenesis as well as its role in maintaining the CSC population in NSCLC. We demonstrate that NQO1 promotes anchorage-independent growth, invasion, reactive oxygen species regulation, anoikis resistance, proliferation, in vivo tumor growth, survival, and ALDH activity. Secondly, we demonstrate that NQO1 also promotes spheroid formation, both in initial and serial contexts, enhances the CSC frequency, and protects spheroid-cultured cells from chemotherapy. Finally, we provide preliminary data that indicates that NQO1 mRNA may be playing an important signaling role in the promotion of the CSC phenotype. This was demonstrated by CRISPR-Cas9 genetic knockout of NQO1 that resulted in a reemergence of the CSC phenotype that can be reversed with transient knockdown of NQO1 mRNA. In summary, our data demonstrate that NQO1 is playing a vital role in the promotion of NSCLC tumorigenesis, as well as supporting the cancer stem cell population. Interestingly, these results may be due to a novel signaling mechanism by NQO1 mRNA, and not the enzyme itself. Further research will be needed to completely understand the role of NQO1 mRNA in NSCLC tumorigenesis and the CSC phenotype.