Quantitative proteomic analysis of mitochondrial proteins differentially expressed between small cell lung cancer cells and normal human bronchial epithelial cells

Abstract

BackgroundSmall cell lung cancer (SCLC) is highly aggressive and is associated with a dismal prognosis. However, there are no clinically recognized biomarkers for early diagnosis. In this study, we used quantitative proteomics to build differential mitochondrial protein profiles that may be used for early diagnosis and investigated the pathogenesis of lung cancer.MethodsWe cultured SCLC cells (NCI‐H446) and normal human bronchial epithelial cells (16‐HBE); mitochondria were extracted and purified using differential and Percoll density gradient centrifugation. Subsequently, we used Western blot analysis to validate mitochondrial purity and labeled proteins/peptides from NCI‐H446 and 16‐HBE cells using relative and absolute quantification of ectopic tags. We then analyzed mixed samples and identified proteins using two‐dimensional liquid chromatography‐tandem mass spectrometry. Additionally, we performed subsequent bioinformatic proteome analyses using the programs ExPASy, GOA, and STRING. Finally, the relationship between ornithine aminotransferase expression and clinicopathological features in lung cancer patients was evaluated using immunohistochemistry.ResultsOne hundred and fifty‐three mitochondrial proteins were differentially expressed between 16‐HBE and NCI‐H446 cells. The expression of 30 proteins between 16‐HBE and NCI‐H446 cells increased more than 1.3‐fold. The upregulation of ornithine aminotransferase was associated with pathological grade and clinical tumor node metastasis stage.ConclusionOur experiment represented a promising method for building differential mitochondrial protein profiles between NCI‐H446 and 16‐HBE cells. Such analysis may also help to identify novel biomarkers of lung cancer.

Comparative mitochondrial proteomic analysis of human large cell lung cancer cell lines with different metastasis potential

“…We also demonstrated that transfection of wild type nm23‐H1 complementary DNA (cDNA) into human high‐metastatic large cell lung cancer cells (which exhibit loss of heterozygosity [LOH] of nm23‐H1 ), can regulate the expression of metastatic relative genes and reverse the metastatic phenotype of lung cancer cell lines . Our findings and other reports have provided sufficient evidence to indicate that nm23‐H1 is a metastasis suppressor gene in many tumors . Our studies have also proven that the nm23‐H1 gene is a key and upstream regulative gene in the “lung cancer metastatic suppressive cascade.” However, the exact molecular mechanism by which nm23‐H1 suppresses or reverses lung cancer metastasis is unclear.…”

“…Lung cancer metastasis is not only the malignant marker but is the main cause of treatment failure . Metastasis is a complex biological behavior that is correlated with many factors, genes, signal pathways, and biological processes . Therefore, exploration of changes to molecular genetics and cell signal transduction related to invasion and metastasis in lung cancer will not only illuminate the molecular mechanisms of tumor invasion and metastasis, but also provide a new targeting molecule and route for blocking signal transduction and reversing the metastatic phenotype of lung cancer .…”

“…[16][17][18][19][20] Metastasis is a complex biological behavior that is correlated with many factors, genes, signal pathways, and biological processes. [21][22][23][24][25][26] Therefore, exploration of changes to molecular genetics and cell signal transduction related to invasion and metastasis in lung cancer will not only illuminate the molecular mechanisms of tumor invasion and metastasis, but also provide a new targeting molecule and route for blocking signal transduction and reversing the metastatic phenotype of lung cancer. [27][28][29][30][31][32] Our previous studies showed that low expression and hetero-deletion of tumor metastasis suppressor gene nm23-H1 are closely correlated with high metastatic ability and poor prognosis in lung cancer patients.…”

“…[23][24][25][26][27][28][29][30] Our findings and other reports have provided sufficient evidence to indicate that nm23-H1 is a metastasis suppressor gene in many tumors. [21][22][23][24][25][26][27][28][29][30][31][32][33][34][35][36][37] Our studies have also proven that the nm23-H1 gene is a key and upstream regulative gene in the "lung cancer metastatic suppressive cascade." [23][24][25][26][27][28][29][30][31][32][33] However, the exact molecular mechanism by which nm23-H1 suppresses or reverses lung cancer metastasis is unclear.…”

Breast Cancer-Derived Microvesicles Are the Source of Functional Metabolic Enzymes as Potential Targets for Cancer Therapy

“…Moreover, proteomics analysis using two-dimensional differential gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization-time-of-flight-mass spectrometry (MALDI-TOF-MS) of canine mammary tumors, proposed as a model for human breast cancer, identified OAT as one of several upregulated proteins in metastatic carcinomas [ 68 ]. In addition, OAT was found upregulated, 1.3-fold, in small cell lung cancer cell line (NCI-H446) compared to the non-cancerous human bronchial epithelial cell line (16-HBE) [ 69 ]. This enzyme’s expression has been correlated to the pathological grade and clinical tumor metastasis stage in lung cancer patients [ 69 , 70 ].…”