An Approach to Studying Lung Cancer-related Proteins in Human Blood

Abstract

Early stage lung cancer detection is the first step toward successful clinical therapy and increased patient survival. Clinicians monitor cancer progression by profiling tumor cell proteins in the blood plasma of afflicted patients. Blood plasma, however, is a difficult cancer protein assessment medium because it is rich in albumins and heterogeneous protein species. We report herein a method to detect the proteins released into the circulatory system by tumor cells. Initially we analyzed the protein components in the conditioned medium (CM) of lung cancer primary cell or organ cultures and in the adjacent normal bronchus using one-dimensional PAGE and nano-ESI-MS/MS. We identified 299 proteins involved in key cellular process such as cell growth, organogenesis, and signal transduction. We selected 13 interesting proteins from this list and analyzed them in 628 blood plasma samples using ELISA. We detected 11 of these 13 proteins in the plasma of lung cancer patients and non-patient controls. Our results showed that plasma matrix metalloproteinase 1 levels were elevated significantly in late stage lung cancer patients and that the plasma levels of 14-3-3 , ␤, and in the lung cancer patients were significantly lower than those in the control subjects. To our knowledge, this is the first time that fascin, ezrin, CD98, annexin A4, 14-3-3 , 14-3-3 ␤, and 14-3-3 proteins have been detected in human plasma by ELISA. The preliminary results showed that a combination of CD98, fascin, polymeric immunoglobulin receptor/secretory component and 14-3-3 had a higher sensitivity and specificity than any single marker. In conclusion, we report a method to detect proteins released into blood by lung cancer. This pilot approach may lead to the identification of novel protein markers in blood and provide a new method of identifying tumor biomarker profiles for guiding both early detection and

Identification of Five Candidate Lung Cancer Biomarkers by Proteomics Analysis of Conditioned Media of Four Lung Cancer Cell Lines

“…Through our method, we successfully identified proteins that are putative or currently used as biomarkers of lung cancer, including CEA (27,28), pro-GRP (29,31), SCC antigen (47,48), tumor M2-PK (49), NCAM (35)(36)(37), chromogranin A (29), and chromogranin B (30). In addition, we identified putative markers previously reported in lung-related proteomics studies such as member C1 of aldo-keto reductase family 1 identified by Huang et al (25) as dihydrodiol dehydrogenase and matrix metallopeptidase 1 found to be overexpressed in lung cancer patients and especially in late stage (18). Furthermore, 129 of 291 extracellular and 168 of 415 membranous proteins identified in this study were found in the plasma proteome.…”

“…Then, we performed literature searches to ensure that these proteins have not been examined as serological markers for lung cancer and showed biological connections with lung or other cancers. We also compared selected proteins with the proteomes of lung-related diseases (lung cancer (18,25,26) and pleural effusion (11)) or the proteome of a lung-related biological fluid (induced sputum (9, 10)) and serum (Plasma Proteome Database). Finally, potential candidates that had commercially available antibodies or immunoassays were selected.…”

“…Eighty microliters of 95% water, 0.1% formic acid, 5% ACN, and 0.02% TFA (buffer A) were added to this mixture, and 40 l were injected via an autosampler on an Agilent 1100 HPLC system. The peptides were first collected onto a 2-cm C 18 trap column (inner diameter, 200 m) and then eluted onto a resolving 5-cm analytical C 18 column (inner diameter, 75 m) with an 8-m tip (New Objective). The HPLC system was coupled on line to a 2D linear ion trap (LTQ, Thermo Inc.) mass spectrometer using a nano-ESI source in data-dependent mode.…”

Heterogeneity of aberrant immunoglobulin expression in cancer cells

“…[25][26][27][28][29] Additionally, high-output proteomics studies revealed that Ig chain fragments were elevated in cancer patients compared with normal healthy controls. [30][31][32] The elevated serum antibody levels were thought to be produced from autoimmune reactions that were stimulated by the disordered immune system. 25,29 Based on our findings, some of these Ig chains could be inferred to arise from the cancer cell itself.…”

Secretory/releasing proteome-based identification of plasma biomarkers in HBV-associated hepatocellular carcinoma

“…Nevertheless, the discrepancy between immortalized cancer cells and clinical samples may be considerable. Therefore, we previously developed a novel approach based on the primary culture of tumor tissues and proteomic analysis of serum-free CM [11,12]. In this study, we collected serum-free CM from cultures of cancerous and surrounding noncancerous tissues from six HCC patients with HBV infection.…”