A New Approach Of Detecting Tumor-related Free Protein In Blood

Detection of lung cancer at early stages is the first step towards successful clinical therapy and increased survival. Blood plasma is a most difficult protein-containing sample to characterize on account of the large abundance of albumin and other proteins. Significantly we report here a novel method to detect free proteins released into blood plasma by tumor cells. By this system, the in vivo tumor microenvironment was partially but accurately assessed. Initially the protein components in conditional medium (CM) of primary cell culture or organ culture of lung cancer and adjacent normal bronchus were analyzed by one-dimensional polyacrylamide gel electrophoresis in combination with nano-ESI-MS/MS. In the CM we identified 299 proteins which are involved in cell growth, organogenesis, signal transduction, etc. We selected 13 interesting proteins from this list, demonstrated that they were detectable using ELISA, and further analyzed a series of 628 blood plasma samples. 11 of these 13 proteins were detectable in the plasma of lung cancer patients and controls. Our results showed that plasma levels of MMP1 were significantly elevated in late-stage lung cancer patients. The plasma levels of 14-3-3 sigma, beta and eta in the lung cancer patients were significantly down-regulated compared to control subjects. To our knowledge, this is the first time that fascin, ezrin, CD98, annexin A4, 14-3-3 sigma, 14-3-3 beta, and 14-3-3 eta have been detected in human plasma by ELISA. The preliminary results showed that combination of CD98, fascin, PIGR/SC, and 14-3-3 eta had a higher sensitivity and specificity than any single marker. In conclusion, we report a novel method to detect free proteins released into blood by lung cancer. This 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 therapy of human cancer.