Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins

Abstract

Background: A limiting factor in performing proteomics analysis on cancerous cells is the difficulty in obtaining sufficient amounts of starting material. Cell lines can be used as a simplified model system for studying changes that accompany tumorigenesis. This study used two-dimensional gel electrophoresis (2DE) to compare the whole cell proteome of oral cancer cell lines vs normal cells in an attempt to identify cancer associated proteins.

Results: Three primary cell cultures of normal cells with a limited lifespan without hTERT immortalization have been successfully established. 2DE was used to compare the whole cell proteome of these cells with that of three oral cancer cell lines. Twenty four protein spots were found to have changed in abundance. MALDI TOF/TOF was then used to determine the identity of these proteins. Identified proteins were classified into seven functional categories - structural proteins, enzymes, regulatory proteins, chaperones and others. IPA core analysis predicted that 18 proteins were related to cancer with involvements in hyperplasia, metastasis, invasion, growth and tumorigenesis. The mRNA expressions of two proteins - 14-3-3 protein sigma and Stress-induced-phosphoprotein 1 - were found to correlate with the corresponding proteins' abundance.

Conclusions: The outcome of this analysis demonstrated that a comparative study of whole cell proteome of cancer versus normal cell lines can be used to identify cancer associated proteins.

Figure 1

Representative 2DE gel of whole cell proteome of cell lines. The whole cell protein of normal primary cultures and cancer cell lines were resolved by 2DE pI 4–7, 11% second dimension polyacrylamide gel. Proteins with different abundance are shown as numbered spots. The numbers correspond to Spot Number in Table 1.

Figure 2

Top-scored molecular network with identified proteins implicated in cancer progression according to the IPA software. Proteins in the network are represented by their gene symbols. Green colored shapes denote proteins with lower abundance in oral cancer cell lines, while red colored shapes denote proteins with higher abundance in oral cancer cell lines A) molecule types, B) relationship types.

Figure 3

Results of quantitative RT-PCR showing relative expressions of STMN1, 1433S, STIP1 and GSTP1. Total mRNA was extracted from normal and oral cancer cell lines. Quantitative RT-PCR was then performed as described. A: Relative expression of STMN1, B: Relative expression of 1433S, C: Relative expression of STIP1, D: Relative expression of GSTP1. Relative intensities of all genes of interest were determined by using β-actin as an internal standard. Results represent the mean ± SD for three experiments. *p < 0.05.