MALDI Imaging and LCMS Identification of Colon Cancer Biomarkers in Benign Polyps and Normal Tandem Mucosa

Abstract

Purpose: MALDI imaging (IMS) demonstrated the same colon tumor proteins, gi| 119592539 and gi| 119592490, in consecutive patients. These proteins were present in the tumors and in normal satellite tissue. Similarly, putative colon cancer proteins were demonstrated in benign polyps. The proteome of normal tandem colon mucosa in patients with polyps or carcinomas is significantly different from that of patients without polyps or carcinomas. This raised several questions, is this evidence of metastatic disease or spread of tumor into normal satellite tissue and does the use of histopathology alone underestimate the extent of potential malignant disease? We hypothesized that histopathology in combination with IMS, may not only identify malignant disease in normal tissue, but also identify patients with field defects that are at risk for polyps and carcinomas. Methods: To test this hypothesis we examined tissue from 25 patients with normal colons at colonoscopy, and compared it with tissue from 25 patients with polyps and carcinomas respectively. Tissue included the tumor or polyp, and tandem proximal and distal normal control tissue. Contiguous histologic sections were obtained for histology (stained with H&E), IMS and protein extraction. MALDI images and protein masses were obtained on a Shimadzu Axima mass spectrometer. The third section was used for high pressure protein extraction and the extract was separated with nanoflow liquid chromatography, trypsinized and processed for protein identification with a nanoflow LCMS, Hitachi NanoFrontier. Results: IMS displayed the loci of proteins in the normal tissue of normal patients, normal tissue in patients with polyps or tumors, and in the polyps and tumors. The extraction experiments confirmed the proteins identified on IMS. There were clear differences in the normal mucosa proteome of the normal patients compared with the patients that had tumors or polyps. Putative carcinoma proteins were again identified in normal polyps. Conclusion: IMS and LCMS can identify protein biomarkers of mucosa at risk, putative carcinoma proteins in histologically normal polyps, and identify those tumors with concordant proteins in carcinomas and tandem normal mucosa.