Use of Tissue Microarray and Automated Quantitative Analysis for Screening and Validation of Potential Biomarkers in Mantle Cell Lymphoma

Abstract

Cancer biomarker studies using the combination of tissue microarray and automated quantitative assessment of immunofluorescence (TMA-AQUA) have been successfully performed for various types of human carcinoma, but its performance characteristics have yet to be evaluated in human lymphoma. A pilot TMA was constructed containing duplicate 1.5 mm cores from 15 cases of mantle cell lymphoma (MCL), 3 cases of low-grade B-cell lymphoma, and 3 cases of benign lymphoid tissue. Protein expression of c-Myc, Cdc2, Cyclin D1, Ki-67, Mcm2, and p27 by immunofluorescence and chromagenic staining were evaluated by AQUA and visual scoring, respectively. Gene expression of cMYC, CDC2, and CCND1 was determined by quantitative nuclease protection assay. Protein expression between duplicate cores determined by AQUA showed excellent correlation for all markers [correlation coefficient (R)=0.79 to 0.94] and Cyclin D1 expression was significantly higher in MCL cases compared with non-MCL cases (P=0.00019). Overall correlation of AQUA with scoring of chromagenic staining by 2 pathologists was good for all markers (R=0.56 to 0.90), except Cdc2 (R=0.25). Localization of expression to cytoplasmic and/or nuclear compartments was comparable with chromagenic staining patterns for all markers except Ki-67 and Mcm2, where a significant difference between nuclear and cytoplasmic expression could not be appreciated by AQUA, despite clear nuclear localization by chromagenic staining. Correlation of gene expression with protein expression was variable for CDC2, cMYC, and CCND1 (R=0.32, 0.35, and 0.69). TMA-AQUA has the potential to be successfully used as a high-throughput protein biomarker screening platform for MCL; however, appropriate target protein selection and antibody performance validation are factors that need to be considered.