RT-qPCR analysis of the tumor antigens TOMM34 and RNF43 in samples extracted from paraffin-embedded specimens of colorectal cancer

Abstract

Comprehensive genetic analysis of colorectal malignant tumors by microarrays has identified translocase of the outer mitochondrial membrane 34 (TOMM34) and ring finger protein 43 (RNF43) as highly expressed oncogenes in malignant colorectal tumors. Vaccine therapy using cancer peptides synthesized using the amino acid sequences of tumor antigens is currently undergoing clinical trials. Since it is important to perform vaccine therapy based on the oncogene expression levels in individual tumors, analysis of tumor antigen expression is necessary for this therapy. However, the quality of the messenger RNA extracted from formalin-fixed and paraffin-embedded specimens is generally considered insufficient for gene quantification. The present study examined whether it could be possible to quantify the expression of TOMM34 and RNF43 in colorectal cancer and liver metastasis samples prepared from paraffin blocks. The formalin-fixed and paraffin-embedded specimens were sliced for slides and the colorectal cancer and normal mucosal tissues were obtained from the slides. Total RNA was extracted from the tissue samples, and quantitative polymerase chain reaction (qPCR) was performed using the Universal ProbeLibrary as a PCR probe. Quantification of TOMM34 and RNF43 gene expression in several-year-old paraffin-embedded colorectal cancer specimens was possible by qPCR using the Universal ProbeLibrary. qPCR revealed that TOMM34 expression was elevated in 78.9% (15 of 19 cases) of the primary tumors and in 73.7% (14 of 19 cases) of the liver metastasis samples. RNF43 expression was elevated in 63.2% (12 of 19 cases) of the primary tumors and in 73.7% (14 of 19 cases) of the liver metastasis samples.