Systemic identification of mutated proteins with frame-shifted from MSI positive colon carcinoma

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Colorectal cancers with microsatellite instability (MSI+ CRCs) caused by dysfunction of DNA mismatch repair have unique clinicopathological characteristics including good prognosis with T-cell infiltration in tumor. Previously we demonstrated frame-shifted mutated CDX2, as tumor antigens that induce immune response against MSI+ CRC, by SEREX (serological analysis of recombinant cDNA expression cloning). In this study, the immunogenic proteins with frameshift mutated in the microsatellite of the coding region were screened. We isolated 26 genes from SEREX with two libraries of MSI positive colon cancer tissues and autologus sera, but found no genes reported as MSI target genes and tumor specific genes by RT-PCR. We made recombinant proteins from 8 target genes (TGFBRII, BAX, TCF-4, CASP-5, SEC63, AIM2, OGT, RAD50), which make frame-shifted mutated peptide caused by mismatch repair genes. The presence of IgG antibody against recombinant proteins examined by MSI+ CRCs sera of 13 patients by western blotting and ELISA, but none of them showed positive. However, the tissues with patients had some mutation in the target genes. We reconstructed recombinant protein of TGFBRII, and we performed Western blot analysis using another 4 MSI+ CRCs sera. One serum from patient with TGFBRII mutation reacted with normal and mutated recombinant protein. Therefore, tumor specific peptides generated by MSI may be involved in anti-tumor immune responses, but the abilities of immuno response to proteins with frameshift mutated in MSI may be relatively weak. Further investigation is required to identify the proteins with frameshift mutated in MSI+ CRCs.