Identification and Selective Degradation of Neopeptide-Containing Truncated Mutant Proteins in the Tumors with High Microsatellite Instability

• Published in: Clinical cancer research Vol. 19; no. 13; pp. 3369 - 3382
• Main Authors: Kim, Won Kyu, Park, Misun, Park, Minhee, Kim, Yun Ji, Shin, Nara, Kim, Hyun Ki, You, Kwon Tae, Kim, Hoguen
• Format: Journal Article
• Language: English
• Published: Philadelphia, PA American Association for Cancer Research 01.07.2013
• Subjects: Adult; Aged; Aged, 80 and over; Antineoplastic agents; Biological and medical sciences; Cell Cycle Proteins - genetics; Cell Cycle Proteins - metabolism; Cell Line, Tumor; Centrosome - metabolism; Codon, Nonsense; Colorectal Neoplasms - genetics; Colorectal Neoplasms - metabolism; Colorectal Neoplasms - pathology; Exons; Female; Frameshift Mutation; Gene Expression; Gene Expression Profiling; Humans; Intracellular Signaling Peptides and Proteins - genetics; Intracellular Signaling Peptides and Proteins - metabolism; Male; Medical sciences; Membrane Proteins - genetics; Membrane Proteins - metabolism; Microsatellite Instability; Middle Aged; Multiple tumors. Solid tumors. Tumors in childhood (general aspects); Mutant Proteins - genetics; Mutant Proteins - metabolism; Myristoylated Alanine-Rich C Kinase Substrate; Neoplasm Grading; Neoplasm Staging; Neoplasms - genetics; Neoplasms - metabolism; Pharmacology. Drug treatments; Polyribosomes - metabolism; Proteasome Endopeptidase Complex - metabolism; Protein Transport; Protein-Serine-Threonine Kinases - genetics; Protein-Serine-Threonine Kinases - metabolism; Protein-Tyrosine Kinases - genetics; Protein-Tyrosine Kinases - metabolism; Proteolysis; RNA, Messenger - genetics; RNA, Messenger - metabolism; Transcription Factor 7-Like 2 Protein - genetics; Transcription Factor 7-Like 2 Protein - metabolism; Tumors; Ubiquitination; Degradation; Microsatellite DNA; Truncated shape; Instability; Identification; Mutation; Malignant tumor; Protein; Cancer
• DOI: 10.1158/1078-0432.CCR-13-0684
• ISSN: 1078-0432; 1557-3265

Purpose: Frameshift mutations in coding mononucleotide repeats (cMNR) are common in tumors with high microsatellite instability (MSI-H). These mutations generate mRNAs containing abnormal coding sequences and premature termination codons (PTC). Normally, mRNAs containing PTCs are degraded by nonsense-mediated mRNA decay (NMD). However, mRNAs containing PTCs located in the last exon are not subject to degradation by NMD (NMD-irrelevant). This study aimed to discover whether genes with frameshift mutations in the last exon generate truncated mutant proteins. Experimental Design: We identified 66 genes containing cMNRs in the last exon by bioinformatic analysis. We found frequent insertion/deletion mutations in the cMNRs of 29 genes in 10 MSI-H cancer cell lines and in the cMNRs of 3 genes in 19 MSI-H cancer tissues. We selected 7 genes (TTK, TCF7L2, MARCKS, ASTE1, INO80E, CYHR1, and EBPL) for mutant mRNA expression analysis and 3 genes (TTK, TCF7L2, and MARCKS) for mutant protein expression analysis. Results: The PTC-containing NMD-irrelevant mRNAs from mutated genes were not degraded. However, only faint amounts of endogenous mutant TTK and TCF7L2 were detected, and we failed to detect endogenous mutant MARCKS. By polysome analysis, we showed that mRNAs from genomic mutant MARCKS constructs are normally translated. After inhibiting 3 protein degradation pathways, we found that only inhibition of the proteasomal pathway facilitated the rescue of endogenous mutant TTK, TCF7L2, and MARCKS. Conclusions: Our findings indicate that cancer cells scavenge potentially harmful neopeptide-containing mutant proteins derived from NMD-irrelevant abnormal mRNAs via the ubiquitin–proteasome system, and these mutant proteins may be important substrates for tumor-specific antigens. Clin Cancer Res; 19(13); 3369–82. ©2013 AACR.