Identification of Binding Targets of a Pyrrole-Imidazole Polyamide KR12 in the LS180 Colorectal Cancer Genome

• Published in: PloS one 2016-10, Vol.11 (10), p.e0165581-e0165581
• Main Authors: Lin, Jason, Hiraoka, Kiriko, Watanabe, Takayoshi, Kuo, Tony, Shinozaki, Yoshinao, Takatori, Atsushi, Koshikawa, Nobuko, Chandran, Anandhakumar, Otsuki, Joe, Sugiyama, Hiroshi, Horton, Paul, Nagase, Hiroki
• Format: Article
• Language: English
• Subjects: Alkylation; Analysis; Anticancer properties; Antifungal agents; Antineoplastic Agents, Alkylating - chemistry; Antineoplastic Agents, Alkylating - metabolism; Antineoplastic Agents, Alkylating - pharmacology; Artificial intelligence; Binders; Binding Sites; Biological effects; Biology and Life Sciences; Cancer; Cancer genetics; Care and treatment; Cell Line, Tumor; Cell Survival - drug effects; Chemistry; Chromatin; Chromatin Immunoprecipitation; Colorectal cancer; Colorectal carcinoma; Colorectal Neoplasms - genetics; Colorectal Neoplasms - metabolism; Comparative analysis; Computer applications; Deoxyribonucleic acid; DNA; DNA - chemistry; DNA - genetics; DNA - metabolism; DNA microarrays; Gene expression; Gene Expression Regulation, Neoplastic - drug effects; Gene sequencing; Genes; Genetic aspects; Genome, Human; Genomes; Genomics; Health aspects; High-Throughput Nucleotide Sequencing; Histones - metabolism; Humans; Imidazole; Imidazoles - chemistry; Imidazoles - metabolism; Imidazoles - pharmacology; Laboratories; Medicine and Health Sciences; Mutation; Nucleosomes - metabolism; Nucleotide Motifs; Nylons - chemistry; Nylons - metabolism; Nylons - pharmacology; Polyamide resins; Polyamides; Protein Binding; Proteins; Proto-Oncogene Proteins p21(ras) - genetics; Research and Analysis Methods; Sequence Analysis, DNA; Target recognition; Thiophene; Workflow
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0165581

Pyrrole-imidazole polyamides are versatile DNA minor groove binders and attractive therapeutic options against oncological targets, especially upon functionalization with an alkylating agent such as seco-CBI. These molecules also provide an alternative for oncogenes deemed "undruggable" at the protein level, where the absence of solvent-accessible pockets or structural crevices prevent the formation of protein-inhibitor ligands; nevertheless, the genome-wide effect of pyrrole-imidazole polyamide binding remain largely unclear to-date. Here we propose a next-generation sequencing-based workflow combined with whole genome expression arrays to address such issue using a candidate anti-cancer alkylating agent, KR12, against codon 12 mutant KRAS. Biotinylating KR12 enables the means to identify its genome-wide effects in living cells and possible biological implications via a coupled workflow of enrichment-based sequencing and expression microarrays. The subsequent computational pathway and expression analyses allow the identification of its genomic binding sites, as well as a route to explore a polyamide's possible genome-wide effects. Among the 3,343 KR12 binding sites identified in the human LS180 colorectal cancer genome, the reduction of KR12-bound gene expressions was also observed. Additionally, the coupled microarray-sequencing analysis also revealed some insights about the effect of local chromatin structure on pyrrole-imidazole polyamide, which had not been fully understood to-date. A comparative analysis with KR12 in a different human colorectal cancer genome SW480 also showed agreeable agreements of KR12 binding affecting gene expressions. Combination of these analyses thus suggested the possibility of applying this approach to other pyrrole-imidazole polyamides to reveal further biological details about the effect of polyamide binding in a genome.