Effect of Wnt inhibition drug treatment on chemoresistant colorectal cancer

Abstract

Colorectal cancer (CRC) is one of the most commonly diagnosed cancers worldwide, in addition to being responsible for a significant portion of cancer-related deaths. It arises as a result of mutations to genes such as the Adenomatous Polyposis Coli (APC), which aberrantly activate the canonical and non-canonical Wnt signaling pathways, resulting in promoted cell growth. In addition to this, chemoresistance is an additional factor which affects cellular growth by making the cells unresponsive to treatment, allowing them to continue proliferating. Thus, there is a need to investigate potential treatments which can counteract these adverse effects. In this study, two PORCN inhibitors, LGK974 and ETC-159, at concentrations of 10M and 0.2M, respectively, were used to treat parental and 5-FU resistant CRC cell lines bearing either wild-type APC or an APC truncation mutation. Presto Blue viability assays were conducted over 72 hours to assess cell viability in response to treatment. Scratch migration and transwell invasion assays were performed to study the drugs' effects on migration and invasion. Western blotting was then carried out for - catenin, CDC42, EGFR, and mono- and tri-methyl lysine. Lastly, an ELISA was used to quantify total and phosphorylated EGFR in response to treatment. Results indicate that daily treatment with the Wnt inhibitors can reduce CRC cell viability, irrespective of APC mutation or chemoresistance status. Results from the migration and invasion assays continue to reveal the complexity of the canonical and non-canonical Wnt pathways with decreases and increases in migration and invasion rate observed. While Western blotting did not reveal any significant differences in the protein expression of -catenin, CDC42 and EGFR, results for mono- and tri-methyl lysine were more interesting and suggest that APC truncation and chemoresistance status affect these modifications to some extent. This revealed lysine methylation as a potential diagnostic or prognostic biomarker for CRC treatment using LGK974 and ETC-159, although further studies are warranted to properly reveal the underlying mechanisms which link these characteristics. APC truncation status and chemoresistance were also revealed to potentially be playing some role in the expression of total and phosphorylated EGFR from the results obtained by the ELISA.