# Inosine alleviates colorectal cancer liver metastasis by promoting M1 macrophage polarization and modulating the PI3K/AKT signaling pathway

## TL;DR

Inosine reduces colorectal cancer liver metastasis by promoting M1 macrophage polarization and affecting the PI3K/AKT pathway.

## Contribution

This study identifies inosine as a novel agent that inhibits liver metastasis by modulating macrophage polarization and the PI3K/AKT pathway.

## Key findings

- Inosine suppresses CRC cell invasion and enhances M1 macrophage polarization.
- Inosine reduces hepatic metastasis and modulates cytokine levels in a mouse model.
- Transcriptomic analysis reveals inosine's effect on genes and the PI3K/AKT pathway.

## Abstract

Colorectal cancer (CRC) represents one of the most common malignancies of the digestive tract, with colorectal cancer liver metastasis (CRLM) constituting a major cause of CRC-related mortality, for which therapeutic options remain limited.

This study examined the inhibitory role of inosine in CRLM and its molecular mechanisms using in vitro and in vivo approaches. CRC cell invasion was measured in transwell assays following direct inosine treatment or incubation with conditioned medium from inosine-exposed macrophage co-cultures. A CRLM mouse model generated by intrasplenic MC-38 injection was evaluated for hepatic metastasis. Histopathological examination of liver tissues was performed using HE staining to evaluate metastatic infiltration. Furthermore, flow cytometry and ELISA were utilized to analyze macrophage polarization markers and cytokine expression levels. Through transcriptomic sequencing and analysis, potential target genes and signaling pathways were predicted and subsequently validated by RT-qPCR.

Inosine directly suppressed the invasive abilities of MC-38, CT-26, and HT-29 cells. This inhibitory effect on MC-38 cells was further enhanced under co-culture conditions with M1-polarized macrophages. In vivo, inosine reduced the formation of hepatic metastatic nodules, downregulated TNF-α and IL-1β, increased M1 macrophage proportions, and suppressed M2 polarization. Transcriptomics and RT-qPCR indicated that inosine upregulates CYP26A1 and CYP39A1, downregulates BCL-2, PTGS2, and OLFM1, and modulates PI3K/AKT signaling pathway. Inosine inhibits the activation of the PI3K/AKT signaling pathway.

Inosine exerted an inhibitory effect on colorectal cancer liver metastasis by skewing macrophages toward an M1 phenotype, dampening pro-inflammatory cytokine release, and regulating key genes via the PI3K/AKT pathway.