5-hydroxymethylcytosine features of portal venous blood predict metachronous liver metastases of colorectal cancer and reveal phosphodiesterase 4 as a therapeutic target

Abstract Metachronous liver metastases (MLM) are characterized by high incidence and high mortality in clinical colorectal cancer treatment. Currently traditional clinical methods cannot effectively predict and prevent the occurrence of metachronous liver metastasis in colorectal cancer. Based on 5hmC-Seal analysis of blood and tissue samples, this study found that portal venous blood was more relevant to tumor gDNA than peripheral blood. We performed a novel epigenetic liquid biopsy strategy using the 10 5hmC epigenetic alterations, to accurately distinguish MLM patients from patients without metastases. Among these epigenetic alterations, phosphodiesterase 4 (PDE4D) was highly increased in MLM patients and correlated with poor survival. Moreover, our studies demonstrated that PDE4D was a key metastasis-driven target for drug development. Interfering with the function of PDE4D significantly repressed liver metastases. Similarly, roflumilast, a PDE4 inhibitor approved for Chronic obstructive pulmonary disease (COPD) also inhibits liver metastases. Roflumilast treatment significantly inhibited the protein levels of HIF-1α, CCN2, p-AKT and p-ERK, thus suppressing metastases. To develop a more efficient PDE4D inhibitor and reduce the occurrence of adverse events, we also designed several new compounds based on 2-arylbenzofurans and discovered lead L11 with potent affinity for PDE4D and significant suppression of liver metastases. Interestingly, we further found that compound L11 was better tolerated in terms of emetic activity on beagle dogs than roflumilast. Summarily, our study provides a promising strategy for predicting metachronous liver metastasis and discovers roflumilast as a potential repurposed drug for inhibiting liver metastasis, which have the potential to benefit patients with CRC in the future.