Tumor-associated macrophages (TAMs) and cancer-associated fibroblasts (CAFs) are known to play supportive roles in tumor development and progression, but their interactions in colorectal cancer (CRC) remain unclear. Here, we investigated the effects of colon-cancer-derived CAFs on TAM differentiation, migration, and tumor immunity, both in vitro and in vivo. When co-cultured with monocytes, CAFs attracted monocytes and induced their differentiation into M2 macrophages. Immunohistology of surgically resected human CRC specimens and orthotopically transplanted mouse tumors revealed a correlation between numbers of CAFs and numbers of M2 macrophages. In a mouse model of CRC orthotopic transplantation, treatment with an inhibitor of the colony-stimulating factor-1 receptor (PLX3397) depleted M2 macrophages and increased CD8-positive T cells infiltrating the tumor nest. While this treatment had a minor effect on tumor growth, combining PLX3397 with anti-PD-1 antibody significantly reduced tumor growth. RNA-seq following combination therapy showed activation of tumor immunity. In summary, CAFs are involved in the induction and mobilization of M2 macrophage differentiation in the CRC tumor immune microenvironment, and the combination of cancer immunotherapy and PLX3397 may represent a novel therapeutic option for CRC.

Inhibition of WEE1, a key regulator of the G2/M checkpoint of the cell cycle, induces apoptosis by initiating mitosis without repairing DNA damage. However, the effects of WEE1 inhibitors on the tumor immune microenvironment in colorectal cancer (CRC) remain unclear. Here, we investigated the association between WEE1 expression and CRC clinicopathological features using surgically resected CRC specimens and assessed the antitumor effects of a WEE1 inhibitor using CRC cell lines and orthotopic transplantation mouse models. WEE1 expression was not correlated with the clinicopathological features of CRC. The WEE1 inhibitor suppressed cell proliferation in a concentration-dependent manner in all CRC cell lines. It also increased the percentage of cells in the G2/M phase and apoptotic cells, especially in cell lines with p53 mutations, but did not alter these cell percentages in most p53 wild-type cell lines. In the orthotopic mouse model of CRC, tumor volume was significantly reduced in the WEE1 inhibitor-treated group compared to that in the control group. RNA sequencing and immunohistochemistry analyses of mouse tumors revealed that treatment with the WEE1 inhibitor activated tumor immunity and suppressed stromal reactions. These results demonstrate the potential antitumor effects of WEE1 inhibitors in CRC, particularly in patients with p53 mutations.

Background/Objectives: Colorectal neoplasia developing from ulcerative colitis mucosa (CRNUC) can be divided into ulcerative colitis-associated neoplasia (UCAN) and non-UCAN; however, it is often difficult to distinguish UCAN from non-UCAN during a biopsy diagnosis. We investigated whether a genomic analysis could improve the diagnostic accuracy of UCAN using biopsy specimens. Methods: In step 1, 14 CRNUCs were used to examine whether the genomic landscape of biopsy and resection specimens matched. In step 2, we investigated the relationship between the genomic landscapes and the pathological diagnosis of 26 CRNUCs. The cancer genome was analyzed by deep sequencing using a custom panel of 27 genes found to be mutated in our previous CRNUC analysis. Results: In step 1, of the 27 candidate genes, 14 were mutated. The concordance rate of the pathogenic mutations in these 14 genes between the biopsy and resection specimens was 29% (4/14), while that of the pathogenic mutations in TP53 and KRAS was 79% (11/14). In step 2, the pathological diagnosis of biopsy specimens using only hematoxylin and eosin (HE) staining had a sensitivity of 33% and an accuracy of 38% for UCAN diagnosis. On the other hand, the combination of the HE pathology and p53 immunohistochemical staining had a sensitivity of 73% and an accuracy of 85% for UCAN diagnosis, while the combination of HE staining and a TP53 mutation had a sensitivity of 87% and an accuracy of 88% for UCAN diagnosis. Conclusions: An evaluation of TP53 mutations in biopsy specimens may be useful for diagnosing UCAN. However, further studies with larger sample sizes are required before this can be applied in clinical practice.