Cell-cell interactions maintain tissue homeostasis and contribute to dynamic alteration of the tumor microenvironment (TME). Communication between cancer and host cells not only promotes advanced disease aggression but also determines therapeutic response in cancer patients. Despite accumulating evidence supporting the role of tumor-infiltrating immunocytes in modulating tumor immunity, the interplay between heterogeneous tumor subpopulations and immunocytes is elusive. We expanded colorectal cancer stem cells (CRCSCs) as cancer spheroids from the murine colorectal cancer (CRC) cell line CT26 to interrogate tumor-host interactions using a syngeneic tumor model. RNA-sequencing analysis of host cells and tumor exosomes was performed to identify molecular determinants that mediate the crosstalk between CRCSCs and immunocytes. The Cancer Genome Atlas (TCGA) database was used to validate the clinical significance in CRC patients. The expanded CT26 cancer spheroids showed increased stemness gene expression, enhanced spheroid and clonogenicity potential, and an elevated tumor-initiating ability, characteristic of CRCSCs. By examining immune cell composition in syngeneic tumor-bearing mice, a systemic increase in CD11b+/Ly6GHigh/Ly6CLow neutrophils was observed in mice bearing CRCSC-derived tumors. An increased secretion of CRCSC exosomes was observed in vitro, and through in vivo tracking, CRCSC exosomes were found to be transported to the bone marrow. Moreover, CRCSC exosomes prolonged the survival of bone marrow-derived neutrophils and engendered a protumoral phenotype in neutrophils. Mechanistically, tumor exosomal tri-phosphate RNAs induced the expression of interleukin-1β (IL-1β) through a pattern recognition-NF-κB signaling axis to sustain neutrophil survival. CRCSC-secreted CXCL1 and CXCL2 then attracted CRCSC-primed neutrophils to promote tumorigenesis of CRC cells via IL-1β. Moreover, neutrophil depletion using a Ly6G-specific antibody (clone 1A8) attenuated the tumorigenicity of CRCSCs. In human specimens, CRC patients exhibiting an active CRCSC signal (Snail+IL8+) showed elevated tumor infiltration of MPO+ neutrophils, and high (in the top 10%) MPO expression predicted poor survival of CRC patients. This study elucidates a multistep CRCSC-neutrophil interaction during advanced cancer progression. Strategies targeting aberrant neutrophil activation may be developed for combating CSC-related malignancy.

Let-7 is crucial for both stem cell differentiation and tumor suppression. Here, we demonstrate a chromatin-dependent mechanism of let-7 in regulating target gene expression in cancer cells. Let-7 directly represses the expression of AT-rich interacting domain 3B (ARID3B), ARID3A, and importin-9. In the absence of let-7, importin-9 facilitates the nuclear import of ARID3A, which then forms a complex with ARID3B. The nuclear ARID3B complex recruits histone demethylase 4C to reduce histone 3 lysine 9 trimethylation and promotes the transcription of stemness factors. Functionally, expression of ARID3B is critical for the tumor initiation in let-7-depleted cancer cells. An inverse association between let-7 and ARID3A/ARID3B and prognostic significance is demonstrated in head and neck cancer patients. These results highlight a chromatin-dependent mechanism where let-7 regulates cancer stemness through ARID3B.

Insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) is an RNA-binding protein and serves as a post-transcriptional fine-tuner regulating the expression of mRNA targets. However, the clinicopathological roles of IGF2BP1 in colorectal cancer (CRC) remains limited. Thus, we aimed to elucidate the clinical significance and biomarker potentials of IGF2BP1 in CRC. A total of 266 specimens from two sets of CRC patients were collected. IGF2BP1 expression was studied by immunohistochemical (IHC) staining. The Kaplan-Meier survival plot and a log-rank test were used for survival analysis. The Cox proportional hazards model was applied to determine the survival impact of IGF2BP1. Public datasets sets from The Cancer Genome Atlas (TCGA) and Human Cancer Metastasis Database (HCMDB), receiver operating characteristic (ROC) plotter, and two CRC cell lines, HCT-116 and DLD-1, were used for validating our findings. We showed that IGF2BP1 was overexpressed in tumor specimens compared to 13 paired normal parts by examining the immunoreactivity of IGF2BP1 (p = 0.045). The increased expression of IGF2BP1 in primary tumor parts was observed regardless of metastatic status (p < 0.001) in HCMDB analysis. IGF2BP1 expression was significantly associated with young age (59.6% vs. 46.7%, p-value = 0.043) and advanced stage (61.3% vs. 40.0%, p-value = 0.001). After controlling for confounding factors, IGF2BP1 remained an independent prognostic factor (HR = 1.705, p-value = 0.005). TCGA datasets analysis indicated that high IGF2BP1 expression showed a lower 5-year survival rate (58% vs. 65%) in CRC patients. The increased expression of IGF2BP1 in chemotherapy non-responder rectal cancer patients was observed using a ROC plotter. Overexpression of IGF2BP1 promoted the colony-forming capacity and 5-fluorouracil and etoposide resistance in CRC cells. Here, IGF2BP1 was an independent poor prognostic marker in CRC patients and contributed to aggressive phenotypes in CRC cell lines.