Background <p>Colorectal cancer (CRC) is one of the most common cancers worldwide and is often treated with chemotherapy. However, systemic toxicity, non-specificity, and drug resistance are major challenges associated with chemotherapeutic drugs. Nanocarriers such as niosomes (NIOs) can enhance drug accumulation at the tumor site while minimizing systemic side effects.</p> Methods <p>Doxorubicin (DOX) was loaded into NIOs using the thin-film hydration method and characterized by dynamic light scattering (DLS), scanning electron microscopy (SEM), and Fourier transform infrared spectroscopy (FTIR). HCT-116 colorectal cancer cells were treated with DOX and NIOs-DOX. Cell viability was assessed using the MTT assay. Additionally, apoptosis and cell cycle distribution were evaluated by flow cytometry, and the expression levels of DLGAP1-AS1 and AFAP1-AS1 were determined by quantitative real-time PCR (qRT-PCR).</p> Results <p>NIOs-DOX were synthesized with a size of 242.8&#xa0;nm and a zeta potential of − 15.4 mV. The IC50 values of NIOs-DOX and DOX against HCT-116 cells were determined to be 0.974 µM and 4.18 µM, respectively. Flow cytometric analysis revealed a substantial increase in apoptotic cells in HCT-116 following treatment with NIOs-DOX compared to DOX. The expression levels of DLGAP1-AS1 and AFAP1-AS1 were significantly higher in tumor tissue samples than adjacent non-tumor samples. Furthermore, the overexpression of AFAP1-AS1 significantly correlated with the adenocarcinoma subtype. Based on the ROC curve analysis, DLGAP1-AS1 and AFAP1-AS1 expression showed poor diagnostic value.</p> Conclusion <p>NIOs-DOX demonstrated significant potential to enhance the therapeutic effects of DOX. Additionally, DLGAP1-AS1 and AFAP1-AS1 expression showed limited potential as diagnostic biomarkers in CRC.</p>