Synergisitic anti-colorectal cancer effects of WNT974 combined with artesunate via cooperative regulation of p21, p27, and AKT

Abstract

Objective

To evaluate the synergistic effects and underlying mechanisms of WNT974 and artesunate (ART) combination therapy in the treatment of colorectal cancer (CRC).

Methods

We conducted an initial assessment of the synergistic impact of WNT974 and ART on cell viability using the MTT assay across eight CRC cell lines with diverse mutation statuses.Subsequently, a 3D in vitro model was utilized to confirm the synergistic effect of the combination. Flow cytometry was employed to analyze cell cycle distribution. The molecular mechanisms were further investigated by examining the expression levels of cell cycle-related proteins (p21, p27, and CDK2) and AKT activity through PCR and Western blot analyses. The in vivo efficacy of the combination therapy was evaluated using a xenograft mouse model of CRC.

Results

The combination of WNT974 and ART significantly reduced cell viability in all eight CRC cell lines, demonstrating a synergistic effect. Additionally, the combination exhibited synergy in inhibiting 3D spheroid growth. Cell cycle analysis revealed that the combination therapy induced a marked G1/G0 arrest through CDK2 inhibition, surpassing the efficacy of individual treatments. Molecular analysis indicated that the combination therapy upregulated p21 and p27 expression and reduced AKT activity. In vivo, the combination therapy significantly inhibited tumor growth in the xenograft mouse model, notably outperforming 5FU, the standard first-line drug for CRC.

Conclusion

The study highlights the synergistic anti-CRC effects of combining WNT974 and ART. This combination induces G1/G0 phase cell cycle arrest by cooperatively regulating p21, p27, and AKT, presenting a promising alternative to current CRC treatments. These findings elucidate the molecular basis of this interaction and provide a scientific foundation for advancing ART combined with WNT94 as a novel therapy for CRC, offering a potential new avenue.