Investigating the Antagonistic Effect of Ceftazidime On 5-Fluorouracil Efficacy in Colorectal Cancer

Abstract

Background: 5-Fluorouracil (5-FU) is a backbone chemotherapeutic agent for colorectal cancer (CRC) whose cytotoxic efficacy depends on activation of the DNA damage response and coordinated cell-cycle checkpoint signaling. Clinical variability in response to 5-FU is attributed to tumor heterogeneity, treatment timing and resistance, cellular stress adaptation mechanisms, and co-administered medications. Among cellular stress adaptation mechanisms, the p53-p21 axis governs the cell's response to oncogene activation, DNA damage, and stress by activating downstream pathways such as DNA repair, cell cycle arrest, senescence, and apoptosis. Due to the immunocompromised state of cancer patients under 5-FU treatment, broad-spectrum antibiotics, including ceftazidime (CFT), are commonly prescribed to cancer patients for prophylaxis or treatment of infection. This raises the concern that antibiotic-chemotherapy interactions may alter therapeutic outcomes. CFT has been shown to antagonize the cytotoxic effect of 5-FU in CRC. While the molecular pathways governing this interaction remain poorly understood, increasing evidence suggests that such drug–drug interactions (DDIs) are attributed to cellular stress adaptation mechanisms and cell cycle checkpoint conditions (p53-p21 axis) as well as transporter mediation solute carrier transporters such as organic anion transporter 2 (OAT2) and organic carnitine/cation transporter 2 (OCTN2) which 5-FU and CFT bind to.

Aims: This study aimed to determine whether CFT antagonizes 5-FU cytotoxicity in colorectal cancer cells and to define the cellular state under which such antagonism occurs, with emphasis on the p53-p21 checkpoint axis and the transporters OAT2 and OCTN2.

Materials and methods: Human HCT-116 CRC cell lines (wild-type, p21-/-, p53-/-, and 5-FU resistant) were employed. Metabolic activity was assessed using MTT assays following exposure to 5-FU, CFT, or their combination for 48h and 72h, with or without pharmacological attenuation of p21 using UC2288. Western blotting was used to evaluate expressions of p53, p21, OAT2, and OCTN2 across genotypes, time points, and treatment conditions.

Results: Antagonism between 5-FU and CFT was observed exclusively in wild-type HCT-116 cells at 48h, where combination treatment resulted in higher metabolic activity than 5-FU alone. This antagonism was absent at 72h and in all cell cycle checkpoint-disrupted models, including HCT-116 p21-/-, HCT-116 p53-/-, and HCT-116 5-FU–resistant cells. Western blot analysis revealed that antagonism coincided with a transient checkpoint state characterized by strong p53 activation in the absence of detectable p21 expression. However, pharmacological attenuation of p21 reversed the seen antagonism despite amplifying early p53 accumulation. Expression of OAT2 and OCTN2 in wild-type cells showed treatment- and time-dependent modulation during this early window of antagonism. In non-antagonism states, their expression varied, increasing and decreasing with minimal treatment-specific trends across cell lines or timepoints. This indicates loss of cellular plasticity rather than loss of transporter abundance.

Conclusion: This study demonstrates that antagonism between 5-FU and CFT is a state-dependent phenomenon that emerges only during an early, plastic checkpoint phase in wild-type CRC cells. Transporter expression and coordinated progression of the p53–p21 axis govern the presence of the antagonism. An alteration in the stage of p53-p21 axis progression, and consequently in treatment-specific transporter expression, away from that seen in cells with antagonism resulted in its ablation. These findings highlight the importance of cellular state and timing in evaluating antibiotic-chemotherapy interactions and suggest that checkpoint disruption or enforcement eliminates the window for antagonistic effects.