[6][7] How it works is not entirely clear, but it is believed to involve blocking the action of thymidylate synthase and thus stopping the production of DNA.

[13] Other uses include ocular injections into a previously created trabeculectomy bleb to inhibit healing and cause scarring of tissue, thus allowing adequate aqueous humor flow to reduce intraocular pressure.

[14] in 2020, the EU and UK license was updated to state that fluorouracil was contra-indicated in patients that "have a known complete absence of dihydropyrimidine dehydrogenase (DPD) activity".

[15] In US, as of 2024, there is no specific contraindication on the package inserts however, there is a cautionary warning: "Increased risk of serious or fatal adverse reactions in patients with low or absent Dipyrimidine Dehydrogenase activity: withhold or permanently discontinue fluorouracil in patients with evidence of acute early-onset or unusually severe toxicity, which may indicate near complete or total absence of dipyrimidine dehydrogenase (DPD) activity.

No fluorouracil dose has been proven safe in patients with absent DPD activity"[16] Within the UK, DPYD testing to check for this contraindication is now routine practice,[17] this is not the case in the US.

[25] Life-threatening cardiotoxicity includes: arrhythmias, ventricular tachycardia and cardiac arrest, secondary to transmural ischaemia.

[31][32][33][34] The limitations of BSA-based dosing prevent oncologists from being able to accurately titer the dosage of 5-FU for the majority of individual patients, which results in sub-optimal treatment efficacy or excessive toxicity.

[31][32] Numerous studies have found significant relationships between concentrations of 5-FU in blood plasma and both desirable or undesirable effects on patients.

[35][36] Studies have also shown that dosing based on the concentration of 5-FU in plasma can greatly increase desirable outcomes while minimizing negative side effects of 5-FU therapy.

[43] The dihydropyrimidine dehydrogenase (DPD) enzyme is responsible for the detoxifying metabolism of fluoropyrimidines, a class of drugs that includes 5-fluorouracil, capecitabine, and tegafur.

Interrupting the action of this enzyme blocks synthesis of the pyrimidine thymidylate (dTMP), which is a nucleotide required for DNA replication.