Fludarabine Phosphate (NSC 312887 Phosphate) is the phosphate salt of a fluorinated nucleotide antimetabolite analog of the antiviral agent vidarabine (ara-A) with antineoplastic activity. Fludarabine phosphate is rapidly dephosphorylated to 2-fluoro-ara-A and then phosphorylated intracellularly by deoxycytidine kinase to the active triphosphate, 2-fluoro-ara-ATP. This metabolite may inhibit DNA polymerase alpha, ribonucleotide reductase, and DNA primase, thereby interrupting DNA synthesis and inhibiting tumor cell growth.

DNA polymerase δ:1.3 μM(Ki), DNA polymerase α:1.1 μM(Ki )

The maximum tolerable dose of Fludarabine Phosphate is determined to be 234 mg/kg, with its LD50 (lethal dose 50%) for mice being 375 mg/kg. Notably, Fludarabine Phosphate exhibits toxicity in tumor-free mice. However, in mice carrying P388 leukemia, treatment with Fludarabine Phosphate leads to a reduction in the number of surviving cells geared towards therapy, significantly enhancing the lifespan by 110% and increasing the average survival time.

Fludarabine Phosphate effectively inhibits DNA polymerase α/δ (Ki: 1.1/1.3 μM), demonstrating potent inhibitory activity in vitro. It is capable of being excised from bound DNA by DNA polymerase δ. Competitively binding with deoxyadenosine triphosphate at the A site for DNA chain elongation, Fludarabine Phosphate leads to the termination of DNA strand extension. Notably, human DNA polymerase α is more adept at incorporating Fludarabine Phosphate into DNA than polymerase δ.

Cells are incubated with Fludarabine Phosphate for 5 hr and washed twice with drug-free warm medium. 800 cells are mixed with 1.3 mL of 0.25% soft agar in Dulbecco's medium supplemented with 20% fetal bovine serum (pre-warmed to 37 ℃) and incubated in a tissue culture dish for 10 days (humidified 5% CO2 , 37 ℃). At the end of the incubation period, colonies of more than 40 cells are scored under a microscope. The cytotoxic effect of the drugs is expressed as a percentage of survival relative to that of untreated control cells.(Only for Reference)