Amiloride is capable of inhibiting DNA synthesis in mammalian cells in culture. 
Recent evidence indicates that the enzyme, DNA topoisomerase II, is probably 
required for DNA synthesis to occur in situ. In experiments to determine the 
mechanism of inhibition of DNA synthesis by amiloride, we observed that 
amiloride inhibited both the catalytic activity of purified DNA topoisomerase II 
in vitro and DNA topoisomerase II-dependent cell functions in vivo. Many 
compounds capable of inhibiting DNA topoisomerase II are DNA intercalators. 
Thus, we performed studies to determine if and how amiloride bound to DNA. 
Results indicated that amiloride 1) shifted the thermal denaturation profile of 
DNA, 2) increased the viscosity of linear DNA, and 3) unwound circular DNA, all 
behavior consistent with a DNA intercalation mechanism. Furthermore, 
quantitative and qualitative measurements of amiloride fluorescence indicated 
that amiloride (a) bound reversibly to purified DNA under conditions of 
physiologic ionic strength, and (b) bound to purified nuclei in a highly 
cooperative manner. Lastly, amiloride did not promote the cleavage of DNA in the 
presence of DNA topoisomerase II, indicating that the mechanism by which 
amiloride inhibited DNA topoisomerase II was not through the stabilization of a 
"cleavable complex" formed between DNA topoisomerase II, DNA, and amiloride. The 
ability of amiloride to intercalate with DNA and inhibit topoisomerase II is 
consistent with the proposed planar, hydrogen-bonded, tricyclic nature of 
amiloride's most stable conformation. Thus, DNA and DNA topoisomerase II must be 
considered as new cellular targets of amiloride action.