Amsacrine (m-AMSA) is an anticancer agent that displays activity against 
refractory acute leukemias as well as Hodgkin's and non-Hodgkin's lymphomas. The 
drug is comprised of an intercalative acridine moiety coupled to a 
4'-amino-methanesulfon-m-anisidide headgroup. m-AMSA is historically significant 
in that it was the first drug demonstrated to function as a topoisomerase II 
poison. Although m-AMSA was designed as a DNA binding agent, the ability to 
intercalate does not appear to be the sole determinant of drug activity. 
Therefore, to more fully analyze structure-function relationships and the role 
of DNA binding in the action of m-AMSA, we analyzed a series of derivatives for 
the ability to enhance DNA cleavage mediated by human topoisomerase IIα and 
topoisomerase IIβ and to intercalate DNA. Results indicate that the 3'-methoxy 
(m-AMSA) positively affects drug function, potentially by restricting the 
rotation of the headgroup in a favorable orientation. Shifting the methoxy to 
the 2'-position (o-AMSA), which abrogates drug function, appears to increase the 
degree of rotational freedom of the headgroup and may impair interactions of the 
1'-substituent or other portions of the headgroup within the ternary complex. 
Finally, the nonintercalative m-AMSA headgroup enhanced enzyme-mediated DNA 
cleavage when it was detached from the acridine moiety, albeit with 100-fold 
lower affinity. Taken together, our results suggest that much of the activity 
and specificity of m-AMSA as a topoisomerase II poison is embodied in the 
headgroup, while DNA intercalation is used primarily to increase the affinity of 
m-AMSA for the topoisomerase II-DNA cleavage complex.