Derivatives of the anthraquinone (anthracene-9,10-dione) such as doxorubicin, mitoxantrone and others have proved great clinical efficacy for decades. Currently the search in this exceptionally productive chemical class is aimed at optimization of antitumor properties including circumvention of drug resistance. Previously we have reported that heteroarene-fused anthraquinones fused to a 5-membered heterocyclic ring are advantageous in killing drug resistant tumor cells. Herein we present the synthesis and antitumor properties of a series of new anthra[2,3-b]furan-2-carboxamides. Vast majority of new derivatives were similarly cytotoxic to wild type tumor cell lines and their isogenic sublines with P-glycoprotein overexpression and/or p53 inactivation. Comparison of structurally close derivatives varying in their position relative to the furan moiety, that is, furan-3-carboxamide 1vs furan-2-carboxamides 5 and 6, revealed fundamental differences in the cytotoxicity profiles, formation of drug-DNA complexes, efficacy of topoisomerase 1 inhibition and mechanisms of tumor cell death. Together with previous SAR data on the role of individual substituents, these results provide evidence that regioisomerization of anthra[2,3-b]furancarboxamides generates the practically perspective derivatives whose properties may vary significantly.