It also confirms that quinacrine and chloroquine are newly identified members of RF-mimicking competitive ligands. where a sulfur atom (S) has replaced the N at the N(5) position in the isoalloxazine ring. The ITC studies indicate that changes to the N(5) position inhibit the binding of RF antagonists to RfBP. Because the N(5) position is involved in redox reactions, it is possible that changes at this SQSTM1 position affect binding to RfBP.1,11 In addition, the ability of N to serve as the H-bond acceptor may be critical for the tighter binding, while replacement with S can eliminate such ability. Lumiflavin, quinacrine, and chloroquine show nearly 1?3 orders of magnitude in change in (kJ mol?1)(kJ mol?1 K?1)= 1 ? RU[I]/RU[I]=0) as a function of inhibitor (ligand) concentration for each of the competitive inhibitors (RF, quinacrine, and 1). The response unit (RU[I]) for each sensorgram was determined by correcting the bulk contribution as described earlier. The range of inhibition concentrations was lower in the case of quinacrine due to its limited solubility in the SPR running buffer. In summary, the SPR results are in good agreement with the Abscisic Acid findings from the ITC study. It also confirms that quinacrine and chloroquine are newly identified members of RF-mimicking competitive ligands. As drugs traditionally used in the treatment of malaria18 and rheumatoid arthritis,19 these molecules are reported to display diverse biological activities due to the ability to inhibit a number of important biological targets such as DNA topoisomerase II18,20 and metabolic enzymes.21 Recently, quinacrine has generated new attention because of the discovery that it has antitumoral activity.22?25 This activity is attributed to its ability to interfere with cell signaling pathways such as activation of the p53 pathway25 and inhibition of Bcl-xL, an antiapoptotic protein.23 The present study suggests another novel application for the quinacrine class Abscisic Acid of the drug molecules as ligands that can target RfBP, a vitamin B2 uptake receptor, in a manner competitive to RF. Implications for this obtaining are many-fold. First, the present Abscisic Acid study suggests a new additional perspective for the biological activities associated with quinacrine and its analogues. On the basis of the interpretation of our data, it is also conceivable that quinacrine can interfere with receptor-mediated RF uptake outside the cell and/or can block a broad range of flavin cofactor-mediated enzymatic activities after internalization. Second, despite Abscisic Acid using a moderate affinity to RF receptor at the lower micromolar concentration, quinacrine can serve as a targeting ligand for specific delivery of additional therapeutic molecules or imaging brokers to the receptor-overexpressing cancer cells implicated in breast and prostate cancers.2,3 Pertinent to this targeting utility, it would be possible to apply the concept of multivalent ligand design,7,16,26 in which even suboptimal targeting capability can be enhanced through multivalent tight binding. Funding Statement National Institutes of Health, United States Notes K.S. thanks the support of NIH (1F33CA138031-01A1), NSF (CHE-0959681), and HHMI (student support). Part of this work was supported by NCI, NIH under award 1 R01 CA119409 (J.R.B.). Supporting Information Available Experimental details and additional SPR sensorgrams. This material is available free of charge via the Internet at http://pubs.acs.org. Supplementary Material ml100296z_si_001.pdf(736K, pdf).