Characterization of a Novel Bisacridone and Comparison with PSC 833 as a Potent and Poorly Reversible Modulator of P-Glycoprotein JULIE K. HORTON, KUNTEBOMMANAHALLI N. THIMMAIAH, GUILLERMO A. ALTENBERG, ARIEL F. CASTRO, GLEN S. GERMAIN, G. KRISHNE GOWDA, and PETER J. HOUGHTON Sealy Center for Molecular Science (J.K.H.) and Department of Physiology and Biophysics (J.K.H., G.A.A., A.F.C.), University of Texas Medical Branch, Galveston, Texas 77555, Department of Studies in Chemistry, University of Mysore, Mysore-570006, India (K.N.T., G.K.G.), and Department of Molecular Pharmacology, St. Jude Children’s Research Hospital, Memphis, Tennessee 38101 (G.S.G., P.J.H.) Received July 8, 1997; Accepted August 12, 1997 SUMMARY Novel compounds, composed of two acridone moieties con- nected by a propyl or butyl spacer, were synthesized and tested as potential modulators of P-glycoprotein (P-gp)-medi- ated multidrug resistance. The propyl derivative 1,3-bis(9- oxoacridin-10-yl)-propane (PBA) was extremely potent and, at a concentration of 1 M, increased steady state accumulation of vinblastine (VLB) 9-fold in the multidrug-resistant cell line KB8 –5. In contrast to the readily reversible effects of VRP and cyclosporin A on VLB uptake and similar to the effects of the cyclosporin analog PSC 833, this modulation by PBA was not fully reversed 6 – 8 hr after transfer of cells to PBA-free medium. Continuous exposure to 3 M PBA was nontoxic and could completely reverse VLB resistance in KB8 –5 cells. Consistent with its effects on VLB transport, the drug resistance-modulat- ing effect of PSC 833 was significantly more persistent than that of VRP. However, the effect of PBA was, like that of VRP, rapidly reversed once the modulator was removed from the extracellular environment. PBA was able to compete with ra- diolabeled azidopine for binding to P-gp and to stimulate P-gp ATPase activity. However, both the steady state accumulation of PBA and the rate of efflux of PBA were similar in drug- sensitive KB3–1 and drug-resistant KB8 –5 cells, suggesting that this compound is not efficiently transported by P-gp. These results indicate that PBA represents a new class of potent and poorly reversible synthetic modulators of P-gp-mediated VLB transport. Despite advances in the use of chemotherapeutic drugs for the treatment of human cancer, the emergence of resistance to these agents, at either initial presentation or the time of relapse, remains a major problem and has been the subject of numerous investigations. MDR in model cell lines is fre- quently associated with overexpression of the MDR1 gene product, P-gp. One characteristic of the MDR phenotype is that tumor cells selected for resistance to a single agent (e.g., a Vinca alkaloid or an anthracycline antibiotic) simulta- neously become resistant to a large number of structurally and functionally unrelated cytotoxic agents. P-gp is a 170- kDa integral membrane protein proposed to catalyze ATP- dependent drug efflux from cells, and thus effectively reduc- ing intracellular drug accumulation in the resistant cells (1). MDR1 expression has been detected in several human tumors (2), but the relevance of P-gp and the MDR phenom- enon to drug resistance in clinical malignancies is still un- certain (3, 4), and other factors conferring drug resistance are important. There is some evidence to suggest that in child- hood cancers and hematological malignancies, P-gp expres- sion is predictive of poor therapeutic outcome (5, 6), although in another study, P-gp expression at diagnosis did not corre- late with response in childhood rhabdomyosarcoma (7). In addition, as described in patients with myeloma, the selec- tion of tumor cells with a high level of MDR1 expression may result from prior therapy with natural product chemothera- peutic agents (8). There seems to be two basic approaches to circumvention of P-gp-associated MDR; the first is to use other effective chemotherapeutic drugs, such as the alkylat- ing agents, in which transport is not mediated by P-gp (1), This work was supported in part by Searle Research and Development; American Heart Association, Texas Affiliate, Grant 966-1613 (G.A.A.); Na- tional Institutes of Health Grants CA72783 (G.A.A.), CA23099, and CA21675 (Cancer Center Support grant); and American Lebanese Syrian Associated Charities (P.J.H.); and the Department of Atomic Energy, Government of India (K.N.T.). ABBREVIATIONS: MDR, multidrug resistant/resistance; PBA, 1,3-bis(9-oxoacridin-10-yl)-propane; VLB, vinblastine; VRP, verapamil; CsA, cy- closporin A; R123, rhodamine 123; P-gp, P-glycoprotein; DTT, dithiothreitol; HEPES, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; HR, HEPES-Ringer. 0026-895X/97/060948-10$3.00/0 Copyright © by The American Society for Pharmacology and Experimental Therapeutics All rights of reproduction in any form reserved. MOLECULAR PHARMACOLOGY, 52:948 –957 (1997). 948 at ASPET Journals on June 21, 2017 molpharm.aspetjournals.org Downloaded from