Inhibition of ABCG2-mediated transport by protein kinase inhibitors with a bisindolylmaleimide or indolocarbazole structure

Abstract

ABCG2 is a transporter with potential importance in cancer drug resistance, drug oral absorption, and stem cell biology. In an effort to identify novel inhibitors of ABCG2, we examined the ability of commercially available bisindolylmaleimides (BIM) and indolocarbazole protein kinase inhibitors (PKI) to inhibit ABCG2, given the previous demonstration that the indolocarbazole PKI UCN-01 interacted with the transporter. At a concentration of 10 Mmol/L, all of the compounds tested increased intracellular fluorescence of the ABCG2-specific substrate pheophorbide a in ABCG2-transfected HEK-293 cells by 1.3-to 6-fold as measured by flow cytometry; the ABCG2-specific inhibitor fumitremorgin C increased intracellular fluorescence by 6.6-fold. In 4-day cytotoxicity assays, wild-type ABCG2-transfected cells were not more than 2-fold resistant to any of the compounds, suggesting that the PKIs are not significantly transported by ABCG2. BIMs I, II, III, IV, and V, K252c, and arcyriaflavin A were also able to inhibit [ 125 I]iodoarylazidoprazosin labeling of ABCG2 by 65% to 80% at 20 Mmol/L, compared with a 50% to 70% reduction by 20 Mmol/L fumitremorgin C. K252c and arcyriaflavin A were the most potent compounds, with IC 50 values for inhibition of [ 125 I]iodoarylazidoprazosin labeling of 0.37 and 0.23 Mmol/L, respectively. K252c and arcyriaflavin A did not have any effect on the ATPase activity of ABCG2. Four minimally toxic compounds-BIM IV, BIM V, arcyriaflavin A, and K252c-reduced the relative resistance of ABCG2-transfected cells to SN-38 in cytotoxicity assays. We find that indolocarbazole and BIM PKIs directly interact with the ABCG2 protein and may thus increase oral bioavailability of ABCG2 substrates.

Sunitinib (Sutent, SU11248), a Small-Molecule Receptor Tyrosine Kinase Inhibitor, Blocks Function of the ATP-Binding Cassette (ABC) Transporters P-Glycoprotein (ABCB1) and ABCG2

“…We next sought to determine whether the effects of sunitinib were caused by direct interaction with the transporters. UIC2 and 5D3 are P-gp-and ABCG2-specific antibodies, respectively, that recognize extracellular epitope(s) and have been shown to recognize conformation changes in these transporters that result in increased binding in the presence of substrates or inhibitors in the case of UIC2, or inhibitors alone in the case of 5D3 (Mechetner et al, 1997;Ozvegy-Laczka et al, 2005;Robey et al, 2007). Binding of these antibodies to P-gp and ABCG2 in the presence and absence of varying concentrations of sunitinib was examined.…”

Marine Natural Products as Models to Circumvent Multidrug Resistance

“…The naturally occurring arcyriaflavin (63) and indolcarbazole K252c or staurosporin aglycone (64, Figure 12) showed the most potent effects in BCRP inhibition in the BCRP-transferred HEK-293 cell line, with low toxicity in BCRP-transfected cells, and reduced the relative resistance of ABCG2-transfected cells SN-38 [29,146]. Bisindolylmaleimide analogs 65-69 ( Figure 12) were found to be able to inhibit [ 125 I]iodoarylazidoprazosin labeling of BCRP by 65% to 80% at 20 μM [146]. These findings revealed that indolocarbazole and bisindolylmaleimide analogs directly interact with BCRP protein and may increase oral bioavailability of BCRP substrates.…”

“…The mechanisms of their antitumor effects include topoisomerase I poisoning, inhibition of PKC, PKA, pyruvate dehydrogenase kinase (PDK)/cyclin B, and cyclin-dependent kinase 5 (CDK5)/p5 [144,145]. The naturally occurring arcyriaflavin (63) and indolcarbazole K252c or staurosporin aglycone (64, Figure 12) showed the most potent effects in BCRP inhibition in the BCRP-transferred HEK-293 cell line, with low toxicity in BCRP-transfected cells, and reduced the relative resistance of ABCG2-transfected cells SN-38 [29,146]. Bisindolylmaleimide analogs 65-69 ( Figure 12) were found to be able to inhibit [ 125 I]iodoarylazidoprazosin labeling of BCRP by 65% to 80% at 20 µM [146].…”

Design and synthesis of novel 5,6-bisindolylpyrimidin-4-ones structurally related to ruboxistaurin (LY333531)

“…12 It has also been proposed that the bisindolylmaleimide framework constitutes an ABCG2 inhibitory pharmacophore, which may increase the oral bioavailability of co-administered ABCG2 substrate drugs, in order to potentiate an effective therapeutic response and significantly improve subsequent clinical outcome. 13 SAR analysis suggests that application of precise structural features to probe interactions between the maleimide head group and essential residues within the conserved enzyme ATP-binding pocket is still challenging, requiring further elucidation of exact mechanisms of inhibition. 14 Accomplishing distinct kinase selectivity with sufficient potency (in the presence of intracellular ATP concentrations) represents the ultimate goal of vast research within this area; however, little work to date has assessed the protein backboneinteracting pyrrole-2,5-dione molecular domain as a viable target for inhibitory optimisation in order to achieve this overall aim.…”