In Vitro and In Vivo Inhibition of P-Glycoprotein (ABCB1) by a Novel Non-Substrate Compound: HG-829

P-glycoprotein (P-gp or ABCB1) is a common cause of multidrug resistance (MDR) in cancer and leukemia that serves to extrude amphipathic drugs from the plasma membrane. P-gp is an ATP-binding cassette (ABC) transporter that exports a wide range of anti-neoplastics that are structurally or functionally unrelated. The vast majority of P-gp inhibitors tested clinically act as competitive export channel inhibitors to promote antineoplastic retention. We investigated the in vitro and in vivo effects of a novel substituted quinoline P-gp modulator, HG-829, in MDR cell lines and xenograft models.

In vitro activity of HG-829 was evaluated in the K562-daunomycin-selected (K562-R) cell line and the ABCB1-transfected human embryonic kidney-293 cell line (HEK-293-B1) using a variety of P-gp substrates (daunomycin, doxorubicin, taxol, etoposide, vincristine) in a 72h MTT proliferation assay, and results compared to the effects of cyclosporine-A (CsA). Rhodamine 123 export and retention was assessed by flow cytometry. Flank injections of K562-R and parental K562-S cells in female SCID beige mice were performed for xenograft models. ANOVA and Turkey's multiple comparison tests were used to determine significant differences between groups in the proliferation assay and xenograft studies. Differences in rhodamine intracellular retention and efflux were assessed by Student's t-test.

Treatment with HG-829 at 0.5uM and 1uM completely reversed resistance to each of the antineoplastics studied in both the K562-R and HEK-293-B1 cell lines, but did not sensitize parental cells. HG-829 sensitized K562-R cells to daunomycin 57-fold and 97-fold at concentrations of 0.5uM and 1uM, respectively (p<0.01). Similarly, resistance to taxol (HG-829 0.5uM p<0.01 and 1uM, p<0.001), vincristine (p<0.01), and etoposide (p<0.05) were completely reversed. Comparable results were obtained with the HEK-293-B1 cell line in which HG-829 potentiated doxorubicin cytotoxicity 33- and 25-fold at concentrations 0.5uM (p<0.01) and 1uM (p<0.001), respectively. Compared to CsA, HG-829 was 3-fold more potent at equimolar concentrations. Functional studies showed that HG-829 (2.5uM) completely inhibited P-gp mediated rhodamine efflux in K562-R and the HEK transfected cells (p<0.05), but had no effect in parental cells. Pre-incubation followed by inhibitor removal showed that HG-829 inhibited rhodamine export as long as 24h (p<0.05), whereas equivalent concentration of CsA inhibited export for only 30 min. HG-829 showed no consistent effects on P-gp-ATPase activity in membrane preparations, whereas verapamil, a known P-gp substrate, displayed concentration-dependent stimulation. In mice bearing K562-R and parental xenografts, intraperitoneal administration of HG-829 significantly potentiated the antitumor activity of daunomyicin compared to vehicle without a significant increase in toxicity. Tumor volumes in HG-829 treated mice decreased to levels comparable to that of the K562-parental cohort (p<0.01 vs. vehicle).

HG-829 is a potent non-substrate inhibitor of P-glycoprotein with a prolonged duration of action. Clinical testing of HG-829 is warranted.

No relevant conflicts of interest to declare.