Abstract 3601: Antibody-drug conjugates (ADCs) with tubulysin and PBD warheads, maintain potent in vitro cytotoxicity against multidrug-resistant tumor cells expressing P-glycoprotein (P-gp)

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA Development of resistance to initially-effective therapies remains a major challenge in the treatment of cancer. Resistance to antibody-drug conjugates (ADCs) can arise due to many factors including down-regulation of the target, modified internalization or trafficking of the internalized ADC/antigen complexes, decreased sensitivity to the warhead conjugated to the ADC, or combinations of these. One of the common mechanisms of resistance to ADCs is increased expression of efflux drug pumps such as P-glycoprotein (P-gp) which can then transport small molecule warheads out of the cell leading to multi-drug resistance (MDR). In fact, the warheads used in the two clinically-approved ADCs, Adcetris® and Kadcyla™, have been reported to be P-gp substrates and P-gp overexpression can lead to acquired resistance against these ADCs. Studies were conducted to determine if pyrrolobenzodiazepine (PBD) dimer or tubulysin warheads, and/or ADCs conjugated with these warheads were susceptible to P-gp-mediated resistance. The P-gp susceptibility of these warheads and associated ADCs was evaluated by comparing the relative cytotoxicity of warheads/ADCs in parental tumor cell lines compared to the same cell lines that have been manipulated to overexpress P-gp. Verapamil, an inhibitor of P-gp-mediated efflux, was used to confirm whether decreased sensitivity was due to P-gp activity. Unlike known P-gp substrates MMAE, paclitaxel, and vinblastine, the majority of PBD and tubulysin warheads tested were not significant P-gp substrates. These warheads and ADC's conjugated with these warheads potently induced cytotoxicity of the parental cancer cell lines, and maintained this potency in the same cell lines overexpressing P-gp. Interestingly, certain tubulysins had differential susceptibility to P-gp depending on whether they were conjugated or depending on the linker that was used. For example, one tubulysin warhead showed moderate susceptibility to P-gp-mediated efflux as a naked warhead, but ADCs conjugated with this warhead had equivocal efficacy in parental and P-gp-overexpressing cell lines. An alternative tubulysin warhead was equipotent as an unconjugated small molecule in either parental or P-gp-overexpressing cell lines, however an ADC conjugated with this warhead using a cleavable linker had no activity in the resistant cells; an effect that was reversible with verapamil treatment, confirming the role of P-gp. These data suggest that ADCs conjugated with either PBD or tubulysin warheads may be active in MDR settings where resistance is mediated by P-gp expression, however activity against MDR cancers may be dependent on the particular warhead that was used and/or the particular linker that is used to conjugate the warhead to the antibody. Citation Format: Shenlan Mao, Ryan Fleming, Binyam Bezabeh, Nazzareno Dimasi, Dorin Toader, Thais Cailleau, Philip Howard, Changshou Gao, Bob Hollingsworth, Adeela Kamal, Jay Harper. Antibody-drug conjugates (ADCs) with tubulysin and PBD warheads, maintain potent in vitro cytotoxicity against multidrug-resistant tumor cells expressing P-glycoprotein (P-gp). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3601. doi:10.1158/1538-7445.AM2015-3601