ABCB1 detoxifies cells by exporting diverse xenobiotic compounds, thereby limiting drug disposition and contributing to multidrug resistance in cancer cells. Multiple small-molecule inhibitors and inhibitory antibodies have been developed for therapeutic applications, but the structural basis of their activity is insufficiently understood. We determined cryo-EM structures of nanodisc-reconstituted, human ABCB1 in complex with the Fab fragment of the inhibitory, monoclonal antibody MRK16 and bound to a substrate (the antitumor drug vincristine) or to the potent inhibitors elacridar, tariquidar, or zosuquidar. We found that inhibitors bound in pairs, with one molecule lodged in the central drug-binding pocket and a second extending into a phenylalanine-rich cavity that we termed the “access tunnel.” This finding explains how inhibitors can act as substrates at low concentration, but interfere with the early steps of the peristaltic extrusion mechanism at higher concentration. Our structural data will also help the development of more potent and selective ABCB1 inhibitors.

ABCB1通过输出各种异种生物化合物来排毒细胞，从而限制了药物的分布并促进了癌细胞的多药耐药性。已经开发出多种小分子抑制剂和抑制性抗体用于治疗应用，但是对其活性的结构基础还知之甚少。我们确定了纳米盘重构的人ABCB1与抑制性单克隆抗体MRK16的Fab片段复合并结合至底物（抗肿瘤药物长春新碱）或强效抑制剂elacridar，tariquidar或zosuquidar的冷冻EM结构。我们发现抑制剂成对结合，一个分子滞留在中心药物结合口袋中，另一个分子伸入富含苯丙氨酸的腔中，我们称之为“通道”。这一发现解释了抑制剂在低浓度下如何能充当底物，但在高浓度下会干扰蠕动挤出机理的早期步骤。我们的结构数据也将有助于开发更有效和更具选择性的ABCB1抑制剂。