ABCG2 transports anticancer drugs via a closed-to-open switch

Benjamin J Orlando,Maofu Liao

doi:10.1038/s41467-020-16155-2

Abstract

ABCG2 is an ABC transporter that extrudes a variety of compounds from cells, and presents an obstacle in treating chemotherapy-resistant cancers. Despite recent structural insights, no anticancer drug bound to ABCG2 has been resolved, and the mechanisms of multidrug transport remain obscure. Such a gap of knowledge limits the development of novel compounds that block or evade this critical molecular pump. Here we present single-particle cryo-EM studies of ABCG2 in the apo state, and bound to the three structurally distinct chemotherapeutics. Without the binding of conformation-selective antibody fragments or inhibitors, the resting ABCG2 adopts a closed conformation. Our cryo-EM, biochemical, and functional analyses reveal the binding mode of three chemotherapeutic compounds, demonstrate how these molecules open the closed conformation of the transporter, and establish that imatinib is particularly effective in stabilizing the inward facing conformation of ABCG2. Together these studies reveal the previously unrecognized conformational cycle of ABCG2.

Highlights

ABCG2 is an ATP-binding cassette (ABC) transporter that extrudes a variety of compounds from cells, and presents an obstacle in treating chemotherapy-resistant cancers
Previous structures of ABCG2 in the inward facing conformation have been determined with the aid of a conformation specific 5D3 antibody fragment or high-affinity inhibitors, both of which are known to bias the conformational landscape of the transporter and potentially hinder analysis of important functional states in the transport cycle[25]
MXN and SN38 did not display any significant effect on the thermostability of ABCG2. Together these results demonstrate that unlike molecules such as Ko143 or imatinib that effectively stabilize certain conformations of ABCG2, transport substrates such as MXN and SN38 have less of an effect on the conformational landscape of the transporter

Summary

Introduction

ABCG2 is an ABC transporter that extrudes a variety of compounds from cells, and presents an obstacle in treating chemotherapy-resistant cancers. Our cryo-EM, biochemical, and functional analyses reveal the binding mode of three chemotherapeutic compounds, demonstrate how these molecules open the closed conformation of the transporter, and establish that imatinib is effective in stabilizing the inward facing conformation of ABCG2. ABCG2 transports uric acid and plays an essential role in the pathology of gout, and confers resistance to a variety of chemotherapeutics including topoisomerase inhibitors (i.e., mitoxantrone (MXN), etoposide, topotecan, SN38), anthracyclines (i.e., doxorubicin and daunorubicin), antimetabolites (i.e., methotrexate and 5-fluorouracil), and photosensitizers (i.e., pheophorbide A)[5] Transport of these compounds by ABCG2 is a limiting factor in their accumulation in cancerous cells, diffusion across the blood–brain barrier, and absorption through the intestine upon ingestion[6,7,8,9]. Our results delineate a new paradigm for the conformational transition cycle of ABCG2, and reveal how different chemotherapy compounds distinctively alter the conformational space of the transporter

Methods

Results

Conclusion