Functions of the ABC transporter breast cancer resistance protein (BCRP/ABCG2)

SY31-1 Our research focuses on genes and proteins that cause drug resistance in tumors, and/or influence the pharmacological and toxicological behavior of anticancer and many other drugs and toxins, including carcinogens. Insight into these systems may improve chemotherapy approaches for cancer as well as pharmacotherapy in a broader sense, and increase insights in factors determining susceptibility to carcinogens. To study the physiological, pharmacological and toxicological roles of the proteins involved, and their interactions, we generate and analyze knockout or transgenic mice lacking or overexpressing the relevant genes. Cell lines obtained from these mice are further used as tools to identify and characterize drug resistance genes. To date, we have primarily focused on plasma membrane proteins of the ATP binding cassette (ABC) transporter family, encompassing amongst others P-glycoprotein (P-gp or ABCB1), MRP2 (ABCC2) and BCRP (ABCG2). These proteins actively export a wide range of anticancer, anti-HIV/AIDS, and many other drugs from cells. This ATP-dependent drug extrusion can cause multidrug resistance (MDR) in tumor cells (1). P-gp, MRP2 and BCRP all localize to the apical membrane of polarized epithelial cells, resulting in vectorial transport of drug substrates, and there is considerable (albeit not complete) overlap in substrate specificity between these transporters. Expression of these transporters by cDNA transfection into polarized epithelial cell lines offers excellent tools to study their transport characteristics in vitro. In the past, we and others have generated knockout mice deficient for P-gp and Bcrp1, the murine homologue of BCRP. Experiments in P-gp and Bcrp1 knockout mice indicated that these transporters can variously protect an organism against exogenous toxins and drugs by limiting penetration of substrates into brain, testis, and fetus, by restricting uptake of orally administered substrates, and by mediating excretion of substrates via liver and intestine. Mice lacking Bcrp1 look overall normal, but they are extremely sensitive to the dietary chlorophyll breakdown product pheophorbide a, resulting in severe, sometimes lethal phototoxic lesions on light-exposed skin (2). Pheophorbide a is a porphyrin that occurs in various plant-derived foods and food supplements. Bcrp1 transports pheophorbide a and is highly efficient in limiting its uptake from ingested food. Furthermore, as predicted from previous in vivo inhibition experiments, Bcrp1 knockout mice demonstrated a 6-fold increased oral uptake, and 2-fold increased fetal penetration of topotecan. Bcrp1 knockout mice also displayed a novel type of protoporphyria: Erythrocyte levels of the heme precursor and phototoxin protoporphyrin IX were 10-fold increased. These results indicate that humans or animals with low or absent BCRP activity may be at increased risk for developing protoporphyria and diet-dependent phototoxicity, and provide a striking illustration of the importance of drug transporters in protection from toxicity of normal food constituents. The data imply that prolonged inhibition of BCRP by administration of BCRP inhibitors to patients should be monitored carefully for unexpected side effects. We have also identified a pronounced role for Bcrp1 in the pharmacokinetics of, and hence possibly protection from, dietary carcinogens (3). The food carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is the most abundant heterocyclic amine found in various protein-containing foods. PhIP is mutagenic and carcinogenic in rodents, and it has been implicated in human breast carcinogenesis. Humans on a normal western diet are exposed to PhIP on a daily basis. We investigated whether Bcrp1 could affect PhIP exposure of the body, as this could implicate BCRP activity in the cancer risk due to PhIP. Using polarized cell lines we found that PhIP is efficiently transported by murine Bcrp1. In vivo pharmacokinetic studies in Bcrp1 knockout mice showed that Bcrp1 effectively restricts the exposure of mice to ingested PhIP, by decreasing its uptake from the gut lumen and by mediating hepatobiliary and intestinal elimination of PhIP. Intra- or interindividual differences in BCRP activity in humans may thus also affect the exposure to PhIP and related food carcinogens, with possible implications for cancer susceptibility. We are currently extending the functional analysis of BCRP and Bcrp1 in the pharmacokinetics of several anticancer drugs and dietary carcinogens. Moreover, by more detailed analysis of expression of Bcrp1 in mice and other species we have identified several additional organs where BCRP/Bcrp1 may have physiological and pharmacologically relevant functions. Some of these newly identified functions and their implications will be discussed.