ABCG2: A perspective

Abstract

ABCG2, or breast cancer resistance protein (BCRP), is an ABC transporter that has been the subject of intense study since its discovery a decade ago. With high normal tissue expression in the brain endothelium, gastrointestinal tract, and placenta, ABCG2 is believed to be important in the protection from xenobiotics, regulating oral bioavailability, forming part of the blood–brain barrier, the blood–testis barrier, and the maternal–fetal barrier. Notably, ABCG2 is often expressed in stem cell populations, where it likely plays a role in xenobiotic protection. However, clues to its epigenetic regulation in various cell populations are only beginning to emerge. While ABCG2 overexpression has been demonstrated in cancer cells after in vitro drug treatment, endogenous ABCG2 expression in certain cancers is likely a reflection of the differentiated phenotype of the cell of origin and likely contributes to intrinsic drug resistance. Notably, research into the transporter's role in cancer drug resistance and its development as a therapeutic target in cancer has lagged. Substrates and inhibitors of the transporter have been described, among them chemotherapy drugs, tyrosine kinase inhibitors, antivirals, HMG-CoA reductase inhibitors, carcinogens, and flavonoids. This broad range of substrates complements the efficiency of ABCG2 as a transporter in laboratory studies and suggests that, while there are redundant mechanisms of xenobiotic protection, the protein is important in normal physiology. Indeed, emerging studies in pharmacology and toxicology assessing polymorphic variants in man, in combination with murine knockout models have confirmed its dynamic role. Work in pharmacology may eventually lead us to a greater understanding of the physiologic role of ABCG2.

Introduction

The efficacy of cancer chemotherapy can be limited by cellular mechanisms of resistance that result in increased drug efflux of chemotherapeutic agents thereby reducing intracellular drug levels and causing drug resistance. The ability of cells to acquire resistance to multiple compounds, termed multidrug resistance (MDR), is often mediated by overexpression of ATP-binding cassette (ABC) transporters that remove substrates out of the cell against a concentration gradient [1]. Of the 48 human ABC transporters, three are most often associated with MDR: the multidrug resistance protein, P-glycoprotein (P-gp), encoded by the ABCB1 (or MDR-1) gene; the multidrug resistance-associated protein-1 (MRP-1) encoded by the ABCC1 (or MRP-1) gene; and the breast cancer resistance protein (BCRP or ABCG2) encoded by the ABCG2 gene [1]. Other ABC transporters have been implicated in drug resistance, but these other transporters play highly specialized roles in normal physiology and are less likely to be usurped to play a role in drug resistance in a cancer cell.

Before the first transporter genes were cloned, it had long been known that incubating cancer cell lines with chemotherapy agents resulted in sublines that were not only resistant to the selecting drug, but also to other, structurally different agents [2], [3], [4]. Juliano and Ling in 1976 were the first to note that a particular 170 kD glycoprotein was associated with this resistance [5] and over a decade later the gene encoding P-gp, then termed mdr1 (and later called MDR-1), was cloned [6]. Early interest in P-gp focused on its role in drug resistance since it was responsible for the transport of a wide variety of chemotherapeutic agents such as anthracyclines, vinca alkaloids, taxanes and etoposide [1]. Today, the importance of P-gp is understood to go well beyond drug resistance, since the high levels of expression in epithelial cells of the gastrointestinal tract and brain capillary endothelium have led to experiments showing that P-gp mediates oral absorption and forms part of the blood–brain barrier (BBB) [7], [8]. P-gp expression in the proximal tubules of the kidney suggests it plays a role in drug excretion [9]. Thus, the significance of P-gp has gone beyond that of a multidrug resistance transporter.

The adriamycin-selected leukemia subline, HL-60/AR, was reported to have a cross-resistance profile slightly different from that observed for cells expressing the MDR-1 gene [10], but was not found to overexpress MDR-1 compared to parental cells [11]. Additionally, a doxorubicin-selected, small-cell lung cancer cell line, H69/AR, and a doxorubicin-selected fibrosarcoma cell line, HT1080/DR4, were also found to exhibit a pattern of drug resistance similar to that of the HL-60/AR cells; a pattern nonetheless distinct from that conferred by expression of P-glycoprotein [12]. A new drug resistance gene, the multidrug resistance-associated protein gene, or MRP (later renamed MRP1), was later cloned by Cole et al. from the H69/AR subline [13]. Later studies revealed MRP1 conferred resistance to drugs that were also transported by P-gp: anthracyclines, vinca alkaloids, mitoxantrone and etoposide [14]. Much like P-gp, the importance of MRP1 is believed to extend beyond conferring drug resistance, as it is also an organic ion transporter, transporting compounds conjugated to glutathione, glucuronide, or sulfate [15].

Still another phenotype, similar but distinct from that found in cells expressing P-gp or MRP1, was reported in cells selected with mitoxantrone [16], [17]. These cells lacked MDR-1 and MRP1 expression and were highly cross resistant to mitoxantrone as well as topotecan, camptothecin, 9-aminocamptothecin, and SN-38, but lacked cross-resistance to vinblastine [18]. A nearly identical phenotype was described in a breast cancer cell line selected by the Fojo lab with doxorubicin in the presence of verapamil to prevent overexpression of P-gp [19]. These cells, MCF-7 Adr/Vp, also displayed ATP-dependent transport of doxorubicin and the fluorescent substrate rhodamine 123 in the absence of P-gp or MRP1 [20].

It was from the MCF-7 Adr/Vp subline that Doyle et al. first cloned the gene responsible for the novel resistance phenotype [21]. They named the gene BCRP for breast cancer resistance protein since it was cloned from a breast cancer subline. Soon after, Allikmets et al. reported a nearly identical transporter, termed ABCP for ABC transporter highly expressed in placenta, after searching an expressed sequence tag database [22]. Our laboratory also cloned a gene from the mitoxantrone-selected colon carcinoma cell line S1-M1-80 [23], derived from the S1-M1-3.2 cell line reported by Rabindran et al. [24]. We called the gene MXR, or mitoxantrone resistance gene, since it appeared to be responsible for the high levels of resistance to mitoxantrone observed in cell lines expressing the gene. When the sequences for the genes became available, they proved to be nearly identical. The BCRP/ABCP/MXR gene was later placed in the “G” subfamily of ABC transporters, which includes only of half-transporters, and was assigned the name ABCG2.