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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptNIH Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
 
Curr Pharm Des. Author manuscript; available in PMC Feb 1, 2012.
Published in final edited form as:
PMCID: PMC3269897
NIHMSID: NIHMS350107
Breast Cancer Resistance Protein and P-glycoprotein in Brain Cancer: Two Gatekeepers Team Up
Sagar Agarwal,1,3 Anika M.S. Hartz,2,3 William F. Elmquist,1,3 and Björn Bauer2,3
1Department of Pharmaceutics, College of Pharmacy, University of Minnesota, Minneapolis, Minnesota, USA
2Department of Pharmaceutical Sciences, College of Pharmacy, University of Minnesota, Duluth, Minnesota, USA
3Brain Barriers Research Center, University of Minnesota, Minnesota, USA
Corresponding Author: Björn Bauer, Ph.D., University of Minnesota, College of Pharmacy, Department of Pharmaceutical Sciences, 1110 Kirby Drive, 232 Life Science, Duluth, MN 55812, USA, Phone: 218-726-6036, Fax: 218-726-6500, bjbauer/at/d.umn.edu
Brain cancer is a devastating disease. Despite extensive research, treatment of brain tumors has been largely ineffective and the diagnosis of brain cancer remains uniformly fatal. Failure of brain cancer treatment may be in part due to limitations in drug delivery, influenced by the ABC drug efflux transporters P-gp and BCRP at the blood-brain and blood-tumor barriers, in brain tumor cells, as well as in brain tumor stem-like cells. P-gp and BCRP limit various anti-cancer drugs from entering the brain and tumor tissues, thus rendering chemotherapy ineffective. To overcome this obstacle, two strategies – targeting transporter regulation and direct transporter inhibition – have been proposed. In this review, we focus on these strategies. We first introduce the latest findings on signaling pathways that could potentially be targeted to down-regulate P-gp and BCRP expression and/or transport activity. We then highlight in detail the new paradigm of P-gp and BCRP working as a “cooperative team of gatekeepers” at the blood-brain barrier, discuss its ramifications for brain cancer therapy, and summarize the latest findings on dual P-gp/BCRP inhibitors. Finally, we provide a brief summary with conclusions and outline the perspectives for future research endeavors in this field.
Keywords: BCRP, P-gp, brain cancer, glioblastoma, multidrug resistance, blood-brain barrier, regulation, inhibition
The number of new cases of malignant brain cancers has significantly increased over the last two decades [1, 2]. In 2010, an estimated 22,000 new patients were expected to be diagnosed and 13,000 patients were expected to die of brain cancer in the United States [1]. Brain cancers can be divided into two categories: primary brain tumors and metastatic brain tumors. Primary brain tumors originate in the brain and usually do not metastasize. These tumors represent only 2% of all cancers but account for a disproportionate rate of morbidity and mortality [3]. Metastatic brain tumors originate outside of the central nervous system (CNS) elsewhere in the body and spread to the brain as metastases [4]. Metastatic brain tumors develop in 10–30% of cancer patients [5], and are the most common type of brain tumors with a more than four times greater annual incidence compared to primary brain tumors [6]. The incidence of brain metastases has increased over the last decade mainly due to improved treatment of primary peripheral cancers resulting in increased patient survival, as well as due the development of newer tools to image and detect tumors of the CNS. Despite extensive research, treatment of metastatic brain tumors has been largely ineffective and the diagnosis of brain cancer remains uniformly fatal [6].
One major challenge researchers face today is effective delivery of anti-cancer drugs to primary and metastatic cancers in the CNS. The primary impediment to successful drug delivery into the CNS is the blood-brain barrier (BBB). The BBB is an endothelial interface that separates the brain from the blood and shields the CNS from exposure to circulating toxins and potentially harmful chemicals [7]. At the same time, this protective barrier excludes therapeutic drugs from entering the brain, and thus, becomes an obstacle for drugs intended to treat CNS diseases, such as brain cancers.
At the molecular level, brain capillary endothelial cells that form the BBB are joined together by tight junctions that limit paracellular passage of solutes into the brain. Circulating solutes can therefore only gain access to the brain by passive diffusion, or uptake transport [8]. The BBB is further fortified by ATP-binding cassette (ABC) efflux transporters that limit xenobiotics, including a large number of therapeutic drugs, from entering the brain. P-glycoprotein (P-gp, ABCB1 or MDR1) and breast cancer resistance protein (BCRP, ABCG2) are two prominent members of the ABC transporter superfamily. Both have broad and partly overlapping substrate specificities that include a variety of structurally diverse drugs currently used in the clinic [911]. These two “gatekeeper” transporters constitute a vital part of the protective defense mechanism at the BBB by limiting drugs from accessing the brain and thereby rendering them ineffective. Moreover, recent literature suggests that P-gp and BCRP team up and work together at the BBB to restrict brain penetration of drugs [1216].
The present review is focused on this phenomenon and the challenge that these two transporters pose to chemotherapeutic delivery into the brain. We review P-gp and BCRP with respect to their roles and regulation at the BBB, and summarize recent findings on the P-gp/BCRP teamwork in restricting brain penetration of anti-cancer drugs.
2.1 History
In 1976, Rudy Juliano and Victor Ling discovered a high molecular weight membrane glycoprotein in mutant cancer cells that appeared to alter membrane permeability for chemotherapeutics, and consequently named it P-glycoprotein (“permeability glycoprotein”; [17]). Shortly afterwards it became clear that P-glycoprotein (P-gp) is a highly potent ATP-driven efflux transporter that actively pumps its substrates out of cells, even against a concentration gradient [18]. This discovery was groundbreaking because it provided the first explanation for treatment failure due to resistance to multiple chemotherapeutics, which is a frequently observed phenomenon in cancer.
Several years later, in 1989, P-gp protein expression was detected at the human BBB [19, 20] and subsequent studies confirmed the presence of P-gp in the luminal (apical) membrane at the BBB of dogfish, killifish, mouse, rat, cat, dog, monkey, pig, and cow [2030]. In addition, P-gp was found in primary brain tumors and is now recognized to be a critical transporter that conveys resistance to a large number of anti-cancer drugs, for example taxanes, vinca-alkaloids, etoposide and analogues, anthracyclines, lanafarnib, imatinib, and topotecan [31, 32]. Thus, P-gp has been a focus of BBB, brain tumor, and drug delivery research for almost two decades.
2.2 P-glycoprotein Inhibition in Brain Cancer
One strategy to improve brain delivery of anti-cancer drugs is to directly block P-gp transport function at the BBB by using transporter inhibitors. The first P-gp inhibitor was found by serendipity in 1981 by Tsuruo et al., who showed that verapamil, a calcium channel blocker, inhibits P-gp-mediated drug efflux in resistant tumor cells, thereby overcoming drug resistance [33]. As a result, over the years numerous chemicals have been screened for their potential to inhibit P-gp, and a variety of inhibitors were developed that differ in potency, selectivity, and side effects [34, 35]. However, only a few compounds have been tested for their potential to enhance drug delivery to the brain. The first proof-of-principle that P-gp inhibition can be used to treat brain cancer came from a study in nude mice with intracerebrally implanted human U-118 MG glioblastoma [36]. In this study, Fellner et al. identified P-gp as the major factor in limiting the anti-cancer therapeutic paclitaxel from crossing the BBB and permeating into the CNS [36]. Consistent with this, treating glioblastoma-bearing mice with paclitaxel had no effect on tumor size but pretreating mice with the P-gp inhibitor PSC833 (valspodar) increased paclitaxel brain levels and reduced tumor size by 90% [36]. Subsequent studies using the P-gp inhibitors cyclosporine A, elacridar (GF120918), tariquidar (XR9576), and zosuquidar (LY335979) confirmed these findings and demonstrated that P-gp inhibition increases paclitaxel brain levels [3740]. It has now been shown that elacridar and tariquidar are not P-gp-specific inhibitors, but at higher concentrations also inhibit BCRP [4143].A recent study also demonstrated that oral, bi-weekly co-administration of the new P-gp inhibitor HM30181A with paclitaxel decreased tumor volume of K1735 melanoma brain metastases and U-87 MG glioblastoma in cancer animal models [44].
Together, direct P-gp inhibition improves brain drug delivery of some anti-cancer drugs and treatment of brain tumors in animal models. Currently, no reports are available on the use of the above mentioned transporter inhibitors in brain cancer patients. However, tariquidar is being tested in an ongoing phase I clinical trial to treat brain cancer among other cancer types in children (www.clinicaltrials.gov; NCT00020514v). Thus, it remains to be demonstrated if the strategy of transporter inhibition can be translated from animal model to patient. Hence, the search for more potent, efficacious, and selective P-gp inhibitors continues.
2.3 Targeting P-gp Regulation
In this review, we will also comment on signaling pathways that affect P-gp and BCRP at the blood-brain and blood-tumor barriers, in brain tumors and brain cancer stem cells and that could potentially be used to improve delivery of chemotherapeutics. In this context, the goal of targeting transporter regulation is to down-regulate transporter expression and/or functional activity, thereby reducing drug efflux and overcoming drug resistance.
The field of BBB transporter regulation is relatively new and only few studies have been conducted. The first study demonstrating P-gp down-regulation at the BBB was published in 2004. This study focused on ET-1 signaling through the ETB receptor, NOS, and PKC, which rapidly decreased P-gp activity in isolated rat brain capillaries [45]. Another report showed that BBB P-gp is regulated by the inflammatory mediators LPS, TNF-α, and ET-1, which activated TLR4, TNFR1, ETB receptor, NOS, and PKC, leading to reduced P-gp activity [46]. In a follow-up study, Rigor et al. identified PKC beta(I) as the responsible PKC isoform for down-regulation of P-gp activity in this pathway [47]. Importantly, this study provides proof-of-principle that targeting PKC beta(I) increases brain uptake of the P-gp substrate verapamil.
In another study, Hawkins et al. made a similar observation using vascular endothelial growth factor (VEGF; [48]). It was shown that VEGF decreased P-gp activity in rat brain capillaries via activation of flk-1 and Src, likely through Src-mediated phosphorylation of caveolin-1. This finding implies that P-gp activity could be acutely diminished in pathological conditions associated with increased brain VEGF expression and that VEGF/Src signaling at the BBB could be targeted to decrease P-gp activity [48].
Together, two signaling pathways have been identified that could potentially be used to down-regulate P-gp transport activity at the BBB; one involves signaling of inflammatory mediators through PKC beta(I), the other one involves VEGF signaling though flk-1 and Src. It remains to be demonstrated if targeting these pathways improves delivery of chemotherapeutics across the BBB and into brain tumors. For more details on signaling pathways that regulate P-gp at the BBB we refer the reader to [49].
3.1 History
In 1998, more than 20 years after the discovery of P-gp, Doyle et al. cloned the ABC transporter breast cancer resistance protein (BCRP, MXR: mitoxantrone resistance protein, ABCP1) from a multidrug-resistant human breast cancer cell line [50]. Four years later, in 2002, two groups found BCRP to be physiologically expressed in brain capillary endothelial cell cultures and BCRP was localized at the luminal membrane of rat and human brain capillaries [5153]. BCRP has also been detected at the human, cow, rat, and mouse BBB [5356]. In addition, BCRP is highly expressed at the plasma membrane of tumor stem cells [57, 58], where it could be involved in stem cell differentiation, protection against xenobiotics, and cancer cell survival under hypoxic conditions [59]. Little is known about BCRP expression in brain cancer. In primary CNS lymphoma, BCRP protein expression and transport activity have been shown to be down-regulated [60]. In contrast, in neuroepithelial tumors such as ependymomas and in glioma tumor stem-like cells BCRP protein and activity are highly up-regulated, causing multidrug resistance [6163].
At the functional level, BCRP is a half transporter that works as a homodimer and possibly as a heterodimer with other ABCG half transporter isoforms [52]. A significant overlap in substrate specificity between P-gp and BCRP has been demonstrated [64, 65], and anti-cancer drugs handled by BCRP include tyrosine kinase inhibitors (imatinib, nilotinib, gefitinib, erlotinib, dasatinib, sorafenib, lapatinib, apatinib, and tandutinib; [12, 13, 6669], topotecan, irinotecan, epirubicin, doxorubicin, daunorubicin, and mitoxantrone [70, 71]. Studies show that BCRP restricts these chemotherapeutics from permeating across the BBB and penetrating into brain tumors. As with P-gp, BCRP-mediated drug resistance in brain tumors is in part due to transporter up-regulation at the blood-tumor barrier contributing to reduced delivery and efficacy of anti-cancer drugs [63].
3.2 BCRP Inhibition in Brain Cancer
Few compounds have been identified that specifically inhibit BCRP. Fumitremorgin C (FTC), a fungal toxin, was the first reported BCRP inhibitor [72], but is not suitable for in vivo studies due to severe neurotoxic side effects. This lead to the development of the FTC-derivatives Ko132, Ko134, and Ko143 that are 2–3-fold more potent, less toxic, and designed for use in vivo [73]. Recent efforts focused on tyrosine kinase inhibitors such as imatinib, nilotinib, gefitinib, and erlotinib that directly interact with BCRP at the substrate binding site and that block ATPase activity of the transporter [68]. These compounds have a unique pharmacologic profile in that they are effective chemotherapeutics, as well as potent BCRP inhibitors and transporter substrates. In this regard, in vivo studies showed that BCRP, together with P-gp, limited brain uptake of imatinib and that BCRP inhibition significantly increased imatinib brain penetration [74]. In a similar study, Breedveld et al. demonstrated that inhibition of BCRP with pantoprazole increased imatinib brain levels 1.8-fold [75]. However, co-administration of the P-gp and BCRP inhibitor elacridar improved brain penetration of imatinib by 4.2-fold [75]. The same group also showed that dual BCRP/P-gp inhibition using elacridar improved oral bioavailability and CNS penetration of anti-cancer drugs [76]. To what extent elacridar inhibits P-gp and/or BCRP depends on the local inhibitor concentration. Consequently, the elacridar dose determines what drug at what amount penetrates through the BBB.
While these studies demonstrate the importance of BBB BCRP for brain uptake of anti-cancer drugs, they also show that for some compounds inhibition of either BCRP or P-gp alone is not sufficient to increase delivery into the brain.
3.3 Targeting BCRP Regulation
Various signaling pathways have been shown to down-regulate BCRP, which is expected to improve anti-cancer drug delivery into brain tumors. In this regard, it was demonstrated that estrogens play a role in BCRP regulation. Imai et al. showed that estrone and 17β-estradiol (E2) reverse BCRP-mediated drug resistance [77] and that E2 triggers post-transcriptional down-regulation of BCRP in human breast cancer cell lines [77]. Ee et al. identified an estrogen response element (ERE) in the BCRP promotor region and showed ERE activation by binding of the E2/estrogen receptor α complex, which up-regulated BCRP mRNA expression [78]. From this study, however, it is unclear if BCRP protein expression and/or transport activity were also affected by ERE activation.
Other studies that were conducted in various tissues reported BCRP regulation through the PI3K/Akt signaling pathway [79, 80]. In these studies, PI3K/Akt signaling triggered BCRP internalization and translocation from the plasma membrane to the cytoplasmic compartment in stem cells and renal epithelial cells and was involved in regulating BCRP expression [79, 80].
With regard to brain cancer, Bleau et al. published a study showing PTEN/PI3K/Akt regulation of BCRP activity in glioma tumor stem-like cells [61, 62]. In these cells, activation of Akt lead to BCRP translocation from the cytoplasm to the plasma membrane and increased BCRP-mediated efflux of anti-cancer drugs, which contributed to drug resistance and tumorigenicity. These findings are interesting considering recent studies where we demonstrated E2-mediated BCRP regulation in isolated brain capillaries and established a link between E2 and PTEN/PI3K/Akt signaling [54, 81]. We showed that E2 signaling through ERβ, PTEN/PI3K/Akt and GSK3 triggered BCRP internalization from the brain capillary plasma membrane, which was followed by proteasomal degradation of the transporter and reduced BCRP functional activity and protein expression [54, 81]. These findings suggest that PTEN/PI3K/Akt-mediated up-regulation of BCRP activity in glioma tumor stem-like cells that Bleau et al. observed [61, 62] could potentially be blocked, which may be one possibility to reduce BCRP-mediated resistance to chemotherapeutics at the level of the blood-brain and blood-tumor barriers. However, despite these studies it remains to be demonstrated if targeting BCRP regulation at the BBB, in brain tumors, and/or in brain tumor stem cells is a valid strategy to improve drug delivery of chemotherapeutics into the CNS. For more details on signaling pathways that regulate BCRP at the BBB we refer the reader to [49].
4.1 Two Gatekeepers Team Up at the Blood-Brain Barrier
The discovery of BCRP in brain endothelial cells changed the long standing opinion that P-gp is the sole important transporter responsible for efflux of drugs at the BBB. However, BCRP expression at the BBB has not been unequivocally correlated to low brain penetration of all BCRP substrates. For example, Lee et al. conducted in situ brain perfusion studies using the BCRP substrates dehydroepiandrosterone sulfate and mitoxantrone and reported that brain penetration of the two compounds was not increased in Bcrp1(−/−) (BCRP knockout) mice [82]. Si milarly, Giri and coworkers showed that BCRP mediated efflux of the antiretroviral drugs abacavir and zidovudine in vitro [83]. However, despite the absence of BCRP, brain uptake of these two compounds was not elevated in Bcrp1(−/−) mice [84]. One conclusion drawn from these studies was that BCRP played a minor role in drug efflux at the BBB and another study showed that interaction of BCRP with substrates in vitro rarely translates to visible effects at the BBB in vivo [85].
In contrast, other studies demonstrated BCRP transport activity at the BBB. Cisternino and colleagues showed that BCRP limits prazosin and mitoxantrone, two prototypical BCRP substrates, from penetrating into the brain [86]. Likewise, Enokizono et al. and Breedveld et al. reported that brain distribution of drugs increased significantly in Bcrp1(−/−) mice [75, 87]. Moreover, we recently reported that sorafenib transport into the brain was significantly increased in Bcrp1(−/−) mice [13].
Taken together, conflicting results on BCRP-mediated drug efflux from the brain initiated a controversy on the role of this transporter at the BBB that lead to further studies. With the development of the P-gp/BCRP knockout mouse (Mdr1a/1b(−/−) Bcrp1(−/−); [88]), researchers have been provided with the opportunity to study the combined impact of these two efflux transporters on the delivery of drugs across the BBB. de Vries et al. showed that brain uptake of topotecan, a substrate for both P-gp and BCRP, was not increased in mice lacking BCRP (Bcrp1(−/−)) [15]. In P-gp knockout mice (Mdr1a/1b(−/−)) topotecan brain levels increased slightly by 1.5-fold. In contrast, in mice lacking both P-gp and BCRP (Mdr1a/1b(−/−)Bcrp1(−/−)), topotecan brain uptake was increased by more than 12-fold. Thus, absence of both P-gp and BCRP resulted in an effect that was significantly larger than the combined effects from the single transporter knockout mice. This finding was confirmed by Polli et al. using lapatinib in P-gp/BCRP knockout mice [16]. We have shown the same with dasatinib [14], gefitinib [12] and sorafenib [13]. Even though these drugs are substrates for both P-gp and BCRP, absence of only one of the transporters did not significantly increase delivery of either drug to the brain, but the greatest enhancement in brain penetration was seen when both transporters were absent or inhibited at the BBB. Several studies now show that this is true for other dual P-gp and BCRP substrates as well (Table 1, [69, 89, 90]). Figure 1 summarizes recent data by Kawamura et al. [91] that demonstrate this phenomenon. These findings suggest that inhibition of either P-gp or BCRP can be compensated by the respective other transporter, and that both transporters “cooperate” with each other in preventing chemotherapeutic drugs from entering the brain.
Table 1
Table 1
Brain Distribution of Dual P-gp and BCRP Substrates
Figure 1
Figure 1
Figure 1a. Transaxial PET images showing [11C] GF120918 in the brain of a (A) wild-type, (B) P-gp knockout, (C) Bcrp knockout and (D) P-gp/Bcrp knockout mouse. GF120918 is a substrate for and inhibits both P-gp and BCRP. The radioactivity level is low (more ...)
P-gp and BCRP cooperation implies that absence of either P-gp or BCRP alone does not result in an appreciable increase in brain penetration of dual substrates. In BCRP knockout mice (where P-gp is present), P-gp alone is sufficient to prevent drugs from penetrating into the brain. Likewise, in P-gp knockout mice (where BCRP is present) BCRP alone is sufficient to limit drug uptake into the brain. The greatest enhancement in brain penetration of dual substrates is always seen when both P-gp and BCRP are absent in the combined P-gp/BCRP knockout mice (Figures 1 and 2).
An insight into the mechanism of P-gp/BCRP cooperation can be gained by looking at relative transporter affinities of substrate drugs, and relative transporter expression levels at the BBB (assuming protein expression correlates with transport capacity for both transporters). In this regard, Kamiie et al. used LC-MS to quantify membrane transporter expression at the mouse BBB and found approximately 5-fold higher P-gp protein levels compared to those of BCRP [92]. Significantly higher protein expression levels at the BBB make P-gp appear to be the dominant efflux transporter for many dual substrates that have similar affinities to both P-gp and BCRP. In comparison, due to lower protein expression levels, BCRP-mediated efflux appears to be minor and becomes apparent only when P-gp or both transporters are absent. For example, for a compound with moderate P-gp affinity, higher P-gp expression levels (higher P-gp transport capacity) will compensate for lower transporter affinity, resulting in a pronounced P-gp effect on the efflux of this compound at the BBB. This is true for almost all anti-cancer drugs mentioned above (Table 1), with the exception of sorafenib and dantrolene. Both these compounds have a significantly higher affinity for BCRP than for P-gp [13, 90]. Therefore, BCRP is the dominant transporter in keeping these drugs out of the brain and an effect of P-gp on drug penetration is only noticeable in BCRP and P-gp/BCRP knockout mice. Kodaira et al. explained P-gp/BCRP cooperation by determining the net contribution of each transporter to the overall efflux of various drugs at the BBB [90]. The authors showed that for many dual substrates, P-gp-mediated efflux out of the brain was greater than that by BCRP. On the other hand, P-gp-mediated efflux of dantrolene (high affinity BCRP substrate) was 10-fold lower than BCRP-mediated dantrolene efflux.
While the above studies have been conducted in animal models, it is now clear that BBB P-gp and BCRP expression is species-dependent. In this regard, Uchida et al. recently reported that at the human BBB, BCRP protein levels are higher compared to P-gp protein levels [93]. Using LC-MS, the authors determined 8 fmol/μg total protein for BCRP vs. 6 fmol/μg total protein for P-gp in human brain capillaries. However, to draw a clear conclusion from these absolute transporter protein levels on the importance of each transporter for brain drug delivery is difficult. LC-MS measures total transporter protein and does not distinguish between transporter protein that is functionally active in the luminal membrane of the brain capillary endothelium and transporter protein that is inactive in intracellular vesicle membranes. For example, LC-MS measures both BCRP monomer and dimer, but only BCRP dimer is the functionally active form [94]. From what we know today only functionally active transporter protein in the luminal membrane of the brain capillary endothelium affects drug delivery across the BBB. Thus, although total BCRP protein expression at the human BBB is higher compared to P-gp, it is impossible to say at this point in time which transporter is more important for brain drug delivery in patients. To make such a statement we will need information on the functional expression of each transporter at the BBB, the local drug concentration, and the drug-transporter affinity.
P-gp/BCRP cooperation at the BBB suggests two fundamental realities. First, these two transporters can significantly affect drug delivery to the brain, thereby influencing drug efficacy. Second, combined inhibition of both P-gp and BCRP is potentially an attractive therapeutic strategy to improve delivery and thus efficacy of substrate drugs in the CNS. Many of the chemotherapeutic drugs mentioned above have been clinically unsuccessful in treating brain cancers. Even though P-gp- and BCRP-mediated cooperative efflux transport is not limited to anti-cancer agents, combined inhibition of both transporters might have the biggest impact in the treatment of brain cancers, where a small increase in drug brain uptake might dramatically improve anti-cancer efficacy.
In summary, absence of either P-gp or BCRP alone does not enhance brain distribution of dual substrates, but genetic or chemical knockout of both transporters is required to significantly increase brain uptake of dual P-gp/BCRP substrate anti-cancer drugs. Thus, current research indicates that P-gp and BCRP team up at the BBB and “cooperate” in preventing dual substrates from entering the brain. This finding has lead to a paradigm shift in the field of BBB transporter research.
4.2 Dual Inhibition of P-gp and BCRP at the BBB
Given the cooperation of P-gp and BCRP at the BBB, developing compounds that are potent inhibitors of both transporters may prove beneficial. Elacridar (GF120918) is a dual P-gp/BCRP inhibitor that has undergone extensive preclinical and clinical evaluation [95]. Elacridar has been used in several preclinical studies to inhibit P-gp and BCRP at the BBB with the purpose of enhancing brain distribution of simultaneously administered compounds [40, 9699]. These studies demonstrated that the greater than additive increase in brain penetration is not restricted to P-gp/BCRP knockout animals, but can also be observed with dual P-gp/BCRP inhibitors. For example, Chen et al. showed that brain penetration of dasatinib increased dramatically with co-administration of elacridar [14]. Likewise, we showed that elacridar significantly enhanced gefitinib and sorafenib brain uptake [12, 13], and de Vries et al. published similar findings for topotecan [15]. Thus, in preclinical studies, elacridar significantly increased brain penetration of drugs that are dual P-gp/BCRP substrates.
Apart from these compounds that were developed for use as multi-drug resistance reversal agents, several studies examined drugs that are dual P-gp/BCRP substrates to competitively inhibit both transporters. These include several anti-cancer tyrosine kinase inhibitors that have been shown to be substrates for both P-gp and BCRP. In vitro studies show that tyrosine kinase inhibitors like erlotinib [100], gefitinib [101], lapatinib [102] and sunitinib [103] inhibit ABC transporters, mainly P-gp and BCRP, and suggest the potential use of these agents as combination therapy to improve drug pharmacokinetics. In 2006, Zhuang et al. showed that concurrent administration of gefitinib results in a significant increase in brain penetration of topotecan [104]. The same group showed that gefitinib also increased intracellular tumor exposure to topotecan in a mouse model of glioma [105]. In a recent clinical trial, Furman and coworkers used gefitinib to inhibit intestinal P-gp and BCRP and showed that it increased oral bioavailability of irinotecan [106]. An interesting study by Nakanishi et al. in 2006 showed that imatinib attenuated its BCRP-mediated resistance by suppressing BCRP expression [107]. The underlying mechanism for these differential responses involved downstream effects of imatinib leading to decreased phosphorylation of Akt, subsequently leading to reduced BCRP expression [107]. Many tyrosine kinase inhibitors have an inhibitory effect on the PTEN/PI3K/Akt signaling pathway. These drugs can thus reduce functional activity and protein expression of ABC transporters, especially BCRP, by blockade of PI3K/Akt signaling. Combination of tyrosine kinase inhibitors with other anti-cancer drugs can therefore have a bimodal effect on ABC transporters, wherein decreased transporter expression/function coupled with competitive inhibition can result in significantly increased drug penetration across the BBB and potentially substantially increased drug levels in brain tumors.
In summary, concurrent treatment with dual P-gp/BCRP inhibitors can improve delivery and thus efficacy of substrate drugs in the CNS. Recent data imply that the use of tyrosine kinase inhibitors to inhibit P-gp/BCRP could have multiple benefits, especially if the anti-cancer agent enhances its own delivery to the brain.
Recent studies indicate that the integrity of the BBB in brain tumors (“blood-brain tumor barrier”) is compromised, questioning its role in limiting delivery of chemotherapeutics into brain tumors. Indeed, reports show that anticancer drug concentrations in resected tumor tissue are remarkably high. In this regard, Pitz et al. provided a summary of anticancer drug concentrations in brain tumors and showed that drug concentrations are high in contrast enhancing tumor areas [108]. Hofer and coworkers demonstrated that gefitinib concentrations in brain tumors were about 10-fold higher compared to gefitinib plasma levels [109]. Likewise, Blakeley et al. showed that local methotrexate levels in brain tumors were significantly greater than methotrexate plasma levels [110]. All these reports suggest that the BBB is disrupted in brain tumor tissue and does not restrict drug delivery to the tumor.
Glioblastomas are an example of a highly invasive brain tumor, with a central core that is a necrotic mass, where the BBB is most likely disrupted. Chemotherapeutics can easily traverse the impaired barrier and reach the tumor, which explains the high tumor drug concentrations that have been reported. However, tissue at the tumor borders that is immediately adjacent to healthy brain parenchyma may have an intact BBB that restricts drug delivery. In this regard, Pitz et al. reported that anti-cancer drug concentrations in non-contrast enhancing brain areas were low compared to drug concentrations in tumor tissue [108]. Blakeley et al. also showed that methotrexate brain penetration was significantly lower in areas adjacent to tumors compared to the tumor core [110].
Thus, the BBB is compromised and disrupted in the tumor core (blood-brain tumor barrier), but may be fully intact at the growing tumor border ([111]; Figure 3). This phenomenon has significant clinical implications for chemotherapeutic treatment after surgical removal of the primary tumor core. Residual tumor cells in the tumor border with intact barrier limit anti-cancer drug uptake; yet, it is these cells that often grow into larger and more aggressive tumors [112]. Therefore, it is important to highlight the need for efficient treatment of residual tumor cells in invasive areas after surgery [113]. A detailed review on this topic has recently been published by Agarwal et al. [114].
Figure 3
Figure 3
Comparison of BBB and Blood-Brain Tumor Barrier
Recent brain cancer research demonstrates that the BBB drug efflux transporter BCRP is, in addition to P-gp, another obstacle for delivering chemotherapeutic drugs into the brain. It is now clear that both BCRP and P-gp are important elements of barrier function and their expression and transport activity are regulated by distinct signaling pathways. For many anti-cancer drugs it was shown that inhibition of one of the two transporters is not sufficient to deliver drugs into the brain because of compensation by the respective other transporter. These findings lead to the currently accepted paradigm that P-gp and BCRP work as a “cooperative team of gatekeepers” at the BBB. Such P-gp/BCRP teamwork efficiently protects the brain, but at the same time prevents effective CNS therapy, which poses a tremendous clinical problem for the treatment of brain cancers. Two strategies have been developed to circumvent P-gp and BCRP at the BBB and improve drug delivery to the brain. One strategy is to target signaling pathways that control P-gp and BCRP with the goal of down-regulating transporter function and/or expression. Several pathways have been identified for P-gp and one has recently been found for BCRP. However, a common pathway for both transporters that could potentially be targeted for therapeutic purposes has not been identified yet. A second strategy is combined inhibition of both P-gp and BCRP at the BBB that has been demonstrated to significantly increase brain uptake of chemotherapeutics that are dual P-gp/BCRP substrates. These findings are remarkable and provide a glimpse of hope, but also raise the question: Where do we go from here?
Future research in this field will have to address several points. First, it will be critical to determine if increases in anti-cancer drug brain levels by combined P-gp/BCRP inhibition or down-regulation of transporter function halts brain tumor growth and reduces tumor size. Second, it will also be important to assess if therapeutic effects on brain tumor growth and size translate into prolonged survival. Third, studies will have to demonstrate if inhibiting or down-regulating P-gp/BCPR is a valid therapeutic strategy that can be used chronically over the long term. Fourth, it will have to be tested if P-gp/BCRP inhibition or down-regulation leads to sustained treatment success or if other drug resistance mechanisms will evolve and undo any therapeutic progress made. Lastly, it will have to be shown if an arrest in tumor growth can be treated as a chronic disease or if brain tumors and brain tumor stem-like cells can be completely eradicated. These challenging questions will have to be answered in brain tumor animal models first before translation to patients can occur.
Figure 2A
Figure 2A
Cooperation of P-gp and BCRP at the Blood-Brain Barrier
Figure 2B
Figure 2B
Impact of P-gp/BCRP Cooperation on Brain Distribution of Three Hypothetical Dual Substrates
Acknowledgements
We thank Britt Johnson for editorial assistance. Financial support for S.A. was provided by the Doctoral Dissertation Fellowship from the University of Minnesota. This work was also supported by a grant from the National Institutes of Health - National Cancer Institute [CA138437] (W.F.E.), a grant from the Children's Cancer Research Fund at the University of Minnesota (W.F.E.), and by a grant from the Whiteside Institute for Clinical Research (A.M.S.H., B.B.).
Footnotes
Conflict of Interest The authors declare no conflict of interest.
[1] Altekruse SF, Kosary CL, Krapcho M, Neyman N, Aminou R, Waldron W, et al. SEER Cancer Statistics Review, 1975–2007. National Cancer Institute; Bethesda, MD: 2010. http://seer.cancer.gov/csr/1975_2007/, based on November 2009 SEER data submission, posted to the SEER web site.
[2] CBTRUS Central Brain Tumor Registry of the United States. Hinsdale, IL: 2010. CBTRUS Statistical Report: Primary Brain and Central Nervous System Tumors Diagnosed in the United States in 2004–2006. website: www.cbtrus.org.
[3] Norden AD, Wen PY, Kesari S. Brain metastases. Curr Opin Neurol. 2005;18(6):654–61. [PubMed]
[4] Palmieri D, Chambers AF, Felding-Habermann B, Huang S, Steeg PS. The biology of metastasis to a sanctuary site. Clin Cancer Res. 2007;13(6):1656–62. [PubMed]
[5] Kamar FG, Posner JB. Brain metastases. Semin Neurol. 2010;30(3):217–35. [PubMed]
[6] Chamberlain MC. Anticancer therapies and CNS relapse: overcoming blood-brain and blood-cerebrospinal fluid barrier impermeability. Expert Rev Neurother. 2010;10(4):547–61. [PubMed]
[7] Hawkins BT, Davis TP. The blood-brain barrier/neurovascular unit in health and disease. Pharmacol Rev. 2005;57(2):173–85. [PubMed]
[8] Pardridge WM. Blood-brain barrier biology and methodology. J Neurovirol. 1999;5(6):556–69. [PubMed]
[9] Begley DJ. ABC transporters and the blood-brain barrier. Curr Pharm Des. 2004;10(12):1295–312. [PubMed]
[10] Loscher W, Potschka H. Blood-brain barrier active efflux transporters: ATP-binding cassette gene family. NeuroRx. 2005;2(1):86–98. [PubMed]
[11] Sun H, Dai H, Shaik N, Elmquist WF. Drug efflux transporters in the CNS. Adv Drug Deliv Rev. 2003;55(1):83–105. [PubMed]
[12] Agarwal S, Sane R, Gallardo JL, Ohlfest JR, Elmquist WF. Distribution of gefitinib to the brain is limited by P-glycoprotein (ABCB1) and breast cancer resistance protein (ABCG2)-mediated active efflux. J Pharmacol Exp Ther. 2010;334(1):147–55. [PubMed]
[13] Agarwal S, Sane R, Ohlfest JR, Elmquist WF. Role of Breast Cancer Resistance Protein (ABCG2/BCRP) in the Distribution of Sorafenib to the Brain. J Pharmacol Exp Ther. 2010 [PubMed]
[14] Chen Y, Agarwal S, Shaik NM, Chen C, Yang Z, Elmquist WF. P-glycoprotein and breast cancer resistance protein influence brain distribution of dasatinib. J Pharmacol Exp Ther. 2009;330(3):956–63. [PubMed]
[15] de Vries NA, Zhao J, Kroon E, Buckle T, Beijnen JH, van Tellingen O. P-glycoprotein and breast cancer re-sistance protein: two dominant transporters working together in limiting the brain penetration of topotecan. Clin Cancer Res. 2007;13(21):6440–9. [PubMed]
[16] Polli JW, Olson KL, Chism JP, John-Williams LS, Yeager RL, Woodard SM, et al. An unexpected synergist role of P-glycoprotein and breast cancer resistance protein on the central nervous system penetration of the tyrosine kinase inhibitor lapatinib (N-{3-chloro-4-[(3-fluorobenzyl)oxy]phenyl}-6-[5-({[2-(methylsulfonyl)ethy l]amino}methyl)-2-furyl]-4-quinazolinamine; GW572016) Drug Metab Dispos. 2009;37(2):439–42. [PubMed]
[17] Juliano RL, Ling V. A surface glycoprotein modulating drug permeability in Chinese hamster ovary cell mutants. Biochim Biophys Acta. 1976;455(1):152–62. [PubMed]
[18] Sharom FJ. The P-glycoprotein efflux pump: how does it transport drugs? J Membr Biol. 1997;160(3):161–75. [PubMed]
[19] Thiebaut F, Tsuruo T, Hamada H, Gottesman MM, Pastan I, Willingham MC. Immunohistochemical localization in normal tissues of different epitopes in the multidrug transport protein P170: evidence for localization in brain capillaries and crossreactivity of one antibody with a muscle protein. J Histochem Cytochem. 1989;37(2):159–64. [PubMed]
[20] Cordon-Cardo C, O'Brien JP, Casals D, Rittman-Grauer L, Biedler JL, Melamed MR, et al. Multidrug-resistance gene (P-glycoprotein) is expressed by endothelial cells at blood-brain barrier sites. Proc Natl Acad Sci U S A. 1989;86(2):695–8. [PubMed]
[21] Jette L, Beliveau R. P-glycoprotein is strongly expressed in brain capillaries. Adv Exp Med Biol. 1993;331:121–5. [PubMed]
[22] Jette L, Tetu B, Beliveau R. High levels of P-glycoprotein detected in isolated brain capillaries. Biochim Biophys Acta. 1993;1150(2):147–54. [PubMed]
[23] Miller DS. Xenobiotic export pumps, endothelin signaling, and tubular nephrotoxicants--a case of molecular hijacking. J Biochem Mol Toxicol. 2002;16(3):121–7. [PubMed]
[24] Nobmann S, Bauer B, Fricker G. Ivermectin excretion by isolated functionally intact brain endothelial capillaries. Br J Pharmacol. 2001;132(3):722–8. [PubMed]
[25] Pekcec A, Schneider EL, Baumgartner W, Stein VM, Tipold A, Potschka H. Age-dependent decline of blood-brain barrier P-glycoprotein expression in the canine brain. Neurobiol Aging. 2009 [PubMed]
[26] Samoto K, Ikezaki K, Yokoyama N, Fukui M. P-glycoprotein expression in brain capillary endothelial cells after focal ischaemia in the rat. Neurol Res. 1994;16(3):217–23. [PubMed]
[27] Schlachetzki F, Pardridge WM. P-glycoprotein and caveolin-1alpha in endothelium and astrocytes of primate brain. Neuroreport. 2003;14(16):2041–6. [PubMed]
[28] Sugawara I. Expression and functions of P-glycoprotein (mdr1 gene product) in normal and malignant tissues. Acta Pathol Jpn. 1990;40(8):545–53. [PubMed]
[29] Tsai CE, Daood MJ, Lane RH, Hansen TW, Gruetzmacher EM, Watchko JF. P-glycoprotein expression in mouse brain increases with maturation. Biol Neonate. 2002;81(1):58–64. [PubMed]
[30] van der Valk P, van Kalken CK, Ketelaars H, Broxterman HJ, Scheffer G, Kuiper CM, et al. Distribution of multi-drug resistance-associated P-glycoprotein in normal and neoplastic human tissues. Analysis with 3 monoclonal antibodies recognizing different epitopes of the P-glycoprotein molecule. Ann Oncol. 1990;1(1):56–64. [PubMed]
[31] Schinkel AH, Smit JJ, van Tellingen O, Beijnen JH, Wagenaar E, van Deemter L, et al. Disruption of the mouse mdr1a P-glycoprotein gene leads to a deficiency in the blood-brain barrier and to increased sensitivity to drugs. Cell. 1994;77(4):491–502. [PubMed]
[32] Tsuji A. P-glycoprotein-mediated efflux transport of anticancer drugs at the blood-brain barrier. Ther Drug Monit. 1998;20(5):588–90. [PubMed]
[33] Tsuruo T, Iida H, Tsukagoshi S, Sakurai Y. Overcoming of vincristine resistance in P388 leukemia in vivo and in vitro through enhanced cytotoxicity of vincristine and vinblastine by verapamil. Cancer Res. 1981;41(5):1967–72. [PubMed]
[34] Avendano C, Menendez JC. Inhibitors of multidrug resistance to antitumor agents (MDR) Curr Med Chem. 2002;9(2):159–93. [PubMed]
[35] Pleban K, Ecker GF. Inhibitors of p-glycoprotein--lead identification and optimisation. Mini Rev Med Chem. 2005;5(2):153–63. [PubMed]
[36] Fellner S, Bauer B, Miller DS, Schaffrik M, Fankhanel M, Spruss T, et al. Transport of paclitaxel (Taxol) across the blood-brain barrier in vitro and in vivo. J Clin Invest. 2002;110(9):1309–18. [PMC free article] [PubMed]
[37] Hubensack M, Muller C, Hocherl P, Fellner S, Spruss T, Bernhardt G, et al. Effect of the ABCB1 modulators elacridar and tariquidar on the distribution of paclitaxel in nude mice. J Cancer Res Clin Oncol. 2008;134(5):597–607. [PubMed]
[38] Kemper EM, Cleypool C, Boogerd W, Beijnen JH, van Tellingen O. The influence of the P-glycoprotein inhibitor zosuquidar trihydrochloride (LY335979) on the brain penetration of paclitaxel in mice. Cancer Che-mother Pharmacol. 2004;53(2):173–8. [PubMed]
[39] Kemper EM, van Zandbergen AE, Cleypool C, Mos HA, Boogerd W, Beijnen JH, et al. Increased penetration of paclitaxel into the brain by inhibition of P-Glycoprotein. Clin Cancer Res. 2003;9(7):2849–55. [PubMed]
[40] Kemper EM, Verheij M, Boogerd W, Beijnen JH, van Tellingen O. Improved penetration of docetaxel into the brain by co-administration of inhibitors of P-glycoprotein. Eur J Cancer. 2004;40(8):1269–74. [PubMed]
[41] Maliepaard M, van Gastelen MA, Tohgo A, Hausheer FH, van Waardenburg RC, de Jong LA, et al. Circumvention of breast cancer resistance protein (BCRP)-mediated resistance to camptothecins in vitro using non-substrate drugs or the BCRP inhibitor GF120918. Clin Cancer Res. 2001;7(4):935–41. [PubMed]
[42] Robey RW, Steadman K, Polgar O, Morisaki K, Blayney M, Mistry P, et al. Pheophorbide a is a specific probe for ABCG2 function and inhibition. Cancer Res. 2004;64(4):1242–6. [PubMed]
[43] Mistry P, Stewart AJ, Dangerfield W, Okiji S, Liddle C, Bootle D, et al. In vitro and in vivo reversal of P-glycoprotein-mediated multidrug resistance by a novel potent modulator, XR9576. Cancer Res. 2001;61(2):749–58. [PubMed]
[44] Joo KM, Park K, Kong DS, Song SY, Kim MH, Lee GS, et al. Oral paclitaxel chemotherapy for brain tumors: ideal combination treatment of paclitaxel and P-glycoprotein inhibitor. Oncol Rep. 2008;19(1):17–23. [PubMed]
[45] Hartz AM, Bauer B, Fricker G, Miller DS. Rapid regulation of P-glycoprotein at the blood-brain barrier by endothelin-1. Mol Pharmacol. 2004;66(3):387–94. [PubMed]
[46] Hartz AM, Bauer B, Fricker G, Miller DS. Rapid modulation of P-glycoprotein-mediated transport at the blood-brain barrier by tumor necrosis factor-alpha and lipopolysaccharide. Mol Pharmacol. 2006;69(2):462–70. [PubMed]
[47] Rigor RR, Hawkins BT, Miller DS. Activation of PKC isoform beta(I) at the blood-brain barrier rapidly de-creases P-glycoprotein activity and enhances drug delivery to the brain. J Cereb Blood Flow Metab. 2010;30(7):1373–83. [PMC free article] [PubMed]
[48] Hawkins BT, Rigor RR, Miller DS. Rapid loss of blood-brain barrier P-glycoprotein activity through transporter internalization demonstrated using a novel in situ proteolysis protection assay. J Cereb Blood Flow Metab. 2010;30(9):1593–7. [PMC free article] [PubMed]
[49] Hartz AM, Bauer B. Regulation of ABC transporters at the blood-brain barrier: new targets for CNS therapy. Mol Interv. 2010;10(5):293–304. [PubMed]
[50] Doyle LA, Yang W, Abruzzo LV, Krogmann T, Gao Y, Rishi AK, et al. A multidrug resistance transporter from human MCF-7 breast cancer cells. Proc Natl Acad Sci U S A. 1998;95(26):15665–70. [PubMed]
[51] Eisenblatter T, Galla HJ. A new multidrug resistance protein at the blood-brain barrier. Biochem Biophys Res Commun. 2002;293(4):1273–8. [PubMed]
[52] Hori S, Ohtsuki S, Tachikawa M, Kimura N, Kondo T, Watanabe M, et al. Functional expression of rat ABCG2 on the luminal side of brain capillaries and its enhancement by astrocyte-derived soluble factor(s) J Neurochem. 2004;90(3):526–36. [PubMed]
[53] Cooray HC, Blackmore CG, Maskell L, Barrand MA. Localisation of breast cancer resistance protein in microvessel endothelium of human brain. Neuroreport. 2002;13(16):2059–63. [PubMed]
[54] Hartz AM, Mahringer A, Miller DS, Bauer B. 17-beta-Estradiol: a powerful modulator of blood-brain barrier BCRP activity. J Cereb Blood Flow Metab. 2010;30(10):1742–55. [PMC free article] [PubMed]
[55] Lee G, Babakhanian K, Ramaswamy M, Prat A, Wosik K, Bendayan R. Expression of the ATP-binding cassette membrane transporter, ABCG2, in human and rodent brain microvessel endothelial and glial cell culture systems. Pharm Res. 2007;24(7):1262–74. [PubMed]
[56] Warren MS, Zerangue N, Woodford K, Roberts LM, Tate EH, Feng B, et al. Comparative gene expression profiles of ABC transporters in brain microvessel endothelial cells and brain in five species including human. Pharmacol Res. 2009;59(6):404–13. [PubMed]
[57] Islam MO, Kanemura Y, Tajria J, Mori H, Kobayashi S, Hara M, et al. Functional expression of ABCG2 transporter in human neural stem/progenitor cells. Neurosci Res. 2005;52(1):75–82. [PubMed]
[58] Zhou S, Schuetz JD, Bunting KD, Colapietro AM, Sampath J, Morris JJ, et al. The ABC transporter Bcrp1/ABCG2 is expressed in a wide variety of stem cells and is a molecular determinant of the side-population phenotype. Nat Med. 2001;7(9):1028–34. [PubMed]
[59] Krishnamurthy P, Ross DD, Nakanishi T, Bailey-Dell K, Zhou S, Mercer KE, et al. The stem cell marker Bcrp/ABCG2 enhances hypoxic cell survival through interactions with heme. J Biol Chem. 2004;279(23):24218–25. [PubMed]
[60] Sakata S, Fujiwara M, Ohtsuka K, Kamma H, Nagane M, Sakamoto A, et al. ATP-binding cassette transporters in primary central nervous system lymphoma: decreased expression of MDR1 P-glycoprotein and breast cancer resistance protein in tumor capillary endothelial cells. Oncol Rep. 2011;25(2):333–9. [PubMed]
[61] Bleau AM, Hambardzumyan D, Ozawa T, Fomchenko EI, Huse JT, Brennan CW, et al. PTEN/PI3K/Akt pathway regulates the side population phenotype and ABCG2 activity in glioma tumor stem-like cells. Cell Stem Cell. 2009;4(3):226–35. [PMC free article] [PubMed]
[62] Bleau AM, Huse JT, Holland EC. The ABCG2 resistance network of glioblastoma. Cell Cycle. 2009;8(18):2936–44. [PubMed]
[63] Ginguene C, Champier J, Maallem S, Strazielle N, Jouvet A, Fevre-Montange M, et al. P-glycoprotein (ABCB1) and breast cancer resistance protein (ABCG2) localize in the microvessels forming the blood-tumor barrier in ependymomas. Brain Pathol. 2010;20(5):926–35. [PubMed]
[64] Allen JD, Brinkhuis RF, Wijnholds J, Schinkel AH. The mouse Bcrp1/Mxr/Abcp gene: amplification and overexpression in cell lines selected for resistance to topotecan, mitoxantrone, or doxorubicin. Cancer Res. 1999;59(17):4237–41. [PubMed]
[65] Sarkadi B, Ozvegy-Laczka C, Nemet K, Varadi A. ABCG2 -- a transporter for all seasons. FEBS Lett. 2004;567(1):116–20. [PubMed]
[66] Mi YJ, Liang YJ, Huang HB, Zhao HY, Wu CP, Wang F, et al. Apatinib (YN968D1) reverses multidrug resistance by inhibiting the efflux function of multiple ATP-binding cassette transporters. Cancer Res. 2010;70(20):7981–91. [PMC free article] [PubMed]
[67] Perry J, Ghazaly E, Kitromilidou C, McGrowder EH, Joel S, Powles T. A synergistic interaction between lapatinib and chemotherapy agents in a panel of cell lines is due to the inhibition of the efflux pump BCRP. Mol Cancer Ther. 2010;9(12):3322–9. [PubMed]
[68] Shukla S, Sauna ZE, Ambudkar SV. Evidence for the interaction of imatinib at the transport-substrate site(s) of the multidrug-resistance-linked ABC drug transporters ABCB1 (P-glycoprotein) and ABCG2. Leukemia. 2008;22(2):445–7. [PubMed]
[69] Yang JJ, Milton MN, Yu S, Liao M, Liu N, Wu JT, et al. P-glycoprotein and breast cancer resistance protein affect disposition of tandutinib, a tyrosine kinase inhibitor. Drug Metab Lett. 2010;4(4):201–12. [PubMed]
[70] Han B, Zhang JT. Multidrug resistance in cancer chemotherapy and xenobiotic protection mediated by the half ATP-binding cassette transporter ABCG2. Curr Med Chem Anticancer Agents. 2004;4(1):31–42. [PubMed]
[71] Schellens JH, Maliepaard M, Scheper RJ, Scheffer GL, Jonker JW, Smit JW, et al. Transport of topoisomerase I inhibitors by the breast cancer resistance protein. Potential clinical implications. Ann N Y Acad Sci. 2000;922:188–94. [PubMed]
[72] Rabindran SK, Ross DD, Doyle LA, Yang W, Greenberger LM. Fumitremorgin C reverses multidrug resistance in cells transfected with the breast cancer resistance protein. Cancer Res. 2000;60(1):47–50. [PubMed]
[73] Allen JD, van Loevezijn A, Lakhai JM, van der Valk M, van Tellingen O, Reid G, et al. Potent and specific inhibition of the breast cancer resistance protein multidrug transporter in vitro and in mouse intestine by a novel analogue of fumitremorgin C. Mol Cancer Ther. 2002;1(6):417–25. [PubMed]
[74] Bihorel S, Camenisch G, Lemaire M, Scherrmann JM. Influence of breast cancer resistance protein (Abcg2) and p-glycoprotein (Abcb1a) on the transport of imatinib mesylate (Gleevec) across the mouse blood-brain barrier. J Neurochem. 2007;102(6):1749–57. [PubMed]
[75] Breedveld P, Pluim D, Cipriani G, Wielinga P, van Tellingen O, Schinkel AH, et al. The effect of Bcrp1 (Abcg2) on the in vivo pharmacokinetics and brain penetration of imatinib mesylate (Gleevec): implications for the use of breast cancer resistance protein and P-glycoprotein inhibitors to enable the brain penetration of imatinib in patients. Cancer Res. 2005;65(7):2577–82. [PubMed]
[76] Breedveld P, Beijnen JH, Schellens JH. Use of P-glycoprotein and BCRP inhibitors to improve oral bioavailability and CNS penetration of anticancer drugs. Trends Pharmacol Sci. 2006;27(1):17–24. [PubMed]
[77] Imai Y, Tsukahara S, Ishikawa E, Tsuruo T, Sugimoto Y. Estrone and 17beta-estradiol reverse breast cancer resistance protein-mediated multidrug resistance. Jpn J Cancer Res. 2002;93(3):231–5. [PubMed]
[78] Ee PL, Kamalakaran S, Tonetti D, He X, Ross DD, Beck WT. Identification of a novel estrogen response element in the breast cancer resistance protein (ABCG2) gene. Cancer Res. 2004;64(4):1247–51. [PubMed]
[79] Mogi M, Yang J, Lambert JF, Colvin GA, Shiojima I, Skurk C, et al. Akt signaling regulates side population cell phenotype via Bcrp1 translocation. J Biol Chem. 2003;278(40):39068–75. [PubMed]
[80] Takada T, Suzuki H, Gotoh Y, Sugiyama Y. Regulation of the cell surface expression of human BCRP/ABCG2 by the phosphorylation state of Akt in polarized cells. Drug Metab Dispos. 2005;33(7):905–9. [PubMed]
[81] Hartz AM, Madole EK, Miller DS, Bauer B. Estrogen receptor beta signaling through phosphatase and tensin homolog/phosphoinositide 3-kinase/Akt/glycogen synthase kinase 3 down-regulates blood-brain barrier breast cancer resistance protein. J Pharmacol Exp Ther. 2010;334(2):467–76. [PubMed]
[82] Lee YJ, Kusuhara H, Jonker JW, Schinkel AH, Sugiyama Y. Investigation of efflux transport of dehydroepiandrosterone sulfate and mitoxantrone at the mouse blood-brain barrier: a minor role of breast cancer resistance protein. J Pharmacol Exp Ther. 2005;312(1):44–52. [PubMed]
[83] Pan G, Giri N, Elmquist WF. Abcg2/Bcrp1 mediates the polarized transport of antiretroviral nucleosides abacavir and zidovudine. Drug Metab Dispos. 2007;35(7):1165–73. [PubMed]
[84] Giri N, Shaik N, Pan G, Terasaki T, Mukai C, Kitagaki S, et al. Investigation of the role of breast cancer resistance protein (Bcrp/Abcg2) on pharmacokinetics and central nervous system penetration of abacavir and zidovudine in the mouse. Drug Metab Dispos. 2008;36(8):1476–84. [PubMed]
[85] Zhao R, Raub TJ, Sawada GA, Kasper SC, Bacon JA, Bridges AS, et al. Breast cancer resistance protein interacts with various compounds in vitro, but plays a minor role in substrate efflux at the blood-brain barrier. Drug Metab Dispos. 2009;37(6):1251–8. [PubMed]
[86] Cisternino S, Mercier C, Bourasset F, Roux F, Scherrmann JM. Expression, up-regulation, and transport activity of the multidrug-resistance protein Abcg2 at the mouse blood-brain barrier. Cancer Res. 2004;64(9):3296–301. [PubMed]
[87] Enokizono J, Kusuhara H, Ose A, Schinkel AH, Sugiyama Y. Quantitative investigation of the role of breast cancer resistance protein (Bcrp/Abcg2) in limiting brain and testis penetration of xenobiotic compounds. Drug Metab Dispos. 2008;36(6):995–1002. [PubMed]
[88] Jonker JW, Freeman J, Bolscher E, Musters S, Alvi AJ, Titley I, et al. Contribution of the ABC transporters Bcrp1 and Mdr1a/1b to the side population phenotype in mammary gland and bone marrow of mice. Stem Cells. 2005;23(8):1059–65. [PubMed]
[89] de Vries NA, Buckle T, Zhao J, Beijnen JH, Schellens JH, van Tellingen O. Restricted brain penetration of the tyrosine kinase inhibitor erlotinib due to the drug transporters P-gp and BCRP. Invest New Drugs. 2010 [PubMed]
[90] Kodaira H, Kusuhara H, Ushiki J, Fuse E, Sugiyama Y. Kinetic analysis of the cooperation of P-glycoprotein (P-gp/Abcb1) and breast cancer resistance protein (Bcrp/Abcg2) in limiting the brain and testis penetration of erlotinib, flavopiridol, and mitoxantrone. J Pharmacol Exp Ther. 2010;333(3):788–96. [PubMed]
[91] Kawamura K, Yamasaki T, Konno F, Yui J, Hatori A, Yanamoto K, et al. Evaluation of Limiting Brain Penetration Related to P-glycoprotein and Breast Cancer Resistance Protein Using [<sup>11</sup>C]GF120918 by PET in Mice. Molecular Imaging and Biology. 2010;13(1):152–60. [PubMed]
[92] Kamiie J, Ohtsuki S, Iwase R, Ohmine K, Katsukura Y, Yanai K, et al. Quantitative atlas of membrane transporter proteins: development and application of a highly sensitive simultaneous LC/MS/MS method combined with novel in-silico peptide selection criteria. Pharm Res. 2008;25(6):1469–83. [PubMed]
[93] Uchida Y, Ohtsuki S, Katsukura Y, Ikeda C, Suzuki T, Kamiie J, et al. Quantitative targeted absolute proteomics of human blood-brain barrier transporters and receptors. J Neurochem. 2011;117(2):333–45. [PubMed]
[94] Xu J, Liu Y, Yang Y, Bates S, Zhang JT. Characterization of oligomeric human half-ABC transporter ATP-binding cassette G2. J Biol Chem. 2004;279(19):19781–9. [PubMed]
[95] Hyafil F, Vergely C, Du Vignaud P, Grand-Perret T. In vitro and in vivo reversal of multidrug resistance by GF120918, an acridonecarboxamide derivative. Cancer Res. 1993;53(19):4595–602. [PubMed]
[96] Cutler L, Howes C, Deeks NJ, Buck TL, Jeffrey P. Development of a P-glycoprotein knockout model in rodents to define species differences in its functional effect at the blood-brain barrier. J Pharm Sci. 2006;95(9):1944–53. [PubMed]
[97] Jin L, Li J, Nation RL, Nicolazzo JA. Impact of p-glycoprotein inhibition and lipopolysaccharide administration on blood-brain barrier transport of colistin in mice. Antimicrob Agents Chemother. 2010;55(2):502–7. [PMC free article] [PubMed]
[98] Oostendorp RL, Buckle T, Beijnen JH, van Tellingen O, Schellens JH. The effect of P-gp (Mdr1a/1b), BCRP (Bcrp1) and P-gp/BCRP inhibitors on the in vivo absorption, distribution, metabolism and excretion of imatinib. Invest New Drugs. 2009;27(1):31–40. [PubMed]
[99] Polli JW, Jarrett JL, Studenberg SD, Humphreys JE, Dennis SW, Brouwer KR, et al. Role of P-glycoprotein on the CNS disposition of amprenavir (141W94), an HIV protease inhibitor. Pharm Res. 1999;16(8):1206–12. [PubMed]
[100] Shi Z, Peng XX, Kim IW, Shukla S, Si QS, Robey RW, et al. Erlotinib (Tarceva, OSI-774) antagonizes ATP-binding cassette subfamily B member 1 and ATP-binding cassette subfamily G member 2-mediated drug resistance. Cancer Res. 2007;67(22):11012–20. [PubMed]
[101] Leggas M, Panetta JC, Zhuang Y, Schuetz JD, Johnston B, Bai F, et al. Gefitinib modulates the function of multiple ATP-binding cassette transporters in vivo. Cancer Res. 2006;66(9):4802–7. [PubMed]
[102] Dai CL, Tiwari AK, Wu CP, Su XD, Wang SR, Liu DG, et al. Lapatinib (Tykerb, GW572016) reverses multidrug resistance in cancer cells by inhibiting the activity of ATP-binding cassette subfamily B member 1 and G member 2. Cancer Res. 2008;68(19):7905–14. [PMC free article] [PubMed]
[103] Dai CL, Liang YJ, Chen LM, Zhang X, Deng WJ, Su XD, et al. Sensitization of ABCB1 overexpressing cells to chemotherapeutic agents by FG020326 via binding to ABCB1 and inhibiting its function. Biochem Pharmacol. 2009;78(4):355–64. [PMC free article] [PubMed]
[104] Zhuang Y, Fraga CH, Hubbard KE, Hagedorn N, Panetta JC, Waters CM, et al. Topotecan central nervous system penetration is altered by a tyrosine kinase inhibitor. Cancer Res. 2006;66(23):11305–13. [PubMed]
[105] Carcaboso AM, Elmeliegy MA, Shen J, Juel SJ, Zhang ZM, Calabrese C, et al. Tyrosine kinase inhibitor gefitinib enhances topotecan penetration of gliomas. Cancer Res. 2010;70(11):4499–508. [PMC free article] [PubMed]
[106] Furman WL, Navid F, Daw NC, McCarville MB, McGregor LM, Spunt SL, et al. Tyrosine kinase inhibitor enhances the bioavailability of oral irinotecan in pediatric patients with refractory solid tumors. J Clin Oncol. 2009;27(27):4599–604. [PMC free article] [PubMed]
[107] Nakanishi T, Shiozawa K, Hassel BA, Ross DD. Complex interaction of BCRP/ABCG2 and imatinib in BCR-ABL-expressing cells: BCRP-mediated resistance to imatinib is attenuated by imatinib-induced reduction of BCRP expression. Blood. 2006;108(2):678–84. [PubMed]
[108] Pitz MW, Desai A, Grossman SA, Blakeley JO. Tissue concentration of systemically administered antineoplastic agents in human brain tumors. J Neurooncol. 2011 [PubMed]
[109] Hofer S, Frei K. Gefitinib concentrations in human glioblastoma tissue. J Neurooncol. 2007;82(2):175–6. [PubMed]
[110] Blakeley JO, Olson J, Grossman SA, He X, Weingart J, Supko JG, et al. Effect of blood brain barrier permeability in recurrent high grade gliomas on the intratumoral pharmacokinetics of methotrexate: a microdialysis study. J Neurooncol. 2009;91(1):51–8. [PubMed]
[111] Deeken JF, Loscher W. The blood-brain barrier and cancer: transporters, treatment, and Trojan horses. Clin Cancer Res. 2007;13(6):1663–74. [PubMed]
[112] Cheshier SH, Kalani MY, Lim M, Ailles L, Huhn SL, Weissman IL. A neurosurgeon's guide to stem cells, cancer stem cells, and brain tumor stem cells. Neurosurgery. 2009;65(2):237–49. discussion 49–50; quiz N6. [PubMed]
[113] Wen PY, Kesari S. Malignant gliomas in adults. N Engl J Med. 2008;359(5):492–507. [PubMed]
[114] Agarwal S, Sane R, Oberoi R, Olhlfest JR, Elmquist WF. Delivery of molecularly targeted therapy to malignant glioma, a disease of the whole brain. Expert Rev Mol Med. 2011 [PubMed]