Although the physiologic functions of ABC transporters are not well known, they are expressed constitutively in not only tumor cells but also normal cells in the digestive system including the small intestine, large intestine, liver, and pancreas; epithelial cells in the kidneys, adrenals, brain, and testes; and endothelial cells (Table 1). From the aspect of the tissue distribution, ABC transporters are thought to participate in the absorption and secretion of endogenous and exogenous substances. Endogenous and exogenous substrates for ABC transporters reported so far are summarized in Table 2. Especially, the ABC transporters have shown to function as an efflux pump for lipid, multiple drugs, natural products and peptides. It is proposed to operate as a hydrophobic vacuum cleaner, expelling non-polar compounds from the membrane bilayer to the exterior, driven by the energy of ATP hydrolysis 143. ATP-dependent transbilayer lipid transporters are classified into cytofacially-directed flippases and exofacially-directed floppases. Floppase activity has been associated with the ABC transporters although not all ABC transporters are floppases 144. Endogenous substrates for Pgp include corticosterone 145, beta-estradiol 17beta-D-glucuronide, an endogenous cholestatic metabolite of estradiol 146, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine (generically platelet-activating factor, PAF) 147, glutamate 148 and endorphin 149. It was also recently reported that Pgp has the function of removing beta-amyloid, which was reported as the causal substance of Alzheimer's disease 150151. MRP1 effluxes various conjugated substrates such as leukotriene C4 conjugates 152, steroid conjugates 153 and the GSH conjugate of aflatoxin B1, which is a mycotoxin 154. Cells can, upon hypoxic demand, use BCRP to reduce heme or porphyrin accumulation, which can be detrimental to cells 155. When cancer originates not only from cells normally expressing efflux pump but also cells having genes but not expressing, gene expression is initiated due to the exposure to anticancer drugs, resulting in resistance to anticancer drugs, eventually interfering with chemotherapy.

Pgp is mainly present in the apical membrane of intestinal mucosal membrane and lowers bioavailability of drugs by preventing the absorption of the drugs. Digoxin, which shows a low bioavailability and is mainly excreted through stool in normal mice due to poor absorption in the mouse intestine, shows a high bioavailability and mainly excreted through urine in mice with mdr1 knocked out156. The bioavailability of the substrate of Pgp, paclitaxel, also increased significantly in mice with mdr1 knocked out and in mice administered with the Pgp inhibitor, PSC-833 157.

Recently, multiple MDR1 polymorphisms including more than 20 single nucleotide polymorphism (SNP) have been identified. The mutations at positions 2677(G→T) and 2995(G →A) of MDR1 in normal cells were firstly reported 158. MDR1 polymorphism could be not only associated with alteration of Pgp expression and/or function, drug disposition and treatment outcome but also increase the risk of diseases such as Parkinson's disease and ulcerative colitis 159. The influence of MDR1 SNP(C3435T and G2677T) on disposition of Pgp substrates or treatment outcome has been examplified in digoxin, phenytoin, fexofenadine, nelfinarvir, cyclosporine, talinolol and loperamide 159. Polymorphisms of other ABC transporters have been reported 160161162163.

If a substance in food affects Pgp, this substance also could affect the bioavailability of substrate drugs for Pgp. It was reported that substances present in grape juice or orange juice could increase the bioavailability of a drug being the substrate of Pgp by inhibiting it. 164. These substances could also affect pharmacokinetics of other drugs 160165. On the contrary, some drugs could increase the expression of Pgp. St John's Wort used as an antidepressant increases the expression of Pgp, so it could significantly lower the serum concentration of indinavir or cyclosporin 166. Digoxin is the substrate of Pgp and induces paclitaxel resistance by increasing Pgp 166. Not only Pgp but also MRP and BCRP could affect the bioavailability of drugs.

One of the important physiological functions of efflux pump present in the cell membrane is to provide a pharmacological sanctuary for tissues present in the blood-tissue barriers such as in the case of blood-brain barrier (BBB), blood-placental barrier and blood-testes barrier. Hydrophilic substances present in blood could not go into tissues when they are not small enough to pass through the tight junction with simple diffusion. Nonetheless, various hydrophobic substances could not enter these tissues because they are effluxed out by efflux pumps. Actually, Pgp effluxes neurotransmitters or neuromodulators such as glutamate 148 and opioids 149167 into blood from the brain. Compared with wild-type mice, drugs beings the substrate of Pgp were significantly increased in the brain of fetus when the mdr1 gene is knocked out in mice 168169170171. When the BCRP inhibitor, GF120918, was introduced to pregnant mice, the topotecan level was increased by two-folds in mouse fetus, suggesting that BCRP would function as the maternal-fetal barrier in the placenta 172. Thus, quantitative and qualitative changes of transporters present in the membrane could affect pharmacokinetics such as the distribution of endogenous and exogenous substances.

Pgp is a 170-kDa membrane protein glycosylated at the first extracellular loop (Fig. 1). Pgp is composed of 12 hydrophobic transmembrane domains (TMDs) and 2 nucleotide-binding domain (NBD). One NBD connects two TMDs with a hydrophilic NBD loop. TDMs form channels for substrate drugs, determine the characteristics of substrate, and efflux substrate drugs whereas NBDs are located in the interior of cytoplasm, and participate in ATP binding and hydrolysis 173. Pgp undergoes conformational changes upon binding of nucleotide to the NBDs 174. Rosenberg et al. have analyzed the three-dimensional structures of Pgp and its conformational change in the presence and absence of nucleotide 175176177. The projection of the protein perpendicular to the membrane is roughly rectangular with a maximum depth of 8 nm, a pore size of 2.5 nm and two 3-nm lobes exposed at the cytoplasmic face of the membrane. The conformational change revealed a major reorganization of the TMDs throughout the entire depth of the membrane upon binding of nucleotide (Fig. 2A). In the absence of nucleotide, the two TMDs form a single barrel 5–6 nm in diameter and about 5 nm deep with a central pore that is open to the extracellular surface and spans much of the membrane depth. Upon binding nucleotide, the TMDs reorganize into three compact domains that are each 2–3 nm in diameter and 5–6 nm deep (Fig. 2B). This reorganization opens the central pore along its length in a manner that could allow access of hydrophobic drugs (transport substrates) directly from the lipid bilayer to the central pore of the transporter 176.

When one of two NBDs of Pgp is inactivated, not only drug transport but also ATP hydrolysis of normal NBD is inhibited. This result indicates that two NBDs would function cooperatively and they could not hydrolyze ATP independently 178. It was recently reported that structural changes of NBDs are brought about when a drug binds to TMD so that the distance between NBDs is changed to affect the activity of ATPase as shown in Fig. 3179. Unlike in Pgp, however, the substrate leukotriene C4 could not be transported once NBD2 is inactivated but the substrate transport could not be inhibited when NBD1 is inactivated in MRP1 180. This result suggests that among ABC transporters, interactions of NBDs are not simple but function differently for every transport. Although the exact site and number of Pgp binding with drugs have not yet been determined, the important binding sites such as TMD 4, 5, 6, 10, 11 and 12 have been determined 181 whereas substrate drugs do not bind to NBDs 182.

One of the causes for the failure of chemotherapy in the treatment of cancer is the emergence of MDR. Once MDR appears, chemotherapy is not effective even when using high doses of drugs enough to overcome resistance, toxic effects are brought about and the resistance mechanism could be further stimulated. These problems could be resolved by the use of the anticancer drugs that could bypass the resistance mechanism. For example, we could use other anticancer drugs such as alkylating drugs (cyclophosphamide), antimetabolites (5-fluorouracil), and the anthracycline modified drugs (annamycin and doxorubicin-peptide) that would not function as the substrates of ABC transporters 183184185. The final method of overcoming resistance is to administer substances inhibiting ABC transporters with anticancer drugs at the same time. These substances would reverse resistance against anticancer drugs to eventually being sensitized for anticancer drugs so they are called chemosensitizers. They are also called MDR modulators and MDR reverters. Chemosensitizers against each transporter are summarized according to the publishing years (Table 3). Of these, some are chemosensitizers against one transporter and some others against more than two transporters.

Many drugs such as the calcium channel blocker verapamil and the immunosuppressant cyclosporin A would inhibit resistance by functioning as competitive substrates of Pgp regardless of their innate pharmacological functions. Different clinical studies also showed that these drugs could reverse resistance to anticancer drugs. Verapamil is the first reported chemosensitizer inhibiting MDR 119 and its effect was also proven in the recent clinical study 186. However, verapamil brings about cardiac toxicity at the concentration inhibiting resistance; thus, in order to resolve this problem, the attempts were made to develop (R)-verapmil 187 and verapamil analogues having lower cardiac toxicity compared with (S)-verapamil 188189. The immunosuppressant cyclosporin A was first reported to reverse resistance by acute leukemia against vincristine and daunorubicin 190. Following cyclosporin A, researchers found that other immunosuppressants including FK506 and rapamycin could inhibit MDR 191. However, when cyclosporin A is applied clinically, researchers placed efforts to develop cyclosporin analogues having few side effects due to their innate immuno suppressant effects and hepatic and renal toxicity with excellent chemosensitizing effects. As a result, PSC-833 (Valspodar), which is the non-immune suppressant analogue of cyclosporin, was developed 110. In addition to non-immunosuppressant effect, its chemosensitivity is about 10 times higher than that by cyclosporin in Pgp-mediated MDR, so clinical studies are being performed on this drug 192. Among those drugs having their innate pharmacological activities such as verapamil and cyclosporin A, those having chemosensitizing effect is called the first-generation chemosensitizers. The problems related with the first-generation chemosensitizers are that they generally show low effects and high toxicity at resistance-inhibiting doses. In order to supplement these problems, the chemosensitizers developed only for chemosensitizing effects are called the second-generation chemosensitizers, which include PSC-833, VX-710, LY335979, XR9051 and XR9576 193. Multi-national companies are pursuing the development of second-generation chemosensitizers by overcoming the problems of existing chemosensitizers (low effects, side effects, and drug-drug interaction), and some of these chemosensitizers are in the process of being tested clinically.

Most chemosensitizers bind with TMD in transporter, but steroid and flavonoid are new recently introduced chemosensitizers, which inhibit transporters by binding with NBD. The binding site of steroid is different from the binding site of ATP but is probably in the vicinity of the ATP binding site 194. On the other hand, the flavonoid, kaempferide, is bifunctional in that it would partially block the binding of the antiprogestin RU-486 in the cytoplasm domain of Pgp and block ATP binding 195 (Fig. 4). Recently, flavonoid chemosensitizers reversing Pgp-mediated MDR have been screening. It is believed that flavonoid chemosensitizers have a significant advantage with respect with a therapeutic index (Table 4). These may be second-generation flavonoid chemosensitizers 33.

The fungal toxin, fumitremorgin C (FTC), is a strong inhibitor of BCRP but its use in vivo has been limited due to its neurotoxicity 196. It was recently reported that the tetracyclic analogue of FTC, Ko143, is the most strong chemosensitizer aginst BCRP having little toxicity 140.

Since ABC transporters can be coexpressed in some types of cancer cells, the development of chemosensitizers against MRP and/or BCRP as well as Pgp has been highly demanding. These include VX-710 against Pgp and MRP 8283, GF120918 against Pgp and BCRP 104141 and tRA98006 against all three transporters 29.