Current Medicinal Chemistry - Anti-Cancer Agents, Vol. 4, No. 1, 2004
Contents
Drug
Resistance in Cancer Therapy
Guest
Editor: Jian-Ting Zhang
Identification and
Characterization of the Binding Sites of P-Glycoprotein for Multidrug
Resistance-Related Drugs and Modulators Pp. 1-17
Ahmad R. Safa
Drug Binding Domains
of MRP1 (ABCC1) as Revealed by Photoaffinity Labeling Pp. 19-30
J.M.
Karwatsky and E. Georges
Multidrug Resistance
in Cancer Chemotherapy and Xenobiotic Protection Mediated by the Half
ATP-Binding Cassette Transporter ABCG2 Pp. 31-42
B.
Han and J.-T. Zhang
Reversing Agents for
ATP-Binding Cassette (ABC) Transporters: Application in Modulating Multidrug
Resistance (MDR) Pp. 43-52
Chow
H. Lee
Structure of
Multidrug-Resistance Proteins of the ATP-Binding Cassette (ABC) Superfamily
Pp. 53-62
Guillermo
A. Altenberg
Abstracts
[Back
to top] Identification and Characterization of the
Binding Sites of P-Glycoprotein for Multidrug Resistance-Related Drugs and
Modulators
Ahmad R. Safa
A major problem in cancer treatment is the development of resistance to multiple chemotherapeutic agents in tumor cells. A major mechanism of this multidrug resistance (MDR) is overexpression of the MDR1 product P-glycoprotein, known to bind to and transport a wide variety of agents. This review concentrates on the progress made toward understanding the role of this protein in MDR, identifying and characterizing the drug binding sites of P-glycoprotein, and modulating MDR by P-glycoprotein-specific inhibitors. Since our initial discovery that P-glycoprotein binds to vinblastine photoaffinity analogs, many P-glycoprotein-specific photoaffinity analogs have been developed and used to identify the particular domains of P-glycoprotein capable of interacting with these analogs and other P-glycoprotein substrates. Furthermore, significant advances have been made in delineating the drug binding sites of this protein by studying mutant P-glycoprotein. Photoaffinity labeling experiments and the use of site-directed antibodies to several domains of this protein have allowed the localization of the general binding domains of some of the cytotoxic agents and MDR modulators on P-glycoprotein. Moreover, site-directed mutagenesis studies have identified the amino acids critical for the binding of some of these agents to P-glycoprotein. Furthermore, equilibrium binding assays using plasma membranes from MDR cells and radioactive drugs have aided our understanding of the modes of drug interactions with P-glycoprotein. Based on the available data, a topological model of P-glycoprotein and the approximate locations of its drug binding sites, as well as a proposed classification of multiple drug binding sites of this protein, is presented in this review.
[Back
to top] Drug Binding Domains of MRP1 (ABCC1) as
Revealed by Photoaffinity Labeling
J.M. Karwatsky and E.
Georges
Drug resistance is a major impediment in the treatment of cancer patients receiving single or multiple drug treatment. Efforts to reverse drug resistance of tumor cells have not been successful. In recent years, considerable emphasis has been placed on understanding the underlying mechanisms that confer drug resistance. The expression of the multidrug resistance protein 1 (MRP1 or ABCC1) in cancer cells has been shown to confer resistance to diverse classes of anti-cancer drugs. MRP1 is a member of the ATP-binding cassette (ABC) family whose function, in tumor cells, is to reduce drug accumulation through energized drug efflux. To learn more about the functions of MRP1 in tumor drug resistance, knowledge of the protein binding characteristics and the location of its binding sites are essential. Photoaffinity labeling (PAL) has emerged as a leading technique that can rapidly shed light on a protein’s drug binding characteristics and ultimately drug binding domains. Several MRP1-specific photoreactive probes have been developed. PAL of MRP1 was first demonstrated with the quinoline-based drug, IAAQ. Other studies showed that the high affinity endogenous substrate of MRP1, LTC4, has intrinsic photoreactive properties and binds within both N- and C-terminal domains of MRP1. LTC4 is conjugated to glutathione (GSH), a property common to several MRP1 substrates. In addition, several unconjugated drugs have been identified that interact with MRP1: [3H]VF-13,159, IAAQ, IACI and IAARh123. Mapping studies showed that IACI and IAARh123 bind two sites within transmembrane (TM) regions 10-11 and 16-17 of MRP1. Interestingly, the GSH-dependent PAL of [125I]azidoAG-A and [125I]LY475776 occurs within, or proximal to TM 16-17. The PAL with several analogs of GSH, IAAGSH and azidophenacyl-[35S]GSH found to interact specifically with MRP1 within TM 10-11 and TM 16-17 in addition to binding two cytoplasmic regions in MRP1, L0 and L1. This review focuses on the use of PAL for studying MRP1 interactions with various drugs and cell metabolites. Furthermore, knowledge of MRP1 drug binding domains, as identified by PAL with various photoreactive drug analogs, provides an important first step towards more detailed analyses of MRP1 binding domains.
[Back
to top] Multidrug Resistance in Cancer Chemotherapy
and Xenobiotic Protection Mediated by the Half ATP-Binding Cassette Transporter
ABCG2
B. Han and J.-T. Zhang
ABCG2, also termed BCRP/MXR/ABCP, is a half ATP-binding cassette (ABC) transporter expressed on plasma membranes. ABCG2 was independently cloned from placenta as well as cell lines selected for resistance to mitoxantrone or anthracyclines. ABCG2 consists of a nucleotide-binding domain (NBD) at the amino terminus and a transmembrane domain (TMD) at the carboxyl terminus and it is postulated to form a homodimer to perform its biological functions. Over-expression of ABCG2 in cell lines confers resistance on a wide variety of anticancer drugs including mitoxantrone, daunorubicin, doxorubicin, topotecan and epirubicin. The expression of ABCG2 has been implicated in multidrug resistance (MDR) of acute myeloid leukemia and some solid tumors. In addition, ABCG2 can transport several fluorescent dyes or toxins. ABCG2 is found to be expressed in epithelial cells of intestine and colon, liver canaliculi, and renal tubules, where it serves to eliminate the plasma level of orally administered anticancer drugs as well as ingested toxins. ABCG2 is found to be highly expressed in placenta and the luminal surface of microvessel endothelium blood-brain barrier where it may play a role in limiting the penetration of drugs, such as topotecan from the maternal plasma into the fetus and from blood to brain. A variety of inhibitors for ABCG2 including GF120918 may prove useful for sensitizing cancer cells to chemotherapy or altering the distribution of orally administered drug substrates of ABCG2. Interestingly, ABCG2 is also expressed highly in hematopoietic stem cells. However, the function of ABCG2 in stem cells is currently unknown, although it may provide protection to stem cells from a variety of xenobiotics.
[Back
to top] Reversing Agents for ATP-Binding Cassette
(ABC) Transporters: Application in Modulating Multidrug Resistance (MDR)
Chow H. Lee
One of the main reasons for the failure in cancer chemotherapy is the existence of multidrug resistance (MDR) mechanisms. One form of MDR phenotype is contributed by a group of plasma membrane proteins that belong to a large superfamily of proteins called the ATP-binding cassette (ABC) transporters. There has been intense search for compounds, which can act at reversing MDR phenotype exhibited by ABC transporters such as P-glycoprotein (P-gp), multidrug-resistance protein (MRP) and breast cancer resistance protein (BCRP). Reversing agents can be designed to target MDR-associated ABC transporters at three levels - the protein, mRNA or DNA level. This review aims at describing, over-viewing and discussing currently known MDR reversing agents, which have been shown to act at either of the three levels against ABC transporters. Other potential agents and strategies, which can be used to reverse the MDR phenotype, are also discussed.
[Back
to top] Structure of Multidrug-Resistance Proteins of
the ATP-Binding Cassette (ABC) Superfamily
Guillermo
A. Altenberg
Multidrug resistance of tumors, characterized by resistance against a variety of chemically unrelated anticancer agents, can be caused by overexpression of ATP-binding cassette (ABC) proteins, such as P-glycoprotein and MRP1. These multidrug-resistance proteins are plasma-membrane proteins that actively extrude chemotherapeutic agents from the cell interior, decreasing drug accumulation and thus, allowing the cells to survive in the presence of toxic levels of anticancer agents. ABC proteins contain multispanning transmembrane domains and nucleotide-binding domains (NBDs). The NBDs are responsible for the ATP binding/hydrolysis that drives drug transport, and their structure is conserved independently of the degree of primary-sequence homology. The transmembrane domains contain the drug-binding sites that are likely located in a flexible internal chamber that is sufficiently large to accommodate different drugs. It has been recently proposed that dimerization of the NBDs induced by ATP binding is a key step for the coupling of ATP hydrolysis to substrate transport. The power stroke for substrate transport can be the formation or the dissociation of the dimers. Since the NBDs and TMDs are tightly associated, association/dissociation of the NBDs may control the “gate” of the translocation pathway, formed by intracellular loops. In the case of P-glycoprotein it seems that the power stroke for transport is ATP binding (and therefore NBD dimerization), and not hydrolysis, because the major conformational and functional changes seem to occur at this step.