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Susan P.C. Cole

 
  Molecular Mechanisms of Drug Resistance and Sensitivity

 
  Contact Information:  
  Canada Research Chair in Cancer Biology
Professor of Pathology & Molecular Medicine and Pharmacology & Toxicology
Senior Scientist, Cancer Care Ontario
B.Sc., Ph.D., F.R.S.C.
Cancer Research Laboratories
3rd Floor, Botterell Hall
Tel: (613) 533-2636
Fax: (613) 533-6830
email: coles@post.queensu.ca
http://meds.queensu.ca
/qcri/cole/ri_spc.htm


 
  One focus of my research is the investigation of the molecular mechanisms of drug sensitivity and resistance that may be relevant to lung cancer and other human solid tumours. By using in vitro selected drug resistant lung cancer cell lines as model systems, we have previously identified two novel forms of resistance. One form of resistance is mediated by the drug efflux pump known as MRP1. The other form of anticancer resistance we investigate is mediated through changes in the biology of the nuclear enzyme topoisomerase II (topo II). My lab is also interested in understanding how novel titanocene containing drugs developed in the lab of our collaborator Dr. Mike Baird (Department of Chemistry) kill tumour cells.

Resistance Mediated by MRP-related Proteins:

In 1992 we cloned a highly overexpressed novel mRNA from doxorubicin-selected multidrug resistant lung cancer cells and subsequently established that the Multidrug Resistance Protein (MRP1) encoded by this mRNA is a 190 kDa ATP-binding phosphoglycoprotein that is capable of conferring resistance on previously sensitive cells. MRP1 is the founding member of subfamily "C" of the ATP-Binding Cassette ("ABC") superfamily of transport proteins (www.gene.ucl.ac.uk/users/hester/abc.html) which now includes six MRP1 homologs as well as the gene responsible for cystic fibrosis (CFTR). In addition to conferring resistance to anticancer drugs, MRP1 and several of its most closely related homologs are efficient energy dependent cellular efflux pumps of glutathione (GSH) and glucuronide conjugated xenobiotics (the so-called Phase III elimination step of drug metabolism). MRP1 also transports certain important physiological molecules such as the cysteinyl leukotriene LTC4. Our ongoing investigations of MRP1 are focused on understanding how its structure is related to recognition of its substrates and its mechanism(s) of transport. Current projects include: (i) structure/function studies using site-directed mutagenesis and epitope mapping techniques, (ii) GSH, drug and drug conjugate transport and binding studies, (iii) biophysical (mass spec., EM, etc.) studies of purified MRP1, (iv) membrane topology studies, (v) identification of compounds in the environment or diet that are substrates or inhibitors of MRP1, and (vi) MRP1 gene regulation by drugs and/or environmental agents.

Investigations of Topoisomerase II and Drug Resistance:

Resistance to drugs that exert their cytotoxicity by stabilizing cleaved complexes of DNA and the essential nuclear enzyme topoisomerase II (topo II) may occur when fewer cleaved complexes are formed. This type of resistance has previously been associated with either a decrease in levels of topo II or by a qualitative change that alters interaction of the enzyme with drug or DNA. We have identified a third mechanism which involves aberrant cytoplasmic localization of a mutant topo II in a VP-16-selected lung cancer cell line site. Our current investigations are aimed at extending our knowledge of the molecular and cellular processes that can result in reduced drug sensitivity by either modulating nuclear localization of the topo II alpha or beta proteins, or by regulating expression of the topo II alpha or beta genes. Thus we are (i) elucidating the pathways by which the two topo II isozymes traffic back and forth between the nucleus and the cytoplasm, and (ii) investigating topo II alpha and beta gene regulation in drug resistant lung cancer cells. These studies have potential therapeutic applications because modulation of topo II can affect the proliferation and drug sensitivity of tumour cells.



 
  Publications:  
  MRP1:
Selected (since 2000):

Q. Mao, R.G. Deeley and S.P.C. Cole. Functional reconstitution of substrate transport by purified multidrug resistance protein MRP1 (ABCC1) in phospholipid vesicles. J. Biol. Chem. 275: 34166-34172 (2000).

M.F. Rosenberg, Q. Mao, A. Holzenburg, R.C. Ford, R.G. Deeley and S.P.C. Cole. The structure of the multidrug resistance protein 1 (MRP1/ABCC1): crystallization and single-particle analysis. J. Biol. Chem. 276: 16076-16082 (2001).

E.M. Leslie, Q. Mao, C.J. Oleschuk, R.G. Deeley and S.P.C. Cole. Modulation of Multidrug Resistance Protein 1 (MRP1/ABCC1) transport and ATPase activities by interaction with dietary flavonoids. Molec. Pharmacol. 59: 1171-1180 (2001).

K. Ito, S.L. Olsen, W. Qiu, R.G. Deeley and S.P.C. Cole. Mutation of a single conserved tryptophan in multidrug resistance protein 1 (MRP1/ABCC1) results in loss of drug resistance and selective loss of organic anion transport. J. Biol. Chem. 276: 15616-15624 (2001).

E.M. Leslie, K. Ito, P. Upadhyaya, S.S. Hecht, R.G. Deeley and S.P.C. Cole. Transport of the b-O-glucuronide conjugate of the tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) by the multidrug resistance protein 1 (MRP1/ABCC1): Requirement for glutathione or a non-sulfur-containing analog. J. Biol. Chem. 276: 27846-27854 (2001).

K. Ito, C.J. Oleschuk, C. Westlake, M.Z. Vasa, R.G. Deeley and S.P.C. Cole. Mutation of Trp1254 in the multispecific organic anion transporter, multidrug resistance protein 2 (MRP2) (ABCC2), alters substrate specificity and results in loss of methotrexate transport activity. J. Biol. Chem. 276: 38108-38114 (2001).

S. Conrad, H-M. Kauffmann, K-i. Ito, E.M. Leslie, R.G. Deeley, D. Schrenk and S.P.C. Cole. A naturally occurring mutation in MRP1 results in a selective decrease in organic anion transport and in increased doxorubicin resistance. Pharmacogenetics 12: 321-330 (2002).

Q. Mao, W. Qiu, K.E. Weigl, P.A. Lander, L.B. Tabas, R.L. Shepard, A.H. Dantzig, R.G. Deeley and S.P.C. Cole. GSH-dependent photolabeling of multidrug resistance protein MRP1 (ABCC1) by [125I]-LY475776: evidence of a major binding site in the COOH-proximal membrane spanning domain. J. Biol. Chem. 277: 28690-28699 (2002).

A. Haimeur, R.G. Deeley and S.P.C. Cole. Charged amino acids in the sixth transmembrane helix of multidrug resistance protein 1 (MRP1/ABCC1) are critical determinants of transport activity. J. Biol. Chem. 277: 41326-41333 (2002).

K. Koike, C.J. Oleschuk, A. Haimeur, S.L. Olsen, R.G. Deeley and S.P.C. Cole. Multiple membrane associated tryptophan residues contribute to the transport activity and substrate specificity of the human multidrug resistance protein, MRP1. J. Biol. Chem. 277: 49495-49503 (2002).

C.J. Oleschuk, R.G. Deeley and S.P.C. Cole. Substitution of Trp1242 of TM17 alters substrate specificity of human multidrug resistance protein, MRP3. Am. J. Physiol. Gastrointest. Liver Physiol. 284: G28-289 (2003).

E.M. Leslie, R.J. Bowers, R.G. Deeley and S.P.C. Cole. Structural requirements for functional interaction of glutathione tripeptide analogs with the human multidrug resistance protein 1 (MRP1). J. Pharmacol. Exp. Ther. 304: 643-653 (2003).

J.D. Campbell, K. Koike, C. Moreau, M.S.P. Sansom, R.G. Deeley and S.P.C. Cole. Molecular modeling correctly predicts the functional importance of Phe594 in transmembrane helix 11 of the multidrug resistance protein, MRP1 (ABCC1). J. Biol. Chem. 279: 463-468 (2004).

K. Koike, G. Conseil, E.M. Leslie, R.G. Deeley and S.P.C. Cole. Identification of proline residues in the core cytoplasmic and transmembrane regions of multidrug resistance protein 1 (MRP1/ABCC1) important for transport function, substrate specificity, and nucleotide interactions. J. Biol. Chem. 279: 12325-12336 (2004).

K. Koike, R.G. Deeley and S.P.C. Cole. Mapping of the MRPm5 epitope to the cytosolic region between transmembrane helices 13 and 14 in the drug and organic transporter, MRP1 (ABCC1). Biochem. Biophys. Res. Commun. 315: 719-725 (2004).

A. Haimeur, G. Conseil, R.G. Deeley and S.P.C. Cole. Mutations of charged amino acids in or proximal to the transmembrane helices of the second membrane spanning domain differentially affect the substrate specificity and transport activity of the multidrug resistance protein, MRP1 (ABCC1). Mol. Pharmacol. 65: 1375-1385 (2004).

D. Situ, A. Haimeur, G. Conseil, K.E. Sparks, D. Zhang, R.G. Deeley and S.P.C. Cole. Mutational analysis of ionizable residues proximal to the cytoplasmic interface of membrane spanning domain 3 of the multidrug resistance protein, MRP1 (ABCC1): Glutamate 1204 is important for both the expression and catalytic activity of the transporter. J. Biol. Chem. 279: 38871-38880 (2004).

S.P.C. Cole and I. Tannock. Drug resistance. In, The Basic Science of Oncology, 4th edition [eds. I. Tannock, R. Hill, R. Bristow and L. Harrington]. Chapter 18, McGraw-Hill. (in press 2004).

G. Conseil, R.G. Deeley and S.P.C. Cole. Role of two adjacent tyrosine residues on MRP1 (ABCC1) transport activity and sensitivity to sulfonylureas. Biochem. Pharmacol. # BP9459 (in press 2004).

Reviews:

A. Haimeur, G. Conseil, R.G. Deeley and S.P.C. Cole. The MRP-related and BCRP/ABCG2 multidrug resistance proteins: biology, substrate specificity and regulation. Curr. Drug Metab. 5: 21-53 (2004). (invited review).

E.M. Leslie, R.G. Deeley and S.P.C. Cole. Multidrug resistance proteins in toxicology: role of p-glycoprotein, MRP1, MRP2 and BCRP (ABCG2) in tissue defense. Toxicol. Appl. Pharmacol. # TAAP-D-04-00889R1 (in press 2004) (invited review).

Topo II:

C-N. Lok, A.J. Lang, S.E.L. Mirski and S.P.C. Cole. Characterization of human topoisomerase IIb (TOP2B) promoter activity: essential roles of the NF-Y and Sp1 binding sites. Biochem. J. 368: 741-751 (2002).

S.E.L. Mirski, J.C. Bielawski and S.P.C. Cole. Identification of functional nuclear export sequences in human topoisomerase II a and b. Biochem. Biophys. Res. Comm. 306: 905-911 (2003).