Abstract

The goal of this project is to characterize the structure and regulation of the """"""""classical"""""""" methotrexate/tetrahydrofolate cofactor membrane transporter in human tumor cells. This system is critical to the antitumor effectiveness of methotrexate and related antifolates. For methotrexate, an agent widely used in the chemotherapy of human cancers, defects in the regulation or biosynthesis of the transport system result in decreased antifolate accumulation within tumor cells and appear to be key factors in the development of drug resistance. In earlier experiments, K562 human erythroleukemia cells with increased uptake capacities for methotrexate and reduced folates were selected and a highly glycosylated transporter protein (GP-MTX) was identified and purified. Most recently, antiserum to GP-MTX was prepared, and several putative GP-MTX cDNAs from a human lymphoblastic leukemia cDNA library were isolated. Based on these initial results, we propose to (i) verify and characterize the putative full length cDNAs for GP-MTX by in vitro transcription/translation, dideoxy sequencing, and eukaryotic expression. In addition, we propose to (ii) identify and characterize the biochemical and molecular bases for impaired methotrexate transport in drug resistant K562 erythroleukemia and CCRF-CEM lymphoblastic leukemia cell variants at the levels of GP-MTX structure, biosynthesis, and/or regulation. These studies provide a framework for the study of regulation of the transport system for methotrexate and tetrahydrofolate cofactors in both drug sensitive and resistant human tumor cells. They should clarify the mechanistic bases for transport-mediated resistance to methotrexate. In this fashion, the identification of membrane-linked biochemical or molecular """"""""markers"""""""" for methotrexate sensitivity or resistance in vitro should promote the more effective clinical use of this and related agents, including the detection and treatment of drug resistant tumors.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
1R01CA053535-01A1
Application #
3198246
Study Section
Experimental Therapeutics Subcommittee 1 (ET)
Project Start
1993-02-12
Project End
1996-01-31
Budget Start
1993-02-12
Budget End
1994-01-31
Support Year
1
Fiscal Year
1993
Total Cost
Indirect Cost

  Institution

Name
Barbara Ann Karmanos Cancer Institute
Department
Type
DUNS #
City
Detroit
State
MI
Country
United States
Zip Code
48201

  Publications

Shah, Khushbu; Raghavan, Sudhir; Hou, Zhanjun et al. (2018) Development and validation of chemical features-based proton-coupled folate transporter/activity and reduced folate carrier/activity models (pharmacophores). J Mol Graph Model 81:125-133
Matherly, Larry H; Hou, Zhanjun; Gangjee, Aleem (2018) The promise and challenges of exploiting the proton-coupled folate transporter for selective therapeutic targeting of cancer. Cancer Chemother Pharmacol 81:1-15
Ravindra, Manasa; Wilson, Mike R; Tong, Nian et al. (2018) Fluorine-Substituted Pyrrolo[2,3- d]Pyrimidine Analogues with Tumor Targeting via Cellular Uptake by Folate Receptor ? and the Proton-Coupled Folate Transporter and Inhibition of de Novo Purine Nucleotide Biosynthesis. J Med Chem 61:4228-4248
Giovannetti, E; Zucali, P A; Assaraf, Y G et al. (2017) Role of proton-coupled folate transporter in pemetrexed resistance of mesothelioma: clinical evidence and new pharmacological tools. Ann Oncol 28:2725-2732
Hou, Zhanjun; Gattoc, Leda; O'Connor, Carrie et al. (2017) Dual Targeting of Epithelial Ovarian Cancer Via Folate Receptor ? and the Proton-Coupled Folate Transporter with 6-Substituted Pyrrolo[2,3-d]pyrimidine Antifolates. Mol Cancer Ther 16:819-830
Wilson, Mike R; Hou, Zhanjun; Yang, Si et al. (2016) Targeting Nonsquamous Nonsmall Cell Lung Cancer via the Proton-Coupled Folate Transporter with 6-Substituted Pyrrolo[2,3-d]Pyrimidine Thienoyl Antifolates. Mol Pharmacol 89:425-34
Golani, Lalit K; Wallace-Povirk, Adrianne; Deis, Siobhan M et al. (2016) Tumor Targeting with Novel 6-Substituted Pyrrolo [2,3-d] Pyrimidine Antifolates with Heteroatom Bridge Substitutions via Cellular Uptake by Folate Receptor ? and the Proton-Coupled Folate Transporter and Inhibition of de Novo Purine Nucleotide Biosynthesi J Med Chem 59:7856-76
Wilson, Mike R; Hou, Zhanjun; Wilson, Lucas J et al. (2016) Functional and mechanistic roles of the human proton-coupled folate transporter transmembrane domain 6-7 linker. Biochem J 473:3545-3562
Wang, Yiqiang; Mitchell-Ryan, Shermaine; Raghavan, Sudhir et al. (2015) Novel 5-substituted pyrrolo[2,3-d]pyrimidines as dual inhibitors of glycinamide ribonucleotide formyltransferase and 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase and as potential antitumor agents. J Med Chem 58:1479-93
Wang, Lei; Wallace, Adrianne; Raghavan, Sudhir et al. (2015) 6-Substituted Pyrrolo[2,3-d]pyrimidine Thienoyl Regioisomers as Targeted Antifolates for Folate Receptor ? and the Proton-Coupled Folate Transporter in Human Tumors. J Med Chem 58:6938-59

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