title: Nicotinamide mononucleotide adenylyltransferase2 overexpression enhances colorectal cancer cell-kill by Tiazofurin.
abstract: Colorectal cancer cells exhibit limited cytotoxicity towards Tiazofurin, a pro-drug metabolized by cytosolic nicotinamide mononucleotide adenylyltransferase2 (NMNAT2) to thiazole-4-carboxamide adenine dinucleotide, a potent inhibitor of inosine 5'-monophosphate dehydrogenase required for cellular guanylate synthesis. We tested the hypothesis that colorectal cancer cells that exhibit low levels of NMNAT2 and are refractory to Tiazofurin can be rendered sensitive to Tiazofurin by overexpressing NMNAT2. Transfection of hNMNAT2 resulted in a six- and threefold cytoplasmic overexpression in Caco2 and HT29 cell lines correlating with Tiazofurin-induced enhanced cell-kill. Folate receptors expressed on the cell surface of 30-50% colorectal carcinomas were exploited for cellular targeting with Tiazofurin encapsulated in folate-tethered nanoparticles. Our results indicated that in wild-type colorectal cancer cells, free Tiazofurin-induced EC50 cell-kill was 1500-2000 μM, which was reduced to 66-156 μM in hNMNAT2-overexpressed cells treated with Tiazofurin encapsulated in non-targeted nanoparticles. This efficacy was improved threefold by encapsulating Tiazofurin in folate-tethered nanoparticles to obtain an EC(50) cell-kill of 22-59 μM, an equivalent of 100-300 mg m(-2) (one-tenth of the approved dose of Tiazofurin in humans), which will result in minimal toxicity leading to cancer cell-kill. This proof-of-principle study suggests that resistance of colorectal cancer cell-kill to Tiazofurin can be overcome by sequentially overexpressing hNMNAT2 and then facilitating the uptake of Tiazofurin by folate-tethered nanoparticles, which enter cells via folate receptors.
authors.affiliation: Research Service, Richard L. Roudebush Veterans Affairs Medical Center, Indianapolis, IN, USA.
authors.forename: P
authors.initials: P
authors.lastname: Kusumanchi
authors.forename: Y
authors.initials: Y
authors.lastname: Zhang
authors.forename: M B
authors.initials: MB
authors.lastname: Jani
authors.forename: N H
authors.initials: NH
authors.lastname: Jayaram
authors.forename: R A
authors.initials: RA
authors.lastname: Khan
authors.forename: Y
authors.initials: Y
authors.lastname: Tang
authors.forename: A C
authors.initials: AC
authors.lastname: Antony
authors.forename: H N
authors.initials: HN
authors.lastname: Jayaram
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000378:metabolism / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D015179:Colorectal Neoplasms; D004337:Drug Carriers / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D005452:Fluoresceins / Q000378:metabolism; D058972:Folic Acid Transporters / Q000378:metabolism; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D007168:IMP Dehydrogenase / Q000037:antagonists & inhibitors; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D053758:Nanoparticles / Q000378:metabolism; D009612:Nicotinamide-Nucleotide Adenylyltransferase / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000235:genetics; D012254:Ribavirin / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D014162:Transfection
chemical_list: D000964:Antimetabolites, Antineoplastic; D004337:Drug Carriers; D005452:Fluoresceins; D058972:Folic Acid Transporters; D007527:Isoenzymes; D034741:RNA, Small Interfering; D012254:Ribavirin; D006160:Guanosine Triphosphate; D007168:IMP Dehydrogenase; C501687:IMPDH2 protein, human; C484008:NMNAT2 protein, human; D009612:Nicotinamide-Nucleotide Adenylyltransferase; C033706:tiazofurin; C007740:fluorexon