New uracil analog U-332 is an inhibitor of NF-κB in 5-fluorouracil-resistant human leukemia HL-60 cell line

Abstract

Background: 5-Fluorouracil (5-FU) is an antimetabolite that interferes with DNA synthesis and has been widely used as a chemotherapeutic drug in various types of cancers. However, the development of drug resistance greatly limits its application. Overexpression of ATP-binding cassette (ABC) transporters in many types of cancer is responsible for the reduction of the cellular uptake of various anticancer drugs causing multidrug resistance (MDR), the major obstacle in cancer chemotherapy. Recently, we have obtained a novel synthetic 5-FU analog, U-332 [(R)-3-(4-bromophenyl)-1-ethyl-5-methylidene-6-phenyldihydrouracil], combining a uracil skeleton with an exo-cyclic methylidene group. U-332 was highly cytotoxic for HL-60 cells and showed similar cytotoxicity in the 5-FU resistant subclone (HL-60/5FU), in which this analog almost completely abolished expression of the ATP-binding cassette (ABC) transporter, multidrug resistance associate protein 1 (ABCC1). The expression of ABC transporters is usually correlated with NF-κB activation. The aim of this study was to determine the level of NF-κB subunits in the resistant HL-60/5-FU cells and to evaluate the potential of U-332 to inhibit activation of NF-κB family members in this cell line. Methods: Anti-proliferative activity of compound U-332 was assessed by the MTT assay. In order to disclose the mechanism of U-332 cytotoxicity, quantitative real-time PCR analysis of the NF-κB family genes, c-Rel, RelA, RelB, NF-κB1, and NF-κB2, was investigated. The ability of U-332 to reduce the activity of NF-κB members was studied by ELISA test. Results: In this report it was demonstrated, using RT-PCR and ELISA assay, that members of the NF-κB family c-Rel, RelA, RelB, NF-κB1, and NF-κB2 were all overexpressed in the 5-FU-resistant HL-60/5FU cells and that U-332 potently reduced the activity of c-Rel, RelA and NF-κB1 subunits in this cell line. Conclusions: This finding indicates that c-Rel, RelA and NF-κB1 subunits are responsible for the resistance of HL-60/5FU cells to 5-FU and that U-332 is able to reverse this resistance. U-332 can be viewed as an important lead compound in the search for novel drug candidates that would not cause multidrug resistance in cancer cells.

Synthesis, Characterization Of Thiazolidin-4-one, Oxazolidin-4-one And Imidazolidin-4-one Derivatives From 6-Amino-1, 3-dimethyluraciland Evaluation Of Their Antioxidant And Antimicrobial Agent

“…FTIR spectra of the synthesized compounds in the (4000-600) cm -1 spectral range were recorded on SHIMAZU FTIR-8400 Fourier transform Infrared spectrophotometer using KBr discs. 1 H-NMR and 13 C-NMR spectra were recorded on BRUKER 500MHz in Iran, instrument using TMS as internal reference and DMSO-d 6 as a solvent. Spectrophotometer were recorded by Apel PD-303-spectrophotometer, japan.…”

“…Using a round bottom flask (50 mL), a mixture of (0.0021 mol) Schiff bases (1)(2)(3)(4)(5)(6) in THF (10 mL) was added to a well-stirred solution of chloroacetic acid (0.2 g, 0.0021 mol) and mixed thoroughly with catalytic few drops of triethylamine in THF solvent. The solution was refluxed for (12)(13)(14)(15) h., after cooling the solution, the crude precipitate was filtered and recrystallization using ethanol. Some of the physical properties and yield of compounds (13)(14)(15)(16)(17)(18) are listed in Table (3.49).…”

“…The solution was refluxed for (12)(13)(14)(15) h., after cooling the solution, the crude precipitate was filtered and recrystallization using ethanol. Some of the physical properties and yield of compounds (13)(14)(15)(16)(17)(18) are listed in Table (3.49). To a mixture of Schiff bases (1-6) (0.0021 mol) was stirred in (9 mL) of THF solvent, 2aminoacetic acid (0.25 g, 0.0021 mol) in (5 mL) of THF was refluxed for (16)(17)(18)(19)(20) h. Then, the resulted mixture was filtered after cooling to room temperature, washed and recrystallized from acetone.…”

“…1 H-NMR spectral data of Schiff's base (3) showed in Table (5). 13 C-NMR spectral data of Schiff's base (3) are listed in Table (6). On the other hand, 1 H-NMR spectrum of Schiff's base (5) showed the disappearance of singlet signal of (-NH 2 ) protons of 6-amino-1,3-dimethyluracil and appearance a singlet signal at δ=(3.16)ppm of (-CH 3 ) protons, a singlet signal at δ=(3.48)ppm for ( ) protons, a singlet signal at δ=(4.79)ppm due to (=CH-) proton, a singlet signal at δ=(6.54)ppm for (N=CH-) proton, multi signals at δ=(6.50-7.43)ppm belonged to aromatic ring protons and a singlet signal at δ=(9.92)ppm is due to (O-H) proton.…”

“…1 H-NMR spectral data of Schiff's base (5) showed in Table (5). 13 C-NMR spectral data of Schiff's base (5) are listed in Table (6). The thiazolidin-4-one derivatives (7-12) were synthesized by the second cyclization method via one-pot synthesis of Schiff bases (1-6) with 2-mercaptoacetic acid in THF as a solvent as in Scheme (1).…”