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Preclinical study of treatment response in HCT-116 cells and xenografts with (1) H-decoupled (31) P MRS.

ABSTRACT: The topoisomerase I inhibitor, irinotecan, and its active metabolite SN-38 have been shown to induce G(2) /M cell cycle arrest without significant cell death in human colon carcinoma cells (HCT-116). Subsequent treatment of these G(2) /M-arrested cells with the cyclin-dependent kinase inhibitor, flavopiridol, induced these cells to undergo apoptosis. The goal of this study was to develop a noninvasive metabolic biomarker for early tumor response and target inhibition of irinotecan followed by flavopiridol treatment in a longitudinal study. A total of eleven mice bearing HCT-116 xenografts were separated into two cohorts where one cohort was administered saline and the other treated with a sequential course of irinotecan followed by flavopiridol. Each mouse xenograft was longitudinally monitored with proton ((1) H)-decoupled phosphorus ((31) P) magnetic resonance spectroscopy (MRS) before and after treatment. A statistically significant decrease in phosphocholine (p?=?0.0004) and inorganic phosphate (p?=?0.0103) levels were observed in HCT-116 xenografts following treatment, which were evidenced within twenty-four hours of treatment completion. Also, a significant growth delay was found in treated xenografts. To discern the underlying mechanism for the treatment response of the xenografts, in vitro HCT-116 cell cultures were investigated with enzymatic assays, cell cycle analysis, and apoptotic assays. Flavopiridol had a direct effect on choline kinase as measured by a 67% reduction in the phosphorylation of choline to phosphocholine. Cells treated with SN-38 alone underwent 83?±?5% G(2) /M cell cycle arrest compared to untreated cells. In cells, flavopiridol alone induced 5?±?1% apoptosis while the sequential treatment (SN-38 then flavopiridol) resulted in 39?±?10% apoptosis. In vivo (1) H-decoupled (31) P MRS indirectly measures choline kinase activity. The decrease in phosphocholine may be a potential indicator of early tumor response to the sequential treatment of irinotecan followed by flavopiridol in noninvasive and/or longitudinal studies.

SUBMITTER: Darpolor MM

PROVIDER: S-EPMC3201722 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Preclinical study of treatment response in HCT-116 cells and xenografts with (1) H-decoupled (31) P MRS.

Darpolor Moses M MM Kennealey Peter T PT Le H Carl HC Zakian Kristen L KL Ackerstaff Ellen E Rizwan Asif A Chen Jin-Hong JH Sambol Elliot B EB Schwartz Gary K GK Singer Samuel S Koutcher Jason A JA

NMR in biomedicine 20110324 9

The topoisomerase I inhibitor, irinotecan, and its active metabolite SN-38 have been shown to induce G(2) /M cell cycle arrest without significant cell death in human colon carcinoma cells (HCT-116). Subsequent treatment of these G(2) /M-arrested cells with the cyclin-dependent kinase inhibitor, flavopiridol, induced these cells to undergo apoptosis. The goal of this study was to develop a noninvasive metabolic biomarker for early tumor response and target inhibition of irinotecan followed by fl ...[more]

PMID: 21994185

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Project description:Curcumin is the main component of the Chinese herbal plant turmeric, which has been demonstrated to possess antitumor and other pharmacological properties. The aim of the present study was to investigate the effects of curcumin on the viability, migration and apoptosis of human colorectal carcinoma HCT?116 cells, and to explore the underlying molecular mechanisms. In addition, it was investigated whether the antitumor effect of curcumin on HCT?116 cells could match that of the chemotherapeutic drug 5?fluorouracil (5?FU). HCT?116 cells were treated with curcumin (10, 20 and 30 µM) and 5?FU (500 µM), and cell viability and proliferation were detected by Cell Counting Kit?8 and colony formation assays, respectively. The migration and invasion of treated cells were determined using Transwell and carboxyfluorescein succinimidyl amino ester fluorescent labeling assays. Cell cycle distribution and apoptosis rates were detected by flow cytometry. Furthermore, cell morphology changes associated with apoptosis were observed by fluorescence microscopy with acridine orange/ethidium bromide dual staining. To investigate the possible underlying molecular mechanisms, the gene and protein levels of Fas, Fas?associated via death domain (FADD), caspase?8, caspase?3, matrix metalloproteinase (MMP)?9, nuclear factor (NF)??B, E?cadherin and claudin?3 were detected using quantitative PCR analysis, zymography and western blotting. The results revealed that curcumin markedly inhibited the viability and proliferation of HCT?116 cells in a dose? and time?dependent manner. The migration, aggregation and invasion of HCT?116 cells into the lungs of mice were decreased by curcumin treatment in a dose?dependent manner. S?phase arrest and gradually increased apoptotic rates of HCT?116 cells were observed with increasing curcumin concentrations. Additionally, the mRNA and protein levels of apoptosis?associated proteins (Fas, FADD, caspase?8 and caspase?3) and E?cadherin in HCT?116 cells were upregulated following treatment with curcumin in a dose?dependent manner. By contrast, the expression of migration?associated proteins, including MMP?9, NF??B and claudin?3, was downregulated with increasing curcumin concentrations. These data suggested that the inhibitory effect of curcumin on HCT?116 cells may match that of 5?FU. Therefore, curcumin induced cell apoptosis and inhibited tumor cell metastasis by regulating the NF??B signaling pathway, and its therapeutic effect may be comparable to that of 5?FU.

Project description:Triethylphosphinegold(I) complexes [Au(HL1)P(CH2CH3)3]PF6 (1), [Au(HL2)P(CH2CH3)3]PF6 (2), and [Au(HL3)P(CH2CH3)3]PF6 (3) were obtained with (E)-2-(1-(2-methyl-5-nitro-1H-imidazol-1-yl)propan-2-ylidene)hydrazinecarbothioamide (HL1), (E)-N-methyl-2-(1-(2-methyl-5-nitro-1H-imidazol-1-yl)propan-2-ylidene)hydrazinecarbothioamide (HL2), and (E)-2-(1-(2-methyl-5-nitro-1H-imidazol-1-yl)propan-2-ylidene)-N-phenylhydrazinecarbothioamide (HL3). All compounds were assayed for their cytotoxic activities against HCT-116 colorectal carcinoma cells under normoxia and hypoxia conditions and against nonmalignant HEK-293 human embryonic kidney cells under normoxia conditions. The thiosemicarbazone ligands HL1-HL3 were inactive against HCT-116 cells under hypoxia but while HL3 was inactive, HL1 and HL2 proved to be cytotoxic to both cell lineages under normoxia conditions. Complexes (1-3) and the triethylphosphinegod(I) precursor proved to be active against both cell lineages in normoxia as well as in hypoxia. While 1 and 3 revealed to be active against HEK-293 and HCT-116 cells, being approximately as active against HCT-116 cells in normoxia as under hypoxia, complex (2) proved to be more active against HCT-116 cells under hypoxia than under normoxia conditions, and more active against HCT-116 cells than against the nonmalignant HEK-293 cells, with the selectivity index, calculated as SI = IC50HEK-293/IC50HCT-116hypoxia, equal to 3.7, similar to the value obtained for the control drug tirapazamine (tirapazamine (TPZ), SI = 4). Although the compounds showed distinct cytotoxic activities, the electrochemical behaviors of HL1-HL3 were very similar, as were the behaviors of complexes (1-3). Complex (2) deserves special interest since it was significantly more active under hypoxia than under normoxia conditions. Hence, in this case, selective reduction of the nitro group in a low oxygen pressure environment, resulting in toxic reactive oxygen species (ROS) and damage to DNA or other biomolecules, might operate, while for the remaining compounds, other modes of action probably occur.

Dataset Information

Preclinical study of treatment response in HCT-116 cells and xenografts with (1) H-decoupled (31) P MRS.

Publications

Preclinical study of treatment response in HCT-116 cells and xenografts with (1) H-decoupled (31) P MRS.

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