Christine Bobin-Dubigeon

Effects of tumour necrosis factor-α synthesis inhibitors on rat trinitrobenzene sulphonic acid-induced chronic colitis

The fact that tumour necrosis factor-alpha (TNF-alpha) is clearly involved in the pathogenesis of intestinal bowel disease, especially Crohns disease, suggests that TNF-alpha synthesis inhibitors could be beneficial for treatment. The present study assessed the effect of chronic oral gavage of two in vitro TNF-alpha synthesis inhibitors, JM 34 maleate or [N-(4,6-dimethylpyridin-2-yl)-furane-2-carboxamide)] maleate and XC 21 or (N-betapicolyl-tetrafluorophtalimide), on colonic inflammation in trinitrobenzene sulphonic acid-induced colitis in rats. Rats received JM 34 maleate (100 mg/kg) and XC 21 (50 mg/kg) 1 h before colitis induction and then daily for 8 days by oral gavage. The colon was removed on day 8 and processed for clinical score, myeloperoxidase activity, and soluble TNF-alpha release. Treatment with XC 21, as well as dexamethasone and sulphasalazine, reduced colonic damage and decreased (except with dexamethasone) the incidence of diarrhoea. JM 34 maleate failed to improve the clinical signs of chronic colitis. After trinitrobenzene sulphonic acid-induced colitis, myeloperoxidase activity and TNF-alpha colonic mucosal production were substantially increased compared to the control (saline instillation). Both of these inflammatory indicators were then significantly decreased (P< or =0.05) after the four chronic treatments (JM 34 maleate, XC 21, sulphasalazine, and dexamethasone). XC 21 appeared to be as efficient as sulphasalazine in improving colonic inflammation.

New anti-inflammatory N-pyridinyl(alkyl)phthalimides acting as tumour necrosis factor-α production inhibitors

This paper describes the synthesis of N-pyridinyl(alkyl)phthalimides related to N-phenyl-4,5,6,7-tetrafluorophthalimides known to be inhibitors of tumour necrosis factor-alpha (TNFalpha) production. Pharmacomodulation at the phthalimidic nitrogen led to the selection of two pharmacophoric fragments (2,4-lutidinyl and beta-picolyl), allowing significant inhibition of TNFalpha production (compounds 12 and 17). Variation of the substituents linked to the homocycle of their phthalimide scaffold indicated that high (TNFalpha production) inhibitory potency could be achieved, notably by 5-fluoro, 4- or 5-nitro, 5-amino and especially tetrafluoro substitution. The most active compound, N-(pyridin-3-ylmethyl)-4,5,6,7-tetrafluorophthalimide (32) (84% inhibition at 10 microM), also produced an anti-oedematous effect in the PMA-induced mouse-ear swelling test. Although less active than dexamethasone, it exerted a marked reduction in ear thickness after oral administration (63% vs. 85% for dexamethasone at 0.2 mMkg(-1)) and remained efficient after topical application (46% vs. 96% for the dexamethasone). It also induced potent inhibition in the rat carrageenan foot oedema test with an ID(50) (0.14 microMkg(-1)) comparable with that of N-(2,6-diisopropylphenyl)phthalimide (4) (0.15 microMkg(-1)).

Paraoxonase 1 (PON1) as a marker of short term death in breast cancer recurrence.

OBJECTIVE To relate paraoxonase (PON1) activity to survival time and short term death in breast cancer recurrence. DESIGN AND METHODS PON1 activity was measured by its rate of hydrolysis of two different substrates, paraoxon (PON) and phenylacetate (ARE) in 50 patients with recurrence of breast cancer. Results were compared between patients surviving more than one year after the analysis (22) and those who died within one year (28). RESULTS In a logistic regression analysis, ARE was negatively associated with early death (OR=0.10 [0.02-0.58], p=0.0109). PON did not reach significance (OR=0.43 [0.17-1.11], p=0.0826). In a multiple logistic regression analysis model, ARE was independently associated with early death (OR=0.12 [0.02-0.98], p=0.0476), besides interval time between diagnosis and recurrence (OR=0.54 [0.27-1.07], p=0.0781) and undernutrition (OR=3.95 [0.81-19.19], p=0.0883). CONCLUSION Paraoxonase is a potential marker of survival in patients with breast cancer recurrence.

A New, Validated Wipe-Sampling Procedure Coupled to LC–MS Analysis for the Simultaneous Determination of 5-Fluorouracil, Doxorubicin and Cyclophosphamide in Surface Contamination

A wipe-sampling procedure followed by a simple liquid chromatography-mass spectrometry method was developed and validated for the simultaneous quantification of three cytotoxic drugs [5-fluorouracil (5FU), doxorubicin and cyclophosphamide (CP)] for the determination of surface contamination. After a solid-phase extraction procedure with wiping filter paper, the separation was performed within 30 min using a gradient mobile phase. The method was validated according to the recommendations of the US Food and Drug Administration. Wiping was performed using Whatman(®) filter paper on different surfaces such as stainless steel, polypropylene and glass. The method was linear, between 10 and 500 ng per wiping sample (i.e., 0.1-5 ng/cm(2)) for 5FU and doxorubicin, and between 1-100 ng per wiping sample (i.e., 0.01-1 ng/cm(2)) for CP. The lower limits of detection and quantification were 5 and 10 ng per wiping sample for 5FU and doxorubicin, and 0.5 and 1 ng per wiping sample for CP. This new sensitive methodology for surface contamination studies was successfully applied on commercial vials and different places in a cancer research hospital. This approach is particularly suitable to assess the risk of occupational exposure to cytotoxic drugs and to optimize the cleaning process, especially for the most toxic molecule studied, CP.

Development and validation of an improved liquid chromatography–mass spectrometry method for the determination of pemetrexed in human plasma

An improved liquid chromatography-mass spectrometry method for the determination of pemetrexed in human plasma was developed and validated using a simple quadrupole LC-MS and a new SPE cartridge (Plexa Bond Elut). The analysis was achieved with a C18 analytical column using a mobile phase consisting of formic acid/acetonitrile and isocratic flow for 7 min. The linear ranges (r(2)>0.99) were found from 5 to 5000 ng/mL. The lower limit of detection was 2.5 ng/mL. Within-day and between-day precisions were less than 7.2% and inaccuracy did not exceed 2.8%. This new method is suitable to support pharmacokinetic studies and drug monitoring.

A new rapid and sensitive LC-MS assay for the determination of sorafenib in plasma: application to a patient undergoing hemodialysis.

A simple liquid chromatography–mass spectrometry method was developed and validated for quantification of sorafenib (Nexavar) in human plasma. After a solid-phase extraction procedure, the separation was performed within 2 minutes using an isocratic flow of a mobile phase consisting of formic acid/acetonitrile applied on a C18 analytical column. The analyte was detected by mass spectrometry in the single-ion monitoring mode. The method was validated according to the recommendations of the US Food and Drug Administration. The method was linear (r2 > 0.99) between 10 and 10,000 ng/mL. The lower limits of detection and quantification were 5 and 10 ng/mL, respectively. Within-day and between-day imprecisions were less than 10.4%, and inaccuracy did not exceed 8.7%. The mean extraction recovery was 92.2%. The method also provided satisfactory results in terms of time stability and dilution integrity. Sorafenib plasma concentrations of the studied patient ranged between 1831 and 3459 ng/mL. This new technique is rapid, sensitive, and was applied to the determination of sorafenib plasma concentrations in a patient undergoing hemodialysis. Our results indicate that sorafenib is not cleared from plasma by hemodialysis, although analysis should be delayed after dialysis to avoid erratic fluctuations.

Paired measurement of serum amyloid A (SAA) and paraoxonase 1 (PON1) as useful markers in breast cancer recurrence.

OBJECTIVES Paraoxonase 1 (PON1) and serum amyloid A (SAA) are carried by HDL. In case of inflammation, SAA and PON1 tend to change in opposite direction. In this study we determined if inflammation leads to altered PON1 activity using three different substrate hydrolysis rates, paraoxonase (PON), arylesterase (ARE) and lactonase (LAC) in breast cancer recurrence. DESIGN AND METHODS 49 patients with a recurrence of breast cancer were analyzed for SAA, CRP, lipids, oxidized LDL, PON, ARE and LAC. Distribution of PON1 activities across the quartiles of CRP and SAA were compared by the Kruskal Wallis test. Non-parametric estimates of the survivor function were computed with Kaplan-Meier method. The association of SAA and ARE with short term death was assessed by logistic regression models. RESULTS HDL and ARE decrease significantly across the quartiles of CRP. No significant differences were observed across SAA quartiles. The survival time was significantly related to the level of SAA (log rank: p<0.001) as well as the level of ARE (log rank: p=0.039). SAA and ARE were independently related to survival time below one year. CONCLUSIONS PON1 does not seem to be directly affected by SAA, for any of the tested substrates, PON, ARE and LAC. The combined measurement of SAA and ARE could be a useful tool in this clinical situation, since they are independently related to short term death.

Intestinal Proportion of Blautia sp. is Associated with Clinical Stage and Histoprognostic Grade in Patients with Early-Stage Breast Cancer

ABSTRACT Improving knowledge about breast cancer etiology is crucial in order to propose prevention strategies for this pathology. Gut microbiota is involved in numerous physiopathological situations including cancers. Although its potential involvement in breast cancer through the alteration of the enterohepatic circulation of estrogens and/or the metabolism of phytoestrogens has been discussed for some time, it remains to be demonstrated. The present study seeks to strengthen this hypothesis by identifying possible links between the fecal microbiota composition and clinical characteristics in breast cancer patients. Bacterial DNA was extracted from the feces of 31 patients with early-stage breast cancer and amplified by real-time polymerase chain reaction (qPCR), targeting 16S rRNA sequences specific to bacterial groups, and then analyzed in relation to clinical characteristics. The absolute numbers of total bacteria and of three bacterial groups (Firmicutes, Faecalibacterium prausnitzii, and Blautia) differed significantly according to the patients body mass index. The percentage and the absolute numbers of certain bacterial groups, namely C. coccoides, F. prausnitzii, and Blautia, differed significantly according to the clinical stages and the histoprognostic grades. Our study highlighted that intestinal microbiota composition in these patients differs according to clinical characteristics and BMI. Further studies are required to clarify the link between breast cancer and intestinal microbiota.

N-Pyridinyl(alkyl)polyhalogenobenzamides Acting as TNF-α Production Inhibitors

A series of N-pyridinyl(methyl)fluorobenzamides issued from 2,4-dimethyl-6-aminopyridine and 3-aminomethylpyridine were synthesized and evaluated as inhibitors of TNF-α production. Although less active than the corresponding phthalimides, several pentafluorobenzamides exhibited significant activity at 10 μM. N-(4,6-dimethylpyridin-2-yl)pentafluorobenzamide was selected for a preliminary in-vivo assay. Although its inhibitory activity against carrageenan-induced rat paw oedema was moderate, it induced a significant reduction in ear thickness in the PMA-induced mouse ear-swelling test (49± 6% inhibition after a dose of 0.1 mM kg−1, p.o.).

Lithocholic bile acid inhibits lipogenesis and induces apoptosis in breast cancer cells

BackgroundIt has amply been documented that mammary tumor cells may exhibit an increased lipogenesis. Biliary acids are currently recognized as signaling molecules in the intestine, in addition to their classical roles in the digestion and absorption of lipids. The aim of our study was to evaluate the impact of lithocholic acid (LCA) on the lipogenesis of breast cancer cells. The putative cytotoxic effects of LCA on these cells were also examined.MethodsThe effects of LCA on breast cancer-derived MCF-7 and MDA-MB-231 cells were studied using MTT viability assays, Annexin-FITC and Akt phosphorylation assays to evaluate anti-proliferative and pro-apoptotic properties, qRT-PCR and Western blotting assays to assess the expression of the bile acid receptor TGR5 and the estrogen receptor ERα, and genes and proteins involved in apoptosis (Bax, Bcl-2, p53) and lipogenesis (SREBP-1c, FASN, ACACA). Intracellular lipid droplets were visualized using Oil Red O staining.ResultsWe found that LCA induces TGR5 expression and exhibits anti-proliferative and pro-apoptotic effects in MCF-7 and MDA-MB-231 cells. Also, an increase in pro-apoptotic p53 protein expression and a decrease in anti-apoptotic Bcl-2 protein expression were observed after LCA treatment of MCF-7 cells. In addition, we found that LCA reduced Akt phosphorylation in MCF-7 cells, but not in MDA-MB-231 cells. We also noted that LCA reduced the expression of SREBP-1c, FASN and ACACA in both breast cancer-derived cell lines and that cells treated with LCA contained low numbers of lipid droplets compared to untreated control cells. Finally, a decrease in ERα expression was observed in MCF-7 cells treated with LCA.ConclusionsOur data suggest a potential therapeutic role of lithocholic acid in breast cancer cells through a reversion of lipid metabolism deregulation.