Marc Fathi

Long-term Angiotensin-converting Enzyme Inhibitor Treatment Attenuates Adrenergic Responsiveness without Altering Hemodynamic Control in Patients Undergoing Cardiac Surgery

Background The sympathoadrenal and the renin-angiotensin systems are involved in blood pressure regulation and are known to be markedly activated during cardiac surgery. Because unexpected hypotensive events have been reported repeatedly during anesthesia in patients chronically treated with angiotensin-converting enzyme (ACE) inhibitors, the authors questioned whether renin-angiotensin system blockade would alter the hemodynamic control through attenuation of the endocrine response to surgery and/or through attenuation of the pressor effects of exogenous catecholamines. Methods Patients with preserved left ventricular function undergoing mitral valve replacement or coronary revascularization were divided into two groups according to preoperative drug therapy: patients receiving ACE inhibitors for at least 3 months (ACEI group, n = 22) and those receiving other cardiovascular drug therapy (control group, n = 19). Anesthesia was induced using fentanyl and midazolam. Systemic hemodynamic variables were recorded before surgery, after anesthesia induction, during sternotomy, after aortic cross-clamping, after aortic unclamping, as well as after separation from cardiopulmonary bypass (CPB) and during skin closure. Blood was sampled repeatedly up to 24 h after surgery for hormone analysis. To test adrenergic responsiveness, incremental doses of norepinephrine were infused intravenously during hypothermic CPB and after separation from CPB. From the dose-response curves, pressor (defined as mean arterial pressure changes), and vasoconstrictor (defined as systemic vascular resistance changes) effects were analyzed, and the slopes and the dose of norepinephrine required to increase mean arterial pressure by 20% were calculated (PD20). Results At no time did the systemic hemodynamics and the need for vasopressor support differ between the two treatment groups. However, for anesthesia induction, significantly less fentanyl and midazolam were given in the ACEI group. Although plasma renin activity was significantly greater in the ACEI group throughout the whole 24-h study period, plasma concentrations of angiotensin II did not differ between the two groups. Similar changes in catecholamines, angiotensin II, and plasma renin activity were found in the two groups in response to surgery and CPB. The pressor and constrictor effects of norepinephrine infusion were attenuated markedly in the ACEI group: the dose-response curves were shifted to the right and the slopes were decreased at the two study periods; PD20 was significantly greater during hypothermic CPB (0.08 micro gram/kg in the ACEI group vs. 0.03 micro gram/kg in the control group; P < 0.05) and after separation from CPB (0.52 micro gram/kg in the ACEI group vs. 0.13 micro gram/kg in the control group; P < 0.05). In both groups, PD20 was significantly less during hypothermic CPB than in the period immediately after CPB. Conclusions Long-term ACE inhibitor treatment in patients with preserved left ventricular function alters neither the endocrine response nor the hemodynamic stability during cardiac surgery. However, a significantly attenuated adrenergic responsiveness associated with incomplete blockade of the plasma renin-angiotensin system supports the hypothesis that inhibition of angiotensin II generation and of bradykinin degradation within the vascular wall mediates some of the vasodilatory effects of ACE inhibitors.

Toxicity, efficacy, plasma drug concentrations and protease mutations in patients with advanced HIV infection treated with ritonavir plus saquinavir

Objective:To assess the safety, efficacy and plasma drug levels of the combination of ritonavir plus saquinavir for the treatment of advanced HIV infection. Design:Multicentre pilot study. Patients:Eighteen protease inhibitor-naive patients, with intolerance or contraindication to reverse transcriptase inhibitors, a median CD4 cell count of 12 × 106/l (range, 1–50 × 106/l), and a median HIV viraemia of 5.25 log10copies/ml (range, 4.00–6.13 log10 copies/ml). Methods:Patients received 600 mg twice daily of both ritonavir and saquinavir. Viraemia was measured at baseline and at weeks 5, 9 and 13. Response was defined as a drop of viraemia of more than 1 log10 at week 5. Plasma drug levels were determined after at least 3 weeks of combined treatment: samples were collected before and 1, 2, and 4 h after the morning ingestion of both drugs. The protease gene was sequenced at baseline and under treatment. Results:Among the 16 patients evaluable at week 5, 11 were responders, and among these patients, six remained responders at week 13 (two with undetectable viraemia). Study discontinuations were due to side-effects (n = 4), patient choice (n = 3), protocol violation (n = 1) and death (n = 1). Responders had higher drug levels than non-responders (P < 0.01 for saquinavir, P = 0.04 for ritonavir). In two non-responders, development of multiple new mutations at positions 10, 20, 48, 82, 84 and 90 was observed after 5–13 weeks. Conclusion:The response to ritonavir plus saquinavir in advanced HIV infection is unpredictable. A minority of patients respond with disappearance of HIV viraemia. In other patients, rapid cumulative emergence of protease mutations conferring resistance to treatment cannot always be prevented by good compliance and relatively high plasma drug levels.

Characterization and classification of matrix effects in biological samples analyses

An exhaustive classification of matrix effects occurring when a sample preparation is performed prior to liquid-chromatography coupled to mass spectrometry (LC-MS) analyses was proposed. A total of eight different situations were identified allowing the recognition of the matrix effect typology via the calculation of four recovery values. A set of 198 compounds was used to evaluate matrix effects after solid phase extraction (SPE) from plasma or urine samples prior to LC-ESI-MS analysis. Matrix effect identification was achieved for all compounds and classified through an organization chart. Only 17% of the tested compounds did not present significant matrix effects.

Influence of ABCB1 genetic polymorphisms on cyclosporine intracellular concentration in transplant recipients.

Objective The expression on lymphocytes of P-glycoprotein, an efflux transporter encoded by the ABCB1 gene, might influence cyclosporine intracellular concentration. Methods ABCB1 genotypes, cyclosporine intracellular and blood concentrations were determined in 64 stable renal, liver or lung transplant recipients. Results Cyclosporine intracellular concentration correlated moderately with blood concentration (r2=0.30, P<0.00005). The ABCB1 1199A carriers presented a 1.8-fold decreased cyclosporine intracellular concentration (P=0.04), whereas the 3435T carriers presented a 1.7-fold increase (P=0.02) as well as a 1.2-fold increased blood concentration (P=0.04). In contrast, ABCB1 61A>G, 1236C>T and 2677G>T polymorphisms did not influence cyclosporine intracellular and blood concentrations. Conclusion This is the first report demonstrating that ABCB1 polymorphisms influence cyclosporine intracellular concentration. Interestingly, its influence on intracellular concentration is significantly higher than on blood concentration (P<0.002). This may therefore modulate cyclosporine immunosuppressive activity.

X-Rank: A Robust Algorithm for Small Molecule Identification Using Tandem Mass Spectrometry

The diversity of experimental workflows involving LC-MS/MS and the extended range of mass spectrometers tend to produce extremely variable spectra. Variability reduces the accuracy of compound identification produced by commonly available software for a spectral library search. We introduce here a new algorithm that successfully matches MS/MS spectra generated by a range of instruments, acquired under different conditions. Our algorithm called X-Rank first sorts peak intensities of a spectrum and second establishes a correlation between two sorted spectra. X-Rank then computes the probability that a rank from an experimental spectrum matches a rank from a reference library spectrum. In a training step, characteristic parameter values are generated for a given data set. We compared the efficiency of the X-Rank algorithm with the dot-product algorithm implemented by MS Search from the National Institute of Standards and Technology (NIST) on two test sets produced with different instruments. Overall the X-Rank algorithm accurately discriminates correct from wrong matches and detects more correct substances than the MS Search. Furthermore, X-Rank could correctly identify and top rank eight chemical compounds in a commercially available test mix. This confirms the ability of the algorithm to perform both a straight single-platform identification and a cross-platform library search in comparison to other tools. It also opens the possibility for efficient general unknown screening (GUS) against large compound libraries.

Urinary serotonin metabolite excretion during cisplatin chemotherapy.

Background. Nausea and vomiting associated with cisplatin chemotherapy is a source of major morbidity that remains difficult to control. Acute phase (0‐24 hours after induction of chemotherapy) nausea and vomiting parallels plasma serotonin release, which explains the effectiveness of 5HT3 antagonists; serotonin release in the delayed phase (24–48 hours after induction), during which consistent antiemetic control remains elusive, has not been investigated. The effect of propofol, a recent addition to the antiemetic armamentarium, on this serotonin release has not been studied.

Simultaneous quantification of cyclosporine, tacrolimus, sirolimus and everolimus in whole blood by liquid chromatography-electrospray mass spectrometry.

OBJECTIVES The aim of this work was to develop a selective method for the simultaneous quantification of cyclosporine, tacrolimus, sirolimus and everolimus in whole blood. DESIGN AND METHODS An automated on-line solid-phase extraction system coupled with liquid chromatography-mass spectrometry (LC-MS) was used. After a simple protein precipitation, the supernatant was load on a C8 column with a mobile phase composed of MeOH/H(2)O (5/95 v/v), supplemented with formic acid 0.02% and sodium formate 1 muM. After column-switching, the analytes were transferred in the back-flush mode on a C18 column with MeOH/H(2)O (65/35). The valve was then commuted to its initial position and the chromatographic separation was performed with a gradient of MeOH/H(2)O (65/35-95/5). The sodium adducts [M+Na](+) were monitored for quantification with an electrospray ionization-single quadrupole MS. RESULTS The LC-MS assay was fully validated on a concentration range of 2.5-30 ng/mL for tacrolimus, sirolimus and everolimus and of 50-1500 ng/mL for cyclosporine, allowing a quantification of cyclosporine 2 h post-dose without sample dilution. Trueness, repeatability and intermediate precision were found to be satisfactory. CONCLUSION This method provided a selective, rapid and automated procedure that can be easily used for routine quantification of immunosuppressive drugs in most clinical laboratories.

Optimized release of dexamethasone and gentamicin from a soluble ocular insert for the treatment of external ophthalmic infections

In the case of external ophthalmic infections, repeated instillations of antibiotics are required to reach therapeutic level, above the minimal inhibitory concentration (MIC). An additional administration of a corticosteroid is often needed, in order to limit the precorneal damages caused by the infection. However, repeated administration of a corticosteroid can increase intraocular pressure and thus lead to glaucoma. To overcome the disadvantages of separated and repeated instillations of two products and to avoid the side effects of dexamethasone, a soluble insert containing gentamicin sulfate and dexamethasone phosphate was developed. The new system ensures the concomitant release of the two drugs during the first 10 h of treatment, followed by an adequate concentration of gentamicin sulfate, above the MIC of 4.0 microgram ml-1, during 50 h, due to a combination of gentamicin sulfate with cellulose acetate phthalate, which reduces the solubility of gentamicin.

Zero-order release formulation of oxprenolol hydrochloride with swelling and erosion control

Zero-order release of oxprenolol hydrochloride was obtained by controlling the swelling and erosion of the matrix. This formulation involves only mixing of drug, hydroxypropylmethylcellulose (HPMC), and sodium carboxymethylcellulose (Na CMC) at the ratio of 1:0.4:1.6, respectively, and compressing the mixture directly into tablets. The in vitro release pattern from this optimized matrix tablet was reproducible. Accelerated stability studies revealed that the optimized formulation remains stable for an approximately 2-year shelf life. This sustained-release (SR) tablet was evaluated in dogs, and for comparison a conventional (CV) formulation was also given at the same dose level. Plasma oxprenolol levels were monitored by a sensitive and specific high-performance liquid chromatographic (HPLC) method. Significant differences in the pharmacokinetic parameters, i.e., lower Cmax, higher values of tmax, MRT, AUC, and plasma concentration at 24 hr, and nearly constant plasma levels over 12 hr, indicated that the SR matrix tablet is superior to the CV rapid-releasing formulation. The in vitro release parameters and in vivo pharmacokinetics correlated well.

A multi-target screening analysis in human plasma using fast liquid chromatography-hybrid tandem mass spectrometry (Part II)

OBJECTIVES Perform a comparison of results obtained with a LC-MS/MS method and a Remedi® instrument on clinical serum samples. DESIGN AND METHODS Results obtained on 146 selected plasma samples were compared between the two methods. RESULTS On the 336 positive identifications, 89% were obtained using the LC-MS/MS technique and 57% by the LC-DAD. Benzodiazepines were well recognized by LC-MS/MS. For some compounds such as antidepressant agents, sensitivity was improved using LC-MS/MS. Moreover, this method extended the panel of drugs detected in clinical toxicology. CONCLUSION The new software platform developed for screening and identification of small molecules (SmileMS) allows an easy and reproducible detection of drugs and toxic compounds in blood for general unknown screening. It offers automated generation of reports, which makes the LC-MS/MS easier to use without having specialised skills in mass spectrometry. This LC-MS/MS screening method will be a reliable alternative to the Remedi® instrument in the global process of screening in emergency clinical toxicology laboratories.