In vivo measurement of tissue oxygenation by time-resolved luminescence spectroscopy: advantageous properties of dichlorotris(1, 10-phenanthroline)-ruthenium(II) hydrate.

Measuring tissue oxygenation in vivo is of interest in fundamental biological as well as medical applications. One minimally invasive approach to assess the oxygen partial pressure in tissue (pO2) is to measure the oxygen-dependent luminescence lifetime of molecular probes. The relation between tissue pO2 and the probes' luminescence lifetime is governed by the Stern-Volmer equation. Unfortunately, virtually all oxygen-sensitive probes based on this principle induce some degree of phototoxicity. For that reason, we studied the oxygen sensitivity and phototoxicity of dichlorotris(1, 10-phenanthroline)-ruthenium(II) hydrate [Ru(Phen)] using a dedicated optical fiber-based, time-resolved spectrometer in the chicken embryo chorioallantoic membrane. We demonstrated that, after intravenous injection, Ru(Phen)'s luminescence lifetime presents an easily detectable pO2 dependence at a low drug dose (1 mg∕kg) and low fluence (120 mJ∕cm2 at 470 nm). The phototoxic threshold was found to be at 10 J∕cm2 with the same wavelength and drug dose, i.e., about two orders of magnitude larger than the fluence necessary to perform a pO2 measurement. Finally, an illustrative application of this pO2 measurement approach in a hypoxic tumor environment is presented.

Evaluation of the impact of a therapeutic drug monitoring program in a Swiss University Hospital

Background and objective: Therapeutic Drug Monitoring (TDM) has been introduced early 1970 in our hospital (CHUV). It represents nowadays an important routine activity of the Division of Clinical Pharmacology and Toxicology (PCL), and its impact and utility for clinicians required assessment. This study thus evaluated the impact of TDM recommendations in terms of dosage regimen adaptation. Design: A prospective observational study was conducted over 5 weeks. The primary objective was to evaluate the application of our TDM recommendations and to identify potential factors associated to variations in their implementation. The secondary objective was to identify pre-analytical problems linked to the collection and processing of blood samples. Setting: Four representative clinical units at CHUV. Main outcome measure: Clinical data, drug related data (intake, collection and processing) and all information regarding the implementation of clinical recommendations were collected and analyzed by descriptive statistics. Results: A total of 241 blood measurement requests were collected, among which 105 triggered a recommendation. 37% of the recommendations delivered were applied, 25 % partially applied and 34% not applied. In 4% it was not applicable. The factors determinant for implementation were the clinical unit and the mode of transmission of the recommendation (written vs oral). No clear difference between types of drugs could be detected. Pre-analytical problems were not uncommon, mostly related to completion of request forms and delays in blood sampling (equilibration or steady-state not reached). We have identified 6% of inappropriate and unusable drug level measurements that could cause a substantial cost for the hospital. Conclusion: This survey highlighted a better implementation of TDM recommendations in clinical units where this routine is well integrated and understood by the medical staff. Our results emphasize the importance of communication with the nurse or the physician in charge, either to transmit clinical recommendations or to establish consensual therapeutic targets in specific conditions. Development of strong partnerships between clinical pharmacists or pharmacologists and clinical units would be beneficial to improve the impact of this clinical activity.

Docking, virtual high throughput screening and in silico fragment-based drug design.

ABSTRACT The drug discovery process has been profoundly changed recently by the adoption of computational methods helping the design of new drug candidates more rapidly and at lower costs. In silico drug design consists of a collection of tools helping to make rational decisions at the different steps of the drug discovery process, such as the identification of a biomolecular target of therapeutical interest, the selection or the design of new lead compounds and their modification to obtain better affinities, as well as pharmacokinetic and pharmacodynamic properties. Among the different tools available, a particular emphasis is placed in this review on molecular docking, virtual high throughput screening and fragment-based ligand design.

Bayesian classification criterion for forensic multivariate data

This study presents a classification criteria for two-class Cannabis seedlings. As the cultivation of drug type cannabis is forbidden in Switzerland, law enforcement authorities regularly ask laboratories to determine cannabis plant's chemotype from seized material in order to ascertain that the plantation is legal or not. In this study, the classification analysis is based on data obtained from the relative proportion of three major leaf compounds measured by gas-chromatography interfaced with mass spectrometry (GC-MS). The aim is to discriminate between drug type (illegal) and fiber type (legal) cannabis at an early stage of the growth. A Bayesian procedure is proposed: a Bayes factor is computed and classification is performed on the basis of the decision maker specifications (i.e. prior probability distributions on cannabis type and consequences of classification measured by losses). Classification rates are computed with two statistical models and results are compared. Sensitivity analysis is then performed to analyze the robustness of classification criteria.

Individual contributions of mutant protease and reverse transcriptase to viral infectivity, replication, and protein maturation of antiretroviral drug-resistant human immunodeficiency virus type 1.

Human immunodeficiency virus type 1 (HIV-1) variants resistant to protease (PR) and reverse transcriptase (RT) inhibitors may display impaired infectivity and replication capacity. The individual contributions of mutated HIV-1 PR and RT to infectivity, replication, RT activity, and protein maturation (herein referred to as "fitness") in recombinant viruses were investigated by separately cloning PR, RT, and PR-RT cassettes from drug-resistant mutant viral isolates into the wild-type NL4-3 background. Both mutant PR and RT contributed to measurable deficits in fitness of viral constructs. In peripheral blood mononuclear cells, replication rates (means +/- standard deviations) of RT recombinants were 72.5% +/- 27.3% and replication rates of PR recombinants were 60.5% +/- 33.6% of the rates of NL4-3. PR mutant deficits were enhanced in CEM T cells, with relative replication rates of PR recombinants decreasing to 15.8% +/- 23.5% of NL4-3 replication rates. Cloning of the cognate RT improved fitness of some PR mutant clones. For a multidrug-resistant virus transmitted through sexual contact, RT constructs displayed a marked infectivity and replication deficit and diminished packaging of Pol proteins (RT content in virions diminished by 56.3% +/- 10.7%, and integrase content diminished by 23.3% +/- 18.4%), a novel mechanism for a decreased-fitness phenotype. Despite the identified impairment of recombinant clones, fitness of two of the three drug-resistant isolates was comparable to that of wild-type, susceptible viruses, suggestive of extensive compensation by genomic regions away from PR and RT. Only limited reversion of mutated positions to wild-type amino acids was observed for the native isolates over 100 viral replication cycles in the absence of drug selective pressure. These data underscore the complex relationship between PR and RT adaptive changes and viral evolution in antiretroviral drug-resistant HIV-1.

Association of mutations in the NALP3/CIAS1/PYPAF1 gene with a broad phenotype including recurrent fever, cold sensitivity, sensorineural deafness, and AA amyloidosis.

OBJECTIVE: Familial cold urticaria (FCU) and Muckle-Wells syndrome (MWS) are dominantly inherited autoinflammatory disorders that cause rashes, fever, arthralgia, and in some subjects, AA amyloidosis, and have been mapped to chromosome 1q44. Sensorineural deafness in MWS, and provocation of symptoms by cold in FCU, are distinctive features. This study was undertaken to characterize the genetic basis of FCU, MWS, and an overlapping disorder in French Canadian, British, and Indian families, respectively. METHODS: Mutations in the candidate gene NALP3, which has also been named CIAS1 and PYPAF1, were sought in the study families, in a British/Spanish patient with apparent sporadic MWS, and in matched population controls. Identified variants were sought in 50 European subjects with uncharacterized, apparently sporadic periodic fever syndromes, 48 subjects with rheumatoid arthritis (RA), and 19 subjects with juvenile idiopathic arthritis (JIA). RESULTS: Point mutations, encoding putative protein variants R262W and L307P, were present in all affected members of the Indian and French Canadian families, respectively, but not in controls. The R262W variant was also present in the subject with sporadic MWS. The V200M variant was present in all affected members of the British family with MWS, in 2 of the 50 subjects with uncharacterized periodic fevers, and in 1 of 130 Caucasian and 2 of 48 Indian healthy controls. No mutations were identified among the subjects with RA or JIA. CONCLUSION: These findings confirm that mutations in the NALP3/CIAS1/PYPAF1 gene are associated with FCU and MWS, and that disease severity and clinical features may differ substantially within and between families. Analysis of this gene will improve classification of patients with inherited or apparently sporadic periodic fever syndromes.

Abnormal sensitivity to negative feedback in late-life depression.

AIMS: The purpose of the present study was to probe sensitivity to potentially misleading negative feedback on cognitive tasks as a possible mechanism of cognitive impairment in elderly patients with mild depression. METHODS: A total of 22 mildly depressed elderly subjects were compared to 22 healthy controls, using a computerized Tower-of-London task. RESULTS: Failure and magnitude of failure were significantly worse after negative but not positive feedback. Depression predicted failure after negative feedback but not the magnitude of failure. Neither failure nor magnitude of failure increased as a consequence of repeated negative feedback. CONCLUSIONS: Altered sensitivity to negative feedback occurs in mild late-life unipolar depression and may represent a subtle context-specific phenomenon.

Médicaments et grossesse : modifications pharmacocinétiques et place du suivi thérapeutique pharmacologique [Pharmacokinetic Alterations in Pregnancy and Use of Therapeutic Drug Monitoring].

Following the thalidomide tragedy, pharmacological research in pregnant women focused primarily on drug safety for the unborn child and remains only limited regarding the efficacy and safety of treatment for the mother. Significant physiological changes during pregnancy may yet affect the pharmacokinetics of drugs and thus compromise its efficacy and/or safety. Therapeutic drug monitoring (TDM) would maximize the potential effectiveness of treatments, while minimizing the potential risk of toxicity for the mother and the fetus. At present, because of the lack of concentration-response relationship studies in pregnant women, TDM can rely only on individual assessment (based on an effective concentration before pregnancy) and remains reserved only to unexpected situations such as signs of toxicity or unexplained inefficiency.

Suivi de l'adhésion thérapeutique des patients hypertendus ou dyslipi- démiques par les cercles de qualité médecins-pharmaciens [Monitoring drug adherence for hypertensive and dyslipidemic patients in networks of community-based pharmacists and physicians].

Patient adherence is often poor for hypertension and dyslipidaemia. A monitoring of drug adherence might improve these risk factors control, but little is known in ambulatory care. We conducted a randomised controlled study in networks of community-based pharmacists and physicians in the canton of Fribourg to examine whether monitoring drug adherence with an electronic monitor (MEMS) would improve risk factor control among treated, but uncontrolled hypertensive and dyslipidemic patients. The results indicate that MEMS achieve a better blood pressure control and lipid profile, although its implementation requires considerable resources. The study also shows the value of collaboration between physicians and pharmacists in the field of patient adherence to improve ambulatory care of patients with cardiovascular risk factors.

The rapid urinary drug toxicology screen in the emergency room: a useful tool ?

Intoxications are a frequent problem in the ER. In the vast majorityof cases, supportive treatment is sufficient. Severe intoxications withunknown agents are considered an indication for a urinary drug screen,and are recommended by several toxicology centers. However, theirusefulness for patient management remains uncertain.Study objectives: Evaluation of the impact of a urinary drug screen(Biosite Triage TOX Drug Screen) testing 11 substances(acetaminophen, amphetamines, methamphetamines, barbiturates,benzodiazepines, cocaïne, methadone, opioids, phencyclidine,cannabis, tricyclic antidepressants) on initial adult patient managementin the emergency department of a university hospital with ~35.000annual admissions.Methods: Observational retrospective analysis of all tests performedbetween 09/2009 and 09/2010. A test utility was defined as useful if itresulted in the administration of a specific antidote (Flumazenil/Naloxone), the use of a quantitative confirmatory toxicologic test, or achange in patient's disposition.Results: 57 tests were performed. Patient age was 32 ± 11 (SD) years;58% were men; 30% were also intoxicated with alcohol. Two patientsdied (3.5%): the first one of a diphenhydramin overdose, the other of ahypertensive intracerebral hemorrhage believed to be caused cocaineabuse but a negative urine test. Test indications were: 54% firstpsychotic episode; 25% acute respiratory failure; 18% coma; 12%seizure; 11% opioids toxidrome; 7% sympathicomimetic toxidrome; 5%hypotension; 4% ventricular arrhythmia (VT, VF, torsades de pointes)or long QT. 75% of tests were positives for >=1 substance (mean 1.7 ±0.9). 47% of results were unexpected by history. 18% of resultsinfluenced patient management: 7% had a negative test that confirmedthe diagnosis of endogenous psychosis in a first psychotic episode, andallowed transfer to psychiatry; 5% received flumazenil/naloxone;2% had an acetaminophen blood level after a positive screen; finally,4% had an unexpected methadone abuse that required prolongationof hospital stay.Conclusions: A rapid urinary toxicologic screen was seldom used inour emergency department, and its impact on patient managementwas marginal: only one in 6 tests influenced treatment decisions.

Microtubule-depolymerizing agents used in antibody-drug conjugates induce antitumor immunity by stimulation of dendritic cells.

Antibody-drug conjugates (ADC) are emerging as powerful treatment strategies with outstanding target-specificity and high therapeutic activity in patients with cancer. Brentuximab vedotin represents a first-in-class ADC directed against CD30(+) malignancies. We hypothesized that its sustained clinical responses could be related to the stimulation of an anticancer immune response. In this study, we demonstrate that the dolastatin family of microtubule inhibitors, from which the cytotoxic component of brentuximab vedotin is derived, comprises potent inducers of phenotypic and functional dendritic cell (DC) maturation. In addition to the direct cytotoxic effect on tumor cells, dolastatins efficiently promoted antigen uptake and migration of tumor-resident DCs to the tumor-draining lymph nodes. Exposure of murine and human DCs to dolastatins significantly increased their capacity to prime T cells. Underlining the requirement of an intact host immune system for the full therapeutic benefit of dolastatins, the antitumor effect was far less pronounced in immunocompromised mice. We observed substantial therapeutic synergies when combining dolastatins with tumor antigen-specific vaccination or blockade of the PD-1-PD-L1 and CTLA-4 coinhibitory pathways. Ultimately, treatment with ADCs using dolastatins induces DC homing and activates cellular antitumor immune responses in patients. Our data reveal a novel mechanism of action for dolastatins and provide a strong rationale for clinical treatment regimens combining dolastatin-based therapies, such as brentuximab vedotin, with immune-based therapies. Cancer Immunol Res; 2(8); 741-55. ©2014 AACR.

Detection of EPO doping and blood doping: the haematological module of the Athlete Biological Passport.

The increase of the body's capacity to transport oxygen is a prime target for doping athletes in all endurance sports. For this pupose, blood transfusions or erythropoiesis stimulating agents (ESA), such as erythropoietin, NESP, and CERA are used. As direct detection of such manipulations is difficult, biomarkers that are connected to the haematopoietic system (haemoglobin concentration, reticulocytes) are monitored over time (Athlete Biological Passport (ABP)) and analyzed using mathematical models to identify patterns suspicious of doping. With this information, athletes can either be sanctioned directly based on their profile or targeted with conventional doping tests. Key issues for the appropriate use of the ABP are correct targeting and use of all available information (e.g. whereabouts, cross sectional population data) in a forensic manner. Future developments of the passport include the correction of all concentration-based variables for shifts in plasma volume, which might considerably increase sensitivity. New passport markers from the genomic, proteomic, and metabolomic level might add further information, but need to be validated before integration into the passport procedure. A first assessment of blood data of federations that have implemented the passport show encouraging signs of a decreased blood-doping prevalence in their athletes, which adds scientific credibility to this innovative concept in the fight against ESA- and blood doping. Copyright © 2012 John Wiley & Sons, Ltd.

Adolescent substance-use assessment: methodological issues in the use of the Adolescent Drug Abuse Diagnosis (ADAD).

During the past twenty years, various instruments have been developed for the assessment of substance use in adolescents, mainly in the United States. However, few of them have been adapted to, and validated in, French-speaking populations. Consequently, although increasing alcohol and drug use among teenagers has become a major concern, the various health and social programs developed in response to this specific problem have received little attention with regard to follow-up and outcome assessment. A standardized multidimensional assessment instrument adapted for adolescents is needed to assess the individual needs of adolescents and assign them to the most appropriate treatment setting, to provide a single measurement within and across health and social systems, and to conduct treatment outcome evaluations. Moreover, having an available instrument makes it possible to develop longitudinal and transcultural research studies. For this reason, a French version of the Adolescent Drug Abuse Diagnosis (ADAD) was developed and validated at the University Child and Adolescent Psychiatric Clinic in Lausanne, Switzerland. This article aims to discuss the methodological issues that we faced when using the ADAD instrument in a 4-year longitudinal study including adolescent substance users. Methodological aspects relating to the content and format of the instrument, the assessment administration and the statistical analyses are discussed.