Improving Treatment Of Urinary Tract Infections In Elderly: Roles And Functions Of Information Technologies To Suggest Antibiotic Prescriptions And To Measure Their Effects OBEZ C1, MONDAIN V2, CHATELIER W1, KOUBI C1, PROTS L3 1Clinique Les Sources Nice France, 2Infectious Dpt CHU de Nice France; 3Lab. Ronchèse, Nice, France
Background:The evolution of Escherichia coli’s resistances and the development of ESBLs call for a better transmission of experts’ knowledge to practising physicians; information technologies can contribute to this purpose.
Methods: To conceive Electronic Antimicrobial Drug Aid Regulation 1] Guidelines adapted to elderly are given to allow a dynamic and interactive consultation. 2] Therapy by antibiotic failures are recorded: they are related either to intolerance or to the occurrence of resistance; broad spectrum molecules prescriptions linked to patients’ details are also recorded. 3] An instrument panel is designed for the follow-up of E. coli sensitivities: in a short stay unit, comparison is made by a chi-square test of 2 series including 283 and 303 colonies, each one corresponding to 12 months of urine sampling respectively in 2006 and 2008.
Results: 1] Suggestion of antibiotic: on the screen, the practitioner determines his diagnosis with signs of gravity and checks for the existence of complications: in case of emergency, a probabilistic treatment is suggested. When bacteriological data are available, suggested antibiotics are divided into 5 classes and developed into 7 tables according to the seat and gravity of the infection and various types of complications: oral or injectable form, dose and posology, length of treatment are specified, renal function is taken into account. 2] In case of adverse drug event, the imputability of the antibiotic is evaluated according to Naranjo's method; if resistance occurs, one records supporting factors related to the patient and those related to care practices. 3] The instrument panel shows a significant increase of resistance to the amoxicillin - ac. clav. (p < 0.05); a sensitive, yet not significant, increase of resistance to acid nalidixic (25.44% versus 32.67%) and ESBLs (2.47% versus 5.61%).
Conclusions:The computer program should make it possible to reduce unnecessary antibiotic prescriptions especially in case of asymptomatic bacteriuria, to deliver an effective treatment in case of severe infection, to prevent adverse drug events and to attenuate ecological risk linked to the increase of bacterial resistances.
New Biomarkers And Targeted Therapies For Breast Cancer O’DRISCOLL L National Institute for Cellular Biotechnlogy, Dublin City University, Dublin 9;
School of Pharmacy & Pharmaceutical Sciences, Trinity College, Dublin 2, Ireland
Background: Using gene expression profiling technologies, improved methods including minimally-invasive, sensitive, specific tests could contribute significantly to cancer diagnosis, on-going monitoring, and treatment. Recently we performed a whole genome microarray study of 104 archived breast tumour and normal biopsies and, in parallel, we optimised methods enabling us to investigate global expression of extracellular mRNAs in serum (which may also be potential diagnostic/prognostic or predictive biomarkers).
Methods: RNA was extracted from 104 tumour and normal breast specimens, as well as from pre-surgery serum specimens from four recently diagnosed breast cancer patients, from their breast tumour/matched normal biopsies, and from serum specimens procured approx. 3 months post-surgery. Serum specimens from healthy age-matched volunteers acted as controls. RNA from each specimen was examined using U133 Plus2.0 arrays. Following normalisation, statistical filters were applied to identify significant differentially-expressed (DE) genes. Using univariate and multivariate methods, individual mRNAs were extensively investigated for associations with patients’ clincopathological characteristics. qRT-PCR was used to validate microarray data.
Results: mRNAs were detected in all specimens. Overall, approx. 8% (of 54,675 probesets representing transcripts on the microarray) were present in serum and approx. 45% were detected in breast tissue. 7448 transcripts were DE (P=0.0068) between tumour and normal breast specimens and 998 (P=0.0009) and 1369 (P=0.0013), respectively, between those that resulted in relapse or death within 5 year compared to those that did not. Clinical statistical analysis identified 36 mRNAs as potential novel biomarkers; some of which tended to be associated with tumours of a basal-like profile. qRT-PCR analysis of 5 transcripts randomly selected from these validated our microarray results.
Conclusion: The implication of these novel findings is that, using microarrays, it may be possible to identify panels of intracellular and extracellular biomarkers that are useful diagnostic, prognostic and/or predictive of outcome for cancer patients.
Acknowledgements: HEA’s PRTLI, Dublin City University’s Research Fellowship and Ireland’s Health Research Board.
A Computationally Designed Mutant Of The Metallo--Lactamase IMP-1 Exhibits Enhanced Catalytic Efficiency OELSCHLAEGER P1, PLEISS J2, MAYO SL3 1California State Polytechnic Univ., Pomona, CA, USA; 2Univ. of Stuttgart, Germany; 3California Institute of Technology, Pasadena, CA, USA.
Background: Metallo--lactamases (MBLs) inactivate a broad range of -lactam antibiotics, they can be transferred horizontally, and there are no clinically useful MBL inhibitors. In addition, the evolutionary potential of MBLs is of great concern. Being able to predict their evolutionary pathways could improve our ability to effectively combat antibiotic resistance.
Methods: Computational protein design was employed to predict improved variants of the MBL IMP-1. The predictions were validated through biochemical characterization and kinetic analysis of purified enzymes overexpressed in E. coli. Subsequently, molecular dynamics simulations of enzyme-substrate intermediate complexes were used to decipher the molecular mechanisms for enhanced catalytic efficiency.
Results: Two mutations (F218Y and S262A) were computationally designed in IMP-1. Experimental validation showed that the single mutant IMP-1-F218Y was superior to the wild-type enzyme IMP-1: overexpression in E. coli yielded a higher amount of soluble protein, the protein folded well and was thermally stable, as monitored by circular dichroism spectroscopy, and it exhibited enhanced catalytic efficiencies toward the following -lactams: nitrocefin, cephalothin, cefotaxime, ceftazidime, benzylpenicillin, ampicillin, and imipenem, mostly due to a decreased KM. Multiple molecular dynamics simulations showed that the F218Y mutation leads to an altered hydrogen bonding pattern and to a movement of a -hairpin loop toward the active site, which could account for the decreased KM.
Conclusions: 1) The IMP-1-F218Y mutant was succesfully predicted to be a more efficient enzyme than the wild-type enzyme IMP-1. 2) The enzyme could easily evolve naturally through a one-nucleotide change and could lead to enhanced antibiotic resistance. 3) Successful prediction of improved MBL variants could assist the design of better antibiotics and MBL inhibitors.
Kinetic Properties Of Recombinant Factor VIIa (rFVIIa) And The Complexity Relating These To Treatment Response OESTERGAARD P, ERHARDTSEN E
Novo Nordisk A/S, Bagsvaerd, Denmark
Background: Haemophilia A (HA) and B (HB) are inherited coagulation disorders caused by deficiency of factor VIII (FVIII) or IX (FIX). These patients are treated with replacement therapy with FVIII or FIX. Approx. 30% of HA1) and 5% of HB2) patients develop inhibitors making replacement therapy ineffective. Pharmacological doses of rFVIIa stop bleeding in inhibitor patients via bypassing FVIII and FIX in the coagulation cascade which requires FVIIa plasma activity significantly higher than physiological levels. However, the therapeutic window of FVIIa activity is unknown due to the complexity of relating pharmacokinetic (PK) /pharmacodynamic (PD) data, especially T½ and clearance, to efficacy and safety.
Methods: Limited rFVIIa data are available from PK studies in non-bleeding and bleeding haemophilia patients (HA/B), in patients with factor VII deficiency (FD) and in healthy subjects (HS). FVII activity has been assessed using two different assays: a FVII coagulation activity assay (detecting both endogenous FVII zymogen and FVII activity), and a FVIIa activity assay specific for activated FVII3).
Results: A comparison between the two assays has shown significantly different PK estimates and a simple conversion is not possible3). The FVIIa activity assay is preferable for PK studies. PK parameters differed slightly but were statistically significant between bleeding and non-bleeding state (HA/B)4). No effect of gender or ethnicity (HS)5) was seen whereas clearance was faster in children than adults6,7). Dose proportionality was seen in HA/B4,7) and HS5). FD patients have a higher clearance than HA/B8).
Conclusions: Haemophilia patients with inhibitors are few (~ 4000 in the Western world) and bleeding episodes are generally treated at home. Conducting PK studies in the bleeding state is therefore extremely difficult. Human subjects may be stratified in two groups based on rFVIIa PK – HS and adult HA/B vs HA/B children and FD. Relating PK profiles to clinical efficacy is complicated by few data and significant inter-patient and inter-bleed variations. Correlating PK parameters to efficacy and safety has so far not been possible.
