Identification of New Drug Candidates Against \u3cem\u3eBorrelia burgdorferi\u3c/em\u3e Using High-Throughput Screening

Lyme disease is the most common zoonotic bacterial disease in North America. It is estimated that .300,000 cases per annum are reported in USA alone. A total of 10%–20% of patients who have been treated with antibiotic therapy report the recrudescence of symptoms, such as muscle and joint pain, psychosocial and cognitive difficulties, and generalized fatigue. This condition is referred to as posttreatment Lyme disease syndrome. While there is no evidence for the presence of viable infectious organisms in individuals with posttreatment Lyme disease syndrome, some researchers found surviving Borrelia burgdorferi population in rodents and primates even after antibiotic treatment. Although such observations need more ratification, there is unmet need for developing the therapeutic agents that focus on removing the persisting bacterial form of B. burgdorferi in rodent and nonhuman primates. For this purpose, high-throughput screening was done using BacTiter-Glo assay for four compound libraries to identify candidates that stop the growth of B. burgdorferi in vitro. The four chemical libraries containing 4,366 compounds (80% Food and Drug Administration [FDA] approved) that were screened are Library of Pharmacologically Active Compounds (LOPAC1280), the National Institutes of Health Clinical Collection, the Microsource Spectrum, and the Biomol FDA. We subsequently identified 150 unique compounds, which inhibited .90% of B. burgdorferi growth at a concentration of ,25 µM. These 150 unique compounds comprise many safe antibiotics, chemical compounds, and also small molecules from plant sources. Of the 150 unique compounds, 101 compounds are FDA approved. We selected the top 20 FDA-approved molecules based on safety and potency and studied their minimum inhibitory concentration and minimum bactericidal concentration. The promising safe FDA-approved candidates that show low minimum inhibitory concentration and minimum bactericidal concentration values can be chosen as lead molecules for further advanced studies

Dizajniranje i vrednovanje okularnih umetaka moksifloksacin hidroklorida

The objective of the present investigation was to prepare and evaluate ocular inserts of moxifloxacin. An ocular insert was made from an aqueous dispersion of moxifloxacin, sodium alginate, polyvinyl alcohol, and dibutyl phthalate by the film casting method. The ocular insert (5.5 mm diameter) was cross-linked by CaCl2 and was coated with Eudragit S-100, RL-100, RS-100, E-100 or Eudragit L-100. The in vitro drug drainage/permeation studies were carried out using an all-glass modified Franz diffusion cell. The drug concentration and mucoadhesion time of the ocular insert were found satisfactory. Cross-linking and coating with polymers extended the drainage from inserts. The cross-linked ocular insert coated with Eudragit RL-100 showed maximum drug permeation compared to other formulations.Cilj rada bio je priprava i evaluacija okularnih umetaka moksifloksacina. Okularni umetak izrađen je od vodene suspenzije moksifloksacina, natrijevog alginata, polivinilnog alkohola i dibutil-ftalata metodom odlijevanja filma. Okularni umetak (promjera 5,5 mm) umrežen je pomoću CaCl2 i obložen Eudragitom S-100, RL-100, RS-100, E-100 ili Eudragit L-100. In vitro drenaža/permeacija lijeka proučavana je koristeći staklenu modificiranu Franzovu difuzijsku ćeliju. Koncentracija lijeka i vrijeme mukoadhezije okularnih umetaka bili su zadovoljavajući. Umrežavanje i oblaganje polimerima produljilo je drenažu iz umetaka. Umreženi okularni umetci obloženi s Eudragit RL-100 pokazali su veću permeaciju lijeka u odnosu na ostale pripravke

Moxifloxacin: Clinically compatible contrast agent for multiphoton imaging

Multiphoton microscopy (MPM) is a nonlinear fluorescence microscopic technique widely used for cellular imaging of thick tissues and live animals in biological studies. However, MPM application to human tissues is limited by weak endogenous fluorescence in tissue and cytotoxicity of exogenous probes. Herein, we describe the applications of moxifloxacin, an FDA-approved antibiotic, as a cell-labeling agent for MPM. Moxifloxacin has bright intrinsic multiphoton fluorescence, good tissue penetration and high intracellular concentration. MPM with moxifloxacin was demonstrated in various cell lines, and animal tissues of cornea, skin, small intestine and bladder. Clinical application is promising since imaging based on moxifloxacin labeling could be 10 times faster than imaging based on endogenous fluorescence.1152sciescopu

Termoreverzibilni mukoadhezivni in situ hidrogel za oftalmičku primjenu: dizajniranje i optimizacija koristeći kombinaciju polimera

The purpose of the study was to develop an optimized thermoreversible in situ gelling ophthalmic drug delivery system based on Pluronic F 127, containing moxifloxacin hydrochloride as a model drug. A 32 full factorial design was employed with two polymers Pluronic F 68 and Gelrite as independent variables used in combination with Pluronic F 127. Gelation temperature, gel strength, bioadhesion force, viscosity and in vitro drug release after 1 and 10 h were selected as dependent variables. Pluronic F 68 loading with Pluronic F 127 was found to have a significant effect on gelation temperature of the formulation and to be of importance for gel formation at temperatures 3336 ºC. Gelrite loading showed a positive effect on bioadhesion force and gel strength and was also found helpful in controling the release rate of the drug. The quadratic mathematical model developed is applicable to predicting formulations with desired gelation temperature, gel strength, bioadhesion force and drug release properties.Cilj rada bio je razvoj i optimizacija termoreverzibilnog sustava za isporuku lijekova koji gelira in situ. Sustav je napravljen na bazi Pluronic F 127, a sadrži moksifloksacin hidroklorid kao modelni lijek. U radu je primjenjeno 32 potpuno faktorijsko dizajniranje s dva polimera, Pluronic F 68 i Gelrite kao nezavisnim varijablama koji su kombinirani s Pluronic F 127. Kao zavisne varijable odabrane su temperatura geliranja, čvrstoća gela, jačina bioadhezije, viskoznost i in vitro oslobađanje lijeka nakon 1 i 10 h. Pronađeno je da Pluronic F 68 u kombinaciji s Pluronic F 127 ima značajan učinak na temperaturu geliranja u rasponu od 33 do 36 C. S druge strane, Gelrite ima povoljan učinak na jačinu bioadhezije, čvrstoću gela i oslobađanje lijeka. Razvijen je kvadratni matematički model pomoću kojeg se može predvidjeti temperatura geliranja, čvrstoća gela, jačina bioadhezije i oslobađanje ljekovite tvari

Termoreverzibilni mukoadhezivni in situ hidrogel za oftalmičku primjenu: dizajniranje i optimizacija koristeći kombinaciju polimera

The purpose of the study was to develop an optimized thermoreversible in situ gelling ophthalmic drug delivery system based on Pluronic F 127, containing moxifloxacin hydrochloride as a model drug. A 32 full factorial design was employed with two polymers Pluronic F 68 and Gelrite as independent variables used in combination with Pluronic F 127. Gelation temperature, gel strength, bioadhesion force, viscosity and in vitro drug release after 1 and 10 h were selected as dependent variables. Pluronic F 68 loading with Pluronic F 127 was found to have a significant effect on gelation temperature of the formulation and to be of importance for gel formation at temperatures 3336 ºC. Gelrite loading showed a positive effect on bioadhesion force and gel strength and was also found helpful in controling the release rate of the drug. The quadratic mathematical model developed is applicable to predicting formulations with desired gelation temperature, gel strength, bioadhesion force and drug release properties.Cilj rada bio je razvoj i optimizacija termoreverzibilnog sustava za isporuku lijekova koji gelira in situ. Sustav je napravljen na bazi Pluronic F 127, a sadrži moksifloksacin hidroklorid kao modelni lijek. U radu je primjenjeno 32 potpuno faktorijsko dizajniranje s dva polimera, Pluronic F 68 i Gelrite kao nezavisnim varijablama koji su kombinirani s Pluronic F 127. Kao zavisne varijable odabrane su temperatura geliranja, čvrstoća gela, jačina bioadhezije, viskoznost i in vitro oslobađanje lijeka nakon 1 i 10 h. Pronađeno je da Pluronic F 68 u kombinaciji s Pluronic F 127 ima značajan učinak na temperaturu geliranja u rasponu od 33 do 36 C. S druge strane, Gelrite ima povoljan učinak na jačinu bioadhezije, čvrstoću gela i oslobađanje lijeka. Razvijen je kvadratni matematički model pomoću kojeg se može predvidjeti temperatura geliranja, čvrstoća gela, jačina bioadhezije i oslobađanje ljekovite tvari

Stability indicating HPLC method for the simultaneous determination of moxifloxacin and prednisolone in pharmaceutical formulations

BACKGROUND: A simple, specific, and fast stability indicating reverse phase liquid chromatographic method was established for instantaneous determination of moxifloxacin and prednisolone in bulk drugs and pharmaceutical formulations. RESULTS: Optimum chromatographic separations among the moxifloxacin, prednisolone and stress-induced degradation products were achieved within 10 minutes by use of BDS Hypersil C8 column (250 X 4.6 mm, 5 μm) as stationary phase with mobile phase consisted of a mixture of phosphate buffer (18 mM) containing 0.1% (v/v) triethylamine, at pH 2.8 (adjusted with dilute phosphoric acid) and methanol (38:62 v/v) at a flow rate of 1.5 mL min(-1). Detection was performed at 254 nm using diode array detector. The method was validated in accordance with ICH guidelines. Response was a linear function of concentrations over the range of 20–80 μg mL(-1) for moxifloxacin (r2 ≥ 0.998) and 40–160 μg mL(-1) for prednisolone (r2 ≥ 0.998). The method was resulted in good separation of both the analytes and degradation products with acceptable tailing and resolution. The peak purity index for both the analytes after all types of stress conditions was ≥ 0.9999 indicated a complete separation of both the analyte peaks from degradation products. The method can therefore, be regarded as stabilityindicating. CONCLUSIONS: The developed method can be applied successfully for simultaneous determination of moxifloxacin and prednisolone in pharmaceutical formulations and their stability studies

In vivo 3D measurement of moxifloxacin and gatifloxacin distributions in the mouse cornea using multiphoton microscopy

Moxifloxacin and gatifloxacin are fourth-generation fluoroquinolone antibiotics used in the clinic to prevent or treat ocular infections. Their pharmacokinetics in the cornea is usually measured from extracted ocular fluids or tissues, and in vivo direct measurement is difficult. In this study multiphoton microscopy (MPM), which is a 3D optical microscopic technique based on multiphoton fluorescence, was applied to the measurement of moxifloxacin and gatifloxacin distribution in the cornea. Intrinsic multiphoton fluorescence properties of moxifloxacin and gatifloxacin were characterized, and their distributions in mouse cornea in vivo were measured by 3D MPM imaging. Both moxifloxacin and gatifloxacin had similar multiphoton spectra, while moxifloxacin had stronger fluorescence than gatifloxacin. MPM imaging of mouse cornea in vivo showed (1) moxifloxacin had good penetration through the superficial corneal epithelium, while gatifloxacin had relatively poor penetration, (2) both ophthalmic solutions had high intracellular distribution. In vivo MPM results were consistent with previous studies. This study demonstrates the feasibility of MPM as a method for in vivo direct measurement of moxifloxacin and gatifloxacin in the cornea.1175Ysciescopu

A signalome screening approach in the autoinflammatory disease TNF Receptor Associated Periodic Syndrome (TRAPS) highlights the anti-inflammatory properties of drugs for repurposing

TNF Receptor Associated Periodic Syndrome (TRAPS) is an autoinflammatory disease caused by mutations in TNF Receptor 1 (TNFR1). Current therapies for TRAPS are limited and do not target the pro-inflammatory signalling pathways that are central to the disease mechanism. Our aim was to identify drugs for repurposing as anti-inflammatories based on their ability to down-regulate molecules associated with inflammatory signalling pathways that are activated in TRAPS. This was achieved using rigorously optimised, high through- put cell culture and reverse phase protein microarray systems to screen compounds for their effects on the TRAPS-associated inflammatory signalome. 1360 approved, publically available, pharmacologically active substances were investigated for their effects on 40 signalling molecules associated with pro-inflammatory signalling pathways that are constitutively upregulated in TRAPS. The drugs were screened at four ten-fold concentrations on cell lines expressing both wild-type (WT) TNFR1 and TRAPS-associated C33Y mutant TNFR1, or WT TNFR1 alone; signalling molecule levels were then determined in cell lysates by the reverse phase protein microarray. A novel mathematical methodology was developed to rank the compounds for their ability to reduce the expression of signalling molecules in the C33Y-TNFR1 transfectants towards the level seen in the WT-TNFR1 transfectants. Seven high-ranking drugs were selected and tested by RPPA for effects on the same 40 signalling molecules in lysates of peripheral blood mononuclear cells (PBMCs) from C33Y-TRAPS patients compared to PBMCs from normal controls. The fluoroquinolone antibiotic lomefloxacin, as well as others from this class of compounds, showed the most significant effects on multiple pro-inflammatory signalling pathways that are constitutively activated in TRAPS; lomefloxacin dose-dependently significantly reduced expression of 7/40 signalling molecules across the Jak/Stat, MAPK, NF-kB and PI3K/AKT pathways. This study demonstrates the power of signalome screening for identifying candidates for drug repurposing

Release of Ciprofloxacin and Moxifloxacin From Daily Disposable Contact Lenses From an In Vitro Eye Model

Bajgrowicz, M., Phan, C.-M., Subbaraman, L. N., & Jones, L. (2015). Release of Ciprofloxacin and Moxifloxacin From Daily Disposable Contact Lenses From an In Vitro Eye Model. Investigative Opthalmology & Visual Science, 56(4), 2234. https://doi.org/10.1167/iovs.15-16379Purpose.: To analyze the release of two fluoroquinolones, ciprofloxacin and moxifloxacin, from conventional hydrogel (CH) and silicone hydrogel (SH) daily disposable contact lenses (CLs), comparing release from a fixed-volume vial and a novel in vitro eye model. Methods.: Four CH CLs (nelfilcon A, omafilcon A, etafilcon A, ocufilcon B) and three SH CLs (somofilcon A, narafilcon A, delefilcon A) were used. The lenses were incubated in drug solutions for 24 hours. After the incubation period, the lenses were placed in two release conditions: (1) a vial containing 4.8 mL PBS for 24 hours and (2) an in vitro eye model with a flow rate at 4.8 mL over 24 hours. Results.: Release in the vial for both drugs was rapid, reaching a plateau between 15 minutes and 2 hours for all lenses. In contrast, under physiological flow conditions, a constant and slow release was observed over 24 hours. The amounts of ciprofloxacin released from the lenses ranged between 49.6 ± 0.7 and 62.8 ± 0.3 μg per lens in the vial, and between 35.0 ± 7.0 and 109.0 ± 5.0 μg per lens in the eye model. Moxifloxacin release ranged from 24.0 ± 4.0 to 226.0 ± 2.0 μg per lens for the vial, and between 13.0 ± 2.0 and 151.0 ± 10.0 μg per lens in the eye model. In both systems and for both drugs, HEMA-based CLs released more drugs than other materials. Conclusions.: The parameters of the release system, in particular the volume and flow rate, have a significant influence on measured release profiles. Under physiological flow, release profiles are significantly slower and constant when compared with release in a vial

An observational study of Donor Ex Vivo Lung Perfusion in UK lung transplantation: DEVELOP-UK

Background: Many patients awaiting lung transplantation die before a donor organ becomes available. Ex vivo lung perfusion (EVLP) allows initially unusable donor lungs to be assessed and reconditioned for clinical use. Objective: The objective of the Donor Ex Vivo Lung Perfusion in UK lung transplantation study was to evaluate the clinical effectiveness and cost-effectiveness of EVLP in increasing UK lung transplant activity. Design: A multicentre, unblinded, non-randomised, non-inferiority observational study to compare transplant outcomes between EVLP-assessed and standard donor lungs. Setting: Multicentre study involving all five UK officially designated NHS adult lung transplant centres. Participants: Patients aged ≥ 18 years with advanced lung disease accepted onto the lung transplant waiting list. Intervention: The study intervention was EVLP assessment of donor lungs before determining suitability for transplantation. Main outcome measures: The primary outcome measure was survival during the first 12 months following lung transplantation. Secondary outcome measures were patient-centred outcomes that are influenced by the effectiveness of lung transplantation and that contribute to the health-care costs. Results: Lungs from 53 donors unsuitable for standard transplant were assessed with EVLP, of which 18 (34%) were subsequently transplanted. A total of 184 participants received standard donor lungs. Owing to the early closure of the study, a non-inferiority analysis was not conducted. The Kaplan–Meier estimate of survival at 12 months was 0.67 [95% confidence interval (CI) 0.40 to 0.83] for the EVLP arm and 0.80 (95% CI 0.74 to 0.85) for the standard arm. The hazard ratio for overall 12-month survival in the EVLP arm relative to the standard arm was 1.96 (95% CI 0.83 to 4.67). Patients in the EVLP arm required ventilation for a longer period and stayed longer in an intensive therapy unit (ITU) than patients in the standard arm, but duration of overall hospital stay was similar in both groups. There was a higher rate of very early grade 3 primary graft dysfunction (PGD) in the EVLP arm, but rates of PGD did not differ between groups after 72 hours. The requirement for extracorporeal membrane oxygenation (ECMO) support was higher in the EVLP arm (7/18, 38.8%) than in the standard arm (6/184, 3.2%). There were no major differences in rates of chest radiograph abnormalities, infection, lung function or rejection by 12 months. The cost of EVLP transplants is approximately £35,000 higher than the cost of standard transplants, as a result of the cost of the EVLP procedure, and the increased ECMO use and ITU stay. Predictors of cost were quality of life on joining the waiting list, type of transplant and number of lungs transplanted. An exploratory model comparing a NHS lung transplant service that includes EVLP and standard lung transplants with one including only standard lung transplants resulted in an incremental cost-effectiveness ratio of £73,000. Interviews showed that patients had a good understanding of the need for, and the processes of, EVLP. If EVLP can increase the number of usable donor lungs and reduce waiting, it is likely to be acceptable to those waiting for lung transplantation. Study limitations include small numbers in the EVLP arm, limiting analysis to descriptive statistics and the EVLP protocol change during the study. Conclusions: Overall, one-third of donor lungs subjected to EVLP were deemed suitable for transplant. Estimated survival over 12 months was lower than in the standard group, but the data were also consistent with no difference in survival between groups. Patients receiving these additional transplants experience a higher rate of early graft injury and need for unplanned ECMO support, at increased cost. The small number of participants in the EVLP arm because of early study termination limits the robustness of these conclusions. The reason for the increased PGD rates, high ECMO requirement and possible differences in lung injury between EVLP protocols needs evaluation