Chagas' disease is responsible for significant mortality and morbidity in Latin America. Current treatments display variable efficacy and have adverse side effects, hence more effective, better ...tolerated drugs are needed. However, recent efforts have proved unsuccessful with failure of the ergosterol biosynthesis inhibitor posaconazole in phase II clinical trials despite promising in vitro and in vivo studies. The lack of translation between laboratory experiments and clinical outcome is a major issue for further drug discovery efforts. Our goal was to identify cell-based assays that could differentiate current nitro-aromatic drugs nifurtimox and benznidazole from posaconazole. Using a panel of T. cruzi strains including the six major lineages (TcI-VI), we found that strain PAH179 (TcV) was markedly less susceptible to posaconazole in vitro. Determination of parasite doubling and cycling times as well as EdU labelling experiments all indicate that this lack of sensitivity is due to the slow doubling and cycling time of strain PAH179. This is in accordance with ergosterol biosynthesis inhibition by posaconazole leading to critically low ergosterol levels only after multiple rounds of division, and is further supported by the lack of effect of posaconazole on the non-replicative trypomastigote form. A washout experiment with prolonged posaconazole treatment showed that, even for more rapidly replicating strains, this compound cannot clear all parasites, indicative of a heterogeneous parasite population in vitro and potentially the presence of quiescent parasites. Benznidazole in contrast was able to kill all parasites. The work presented here shows clear differentiation between the nitro-aromatic drugs and posaconazole in several assays, and suggests that in vitro there may be clinically relevant heterogeneity in the parasite population that can be revealed in long-term washout experiments. Based on these findings we have adjusted our in vitro screening cascade so that only the most promising compounds are progressed to in vivo experiments.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Background
There is increasing interest in using ultrasound for thermal ablation, histotripsy, and thermal or cavitational enhancement of drug delivery for the treatment of pancreatic cancer. ...Ultrasonic and thermal modelling conducted as part of the treatment planning process requires acoustic property values for all constituent tissues, but the literature contains no data for the human pancreas.
Purpose
This study presents the first acoustic property measurements of human pancreatic samples and provides examples of how these properties impact a broad range of ultrasound therapies.
Methods
Data were collected on human pancreatic tissue samples at physiological temperature from 23 consented patients in cooperation with a hospital pathology laboratory. Propagation of ultrasound over the 2.1–4.5 MHz frequency range through samples of various thicknesses and pathologies was measured using a set of custom‐built ultrasonic calipers, with the data processed to estimate sound speed and attenuation. The results were used in acoustic and thermal simulations to illustrate the impacts on extracorporeal ultrasound therapies for mild hyperthermia, thermal ablation, and histotripsy implemented with a CE‐marked clinical system operating at 0.96 MHz.
Results
The mean sound speed and attenuation coefficient values for human samples were well below the range of values in the literature for non‐human pancreata, while the human attenuation power law exponents were substantially higher. The simulated impacts on ultrasound mediated therapies for the pancreas indicated that when using the human data instead of the literature average, there was a 30% reduction in median temperature elevation in the treatment volume for mild hyperthermia and 43% smaller volume within a 60°C contour for thermal ablation, all driven by attenuation. By comparison, impacts on boiling and intrinsic threshold histotripsy were minor, with peak pressures changing by less than 15% (positive) and 1% (negative) as a consequence of the counteracting effects of attenuation and sound speed.
Conclusion
This study provides the most complete set of speed of sound and attenuation data available for the human pancreas, and it reiterates the importance of acoustic material properties in the planning and conduct of ultrasound‐mediated procedures, particularly thermal therapies.
Previous preclinical research has shown that extracorporeal devices can be used to enhance the delivery and distribution of systemically administered anticancer drugs, resulting in increased ...intratumoural concentrations. We aimed to assess the safety and feasibility of targeted release and enhanced delivery of doxorubicin to solid tumours from thermosensitive liposomes triggered by mild hyperthermia, induced non-invasively by focused ultrasound.
We did an open-label, single-centre, phase 1 trial in a single UK hospital. Adult patients (aged ≥18 years) with unresectable and non-ablatable primary or secondary liver tumours of any histological subtype were considered for the study. Patients received a single intravenous infusion (50 mg/m2) of lyso-thermosensitive liposomal doxorubicin (LTLD), followed by extracorporeal focused ultrasound exposure of a single target liver tumour. The trial had two parts: in part I, patients had a real-time thermometry device implanted intratumourally, whereas patients in part II proceeded without thermometry and we used a patient-specific model to predict optimal exposure parameters. We assessed tumour biopsies obtained before and after focused ultrasound exposure for doxorubicin concentration and distribution. The primary endpoint was at least a doubling of total intratumoural doxorubicin concentration in at least half of the patients treated, on an intention-to-treat basis. This study is registered with ClinicalTrials.gov, number NCT02181075, and is now closed to recruitment.
Between March 13, 2015, and March 27, 2017, ten patients were enrolled in the study (six patients in part I and four in part II), and received a dose of LTLD followed by focused ultrasound exposure. The treatment resulted in an average increase of 3·7 times in intratumoural biopsy doxorubicin concentrations, from an estimate of 2·34 μg/g (SD 0·93) immediately after drug infusion to 8·56 μg/g (5·69) after focused ultrasound. Increases of two to ten times were observed in seven (70%) of ten patients, satisfying the primary endpoint. Serious adverse events registered were expected grade 4 transient neutropenia in five patients and prolonged hospital stay due to unexpected grade 1 confusion in one patient. Grade 3–4 adverse events recorded were neutropenia (grade 3 in one patient and grade 4 in five patients), and grade 3 anaemia in one patient. No treatment-related deaths occurred.
The combined treatment of LTLD and non-invasive focused ultrasound hyperthermia in this study seemed to be clinically feasible, safe, and able to enhance intratumoural drug delivery, providing targeted chemo-ablative response in human liver tumours that were refractory to standard chemotherapy.
Oxford Biomedical Research Centre, National Institute for Health Research.
Allosteric proteins bind an effector molecule at one site resulting in a functional change at a second site. We hypothesize that allosteric communication in proteins relies upon networks of ...quaternary (collective, rigid-body) and tertiary (residue-residue contact) motions. We argue that cyclic topology of these networks is necessary for allosteric communication. An automated algorithm identifies rigid bodies from the displacement between the inactive and the active structures and constructs "quaternary networks" from these rigid bodies and the substrate and effector ligands. We then integrate quaternary networks with a coarse-grained representation of contact rearrangements to form "global communication networks" (GCNs). The GCN reveals allosteric communication among all substrate and effector sites in 15 of 18 multidomain and multimeric proteins, while tertiary and quaternary networks exhibit such communication in only 4 and 3 of these proteins, respectively. Furthermore, in 7 of the 15 proteins connected by the GCN, 50% or more of the substrate-effector paths via the GCN are "interdependent" paths that do not exist via either the tertiary or the quaternary network. Substrate-effector "pathways" typically are not linear but rather consist of polycyclic networks of rigid bodies and clusters of rearranging residue contacts. These results argue for broad applicability of allosteric communication based on structural changes and demonstrate the utility of the GCN. Global communication networks may inform a variety of experiments on allosteric proteins as well as the design of allostery into non-allosteric proteins.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
With the increased use of small self-complementary adeno-associated viral (AAV) vectors, the design of compact promoters becomes critical for packaging and expressing larger transgenes under ...ubiquitous or cell-specific control. In a comparative study of commonly used 800-bp cytomegalovirus (CMV) and chicken β-actin (CBA) promoters, we report significant differences in the patterns of cell-specific gene expression in the central and peripheral nervous systems. The CMV promoter provides high initial neural expression that diminishes over time. The CBA promoter displayed mostly ubiquitous and high neural expression, but substantially lower expression in motor neurons (MNs). We report the creation of a novel hybrid form of the CBA promoter (CBh) that provides robust long-term expression in all cells observed with CMV or CBA, including MNs. To develop a short neuronal promoter to package larger transgenes into AAV vectors, we also found that a 229-bp fragment of the mouse methyl-CpG-binding protein-2 (MeCP2) promoter was able to drive neuron-specific expression within the CNS. Thus the 800-bp CBh promoter provides strong, long-term, and ubiquitous CNS expression whereas the MeCP2 promoter allows an extra 570-bp packaging capacity, with low and mostly neuronal expression within the CNS, similar to the MeCP2 transcription factor.
Visceral leishmaniasis causes considerable mortality and morbidity in many parts of the world. There is an urgent need for the development of new, effective treatments for this disease. Here we ...describe the development of an anti-leishmanial drug-like chemical series based on a pyrazolopyrimidine scaffold. The leading compound from this series (7, DDD853651/GSK3186899) is efficacious in a mouse model of visceral leishmaniasis, has suitable physicochemical, pharmacokinetic and toxicological properties for further development, and has been declared a preclinical candidate. Detailed mode-of-action studies indicate that compounds from this series act principally by inhibiting the parasite cdc-2-related kinase 12 (CRK12), thus defining a druggable target for visceral leishmaniasis.
Allosteric kinase inhibitors offer a potentially complementary therapeutic strategy to ATP-competitive kinase inhibitors due to their distinct sites of target binding. In this study, we identify and ...study a mutant-selective EGFR allosteric inhibitor, JBJ-04-125-02, which as a single agent can inhibit cell proliferation and EGFR
signaling
and
. However, increased EGFR dimer formation limits treatment efficacy and leads to drug resistance. Remarkably, osimertinib, an ATP-competitive covalent EGFR inhibitor, uniquely and significantly enhances the binding of JBJ-04-125-02 for mutant EGFR. The combination of osimertinib and JBJ-04-125-02 results in an increase in apoptosis, a more effective inhibition of cellular growth, and an increased efficacy
and
compared with either single agent alone. Collectively, our findings suggest that the combination of a covalent mutant-selective ATP-competitive inhibitor and an allosteric EGFR inhibitor may be an effective therapeutic approach for patients with
-mutant lung cancer. SIGNIFICANCE: The clinical efficacy of EGFR tyrosine kinase inhibitors (TKI) in
-mutant lung cancer is limited by acquired drug resistance, thus highlighting the need for alternative strategies to inhibit EGFR. Here, we identify a mutant EGFR allosteric inhibitor that is effective as a single agent and in combination with the EGFR TKI osimertinib.
.
Segmental maxillary osteotomies require precise occlusal control due to variability in individual segment positioning. The role of maxillomandibular fixation (MMF) technique on occlusal control has ...not been validated.
The purpose is to measure and compare the accuracy of occlusal positioning among MMF techniques.
This was a double-blinded in vitro study on experiment models to simulate a 3-piece LeFort I osteotomy. The models were constricted posteriorly and expanded using 3 different MMF techniques and compared to the unaltered baseline occlusion. Based on sample size calculation, 32 separate attempts were made for each MMF technique.
The predictor variable was MMF technique (brackets, MMF screws, and embrasure wires).
The primary outcome variable was the visual occlusal analysis score, a 1.00 to 4.00 continuous scale measuring the similarity of the achieved occlusion to the planned (control) occlusion assessed by an oral and maxillofacial surgeon and an orthodontist. High visual occlusal analysis score indicated greater occlusal accuracy, with 3.50 defined as the threshold for accuracy. The secondary outcome variable was the linear error of the achieved occlusion at the canine and first molar teeth, with lower error indicating greater accuracy. An a priori accuracy threshold of 0.5 mm was set for this variable.
None.
Kruskal-Wallis test with post hoc testing was used to analyze the difference in the outcome variables of interest. P value < .05 was considered statistically significant.
Thirty-two attempts for each technique showed that brackets had higher VAOS than MMF screws and embrasure wires (median differences 1.49 and 0.48, P < .001), and had lower linear occlusal error (median differences 0.35 to 0.99 mm, P < .001).
MMF technique influences the quality of occlusal control, with greater visual rating scores and lower linear errors seen with brackets than with embrasure wires or MMF screws.
Selective activation of dopamine D1 receptors (D1Rs) has been pursued for 40 years as a therapeutic strategy for neurologic and psychiatric diseases due to the fundamental role of D1Rs in motor ...function, reward processing, and cognition. All known D1R-selective agonists are catechols, which are rapidly metabolized and desensitize the D1R after prolonged exposure, reducing agonist response. As such, drug-like selective D1R agonists have remained elusive. Here we report a novel series of selective, potent non-catechol D1R agonists with promising in vivo pharmacokinetic properties. These ligands stimulate adenylyl cyclase signaling and are efficacious in a rodent model of Parkinson's disease after oral administration. They exhibit distinct binding to the D1R orthosteric site and a novel functional profile including minimal receptor desensitization, reduced recruitment of β-arrestin, and sustained in vivo efficacy. These results reveal a novel class of D1 agonists with favorable drug-like properties, and define the molecular basis for catechol-specific recruitment of β-arrestin to D1Rs.