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Exploring the kinetic selectivity of drugs targeting the beta1-adrenoceptor
No full textIn this study, we report the beta1-adrenoceptor binding kinetics of several clinically relevant beta1/2-adrenoceptor (beta1/2AR) agonists and antagonists. We demonstrate that the physicochemical properties of a molecule directly affect its kinetic association rate (kon) and affinity for the target. In contrast to our findings at the beta2-adrenoceptor, a drug’s immobilized artificial membrane partition coefficient (KIAM), reflecting both hydrophobic and electrostatic interactions of the drug with the charged surface of biological membranes, was no better predictor than simple hydrophobicity measurements such as log P or logD7.4, characterized by a distribution between water and a non-aqueous organic phase (e.g. n-octanol) at predicting association rate. Overall, this suggests that hydrophobic interactions rather than a combination of polar and hydrophobic interactions play a more prominent role in dictating the binding of these ligands to the beta1-adrenoceptor.
Using a combination of kinetic data, detailed structural and physicochemical information we rationalize the above findings and speculate that the association of positively charged ligands at the beta1AR is curtailed somewhat by its predominantly neutral/positive charged extracellular surface. Consequently, hydrophobic interactions in the ligand binding pocket dominate the kinetics of ligand binding. In comparison at the beta2AR, a combination of hydrophobicity and negative charge attracts basic, positively charged ligands to the receptor’s surface promoting the kinetics of ligand binding. Additionally, we reveal the potential role kinetics plays in the on-target and off-target pharmacology of clinically used beta-blockers
Considerations and recommendations for assessment of plasma protein binding and drug-drug interactions for siRNA therapeutics
No full textAt the time of writing, although four siRNA therapeutics have been approved for human use, no official regulatory guidance specific to this modality is available. In the absence of guidance, preclinical development for siRNA followed a hybrid of the small molecule (ICH M3(R2)) and biologics (ICH S6(R1)) guidance documents. However, siRNA differs significantly from small molecules and protein-based biologics in both its physicochemical and absorption, distribution, metabolism and excretion (ADME) properties, and its mechanism of action. Consequently, certain reports typically included in filing packages for small molecule or biologics may benefit from adaption, or even omission, from an siRNA filing. In this white paper, members of the ‘siRNA plasma protein binding and drug-drug interaction working group’ in the IQ consortium discuss the relevance of two such reports---the plasma protein binding (PPB) evaluation and the drug-drug interaction (DDI) risk assessment---to support siRNA regulatory filings. Publicly available siRNA approval packages and the literature were systematically reviewed to examine the role of siRNA PPB and DDIs in ADME, safety and translation. This information has been summarized into two decision trees to help guide industry to decide when siRNA PPB and DDI studies are warranted
A high-throughput drug screen reveals means to differentiate triple-negative breast cancer.
No full textPlasticity delineates cancer subtypes with more or less favourable outcomes. In breast cancer, the subtype triple-negative lacks expression of major differentiation markers, e.g., estrogen receptor α (ERα), and its high cellular plasticity results in greater aggressiveness and poorer prognosis than other subtypes. Whether plasticity itself represents a potential vulnerability of cancer cells is not clear. However, we show here that cancer cell plasticity can be exploited to differentiate triple-negative breast cancer (TNBC). Using a high-throughput imaging-based reporter drug screen with 9 501 compounds, we have identified three polo-like kinase 1 (PLK1) inhibitors as major inducers of ERα protein expression and downstream activity in TNBC cells. PLK1 inhibition upregulates a cell differentiation program characterized by increased DNA damage, mitotic arrest, and ultimately cell death. Furthermore, cells surviving PLK1 inhibition have decreased tumorigenic potential, and targeting PLK1 in already established tumours reduces tumour growth both in cell line- and patient-derived xenograft models. In addition, the upregulation of genes upon PLK1 inhibition correlates with their expression in normal breast tissue and with better overall survival in breast cancer patients. Our results indicate that differentiation therapy based on PLK1 inhibition is a potential alternative strategy to treat TNBC
Editorial for "Evaluation of Dynamic Contrast-Enhanced MRI Measures of Lung Congestion and Endothelial Permeability in Heart Failure: A Prospective Method Validation Study”
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Design of potent antimalarials with generative chemistry
No full textRecent advances in generative modelling allow designing novel compounds through deep neural networks. One such neural network model, JT-VAE (the Junction Tree Variational Auto-Encoder), excels at proposing chemically valid structures. Here, on the basis of JT-VAE, we built a generative modelling approach, JAEGER, for finding novel chemical matter with desired bioactivity. Using JAEGER, we designed compounds to inhibit malaria. To prioritize the compounds for synthesis, we used the in-house pQSAR (Profile-QSAR) program, a massively multitask bioactivity model based on 12,000 Novartis assays. On the basis of pQSAR activity predictions, we selected, synthesized and experimentally profiled two compounds. Both compounds exhibited low nanomolar activity in a malaria proliferation assay as well as a biochemical assay measuring activity against PI(4)K, which is an essential kinase that regulates intracellular development in malaria. The compounds also showed low activity in a cytotoxicity assay. Our findings show that JAEGER is a viable approach for finding novel active compounds for drug discovery
Host succinate inhibits influenza virus infection through succinylation and nuclear retention of the viral nucleoprotein.
No full textInfluenza virus infection causes considerable morbidity and mortality, but current therapies have limited efficacy. We hypothesized that investigating the metabolic signaling during infection may help to design innovative antiviral approaches. Using bronchoalveolar lavages of infected mice, we here demonstrate that influenza virus induces a major reprogramming of lung metabolism. We focused on mitochondria-derived succinate that accumulated both in the respiratory fluids of virus-challenged mice and of patients with influenza pneumonia. Notably, succinate displays a potent antiviral activity in vitro as it inhibits the multiplication of influenza A/H1N1 and A/H3N2 strains and strongly decreases virus-triggered metabolic perturbations and inflammatory responses. Moreover, mice receiving succinate intranasally showed reduced viral loads in lungs and increased survival compared to control animals. The antiviral mechanism involves a succinate-dependent posttranslational modification, that is, succinylation, of the viral nucleoprotein at the highly conserved K87 residue. Succinylation of viral nucleoprotein altered its electrostatic interactions with viral RNA and further impaired the trafficking of viral ribonucleoprotein complexes. The finding that succinate efficiently disrupts the influenza replication cycle opens up new avenues for improved treatment of influenza pneumonia
People of TM: Video of Florencia Segal
No full textThe video will be used for an external social media engagement campaign on platforms like linked-in, facebook etc. featuring stories of people in TM.
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Home cage measures of Alzheimer's disease in the rTg4510 mouse model
No full textAlzheimer's disease affects an array of activities in patients' daily lives but measures
other than memory are rarely evaluated in animal models. Home cage behavior, however, may provide an opportunity to back translate a variety of measures seen in
human disease progression to animal models, providing external and face validity.
The aim of this study was to evaluate if home cage measures could indicate disease
in the rTg4510 mouse model. We hypothesized that sleep, nesting, and smell discrimination would be altered in mutant mice. Thirty-two transgenic mice were used in a
Latin square design of four genotypes x both sexes x two diets. Half the mice
received a doxycycline diet to suppress tauopathy and evaluate tau severity on various measures. At 8-, 12-, and 16-weeks old, 24 h activity/sleep patterns, nest complexity, and odor discrimination were measured. After 16-weeks, tau concentration
in the brain was quantified. Mutant mice had increased tau concentration in brain tissue, but it was reduced by the doxycycline diet. However, only nest complexity was
different between mutant mice and controls. Overall, tauopathy in rTg4510 mice
does seem to affect these commonly observed symptoms in human patients. However, while running this study, a report showed that the rTg4510 mutant phenotype
is not caused by the mutation itself, but confounding factors from transgene insertion. Combined with report findings and our data, the rTg4510 model may not be an
ideal model for all aspects of human Alzheimer's disease
The Proteomic Profile of Interstitial Lung Abnormalities.
No full textRationale Knowledge on biomarkers of interstitial lung disease is incomplete. Interstitial lung abnormalities (ILA) are radiologic changes that may present in its early stages. Objectives To uncover blood proteins associated with ILA using large-scale proteomics methods. Methods Data from two prospective cohort studies, the AGES-Reykjavik study (n=5,259) for biomarker discovery and the Genetic Epidemiology of COPD (COPDGene) study (n=4,899) for replication, were used. Blood proteins were measured using DNA aptamers, targeting over 4,700 protein analytes. The association of proteins with ILA and ILA progression was assessed with regression modelling, as were associations with genetic risk factors. Adaptive LASSO models were applied to bootstrap data samples to discover sets of proteins predictive of ILA and their progression. Measurements and Main Results Of 287 associations, SFTPB (OR 3.71 [95% CI 3.20-4.30], P 4.28×10-67), SCG3AB1 (OR 2.43 [2.13-2.77], P 8.01×10-40) and WFDC2 (OR 2.42 [2.11-2.78], P 4.01×10-36) were most significantly associated with ILA in AGES-Reykjavik and were replicated in COPDGene. In AGES-Reykjavik, levels of SFTPB were associated with the rs35705950 MUC5B promoter polymorphism and SFTPB and WFDC2 had the strongest associations with ILA progression. Multivariate models of ILA in AGES-Reykjavik, ILA in COPDGene and ILA progression in AGES-Reykjavik, had validated areas under the receiver operating characteristic curve of 0.880, 0.826 and 0.824, respectively. Conclusions Novel, replicated associations of ILA, its progression and genetic risk factors with numerous blood proteins are demonstrated as well as machine-learning based models with favourable predictive potential. Several proteins are revealed as potential markers of early fibrotic lung disease
The effect of mAb and excipient cryoconcentration on long-term frozen storage stability - part 2: Aggregate formation and oxidation.
No full textWe examined the impact of monoclonal antibody (mAb) and buffer concentration, mimicking the cryoconcentration found upon freezing in a 2 L bottle, on mAb stability during frozen storage. Upon cryoconcentration, larger protein molecules and small excipient molecules freeze-concentrate differently, resulting in different protein to stabiliser ratios within a container. Understanding the impact of these shifted ratios on protein stability is essential. For two mAbs a set of samples with constant mAb (5 mg/mL) or buffer concentration (medium histidine/adipic acid) was prepared and stored for 6 months at -10 °C. Stability was evaluated via size-exclusion chromatography, flow imaging microscopy, UV/Vis spectroscopy at 350 nm, and protein A chromatography. Dynamic light scattering was used to determine kD values. Soluble aggregate levels were unaffected by mAb concentration, but increased with histidine concentration. No trend in optical density could be identified. In contrast, increasing mAb or buffer concentration facilitated the formation of subvisible particles. A trend towards attractive protein-protein interactions was seen with higher ionic strength. MAb oxidation levels were negatively affected by increasing histidine concentration, but became less with higher mAb concentration. Small changes in mAb and buffer composition had a significant impact on stability during six-month frozen storage. Thus, preventing cryoconcentration effects in larger freezing containers may improve long-term stability