Small molecules inducing protein degradation are important pharmacological tools to interrogate complex biology and are rapidly translating into clinical agents. However, to fully realise the ...potential of these molecules, selectivity remains a limiting challenge. Herein, we addressed the issue of selectivity in the design of CRL4
recruiting PROteolysis TArgeting Chimeras (PROTACs). Thalidomide derivatives used to generate CRL4
recruiting PROTACs have well described intrinsic monovalent degradation profiles by inducing the recruitment of neo-substrates, such as GSPT1, Ikaros and Aiolos. We leveraged structural insights from known CRL4
neo-substrates to attenuate and indeed remove this monovalent degradation function in well-known CRL4
molecular glues degraders, namely CC-885 and Pomalidomide. We then applied these design principles on a previously published BRD9 PROTAC (dBRD9-A) and generated an analogue with improved selectivity profile. Finally, we implemented a computational modelling pipeline to show that our degron blocking design does not impact PROTAC-induced ternary complex formation. We believe that the tools and principles presented in this work will be valuable to support the development of targeted protein degradation.
Macrocyclic drugs can address an increasing range of molecular targets but enabling central nervous system (CNS) access to these drugs has been viewed as an intractable problem. We designed and ...synthesized a series of quinolinium-modified cyclosporine derivatives targeted to the mitochondrial cyclophilin D protein. Modification of the cation to enable greater delocalization was confirmed by x-ray crystallography of the cations. Critically, greater delocalization improved brain concentrations. Assessment of the compounds in preclinical assays and for pharmacokinetics identified a molecule JP1-138 with at least 20 times the brain levels of a non-delocalized compound or those reported for cyclosporine. Levels were maintained over 24 hours together with low hERG potential. The paradigm outlined here could have widespread utility in the treatment of CNS diseases.
Incorporation of a delocalized quinolinium can get macrocycles into the brain.
The mitochondrial permeability transition pore is a recognized drug target for neurodegenerative conditions such as multiple sclerosis and for ischemia-reperfusion injury in the brain and heart. The ...peptidylprolyl isomerase, cyclophilin D (CypD, PPIF), is a positive regulator of the pore, and genetic down-regulation or knock-out improves outcomes in disease models. Current inhibitors of peptidylprolyl isomerases show no selectivity between the tightly conserved cyclophilin paralogs and exhibit significant off-target effects, immunosuppression, and toxicity. We therefore designed and synthesized a new mitochondrially targeted CypD inhibitor, JW47, using a quinolinium cation tethered to cyclosporine. X-ray analysis was used to validate the design concept, and biological evaluation revealed selective cellular inhibition of CypD and the permeability transition pore with reduced cellular toxicity compared with cyclosporine. In an experimental autoimmune encephalomyelitis disease model of neurodegeneration in multiple sclerosis, JW47 demonstrated significant protection of axons and improved motor assessments with minimal immunosuppression. These findings suggest that selective CypD inhibition may represent a viable therapeutic strategy for MS and identify quinolinium as a mitochondrial targeting group for in vivo use.
Readily accessible, chiral complexes of aluminium are effective and enantioselective catalysts for the phospho-aldol reaction under aerobic conditions.
Chiral Schiff's base complexes of aluminium ...such as the methyl complex
1a (opposite) are effective and re-cyclable catalysts for the enantioselective phospho-aldol reaction under aerobic conditions, achieving enantioenrichments up to 50%.
Previous work has shown that pulsatile urea excretion at the gills of the gulf toadfish is due to periodic activation of a facilitated diffusion transport system with molecular and pharmacological ...similarity to the UT-A transport system of the mammalian kidney. In mammals, AVP and glucocorticoids are two important endocrine regulators of this system. The present study focused on the potential role of circulating AVT (the teleost homologue of AVP) and cortisol levels as possible triggers for urea pulses. Long-term (34–84 h) monitoring of plasma levels by repetitive sampling at 2-h intervals from chronic cannulae in individual toadfish demonstrated that circulating AVT concentrations are low (10
−12–10
−11 M), and show no relationship to the occurrence of natural urea pulses. In contrast, plasma cortisol levels decline greatly prior to natural pulses and rise rapidly thereafter. AVT injections into the caudal artery or ventral aorta elicited pulse events, but these were extremely small (1–10%) relative to natural pulses, and occurred only at unphysiological dose levels (10
−9 M in the plasma). AVP was a partial agonist, but isotocin, insulin-like growth factor-1, and atrial natriuretic peptide were without effect at the same concentration. Artificially raising plasma cortisol levels by cortisol injection tended to reduce responsiveness to AVT. Pharmacological reduction of plasma cortisol levels by metyrapone injection elicited small pulses similar to those caused by AVT. Following such pulse events, AVT was ineffective in inducing pulses. We conclude that
decreases in circulating cortisol play an important permissive role in urea pulsing, but that circulating AVT levels are not involved.
Researchers working on environmentally relevant organisms, populations, and communities are increasingly turning to the application of OMICS technologies to answer fundamental questions about the ...natural world, how it changes over time, and how it is influenced by anthropogenic factors. In doing so, the need to capture meta-data that accurately describes the biological "source" material used in such experiments is growing in importance. Here, we provide an overview of the formation of the "Env" community of environmental OMICS researchers and its efforts at considering the meta-data capture needs of those working in environmental OMICS. Specifically, we discuss the development to date of the Env specification, an informal specification including descriptors related to geographic location, environment, organism relationship, and phenotype. We then describe its application to the description of environmental transcriptomic experiments and how we have used it to extend the Minimum Information About a Microarray Experiment (MIAME) data standard to create a domain-specific extension that we have termed MIAME/Env. Finally, we make an open call to the community for participation in the Env Community and its future activities.
Arginine vasotocin (AVT) is a nonapeptide released from the neurohypophysis of fish. Evidence suggests that AVT is involved in the control of osmoregulation and cardiovascular actions suggest that ...AVT may also play a role in blood pressure homeostasis. In this study, flounder were cannulated under MS222 anesthesia in the dorsal aorta and this free swimming model was used to compare the blood pressure effects of AVT with responses from other neurohypophysial peptides in order to to characterize the receptors involved. Peptides were injected through the cannula (50 ng.kg super(-1)), and blood pressure was monitored via a pressure was transducer and recorded on a paper chart recorder (resting dorsal mean arterial pressure 31.0 plus or minus 3.0 mm Hg).
A specific radioimmunoassay was developed and characterised for the measurement of arginine vasotocin (AVT) in teleost fish. Crossreactivity of the antibody with the other teleost neurohypophysial ...peptide, isotocin, was less than 1 % and the sensitivity of the assay 0.25 pg/assay tube. Rapid blood sampling, within 1 minute of capture, by needle puncture of the caudal blood vessels, and collection of blood through chronic arterial cannula 1 - 4 days after surgery were identified as procedures which did not influence plasma ion levels, osmolality or hormone concentrations. These procedures were used in later experimental protocols examining the effects of chronic and acute osmotic stimuli on plasma AVT concentrations. Plasma AVT concentrations in a range of euryhaline and stenohaline teleost fish were measured at 10e-12 to low 10e-11 M (1-20 pg/ml). There was no consistent difference in circulating AVT levels in euryhaline fish (flounder, eel and trout) adapted to fresh water (FW) or seawater (SW) for at least 2 weeks. The flounder was studied in greater detail and seasonal influences were identified as having an effected on osmoregulation. In the winter/spring period plasma osmolality was significantly lower in FW adapted fish than at other times of the year. In SW adapted fish at this time plasma osmolality was elevated and this was accompanied by elevated plasma AVT concentrations. The winter/spring period was associated with the development of the gonads and this factor may affect osmoregulation in the flounder. In SW adapted flounder a positive linear relationship between plasma AVT and plasma osmolality and ion levels was observed, indicating a link between these two parameters but no such relationship was evident in FW adapted fish. Chronically cannulated SW adapted flounder were used to examine the effects of acute osmotic stimuli on plasma AVT concentrations. Intraperitoneal injection of hypertonic saline acutely increased plasma osmolality and was associated with a significant increase in plasma AVT concentration, indicating that AVT secretion was sensitive to increased plasma osmolality. The expansion of extracellar fluid volume via intravascular infusion of isotonic saline reduced plasma AVT concentrations independently of any change in plasma osmolality. This indicated that vascular volume also influences plasma AVT concentrations. Reducing blood volume by haemorrhage and lowering plasma osmolality by infusion of hypotonic saline did not result in any change in plasma AVT concentrations. Plasma AVT concentrations were measured in lamprey at a 10e-12 M concentration, and the transfer of lamprey from FW to Baltic SW caused an initial rise in plasma AVT concentrations, suggesting that AVT is involved in osmoregulation in these fish. AVT immunoreactivity was also demonstrated in two species of elasmobranch fish in a 10e-12 to low 10e-11 M range.
A specific radioimmunoassay was developed and characterised for the measurement of arginine vasotocin (A VT) in teleost fish. Crossreactivity of the antibody with the other teleost neurohypophysial ...peptide, isotocin, was less than 1 % and the sensitivity of the assay 0.25 pglassay tube. Rapid blood sampling, within 1 minute of capture, by needle puncture of the caudal blood vessels, and collection of blood through chronic arterial cannula 1 - 4 days after surgery were identified as procedures which did not influence plasma ion levels, osmolality or hormone concentrations. These procedures were used in later experimental protocols examining the effects of chronic and acute osmotic stimuli on plasma A VT concentrations. Plasma A VT concentrations in a range of euryhaline and stenohaline teleost fish were measured at 10-12 to low 10-11 M (1-20 pglml). There was no consistent difference in circulating A VT levels in euryhaline fish (flounder, eel and trout) adapted to fresh water (FW) or seawater (SW) for at least 2 weeks. The flounder was studied in greater detail and seasonal influences were identified as having an effected on osmoregulation. In the winter/spring period plasma osmolality was significantly lower in FW adapted fish than at other times of the year. In SW adapted fish at this time plasma osmolality was elevated and this was accompanied by elevated plasma A VT concentrations. The winter/spring period was associated with the development of the gonads and this factor may affect osmoregulation in the flounder. In SW adapted flounder a positive linear relationship between plasma A VT and plasma osmolality and ion levels was observed, indicating a link between these two parameters but no such relationship was evident in FW adapted fish. Chronically cannulated SW adapted flounder were used to examine the effects of acute osmotic stimuli on plasma A VT concentrations. Intraperitoneal injection of hypertonic saline acutely increased plasma osmolality and was associated with a significant increase in plasma A VT concentration, indicating that A VT secretion was sensitive to increased plasma osmolality. The expansion of extracellar fluid volume via intravascular infusion of isotonic saline reduced plasma A VT concentrations independently of any change in plasma osmolality. This indicated that vascular volume also influences plasma A VT concentrations. Reducing blood volume by haemorrhage and lowering plasma osmolality by infusion of hypotonic saline did not result in any change in plasma A VT concentrations. Plasma A VT concentrations were measured in lamprey at a 10-12 M concentration, and the transfer of lamprey from FW to Baltic SW caused an initial rise in plasma A VT concentrations, suggesting that A VT is involved in osmoregulation in these fish. A VT immunoreactivity was also demonstrated in two species of elasmobranch fish in a 10-12 to low 10-11 M range.
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