ABSTRACT
This investigation compared the spatial ecology and population dynamics of brown trout Salmo trutta L. between reservoirs with (impact; Langsett Reservoir) and without (control; Grimwith Reservoir) barriers to fish movements into headwater tributaries, and the effectiveness of a fish pass intended to remediate connectivity. Passive integrated transponder (PIT) telemetry revealed that fish that emigrated from Langsett and Grimwith tributaries were 1-3 and 0-2 years old, respectively, and predominantly did so in spring and autumn-early winter in both systems. Weirs at Langsett Reservoir appeared to thwart emigration rate (26%) relative to Grimwith Reservoir (85%). Acoustic telemetry (two-dimensional positions) in the impacted reservoir revealed that the largest home range was in October-December (95% monthly activity space S.D. up to 26.9 ± 6.69 ha in November), activity was influenced by both month and time of day, and fish occupied shallow water depths (relative to reservoir depth), especially at night. Brown trout tagged in Grimwith and Langsett Reservoirs (42.9% and 64.1%, respectively) and fish tagged in the tributaries that emigrated (37.2% and 27.7%, respectively) were detected immigrating into tributaries throughout the year. At both reservoirs, peak immigration for ≥3-year-old trout occurred primarily in autumn-early winter. Overall passage efficiency went from 3% prior to remediation to 14% after and there was no significant increase in fish densities following the construction of the fish pass. Fish were attracted towards and entered the fish pass under a wide range of river levels, but only succeeded in passing upstream during low levels, which are uncommon druing the main migration period. Overall, this investigation significantly furthers our understanding of brown trout spatial ecology and population dynamics in reservoirs and headwater tributaries.
ABSTRACT
A major drawback of cytotoxic chemotherapy is the lack of selectivity toward noncancerous cells. The targeted delivery of cytotoxic drugs to tumor cells is a longstanding goal in cancer research. We proposed that covalent inhibitors could be adapted to deliver cytotoxic agents, conjugated to the ß-position of the Michael acceptor, via an addition-elimination mechanism promoted by covalent binding. Studies on model systems showed that conjugated 5-fluorouracil (5FU) could be released upon thiol addition in relevant time scales. A series of covalent epidermal growth factor receptor (EGFR) inhibitors were synthesized as their 5FU derivatives. Achieving the desired release of 5FU was demonstrated to depend on the electronics and geometry of the compounds. Mass spectrometry and NMR studies demonstrated an anilinoquinazoline acrylate ester conjugate bound to EGFR with the release of 5FU. This work establishes that acrylates can be used to release conjugated molecules upon covalent binding to proteins and could be used to develop targeted therapeutics.
Subject(s)
Cytotoxins , Fluorouracil , Fluorouracil/pharmacology , ErbB Receptors , Esters , Mass SpectrometryABSTRACT
The extracellular signal-regulated kinase 5 (ERK5) signaling pathway is one of four conventional mitogen-activated protein (MAP) kinase pathways. Genetic perturbation of ERK5 has suggested that modulation of ERK5 activity may have therapeutic potential in cancer chemotherapy. This Miniperspective examines the evidence for ERK5 as a drug target in cancer, the structure of ERK5, and the evolution of structurally distinct chemotypes of ERK5 kinase domain inhibitors. The emerging complexities of ERK5 pharmacology are discussed, including the confounding phenomenon of paradoxical ERK5 activation by small-molecule ERK5 inhibitors. The impact of the recent development and biological evaluation of potent and selective bifunctional degraders of ERK5 and future opportunities in ERK modulation are also explored.
Subject(s)
MAP Kinase Signaling System , Signal Transduction , Signal Transduction/physiology , Phosphorylation , Mitogen-Activated Protein Kinase 7 , Protein Processing, Post-TranslationalABSTRACT
BACKGROUND: We evaluated the therapeutic potential of combining the monocarboxylate transporter 1 (MCT1) inhibitor AZD3965 with the mitochondrial respiratory Complex I inhibitor IACS-010759, for the treatment of diffuse large B-cell lymphoma (DLBCL), a potential clinically actionable strategy to target tumour metabolism. METHODS: AZD3965 and IACS-010759 sensitivity were determined in DLBCL cell lines and tumour xenograft models. Lactate concentrations, oxygen consumption rate and metabolomics were examined as mechanistic endpoints. In vivo plasma concentrations of IACS-010759 in mice were determined by LC-MS to select a dose that reflected clinically attainable concentrations. RESULTS: In vitro, the combination of AZD3965 and IACS-010759 is synergistic and induces DLBCL cell death, whereas monotherapy treatments induce a cytostatic response. Significant anti-tumour activity was evident in Toledo and Farage models when the two inhibitors were administered concurrently despite limited or no effect on the growth of DLBCL xenografts as monotherapies. CONCLUSIONS: This is the first study to examine a combination of two distinct approaches to targeting tumour metabolism in DLBCL xenografts. Whilst nanomolar concentrations of either AZD3965 or IACS-010759 monotherapy demonstrate anti-proliferative activity against DLBCL cell lines in vitro, appreciable clinical activity in DLBCL patients may only be realised through their combined use.
Subject(s)
Lymphoma, Large B-Cell, Diffuse , Symporters , Animals , Apoptosis , Cell Line, Tumor , Glycolysis , Humans , Lymphoma, Large B-Cell, Diffuse/pathology , Mice , Monocarboxylic Acid Transporters , Oxidative Phosphorylation , Symporters/metabolismABSTRACT
The nonclassical extracellular signal-related kinase 5 (ERK5) mitogen-activated protein kinase pathway has been implicated in increased cellular proliferation, migration, survival, and angiogenesis; hence, ERK5 inhibition may be an attractive approach for cancer treatment. However, the development of selective ERK5 inhibitors has been challenging. Previously, we described the development of a pyrrole carboxamide high-throughput screening hit into a selective, submicromolar inhibitor of ERK5 kinase activity. Improvement in the ERK5 potency was necessary for the identification of a tool ERK5 inhibitor for target validation studies. Herein, we describe the optimization of this series to identify nanomolar pyrrole carboxamide inhibitors of ERK5 incorporating a basic center, which suffered from poor oral bioavailability. Parallel optimization of potency and in vitro pharmacokinetic parameters led to the identification of a nonbasic pyrazole analogue with an optimal balance of ERK5 inhibition and oral exposure.
Subject(s)
Mitogen-Activated Protein Kinase 7 , Pyrroles , Cell Proliferation , Pyrroles/pharmacologyABSTRACT
MDM2 is a key negative regulator of the p53 tumor suppressor. Direct binding of MDM2 to p53 represses the protein's transcriptional activity and induces its polyubiquitination, targeting it for degradation by the proteasome. Consequently, small molecule inhibitors that antagonize MDM2-p53 binding, such as RG7388, have progressed into clinical development aiming to reactivate p53 function in TP53 wild-type tumors. Here, we describe the design, synthesis, and biological evaluation of a trans-cyclooctene tagged derivative of RG7388, RG7388-TCO, which showed high cellular potency and specificity for MDM2. The in-cell reaction of RG7388-TCO with a tetrazine-tagged BODIPY dye enabled fluorescence imaging of endogenous MDM2 in SJSA-1 and T778 tumor cells. RG7388-TCO was also used to pull down MDM2 by reaction with tetrazine-tagged agarose beads in SJSA-1 lysates. The data presented show that RG733-TCO enables precise imaging of MDM2 in cells and can permit a relative assessment of target engagement and MDM2-p53 antagonism in vitro.
Subject(s)
Boron Compounds/chemistry , Fluorescent Dyes/chemistry , Proto-Oncogene Proteins c-mdm2/analysis , Proto-Oncogene Proteins c-mdm2/metabolism , Pyrrolidines/chemistry , Tumor Suppressor Protein p53/metabolism , para-Aminobenzoates/chemistry , Cell Line, Tumor , Click Chemistry , Cyclooctanes/analogs & derivatives , Cyclooctanes/pharmacology , Humans , Molecular Docking Simulation , Optical Imaging/methods , Protein Binding/drug effects , Protein Interaction Maps/drug effects , Pyrrolidines/pharmacology , para-Aminobenzoates/pharmacologyABSTRACT
Inhibition of monocarboxylate transporter 1 has been proposed as a therapeutic approach to perturb lactate shuttling in tumor cells that lack monocarboxylate transporter 4. We examined the monocarboxylate transporter 1 inhibitor AZD3965, currently in phase I clinical studies, as a potential therapy for diffuse large B-cell lymphoma and Burkitt lymphoma. Whilst extensive monocarboxylate transporter 1 protein was found in 120 diffuse large B-cell lymphoma and 10 Burkitt lymphoma patients' tumors, monocarboxylate transporter 4 protein expression was undetectable in 73% of the diffuse large B-cell lymphoma samples and undetectable or negligible in each Burkitt lymphoma sample. AZD3965 treatment led to a rapid accumulation of intracellular lactate in a panel of lymphoma cell lines with low monocarboxylate transporter 4 protein expression and potently inhibited their proliferation. Metabolic changes induced by AZD3965 in lymphoma cells were consistent with a feedback inhibition of glycolysis. A profound cytostatic response was also observed in vivo: daily oral AZD3965 treatment for 24 days inhibited CA46 Burkitt lymphoma growth by 99%. Continuous exposure of CA46 cells to AZD3965 for 7 weeks in vitro resulted in a greater dependency upon oxidative phosphorylation. Combining AZD3965 with an inhibitor of mitochondrial complex I (central to oxidative phosphorylation) induced significant lymphoma cell death in vitro and reduced CA46 disease burden in vivo These data support clinical examination of AZD3965 in Burkitt lymphoma and diffuse large B-cell lymphoma patients with low tumor monocarboxylate transporter 4 expression and highlight the potential of combination strategies to optimally target the metabolic phenotype of tumors.