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LIM domain kinases 1 and 2 (LIMK1 and LIMK2) regulate actin dynamics and subsequently key cellular functions such as proliferation and migration. LIMK1 and LIMK2 phosphorylate and inactivate cofilin leading to increased actin polymerization. As a result, LIMK inhibitors are emerging as a promising treatment strategy for certain cancers and neurological disorders. High-quality chemical probes are required if the role of these kinases in health and disease is to be understood. To that end, we report the results of a comparative assessment of 17 reported LIMK1/2 inhibitors in a variety of

Lead Optimisation (LO) is a critical part of the drug discovery process as it is the part where all efforts that go into earlier parts of the process (Target Identification, HTS, Hit-to-Lead, Lead Identification) are crystallised into a single compound, the candidate drug. The optimisation phase is like a treasure hunt, and discovery projects can take many paths to discover their treasured molecule. In this chapter we have chosen to focus on the practicalities of how the LO project can be organised, and what different property issues the project may face as it attempts to find an efficient path forward to find a suitable candidate drug for development. This multiparameter optimisation dance can have many issues, often interconnected. We discuss approaches to many of these problems and offer advice on how project members should think about tackling issues in a holistic way. In this way we hope to support projects in understanding the right issues to tackle at the right time. We offer some tips, and examples, of successes, and suggest that identifying the issue to tackle is key to establishing robust hypotheses to test and so drive projects forward. Our goal in this chapter is to aid project thinking about the best way forward.

Dual-specificity tyrosine-regulated kinase 1A (DYRK1A) regulates the proliferation and differentiation of neuronal progenitor cells during brain development. Consequently, DYRK1A has attracted interest as a target for the treatment of neurodegenerative diseases, including Alzheimer's disease (AD) and Down's syndrome. Recently, the inhibition of DYRK1A has been investigated as a potential treatment for diabetes, while DYRK1A's role as a mediator in the cell cycle has garnered interest in oncologic indications. Structure-activity relationship (SAR) analysis in combination with high-resolution X-ray crystallography leads to a series of pyrazolo[1,5-b]pyridazine inhibitors with excellent ligand efficiencies, good physicochemical properties, and a high degree of selectivity over the kinome. Compound 11 exhibited good permeability and cellular activity without P-glycoprotein liability, extending the utility of 11 in an in vivo setting. These pyrazolo[1,5-b]pyridazines are a viable lead series in the discovery of new therapies for the treatment of diseases linked to DYRK1A function.

Ectopic expression in T-cell precursors of LIM only protein 2 (LMO2), a key factor in hematopoietic development, has been linked to the onset of T-cell acute lymphoblastic leukaemia (T-ALL). In the T-ALL context, LMO2 drives oncogenic progression through binding to erythroid-specific transcription factor SCL/TAL1 and sequestration of E-protein transcription factors, normally required for T-cell differentiation. A key requirement for the formation of this oncogenic protein-protein interaction (PPI) is the conformational flexibility of LMO2. Here we identify a small molecule inhibitor of the SCL-LMO2 PPI, which hinders the interaction in vitro through direct binding to LMO2. Biophysical analysis demonstrates that this inhibitor acts through a mechanism of conformational modulation of LMO2. Importantly, this work has led to the identification of a small molecule inhibitor of the SCL-LMO2 PPI, which can provide a starting point for the development of new agents for the treatment of T-ALL. These results suggest that similar approaches, based on the modulation of protein conformation by small molecules, might be used for therapeutic targeting of other oncogenic PPIs.

Graphical abstract, Kinases are signalling proteins which have proven to be successful targets for the treatment of a variety of diseases, predominantly in cancers. However, only a small proportion of kinases (

Purpose: There is considerable interest in positive allosteric modulators (PAMs) of the α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA) subtype of ionotropic glutamate receptors as therapeutic agents for a range of cognitive and mood disorders. However, the challenge is to increase AMPA receptor (AMPAR) function sufficient to enhance cognitive function but not to the extent that there are mechanism-related pro-convulsant or convulsant side effects. In this present study, we report the preclinical pharmacology data for MDI-222, an AMPAR PAM which enhances cognition but has a much reduced side-effect (i.e. convulsant) liability relative to other molecules of this mechanism. Methods: The pharmacological effects of MDI-222 were characterised in in vitro and in vivo preclinical electrophysiology, efficacy (cognition), side-effect (pro-convulsant/convulsant), tolerability and toxicity assays. Results: We demonstrate that MDI-222 is an AMPAR PAM, since it enhanced AMPAR function in vitro at human (hGluA1-4) and rat (rGluA2) homomeric receptors, and potentiated hetero-oligomeric AMPARs in rat neurons. MDI-222 enhanced electrically evoked AMPAR-mediated synaptic transmission in the anaesthetised rat at 10 mg/kg (administered intravenously) and did not significantly lower the seizure threshold in the pro-convulsant maximal electroshock threshold test (MEST) at any dose tested up to a maximum of 30 mg/kg (administered by oral gavage (p.o.)). MDI-222 reversed a delay-induced deficit in novel object recognition (NOR) in rats with a minimum effective dose (MED) of 0.3 mg/kg (p.o.) following acute administration, which was reduced to 0.1 mg/kg following sub-chronic administration, and improved passive avoidance performance in scopolamine-impaired rats with a MED of 10 mg/kg p.o. On the other hand, MDI-222 was not pro-convulsant in the MEST, resulting in a therapeutic window between plasma concentrations that enhanced cognitive performance and those associated with mechanism-related side effects of ⩾1000-fold. Unfortunately, despite the excellent preclinical profile of this compound, further development had to be halted due to non-mechanism-related issues. Conclusions: We conclude that MDI-222 is an AMPAR PAM which enhances cognitive performance in rats and has a significantly improved safety profile in preclinical species.

Simple Summary Liver metastasis remains the major contributor in colorectal cancer-related death. It has become clear that the unique immune features of liver microenvironment take part in many steps of metastatic cascade, from pre-metastatic niche formation, tumor cell colonization to metastatic tumor establishment. Therefore, better understanding of mechanisms orchestrating the formation of a hospitable hepatic metastatic niche is necessary for the development of effective therapies. This review summarizes the current understandings of the critical role of liver immune microenvironment in metastasis development and provides therapeutic perspective on targeting the metastasis-prone microenvironment. Abstract A drastic difference exists between the 5-year survival rates of colorectal cancer patients with localized cancer and distal organ metastasis. The liver is the most favorable organ for cancer metastases from the colorectum. Beyond the liver-colon anatomic relationship, emerging evidence highlights the impact of liver immune microenvironment on colorectal liver metastasis. Prior to cancer cell dissemination, hepatocytes secrete multiple factors to recruit or activate immune cells and stromal cells in the liver to form a favorable premetastatic niche. The liver-resident cells including Kupffer cells, hepatic stellate cells, and liver-sinusoidal endothelial cells are co-opted by the recruited cells, such as myeloid-derived suppressor cells and tumor-associated macrophages, to establish an immunosuppressive liver microenvironment suitable for tumor cell colonization and outgrowth. Current treatments including radical surgery, systemic therapy, and localized therapy have only achieved good clinical outcomes in a minority of colorectal cancer patients with liver metastasis, which is further hampered by high recurrence rate. Better understanding of the mechanisms governing the metastasis-prone liver immune microenvironment should open new immuno-oncology avenues for liver metastasis intervention.

To identify small molecules that shield mammalian sensory hair cells from the ototoxic side effects of aminoglycoside antibiotics we screened 10,240 compounds, selecting those that protected against neomycin and gentamicin in zebrafish lateral-line hair cells and, when retested in mouse cochlear cultures, prevented gentamicin-induced death of outer hair cells (OHCs). Of 64 compounds that protected zebrafish hair cells, 8 protect OHCs from gentamicin in vitro. These hits share structural features and all block, to varying degrees, the OHC’s mechano-electrical transducer (MET) channel, a known route of aminoglycoside entry into hair cells. Further characterisation of one of the strongest MET-channel blockers, UoS-7692, revealed it additionally protects against kanamycin and tobramycin, and does not abrogate the bactericidal activity of gentamicin. UoS-7692 behaves, like the aminoglycosides, as a permeant blocker of the MET channel, significantly reduces gentamicin-Texas Red loading into OHCs, and preserves lateral-line function in neomycin-treated zebrafish. Trans-tympanic injection of UoS-7692 protects mouse OHCs from furosemide-kanamycin exposure in vivo and partially preserves hearing. The results confirm the hair-cell MET channel as a viable target for the identification of compounds that protect the cochlea from aminoglycosides, and provide a series of hit compounds that will inform the design of future otoprotectants.

RV521 is an orally bioavailable inhibitor of respiratory syncytial virus (RSV) fusion that was identified after a lead optimization process based upon hits that originated from a physical property directed hit profiling exercise at Reviral. This exercise encompassed collaborations with a number of contract organizations with collaborative medicinal chemistry and virology during the optimization phase in addition to those utilized as the compound proceeded through preclinical and clinical evaluation. RV521 exhibited a mean IC50 of 1.2 nM against a panel of RSV A and B laboratory strains and clinical isolates with antiviral efficacy in the Balb/C mouse model of RSV infection. Oral bioavailability in preclinical species ranged from 42 to >100% with evidence of highly efficient penetration into lung tissue. In healthy adult human volunteers experimentally infected with RSV, a potent antiviral effect was observed with a significant reduction in viral load and symptoms compared to placebo.

BLM (Bloom syndrome protein) is a RECQ-family helicase involved in the dissolution of complex DNA structures and repair intermediates. Synthetic lethality analysis implicates BLM as a promising target in a range of cancers with defects in the DNA damage response, however selective small molecule inhibitors of defined mechanism are currently lacking. Here we identify and characterise a specific inhibitor of BLM’s ATPase-coupled DNA helicase activity, by allosteric trapping of a DNA-bound translocation intermediate. Crystallographic structures of BLM-DNA-ADP-inhibitor complexes identify a hitherto unknown interdomain interface, whose opening and closing are integral to translocation of ssDNA, and which provides a highly selective pocket for drug discovery. Comparison with structures of other RECQ helicases provides a model for branch migration of Holliday junctions by BLM.

Human serine racemase (hSR) catalyses racemisation of L-serine to D-serine, the latter of which is a co-agonist of the NMDA subtype of glutamate receptors that are important in synaptic plasticity, learning and memory. In a 'closed' hSR structure containing the allosteric activator ATP, the inhibitor malonate is enclosed between the large and small domains while ATP is distal to the active site, residing at the dimer interface with the Tyr121 hydroxyl group contacting the α-phosphate of ATP. In contrast, in 'open' hSR structures, Tyr121 sits in the core of the small domain with its hydroxyl contacting the key catalytic residue Ser84. The ability to regulate SR activity by flipping Tyr121 from the core of the small domain to the dimer interface appears to have evolved in animals with a CNS. Multiple X-ray crystallographic enzyme-fragment structures show Tyr121 flipped out of its pocket in the core of the small domain. Data suggest that this ligandable pocket could be targeted by molecules that inhibit enzyme activity.

Human serine racemase (hSR) catalyses racemisation of L-serine to D-serine, the latter of which is a co-agonist of the NMDA subtype of glutamate receptors that are important in synaptic plasticity, learning and memory. In a ‘closed’ hSR structure containing the allosteric activator ATP, the inhibitor malonate is enclosed between the large and small domains while ATP is distal to the active site, residing at the dimer interface with the Tyr121 hydroxyl group contacting the ATP a-phosphate. In contrast, in ‘open’ hSR structures, Tyr121 sits in the core of the small domain with its hydroxyl contacting the key catalytic residue Ser84. The ability to regulate SR activity by flipping Tyr121 from the core of the small domain to the dimer interface appears to have evolved in animals with a CNS. Multiple X-ray crystallographic enzymefragment structures show that Tyr121 is flipped out of its pocket, suggesting that this pocket is druggable.

Kinases represent one of the most intensively pursued groups of targets in modern-day drug discovery. Often it is desirable to achieve selective inhibition of the kinase of interest over the remaining ∼500 kinases in the human kinome. This is especially true when inhibitors are intended to be used to study the biology of the target of interest. We present a pipeline of open-source software that analyzes public domain data to repurpose compounds that have been used in previous kinase inhibitor development projects. We define the dual-specificity tyrosine-regulated kinase 1A (DYRK1A) as the kinase of interest, and by addition of a single methyl group to the chosen starting point we remove glycogen synthase kinase β (GSK3β) and cyclin-dependent kinase (CDK) inhibition. Thus, in an efficient manner we repurpose a GSK3β/CDK chemotype to deliver 8b, a highly selective DYRK1A inhibitor.

γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter within the central nervous system (CNS) with fast, transsynaptic, and modulatory extrasynaptic effects being mediated by the ionotropic GABA type A receptors (GABAARs). These receptors are of particular interest because they are the molecular target of a number of pharmacological agents, of which the benzodiazepines (BZDs), such as diazepam, are the best described. The anxiolytic, sedating, and myorelaxant effects of BZDs are mediated by separate populations of GABAARs containing either α1, α2, α3, or α5 subunits and the molecular dissection of the pharmacology of BZDs indicates that subtype-selective GABAAR modulators will have novel pharmacological profiles. This is best exemplified by α2/α3-GABAAR positive allosteric modulators (PAMs) and α5-GABAAR negative allosteric modulators (NAMs), which were originally developed as nonsedating anxiolytics and cognition enhancers, respectively. This review aims to summarize the current state of the field of subtype-selective GABAAR modulators acting via the BZD binding site and their potential clinical indications.

Background and Purpose AMPA receptor positive allosteric modulators represent a potential therapeutic strategy to improve cognition in people with schizophrenia. These studies collectively constitute the preclinical pharmacology data package used to build confidence in the pharmacology of this molecule and enable a clinical trial application. Experimental Approach [N-[(2S)-5-(6-fluoro-3-pyridinyl)-2,3-dihydro 1H–inden-2-yl]-2-propanesulfonamide] (UoS12258) was profiled in a number of in vitro and in vivo studies to highlight its suitability as a novel therapeutic agent. Key Results We demonstrated that UoS12258 is a selective, positive allosteric modulator of the AMPA receptor. At rat native hetero-oligomeric AMPA receptors, UoS12258 displayed a minimum effective concentration of approximately 10 nM in vitro and enhanced AMPA receptor-mediated synaptic transmission at an estimated free brain concentration of approximately 15 nM in vivo. UoS12258 reversed a delay-induced deficit in novel object recognition in rats after both acute and sub-chronic dosing. Sub-chronic dosing reduced the minimum effective dose from 0.3 to 0.03 mg·kg−1. UoS12258 was also effective at improving performance in two other cognition models, passive avoidance in scopolamine-impaired rats and water maze learning and retention in aged rats. In side-effect profiling studies, UoS12258 did not produce significant changes in the maximal electroshock threshold test at doses below 10 mg·kg−1. Conclusion and Implications We conclude that UoS12258 is a potent and selective AMPA receptor modulator exhibiting cognition enhancing properties in several rat behavioural models superior to other molecules that have previously entered clinical evaluation.

γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter within the central nervous system (CNS) with fast, transsynaptic, and modulatory extrasynaptic effects being mediated by the ionotropic GABA type A receptors (GABA

Aminoglycosides (AGs) are broad-spectrum antibiotics used for the treatment of serious bacterial infections but have use-limiting side effects including irreversible hearing loss. Here, we assessed the otoprotective profile of carvedilol in mouse cochlear cultures and in vivo zebrafish assays and investigated its mechanism of protection which, we found, may be mediated by a block of the hair cell’s mechanoelectrical transducer (MET) channel, the major entry route for the AGs. To understand the full otoprotective potential of carvedilol, a series of 18 analogues were prepared and evaluated for their effect against AG-induced damage as well as their affinity for the MET channel. One derivative was found to confer greater protection than carvedilol itself in cochlear cultures and also to bind more tightly to the MET channel. At higher concentrations, both carvedilol and this derivative were toxic in cochlear cultures but not in zebrafish, suggesting a good therapeutic window under in vivo conditions.

The neurotransmitter glutamate and its receptors have long been of interest to scientists involved in pharmaceutical research since dysfunction of the glutamatergic signalling pathway has been associated with the pathophysiology of\ud several psychiatric and neurological disorders. The research on AMPAR positive allosteric modulators offers opportunities to modulate fast excitatory synaptic transmission and identify new potential therapeutic agents for a range of neurodiseases. The field of AMPAR modulators continues to be a dynamic area of drug discovery with a pronounced diversification of the chemotypes explored in recent\ud years. This article reviews literature published in this area in the last 6 years, focusing on the new core templates, some derived from high-throughput screens, with an emphasis on structure-activity relationships, drug metabolism and pharmacokinetics properties, and pharmacological profiles of these series.

Cyclin-dependent kinase 20 (CDK20), or more commonly referred to as cell cycle-related kinase (CCRK), is the latest member of CDK family with strong linkage to human cancers. Accumulating studies have reported the consistent overexpression of CCRK in cancers arising from brain, colon, liver, lung and ovary. Such aberrant up-regulation of CCRK is clinically significant as it correlates with tumor staging, shorter patient survival and poor prognosis. Intriguingly, the signalling molecules perturbed by CCRK are divergent and cancer-specific, including the cell cycle regulators CDK2, cyclin D1, cyclin E and RB in glioblastoma, ovarian carcinoma and colorectal cancer, and KEAP1-NRF2 cytoprotective pathway in lung cancer. In hepatocellular carcinoma (HCC), CCRK mediates virus-host interaction to promote hepatitis B virus-associated tumorigenesis. Further mechanistic analyses reveal that CCRK orchestrates a self-reinforcing circuitry comprising of AR, GSK3β, β-catenin, AKT, EZH2, and NF-κB signalling for transcriptional and epigenetic regulation of oncogenes and tumor suppressor genes. Notably, EZH2 and NF-κB in this circuit have been recently shown to induce IL-6 production to facilitate tumor immune evasion. Concordantly, in a hepatoma preclinical model, ablation of Ccrk disrupts the immunosuppressive tumor microenvironment and enhances the therapeutic efficacy of immune checkpoint blockade via potentiation of anti-tumor T cell responses. In this review, we summarized the multifaceted tumor-intrinsic and -extrinsic functions of CCRK, which represents a novel signalling hub exploitable in cancer immunotherapy.

The halodecarboxylation of heteroarene carboxylic acids by treatment with N -bromosuccinimide or N -chlorosuccinimide was performed. This procedure provides a convenient route to synthetically useful mono-halogenated heteroarene intermediates such as halo-indoles, -aza-indoles, -indazoles and -aza-indazoles. The mild conditions employed and simple protocol provides an advantage over traditional halodecarboxylation procedures that require expensive and toxic metal catalysts, basic conditions, time-consuming intermediate isolation and elevated reaction temperatures.

New tools are required to ensure the adequate control of the neglected tropical disease human African trypanosomiasis. Annual reports of infection have recently fallen to fewer than 5000 cases per year; however, current therapies are hard to administer and have safety concerns and, hence, are far from ideal. Trypanosome alternative oxidase is an exciting target for controlling the infection; it is unique to the parasite, and inhibition of this enzyme with the natural product ascofuranone has shown to clear in vivo infections. We report the synthesis and associated structure activity relationships of inhibitors based upon this natural product with correlation to T. b. brucei growth inhibition in an attempt to generate molecules that possess improved physicochemical properties and potential for use as new treatments for human African trypanosomiasis.

Aminoglycoside antibiotics are used to treat life-threatening bacterial infections but can cause deafness due to hair cell death in the inner ear. Compounds have been described that protect zebrafish lateral line hair cells from aminoglycosides, but few are effective in the cochlea. As the aminoglycosides interact with several ion channels, including the mechanoelectrical transducer (MET) channels by which they can enter hair cells, we screened 160 ion-channel modulators, seeking compounds that protect cochlear outer hair cells (OHCs) from aminoglycoside-induced death in vitro. Using zebrafish, 72 compounds were identified that either reduced loading of the MET-channel blocker FM 1-43FX, decreased Texas red–conjugated neomycin labeling, or reduced neomycin-induced hair cell death. After testing these 72 compounds, and 6 structurally similar compounds that failed in zebrafish, 13 were found that protected against gentamicin-induced death of OHCs in mouse cochlear cultures, 6 of which are permeant blockers of the hair cell MET channel. None of these compounds abrogated aminoglycoside antibacterial efficacy. By selecting those without adverse effects at high concentrations, 5 emerged as leads for developing pharmaceutical otoprotectants to alleviate an increasing clinical problem.

The role of glutamate and its receptors in central nervous system biology and disease has long been of interest to scientists involved in both fundamental research and drug discovery, however the complex pharmacology and lack of highly selective compounds has severely hampered drug discovery efforts in this area. Recent advances in the identification and profiling of positive allosteric modulators of the AMPA receptor offer a potential way forward and the hope of a new treatment for schizophrenia. This article will review recent patent applications published in this area.

Colorectal cancer is the third most common cancer in the world. Despite surgery, up to 50% of patients relapse with incurable disease. First-line chemotherapy uses the topoisomerase 1 (TOP1) poison irinotecan, which triggers cell death by trapping TOP1 on DNA. The removal of TOP1 peptide from TOP1–DNA breaks is conducted by tyrosyl-DNA phosphodiesterase 1 (TDP1). Despite putative roles for TDP1 and TOP1 in colorectal cancer, their role in cellular and clinical responses to TOP1-targeting therapies remains unclear. Here, we show varying expression levels of TOP1 and TDP1 polypeptides in multiple colorectal cancer cell lines and in clinical colorectal cancer samples. TDP1 overexpression or TOP1 depletion is protective. Conversely, TDP1 depletion increases DNA-strand breakage and hypersensitivity to irinotecan in a TOP1-dependent manner, presenting a potential therapeutic opportunity in colorectal cancer. TDP1 protein levels correlate well with mRNA and with TDP1 catalytic activity. However, no correlation is observed between inherent TDP1 or TOP1 levels alone and irinotecan sensitivity, pointing at their limited utility as predictive biomarkers in colorectal cancer. These findings establish TDP1 as a potential therapeutic target for the treatment of colorectal cancer and question the validity of TOP1 or TDP1 on their own as predictive biomarkers for irinotecan response. Mol Cancer Ther; 14(2); 575–85. ©2014 AACR.

African trypanosomiasis is a parasitic disease affecting 5000 humans and millions of livestock animals in sub-Saharan Africa every year. Current treatments are limited, difficult to administer and often toxic causing long term injury or death in many patients. Trypanosome alternative oxidase is a parasite specific enzyme whose inhibition by the natural product ascofuranone (AF) has been shown to be curative in murine models. Until now synthetic methods to AF analogues have been limited, this has restricted both understanding of the key structural features required for binding and also how this chemotype could be developed to an effective therapeutic agent. The development of 3 amenable novel synthetic routes to ascofuranone-like compounds is described. The SAR generated around the AF chemotype is reported with correlation to the inhibition of T. b. brucei growth and corresponding selectivity in cytotoxic assessment in mammalian HepG2 cell lines. These methods allow access to greater synthetic diversification and have enabled the synthesis of compounds that have and will continue to facilitate further optimisation of the AF chemotype into a drug-like lead., Graphical abstract Image 1, Highlights • Synthesis of ascofuranone like inhibitors of trypanosome alternative oxidase. • Structure activity relationships of trypanosome alternative oxidase inhibitors. • Correlation of trypanosome alternative oxidase inhibition and T. b. brucei growth.

Aminoglycoside antibiotics are widely used for the treatment of life-threatening bacterial infections, but cause permanent hearing loss in a substantial proportion of treated patients. The sensory hair cells of the inner ear are damaged following entry of these antibiotics via the mechano-electrical transducer (MET) channels located at the tips of the hair cell’s stereocilia. d-Tubocurarine (dTC) is a MET channel blocker that reduces the loading of gentamicin-Texas Red (GTTR) into rat cochlear hair cells and protects them from gentamicin treatment. Berbamine is a structurally related alkaloid that reduces GTTR labeling of zebrafish lateral-line hair cells and protects them from aminoglycoside-induced cell death. Both compounds are thought to reduce aminoglycoside entry into hair cells through the MET channels. Here we show that dTC (≥6.25 µM) or berbamine (≥1.55 µM) protect zebrafish hair cells in vivo from neomycin (6.25 µM, 1 h). Protection of zebrafish hair cells against gentamicin (10 µM, 6 h) was provided by ≥25 µM dTC or ≥12.5 µM berbamine. Hair cells in mouse cochlear cultures are protected from longer-term exposure to gentamicin (5 µM, 48 h) by 20 µM berbamine or 25 µM dTC. Berbamine is, however, highly toxic to mouse cochlear hair cells at higher concentrations (≥30 µM) whilst dTC is not. The absence of toxicity in the zebrafish assays prompts caution in extrapolating results from zebrafish neuromasts to mammalian cochlear hair cells. MET current recordings from mouse outer hair cells (OHCs) show that both compounds are permeant open-channel blockers, rapidly and reversibly blocking the MET channel with half-blocking concentrations of 2.2 µM (dTC) and 2.8 µM (berbamine) in the presence of 1.3 mM Ca2+ at −104 mV. Berbamine, but not dTC, also blocks the hair cell’s basolateral K + current, IK,neo, and modeling studies indicate that berbamine permeates the MET channel more readily than dTC. These studies reveal key properties of MET-channel blockers required for the future design of successful otoprotectants.

Glutamate receptors and the associated neurotransmission have been of interest to the pharmaceutical research community for many years since dysfunction of the glutamatergic signaling pathway has been associated with the pathophysiology of several psychiatric and neurological disorders. The research on ligands modulating the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR), one subtype of glutamate ionotropic receptor, offers opportunities to modulate fast excitatory synaptic transmission, and to identify new potential therapeutic agents for a range of neurodisease, such as schizophrenia and epilepsy. The field of AMPAR ligands continues to be a dynamic area of drug discovery, with a pronounced diversification of the chemotypes explored as positive allosteric modulators in recent years. This article reviews literature published in this area in the last decade.

Topoisomerase 1 (TOP1) generates transient nicks in the DNA to relieve torsional stress encountered during the cellular processes of transcription, replication, and recombination. At the site of the nick there is a covalent linkage of TOP1 with DNA via a tyrosine residue. This reversible TOP1-cleavage complex intermediate can become trapped on DNA by TOP1 poisons such as camptothecin, or by collision with replication or transcription machinery, thereby causing protein-linked DNA single- or double-strand breaks and resulting in cell death. Tyrosyl-DNA phosphodiesterase 1 (TDP1) is a key enzyme involved in the repair of TOP1-associated DNA breaks via hydrolysis of 3′-phosphotyrosine bonds. Inhibition of TDP1 is therefore an attractive strategy for targeting cancer cells in conjunction with TOP1 poisons. Existing methods for monitoring the phosphodiesterase activity of TDP1 are generally gel based or of high cost. Here we report a novel, oligonucleotide-based fluorescence assay that is robust, sensitive, and suitable for high-throughput screening of both fragment and small compound libraries for the detection of TDP1 inhibitors. We further validated the assay using whole cell extracts, extending its potential application to determine of TDP1 activity in clinical samples from patients undergoing chemotherapy.

The topoisomerase I (TOP1) inhibitor irinotecan triggers cell death by trapping TOP1 on DNA, generating cytotoxic protein-linked DNA breaks (PDBs). Despite its wide application in a variety of solid tumors, the mechanisms of cancer cell resistance to irinotecan remains poorly understood. Here, we generated colorectal cancer (CRC) cell models for irinotecan resistance and report that resistance is neither due to downregulation of the main cellular target of irinotecan TOP1 nor upregulation of the key TOP1 PDB repair factor TDP1. Instead, the faster repair of PDBs underlies resistance, which is associated with perturbed histone H4K16 acetylation. Subsequent treatment of irinotecan-resistant, but not parental, CRC cells with histone deacetylase (HDAC) inhibitors can effectively overcome resistance. Immunohistochemical analyses of CRC tissues further corroborate the importance of histone H4K16 acetylation in CRC. Finally, the resistant clones exhibit cross-resistance with oxaliplatin but not with ionising radiation or 5-fluoruracil, suggesting that the latter two could be employed following loss of irinotecan response. These findings identify perturbed chromatin acetylation in irinotecan resistance and establish HDAC inhibitors as potential therapeutic means to overcome resistance.

AMPA receptor positive allosteric modulators represent a potential therapeutic strategy to improve cognition in people with schizophrenia. These studies collectively constitute the preclinical pharmacology data package used to build confidence in the pharmacology of this molecule and enable a clinical trial application.[N-[(2S)-5-(6-fluoro-3-pyridinyl)-2,3-dihydro 1H-inden-2-yl]-2-propanesulfonamide] (UoS12258) was profiled in a number of in vitro and in vivo studies to highlight its suitability as a novel therapeutic agent.We demonstrated that UoS12258 is a selective, positive allosteric modulator of the AMPA receptor. At rat native hetero-oligomeric AMPA receptors, UoS12258 displayed a minimum effective concentration of approximately 10 nM in vitro and enhanced AMPA receptor-mediated synaptic transmission at an estimated free brain concentration of approximately 15 nM in vivo. UoS12258 reversed a delay-induced deficit in novel object recognition in rats after both acute and sub-chronic dosing. Sub-chronic dosing reduced the minimum effective dose from 0.3 to 0.03 mg·kgWe conclude that UoS12258 is a potent and selective AMPA receptor modulator exhibiting cognition enhancing properties in several rat behavioural models superior to other molecules that have previously entered clinical evaluation.

A novel approach of the convergent functionalisation of aryl dibromides to form NS5A type inhibitors using C–H activation is reported. The focus of investigation was to reduce the formation of homodimeric side product, as well as to investigate the scope of different aryl dibromides that were tolerated under the reaction conditions. The C–H activation methodology was found to give a viable synthetic route to NS5A inhibitors, with late stage functionalisation of the core portion of the molecule, albeit with some chemical functionalities not tolerated.

A series of novel AMPA receptor positive modulators displaying CNS penetration have been discovered with sub-micromolar activity and good selectivity over the cardiac channel receptor, hERG. We describe here the synthesis of these compounds which are biaryl pyrrolidine and tetrahydrofuran sulfonamides and disclose their activities against the human GluA2 flip isoform homotetrameric receptor.

Gene expression frequently requires the dynamic remodelling of the chromatin architecture to orchestrate the assembly of transcriptional protein complexes at the promoter sites of DNA. Chromatin remodelling complexes are multicomponent proteins that facilitate reorganisation of chromatin structure using the energy derived from ATP hydrolysis, and as a result play a pivotal role in controlling gene expression. Mammalian SWI/SNF complexes are examples of chromatin remodelling complexes with multiple roles in maintaining genome stability. It is estimated that 19.6% of all human tumours have mutations in SWI/SNF complexes, leading to the deregulation of gene expression as an early driver of carcinogenesis. Our interest in developing novel treatments for renal cancer focused on the role of the chromatin remodelling complex, PBAF. The BAF180 component of PBAF is mutated in 40 % of clear cell renal carcinomas and its loss is an early event in renal cell tumorigenesis. We performed an siRNA based high throughput screen to identify genes associated with genome stability that were synthetically lethal with the loss of BAF180 and identified KAT2a as a potential druggable target. KAT2a is a protein that also functions in the regulation of gene expression as a component of the SAGA histone modifying complex. It is a multifunctional protein that can both ‘read' and ‘write' acetylation marks on histone proteins through its bromodomain and a histone acetyltransferase domain respectively. We will describe our efforts to establish suitable assays to support the discovery of selective inhibitors of the bromodomain of KAT2a. The work will include the establishment of a high throughput assay to measure compound binding, crystallographic characterisation of small molecule inhibitors of the bromodomain, and our initial studies towards the target validation of the bromodomain to recapitulate the synthetic lethality observed with depletion of BAF180. Citation Format: Andrea Gunnell, Jessica Downs, Lewis Pennicott, Kay Osborn, Darren Le Grand, Katie Duffell, Hitesh Patel, Jessica Hudson, Jessica R. Booth, Simon Ward. Targeting the epigenetic regulator KAT2a in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2907.

This article highlights recent advances in the discovery of new agonists, antagonists and partial agonists of the 5-HT(1D) receptor. The field of 5-HT(1D) agonists continues to deliver a number of new potential therapeutic agents, although advances in this field are now more focussed on the clinical evaluation phase. The identification of novel compounds is greater for the 5-HT(1D) receptor antagonists, and whilst few truly selective ligands have been identified, a number of approaches are discussed towards defined mixed-pharmacology profiles. An overview is also given of recent advances in biological and clinical understanding of the receptor.

AMPA receptors consist of a family of hetero-oligomeric (tetrameric) receptors arising from four genes, each of which encodes a distinct receptor subunit (GluA1-4). Recombinant homo-tetrameric AMPA receptors, comprising four identical subunits, are functionally active and have been used in in vitro assays. However, the many different subunit permutations make possible the functional and anatomical diversity of AMPA receptors throughout the CNS. Furthermore, AMPA receptor subunit stoichiometry influences the biophysical and functional properties of the receptor. A number of chemically diverse positive modulators of AMPA receptor have been identified which potentiate AMPA receptor-mediated activity in vitro as well as improving cognitive performance in rodents and non-human primates with several being taken further in the clinic. This review article summarizes the current status in the research on positive allosteric modulation of AMPA receptors and outlines the challenges involved in identifying a chemically distinct series of AMPA receptor positive modulators, addressing the challenges created by the heterogeneity of the AMPA receptor populations and the development of structure-activity relationships driven by homomeric, recombinant systems on high-throughput platforms. We also review the role of X-ray crystallography in the selection and prioritization of targets for lead optimization for AMPA receptor positive modulators.

The (-)-(11R,2'S)-enantiomer of the antimalarial drug mefloquine has been found to be a reasonably potent and moderately selective adenosine A(2A) receptor antagonist. Further investigation of this compound has led to the discovery of a series of keto-aryl thieno[3,2-d]pyrimidine derivatives, which are potent and selective antagonists of the adenosine A(2A) receptor. These derivatives show selectivity against the A(1) receptor. Furthermore, some of these compounds have been shown to have in vivo activity in a commonly used model, suggesting the potential for the treatment of Parkinson's disease.

The DNA damage response (DDR) is essential for maintaining the genomic integrity of the cell, and its disruption is one of the hallmarks of cancer. Classically, defects in the DDR have been exploited therapeutically in the treatment of cancer with radiation therapies or genotoxic chemotherapies. More recently, protein components of the DDR systems have been identified as promising avenues for targeted cancer therapeutics. Here, we present an in-depth analysis of the function, role in cancer and therapeutic potential of 450 expert-curated human DDR genes. We discuss the DDR drugs that have been approved by the US Food and Drug Administration (FDA) or that are under clinical investigation. We examine large-scale genomic and expression data for 15 cancers to identify deregulated components of the DDR, and we apply systematic computational analysis to identify DDR proteins that are amenable to modulation by small molecules, highlighting potential novel therapeutic targets.

Lead Optimisation (LO) is a critical part of the drug discovery process as it is the part where all efforts that go into earlier parts of the process (Target Identification, HTS, Hit-to-Lead, Lead Identification) are crystallised into a single compound, the candidate drug. The optimization phase is like a treasure hunt, and discovery projects can take many paths to discover their treasured molecule. In this Chapter we have chosen to focus on the practicalities of how the LO project can be organized, and what different property issues the project may face as it attempts to find an efficient path forward to find a suitable candidate drug for Development. This multiparameter optimization dance can have many issues, often interconnected. We discuss approaches to many of these problems, and offer advice on how project members should think about tackling issues in a holistic way. In this way we hope to support projects in understanding the right issues to tackle at the right time. We offer some tips, and examples, of successes, and suggest that identifying the issue to tackle is key to establishing robust hypotheses to test and so drive projects forwards. Our goal in this Chapter is to aid project thinking about the best way forward.

Drug discovery is a constantly developing and expanding area of research. Developed to provide a comprehensive guide, the Handbook of Medicinal Chemistry covers the past, present and future of the entire drug development process. Highlighting the recent successes and failures in drug discovery, the book will help readers to understand the factors governing modern drug discovery from the initial concept through to a marketed medicine. With chapters covering a wide range of topics from drug discovery processes and optimization, development of synthetic routes, pharmaceutical properties and computational biology, the handbook aims to help medicinal chemists to apply their academic understanding to every aspect of drug discovery. Each chapter includes expert advice to not only provide a rigorous understanding of the principles being discussed, but to provide useful hints and tips gained from within the pharmaceutical industry. This expertise, combined with project case studies, highlighting and discussing all areas of successful projects, make this an essential handbook for all those involved in pharmaceutical development. A free app has been created in collaboration with the editors of the book. The Medicinal Chemistry Toolkit provides a suite of resources to support the day to day work of a medicinal chemist.