Synthesis of 3,4-dihydroxy-1,8-naphthalimides with inbuilt catechol and crown ether functionalities.

Herein the synthesis of 1,8-naphthalimides functionalised as the 3,4-dihydroxy-1,8-naphthalimide (catechol, Nap-Cat) and the corresponding 15-crown-5 (Nap-Crown) is reported. These compounds represent the first examples where these two recognition groups are directly incorporated into the 1,8-naphthalimide ring system. Both Nap-Cat and Nap-Crown were evaluated for their capacity to respond to analytes such as H2O2 (a mimic for cellular oxidation) and metal ions (as elements of environmental and physiological interest). While slow oxidation was observed for Nap-Cat upon prolonged exposure to H2O2, no significant changes in photophysical properties were observed upon treatment of Nap-Crown with metal ions.

Nickel-Catalyzed C-N Cross-Coupling of 4-Chloro-1,8-naphthalimides and Bulky, Primary Alkylamines at Room Temperature.

4-Amino-1,8-naphthalimides, potentially useful fluorescent probes in biological applications, are prepared via Ni(cod)2/IPr-catalyzed cross-couplings between 4-chloro-1,8-naphthalimide electrophiles and α,α,α-trisubstituted, primary alkylamines at room temperature. This method represents the first synthesis of 4-amino-1,8-naphthalimides using Ni-catalyzed C-N cross-coupling and provides the first examples of 4-amino-1,8-naphthalimides incorporating such bulky primary alkylamines, thereby highlighting the utility of Ni-catalyzed processes in synthesizing naphthalimide scaffolds that were inaccessible using established methods (SNAr; Pd or Cu catalysis).

Effect of Triton X-100 on the Activity and Selectivity of Lipase Immobilized on Chemically Reduced Graphene Oxides.

The effect of support hydrophobicity on lipase activity and substrate selectivity was investigated with and without Triton X-100 (TX-100). Lipases from Thermomyces lanuginosa (TL) and Alcaligenes sp. (QLM) were immobilized on graphene oxide (GO) and a range of chemically reduced graphene oxides (CRGOs) with different levels of surface hydrophobicity. Activity assays using 4-hydroxy-N-propyl-1,8-naphthalimide (NAP) esters of varying chain lengths (NAP-butyrate (NAP-B), NAP-octanoate (NAP-O), and NAP-palmitate (NAP-P)) showed that the activity of immobilized QLM and TL decreased by more than 60% on GO and 80% on CRGO (2 h), with activity decreasing further as surface hydrophobicity of the CRGOs increased. Across the hydrophobicity range of GO/CRGOs, the substrate selectivity of QLM shifted from more readily hydrolyzing NAP-P to NAP-B, while TL retained its substrate selectivity for NAP-O. Lipase TL was also shown to desorb from GO and 2 h CRGO when mixed with NAP-O and NAP-P, whereas QLM did not. Circular dichroism analyses of the lipase α-helix content correlate to the observed activity data, with decreases in the α-helical content (40% in TL and 20% in QLM relative to free lipase) consistent with decreases in activity after immobilization on GO. α-Helical content decreased even further as the surface hydrophobicity of CRGOs increased. Attenuated total reflectance-Fourier transform infrared spectroscopy also showed significant changes to the lipase secondary structure upon immobilization. The addition of TX-100 into the activity assay modified the substrate selectivity of immobilized QLM, improving the activity against NAP-O (90%) and NAP-P (67%) compared to the activity measured without TX-100. It was shown that TX-100 primarily affected the activity of QLM by interacting with the ester substrate and the lipase itself. This study provides an improved understanding of how support hydrophobicity and the presence of TX-100 can affect activity/selectivity of lipases immobilized on hydrophobic supports.

Property activity refinement of 2-anilino 4-amino substituted quinazolines as antimalarials with fast acting asexual parasite activity.

Malaria is a devastating disease caused by Plasmodium parasites. Emerging resistance against current antimalarial therapeutics has engendered the need to develop antimalarials with novel structural classes. We recently described the identification and initial optimization of the 2-anilino quinazoline antimalarial class. Here, we refine the physicochemical properties of this antimalarial class with the aim to improve aqueous solubility and metabolism and to reduce adverse promiscuity. We show the physicochemical properties of this class are intricately balanced with asexual parasite activity and human cell cytotoxicity. Structural modifications we have implemented improved LipE, aqueous solubility and in vitro metabolism while preserving fast acting P. falciparum asexual stage activity. The lead compounds demonstrated equipotent activity against P. knowlesi parasites and were not predisposed to resistance mechanisms of clinically used antimalarials. The optimized compounds exhibited modest activity against early-stage gametocytes, but no activity against pre-erythrocytic liver parasites. Confoundingly, the refined physicochemical properties installed in the compounds did not engender improved oral efficacy in a P. berghei mouse model of malaria compared to earlier studies on the 2-anilino quinazoline class. This study provides the framework for further development of this antimalarial class.

Cross-Coupling of Amide and Amide Derivatives to Umbelliferone Nonaflates: Synthesis of Coumarin Derivatives and Fluorescent Materials.

The Buchwald-Hartwig cross-coupling reaction between 4-methylumbelliferone-derived nonaflates with amides, carbamates, and sulfonamides is described. A wide variety of N-substituted 7-amino coumarin analogues was prepared in good to excellent yields. The photophysical properties of aqueous-soluble derivatives were determined, and they displayed auxochrome-based variations. Gram-scale synthesis provided an acrylamide analogue, which was used to fabricate a fluorescent poly(2-hydroxylethyl methacrylate) (pHEMA) hydrogel that was resistant to leaching in ultrapure H2O. We envisage that our reported protocol to access 7-amino-4-methylcoumarin derivatives will find use toward the development of new fluorescent coumarin-based probes by researchers in the field.

Evaluation of 4-Amino 2-Anilinoquinazolines against Plasmodium and Other Apicomplexan Parasites In Vitro and in a P. falciparum Humanized NOD-scid IL2Rγnull Mouse Model of Malaria.

A series of 4-amino 2-anilinoquinazolines optimized for activity against the most lethal malaria parasite of humans, Plasmodium falciparum, was evaluated for activity against other human Plasmodium parasites and related apicomplexans that infect humans and animals. Four of the most promising compounds from the 4-amino 2-anilinoquinazoline series were equally as effective against the asexual blood stages of the zoonotic P. knowlesi, suggesting that they could also be effective against the closely related P. vivax, another important human pathogen. The 2-anilinoquinazoline compounds were also potent against an array of P. falciparum parasites resistant to clinically available antimalarial compounds, although slightly less so than against the drug-sensitive 3D7 parasite line. The apicomplexan parasites Toxoplasma gondii, Babesia bovis, and Cryptosporidium parvum were less sensitive to the 2-anilinoquinazoline series with a 50% effective concentration generally in the low micromolar range, suggesting that the yet to be discovered target of these compounds is absent or highly divergent in non-Plasmodium parasites. The 2-anilinoquinazoline compounds act as rapidly as chloroquine in vitro and when tested in rodents displayed a half-life that contributed to the compound's capacity to clear P. falciparum blood stages in a humanized mouse model. At a dose of 50 mg/kg of body weight, adverse effects to the humanized mice were noted, and evaluation against a panel of experimental high-risk off targets indicated some potential off-target activity. Further optimization of the 2-anilinoquinazoline antimalarial class will concentrate on improving in vivo efficacy and addressing adverse risk.

Luminescent probes for the bioimaging of small anionic species in vitro and in vivo.

The ability to spatiotemporally identify the formation of specific anionic species, or track changes in their concentration inside living systems, is of critical importance in deciphering their exact biological roles and effects. The development of probes (also called bioimaging agents and intracellular sensors) to achieve this goal has become a rapidly growing branch of supramolecular chemistry. In this critical review the challenges specific to the task are identified and for a select range of small anions of environmental and biological relevance (fluoride, chloride, iodide, cyanide, pyrophosphate, bicarbonate, hydrosulphide, peroxynitrite, hypochlorite and hypobromite) a comprehensive overview of the currently available in vitro and in vivo probes is provided.

Blue Electrogenerated Chemiluminescence from Water-Soluble Iridium Complexes Containing Sulfonated Phenylpyridine or Tetraethylene Glycol Derivatized Triazolylpyridine Ligands.

Incorporating phenylpyridine- and triazolylpyridine-based ligands decorated with methylsulfonate or tetraethylene glycol (TEG) groups, a series of iridium(III) complexes has been created for green and blue electrogenerated chemiluminescence under analytically useful aqueous conditions, with tri-n-propylamine as a coreactant. The relative electrochemiluminescence (ECL) intensities of the complexes were dependent on the sensitivity of the photodetector over the wavelength range and the pulse time of the applied electrochemical potential. In terms of the integrated area of corrected ECL spectra, with a pulse time of 0.5 s, the intensities of the Ir(III) complexes were between 18 and 102 % that of [Ru(bpy)3 ](2+) (bpy=2,2'-bipyridine). However, when the intensities were measured with a typical bialkali photomultiplier tube, the signal of the most effective blue emitter, [Ir(df-ppy)2 (pt-TEG)](+) (df-ppy=2-(2,4-difluorophenyl)pyridine anion, pt-TEG=1-(2-(2-(2-(2-hydroxyethoxy)ethoxy)ethoxy)ethyl)-4-(2-pyridyl)-1,2,3-triazole), was over 1200 % that of the orange-red emitter [Ru(bpy)3 ](2+) . A combined experimental and theoretical investigation of the electrochemical and spectroscopic properties of the Ir(III) complexes indicated that the greater intensity from [Ir(df-ppy)2 (pt-TEG)](+) relative to those of the other Ir(III) complexes resulted from a combination of many factors, rather than being significantly favored in one area.

Synthesis and evaluation of cationic norbornanes as peptidomimetic antibacterial agents.

A series of structurally amphiphilic biscationic norbornanes have been synthesised as rigidified, low molecular weight peptidomimetics of cationic antimicrobial peptides. A variety of charged hydrophilic functionalities were attached to the norbornane scaffold including aminium, guanidinium, imidazolium and pyridinium moieties. Additionally, a range of hydrophobic groups of differing sizes were incorporated through an acetal linkage. The compounds were evaluated for antibacterial activity against both Gram-negative and Gram-positive bacteria. Activity was observed across the series; the most potent of which exhibited an MIC's ≤ 1 µg mL(-1) against Streptococcus pneumoniae, Enterococcus faecalis and several strains of Staphylococcus aureus, including multi-resistant methicillin resistant (mMRSA), glycopeptide-intermediate (GISA) and vancomycin-intermediate (VISA) S. aureus.

In silico identification of an aryl hydrocarbon receptor antagonist with biological activity in vitro and in vivo.

The aryl hydrocarbon receptor (AHR) is critically involved in several physiologic processes, including cancer progression and multiple immune system activities. We, and others, have hypothesized that AHR modulators represent an important new class of targeted therapeutics. Here, ligand shape-based virtual modeling techniques were used to identify novel AHR ligands on the basis of previously identified chemotypes. Four structurally unique compounds were identified. One lead compound, 2-((2-(5-bromofuran-2-yl)-4-oxo-4H-chromen-3-yl)oxy)acetamide (CB7993113), was further tested for its ability to block three AHR-dependent biologic activities: triple-negative breast cancer cell invasion or migration in vitro and AHR ligand-induced bone marrow toxicity in vivo. CB7993113 directly bound both murine and human AHR and inhibited polycyclic aromatic hydrocarbon (PAH)- and TCDD-induced reporter activity by 75% and 90% respectively. A novel homology model, comprehensive agonist and inhibitor titration experiments, and AHR localization studies were consistent with competitive antagonism and blockade of nuclear translocation as the primary mechanism of action. CB7993113 (IC50 3.3 × 10(-7) M) effectively reduced invasion of human breast cancer cells in three-dimensional cultures and blocked tumor cell migration in two-dimensional cultures without significantly affecting cell viability or proliferation. Finally, CB7993113 effectively inhibited the bone marrow ablative effects of 7,12-dimethylbenz[a]anthracene in vivo, demonstrating drug absorption and tissue distribution leading to pharmacological efficacy. These experiments suggest that AHR antagonists such as CB7993113 may represent a new class of targeted therapeutics for immunomodulation and/or cancer therapy.

Optimization of pyrazolopyridine 4-carboxamides with potent antimalarial activity for which resistance is associated with the P. falciparum transporter ABCI3.

Emerging resistance to current antimalarials is reducing their effectiveness and therefore there is a need to develop new antimalarial therapies. Toward this goal, high throughput screens against the P. falciparum asexual parasite identified the pyrazolopyridine 4-carboxamide scaffold. Structure-activity relationship analysis of this chemotype defined that the N1-tert-butyl group and aliphatic foliage in the 3- and 6-positions were necessary for activity, while the inclusion of a 7'-aza-benzomorpholine on the 4-carboxamide motif resulted in potent anti-parasitic activity and increased aqueous solubility. A previous report that resistance to the pyrazolopyridine class is associated with the ABCI3 transporter was confirmed, with pyrazolopyridine 4-carboxamides showing an increase in potency against parasites when the ABCI3 transporter was knocked down. The low metabolic stability intrinsic to the pyrazolopyridine scaffold and the slow rate by which the compounds kill asexual parasites resulted in poor performance in a P. berghei asexual blood stage mouse model. Lowering the risk of resistance and mitigating the metabolic stability and cytochrome P450 inhibition will be challenges in the future development of the pyrazolopyrimidine antimalarial class.

Lactam Truncation Yields a Dihydroquinazolinone Scaffold with Potent Antimalarial Activity that Targets PfATP4.

The emergence of resistance against current antimalarial treatments has necessitated the need for the development of novel antimalarial chemotypes. Toward this goal, we recently optimised the antimalarial activity of the dihydroquinazolinone scaffold and showed it targeted PfATP4. Here, we deconstruct the lactam moiety of the tricyclic dihydroquinazolinone scaffold and investigate the structure-activity relationship of the truncated scaffold. It was shown that SAR between scaffolds was largely transferrable and generated analogues with potent asexual stage activity. Evaluation of the truncated analogues against PfATP4 mutant drug resistant parasite strains and in assays measuring PfATP4-associated ATPase activity demonstrated retention of PfATP4 as the molecular target. Analogues exhibited activity against both male and female gametes and multidrug resistant parasites. Limited efficacy of analogues in a P. berghei asexual stage mouse model was attributed to their moderate metabolic stability and low aqueous stability. Further development is required to address these attributes toward the potential use of the dihydroquinazolinone class in a curative and transmission blocking combination antimalarial therapy.

Property and Activity Refinement of Dihydroquinazolinone-3-carboxamides as Orally Efficacious Antimalarials that Target PfATP4.

To contribute to the global effort to develop new antimalarial therapies, we previously disclosed initial findings on the optimization of the dihydroquinazolinone-3-carboxamide class that targets PfATP4. Here we report on refining the aqueous solubility and metabolic stability to improve the pharmacokinetic profile and consequently in vivo efficacy. We show that the incorporation of heterocycle systems in the 8-position of the scaffold was found to provide the greatest attainable balance between parasite activity, aqueous solubility, and metabolic stability. Optimized analogs, including the frontrunner compound S-WJM992, were shown to inhibit PfATP4-associated Na+-ATPase activity, gave rise to a metabolic signature consistent with PfATP4 inhibition, and displayed altered activities against parasites with mutations in PfATP4. Finally, S-WJM992 showed appreciable efficacy in a malaria mouse model and blocked gamete development preventing transmission to mosquitoes. Importantly, further optimization of the dihydroquinazolinone class is required to deliver a candidate with improved pharmacokinetic and risk of resistance profiles.

Optimization of 2,3-Dihydroquinazolinone-3-carboxamides as Antimalarials Targeting PfATP4.

There is an urgent need to populate the antimalarial clinical portfolio with new candidates because of resistance against frontline antimalarials. To discover new antimalarial chemotypes, we performed a high-throughput screen of the Janssen Jumpstarter library against the Plasmodium falciparum asexual blood-stage parasite and identified the 2,3-dihydroquinazolinone-3-carboxamide scaffold. We defined the SAR and found that 8-substitution on the tricyclic ring system and 3-substitution of the exocyclic arene produced analogues with potent activity against asexual parasites equivalent to clinically used antimalarials. Resistance selection and profiling against drug-resistant parasite strains revealed that this antimalarial chemotype targets PfATP4. Dihydroquinazolinone analogues were shown to disrupt parasite Na+ homeostasis and affect parasite pH, exhibited a fast-to-moderate rate of asexual kill, and blocked gametogenesis, consistent with the phenotype of clinically used PfATP4 inhibitors. Finally, we observed that optimized frontrunner analogue WJM-921 demonstrates oral efficacy in a mouse model of malaria.

Synthesis and evaluation of human phosphodiesterases (PDE) 5 inhibitor analogs as trypanosomal PDE inhibitors. Part 1. Sildenafil analogs.

Parasitic diseases, such as African sleeping sickness, have a significant impact on the health and well-being in the poorest regions of the world. Pragmatic drug discovery efforts are needed to find new therapeutic agents. In this Letter we describe target repurposing efforts focused on trypanosomal phosphodiesterases. We outline the synthesis and biological evaluation of analogs of sildenafil (1), a human PDE5 inhibitor, for activities against trypanosomal PDEB1 (TbrPDEB1). We find that, while low potency analogs can be prepared, this chemical class is a sub-optimal starting point for further development of TbrPDE inhibitors.

Substrate Peptidomimetic Inhibitors of P. falciparum Plasmepsin X with Potent Antimalarial Activity.

Plasmepsin X (PMX) is an aspartyl protease that processes proteins essential for Plasmodium parasites to invade and egress from host erythrocytes during the symptomatic asexual stage of malaria. PMX substrates possess a conserved cleavage region denoted by the consensus motif, SFhE (h=hydrophobic amino acid). Peptidomimetics reflecting the P3 -P1 positions of the consensus motif were designed and showed potent and selective inhibition of PMX. It was established that PMX prefers Phe in the P1 position, di-substitution at the ß-carbon of the P2 moiety and a hydrophobic P3 group which was supported by modelling of the peptidomimetics in complex with PMX. The peptidomimetics were shown to arrest asexual P. falciparum parasites at the schizont stage by impairing PMX substrate processing. Overall, the peptidomimetics described will assist in further understanding PMX substrate specificity and have the potential to act as a template for future antimalarial design.

Direct Amidation to Access 3-Amido-1,8-Naphthalimides Including Fluorescent Scriptaid Analogues as HDAC Inhibitors.

Methodology to access fluorescent 3-amido-1,8-naphthalimides using direct Buchwald-Hartwig amidation is described. The protocol was successfully used to couple a number of substrates (including an alkylamide, an arylamide, a lactam and a carbamate) to 3-bromo-1,8-naphthalimide in good yield. To further exemplify the approach, a set of scriptaid analogues with amide substituents at the 3-position were prepared. The new compounds were more potent than scriptaid at a number of histone deacetylase (HDAC) isoforms including HDAC6. Activity was further confirmed in a whole cell tubulin deacetylation assay where the inhibitors were more active than the established HDAC6 selective inhibitor Tubastatin. The optical properties of these new, highly active, compounds make them amenable to cellular imaging studies and theranostic applications.

NMR studies of dextromethorphan in both isotropic and anisotropic states.

Herein, we present the solution-state NMR studies on dextromethorphan (1) under both isotropic and anisotropic conditions. From the measurement of 22 residual dipolar couplings using a stretched polystyrene gel (PS), we show that accurate and detailed structural information is readily determined including the relative stereochemical assignments of chiral centers, validation of diastereomer configuration, and the stereospecific assignment of the seven pairs of prochiral protons. This utility of PS gels is thus showcased to obtain rapid, accurate conformational, and relative configuration information in this important class of compounds without recourse to X-ray analysis.

Mixed alkoxy/hydroxy 1,8-naphthalimides: expanded fluorescence colour palette and in vitro bioactivity.

An efficient and functional group tolerant route to access hydroxy 1,8-naphthalimides has been used to synthesise a range of mono- and disubstituted hydroxy-1,8-naphthalimides with fluorescence emissions covering the visible spectrum. The dialkoxy substituted compounds prepared possess high quantum yields (up to 0.95) and long fluorescent lifetimes (up to 14 ns). The method has been used to generate scriptaid analogues that successfully inhibit HDAC6 in vitro with tubulin acetylation assays confirming that these compounds are more effective than tubastatin.

The Development Process for Discovery and Clinical Advancement of Modern Antimalarials.

Malaria is a devastating disease caused by Plasmodium parasites, resulting in approximately 435000 deaths in 2018. The impact of malaria is compounded by the emergence of widespread resistance to current antimalarial therapies. Recently, a new strategy was initiated to screen small molecule collections against the Plasmodium parasite enabling the identification of new antimalarial chemotypes with novel modes of action. This initiative ushered in the modern era of antimalarial drug development, and as a result, numerous lead candidates are advancing toward or are currently in human clinical trials. In this Perspective, we describe the development pathway of four of the most clinically advanced modern antimalarials, KAE609, KAF156, DSM265, and MMV048. Additionally, the mechanism of action and life-cycle stage specificity of the four antimalarials is discussed in relation to aligning with global strategies to treat and eliminate malaria. This perspective serves as a guide to the expectations of modern antimalarial drug development.