The Rise of Boron-Containing Compounds: Advancements in Synthesis, Medicinal Chemistry, and Emerging Pharmacology.

Boron-containing compounds (BCC) have emerged as important pharmacophores. To date, five BCC drugs (including boronic acids and boroles) have been approved by the FDA for the treatment of cancer, infections, and atopic dermatitis, while some natural BCC are included in dietary supplements. Boron's Lewis acidity facilitates a mechanism of action via formation of reversible covalent bonds within the active site of target proteins. Boron has also been employed in the development of fluorophores, such as BODIPY for imaging, and in carboranes that are potential neutron capture therapy agents as well as novel agents in diagnostics and therapy. The utility of natural and synthetic BCC has become multifaceted, and the breadth of their applications continues to expand. This review covers the many uses and targets of boron in medicinal chemistry.

A Potential Boron Neutron Capture Therapy Agent Selectively Suppresses High-Grade Glioma: In Vitro and in Vivo Exploration.

Glioblastoma (GBM), as the most central nervous system (CNS) intractable disease, has spoiled millions of lives due to its high mortality. Even though several efforts have been made, the existing treatments have had limited success. In this sense, we studied a lead compound, the boron-rich selective epidermal growth factor receptor (EGFR)-inhibitor hybrid 1, as a potential drug for GBM treatment. For this end, we analyzed the in vitro activity of hybrid 1 in a glioma/primary astrocytes coculture, studying cellular death types triggered by treatment with this compound and its cellular localizations. Additionally, hybrid 1 concentrated boron in glioma cells selectively and more effectively than the boron neutron capture therapy (BNCT)-clinical agent 10B-l-boronophenylalanine and thus displayed a better in vitro-BNCT effect. This encouraged us to analyze hybrid 1 in vivo. Therefore, immunosuppressed mice bearing U87 MG human GBM were treated with both 1 and 1 encapsulated in a modified liposome (recognized by brain-blood barrier peptide transporters), and we observed a potent in vivo per se antitumor activity (tumor size decrease and animal survival increase). These data demonstrate that 1 could be a promising new targeted therapy for GBM.

Tautomerism and Rotamerism of Favipiravir and Halogenated Analogues in Solution and in the Solid State.

Favipiravir is an important selective antiviral against RNA-based viruses, and currently, it is being repurposed as a potential drug for the treatment of COVID-19. This type of chemical system presents different carboxamide-rotameric and hydroxyl-tautomeric states, which could be essential for interpreting its selective antiviral activity. Herein, the tautomeric 3-hydroxypyrazine/3-pyrazinone pair of favipiravir and its 6-substituted analogues, 6-Cl, 6-Br, 6-I, and 6-H, were fully investigated in solution and in the solid state through ultraviolet-visible, 1H nuclear magnetic resonance, infrared spectroscopy, and X-ray diffraction techniques. Also, a study of the gas phase was performed using density functional theory calculations. In general, the keto-enol balance in these 3-hydroxy-2-pyrazinecarboxamides is finely modulated by external and internal electrical variations via changes in solvent polarity or by replacement of substituents at position 6. The enol tautomer was prevalent in an apolar environment, whereas an increase in the level of the keto tautomer was favored by an increase in solvent polarity and, even moreso, with a strong hydrogen-donor solvent. Keto tautomerization was favored either in solution or in the solid state with a decrease in 6-substituent electronegativity as follows: H ≫ I ≈ Br > Cl ≥ F. Specific rotameric states based on carboxamide, "cisoide" and "transoide", were identified for the enol and keto tautomer, respectively; their rotamerism is dependent on the tautomerism and not the aggregation state.

One-step synthesis of favipiravir from Selectfluor® and 3-hydroxy-2-pyrazinecarboxamide in an ionic liquid.

Favipiravir is an important selective antiviral that emerged as an alternative against COVID-19 during the pandemic. Its synthesis has gained great interest and the conventional strategies proceed through multiple-step protocols (6-7 reaction steps), which involve, in addition, several drawbacks with global yields, lower than 34%. Herein, a simple, economical, eco-friendly and scalable (1 g) one-step protocol for the synthesis of favipiravir from the direct fluorination of the available 3-hydroxy-2-pyrazinecarboxamide with Selectfluor® is reported. The reaction proceeds easily in BF4-BMIM through a simple operational work-up, affording the favipiravir with a yield of 50% without the need of a special catalyst/additive. The key point of the present strategy was the use of the ionic liquid of BF4-BMIM, which helps to minimize the several chemical limitations derived from 3-hydroxy-2-pyrazinecarboxamide as a substrate for the direct Selectfluor-mediated fluorination. All these chemical reactivity aspects are also discussed in detail.

Exploring binding chemistry of alkali/alkaline earth cations in solution through modulation of intramolecular charge-transfer in an excited ambidentate organic fluorophore.

Studying the metal-ligand monoligation of alkali/alkaline earth metals (AMs) in solution represents a significant challenge due to the low stabilization of their complexes and the absence of an effective strategy to identify this type of weak binding. Herein, we show that the modulation of the intramolecular charge-transfer (ICT) in an excited ambidentate organic fluorophore is a convenient strategy to characterize the binding chemistry of AM cations in solution through simple steady-state fluorescence and fluorescence lifetime measurements. The key points of the fluorophore as a metal-binding probe were the location of diverse coordination functionalities with different binding abilities (ionic-, pseudo-covalent- and non-covalent-probes) along the donor-acceptor (D-A) chain and the occurrence of an intramolecular charge-transfer (ICT) mechanism upon excitation. The binding of these functionalities with AM-cations generated selective and specific fluorescence responses, which were quantifiable and allowed us to recognize the primary, secondary and tertiary interactions for all the AM cations in the solution. The relative binding affinities for each one of the functionalities with AM cations was estimated, and a general and consistent perspective of the binding of AMs as a function of their location in the Periodic Table, hardness property and ionic radius was established. The binding preferences of the AM cations were supported by DFT calculations. Our strategy allowed us to validate the binding dynamics of AMs in solution for three types of key ligations, which opens a new perspective to recognize weak intermolecular interactions derived from acidic species and rationally design selective AM-cation probes using an ICT-based ambidentate organic fluorophore.

In vitro and in vivo studies on a group of chalcones find promising results as potential drugs against fascioliasis.

About a third of the world population is infected by helminth parasites implicated in foodborne trematodiasis. Fascioliasis is a worldwide disease caused by trematodes of the genus Fasciola spp. It generates huge economic losses to the agri-food industry and is currently considered an emerging zoonosis by the World Health Organization (WHO). The only available treatment relies on anthelmintic drugs, being triclabendazole (TCBZ) the drug of choice to control human infections. The emergence of TCBZ resistance in several countries and the lack of an effective vaccine to prevent infection highlights the need to develop new drugs to control this parasitosis. We have previously identified a group of benzochalcones as inhibitors of cathepsins, which have fasciolicidal activity in vitro and are potential new drugs for the control of fascioliasis. We selected the four most active compounds of this group to perform further preclinical studies. The compound's stability was determined against a liver microsomal enzyme fraction, obtaining half-lives of 34-169 min and low intrinsic clearance values (<13 µL/min/mg), as desirable for potential new drugs. None of the compounds were mutagenic or genotoxic and no in vitro cytotoxic effects were seen. Compounds C31 and C34 showed the highest selectivity index against liver fluke cathepsins when compared to human cathepsin L. They were selected for in vivo efficacy studies observing a protective effect, similar to TCBZ, in a mouse model of infection. Our findings strongly encourage us to continue the drug development pipeline for these molecules.

High CT-Fluorophore Featuring a Basic Moiety into D-A Chain as a pKa Probe.

The determination of acidity represents a significant challenge within fluorometry, and no effective strategy has been developed successfully yet. It is attributed to the fact that acidity tends to be enhanced upon excitation, giving, in general, an overestimation of the ionization constant, pKa. Herein, we developed a strategy for pKa estimation of Brønsted acids in solution through fluorometry by using a convenient pKa probe, N1-aryl-7-methoxy-2-(trifluoromethyl)benzo[b][1,8]naphthyridin-4(1H)-one. It allowed us to obtain a linear log KSV versus pKa correlation derived from the selective quenching response of the probe by an interaction with different Brønsted acids. The key points of N1-aryl-7-methoxy-2-(trifluoromethyl)benzo[b][1,8]naphthyridin-4(1H)-one as a pKa probe were (i) the location of a weak basic moiety in the donor-acceptor chain of the fluorophore, which favors a selective quenching of the intramolecular charge-transfer process according to the acidity of acid, and (ii) the high CT character upon excitation that promotes higher quenching magnitudes and favors a wider pKa range (19.5pKa) for the log KSV versus pKa correlation. Other key principles were to delimit the study to pure proton transfer and nonfluorescent acids, which allowed restricting the quenching response to a process dependent mainly on the acid-base equilibrium. All these findings open a new perspective as a proof of concept to design effective fluorescent pKa probes.

Bimodal Therapeutic Agents Against Glioblastoma, One of the Most Lethal Forms of Cancer.

About 95 % of people diagnosed with glioblastoma die within five years. Glioblastoma is the most aggressive central nervous system tumour. It is necessary to make progress in the glioblastoma treatment so that advanced chemotherapy drugs or radiation therapy or, ideally, two-in-one hybrid systems should be implemented. Tyrosine kinase receptors-inhibitors and boron neutron capture therapy (BNCT), together, could provide a therapeutic strategy. In this work, sunitinib decorated-carborane hybrids were prepared and biologically evaluated identifying excellent antitumoral- and BNCT-agents. One of the selected hybrids was studied against glioma-cells and found to be 4 times more cytotoxic than sunitinib and 1.7 times more effective than 10 B-boronophenylalanine fructose complex when the cells were irradiated with neutrons.

Identification and characterization of human interferon alpha inhibitors through a WISH cell line-based reporter gene assay.

Interferons (IFNs) are important glycoproteins which can stimulate or inhibit up to three hundred different genes encoding proteins involved in antiviral defense mechanisms, inflammation, adaptive immunity, angiogenesis and among other processes. Nevertheless, different genetic alterations may lead to interferon alpha (IFN-α) overproduction in human autoimmune diseases like systemic lupus erythematosus. As a consequence, IFN-α is a central molecule whose activity must be regulated to block their harmful effect on those disorders where the endogenous cytokine production constitutes the etiology of the illnesses. In this work, we evaluate the biological activity of eighty-eight compounds, from our own chemo-library, to find potential IFN-α inhibitors by using a reporter gene assay (RGA) WISH-Mx2/EGFP. We identified some compounds able to modulate negatively the IFN-α activity. The most active IFN-α inhibitors were further studied achieving promising results. In addition, some combinations of the most active compounds were analyzed accomplishing a stronger effect to decrease the IFN-α activity than each compound alone. Furthermore, the complete inhibition of the cytokine activity was reached with some combinations of compounds.

New aryloxy-quinone derivatives with promising activity on Trypanosoma cruzi.

Continuing with a program to develop new quinone derivatives as biologically active compounds, we designed and synthesized a new series of aryloxy-quinones, which were evaluated in vitro against Trypanosoma cruzi in epimastigote form. Chemical modifications in three specific moieties on the aryloxy-quinone core were considered for developing new anti-T. cruzi agents. The majority of our new quinones showed higher potency (IC50 values of <0.70 µM) than nifurtimox, a known pharmaceutical used as a baseline drug (IC50 values of 7.00 µM); however, only two of them elicited higher selectivity than nifurtimox against Vero cells. A structure-activity relationship analysis provided information about the stereoelectronic features of these compounds, which are responsible for an increase in trypanosomicidal activity. Using a pharmacophore model, we mapped the substitution patterns of the five pharmacophoric features of trypanosomicidal activity. We chose the Epc1 compounds and found no relationship with the trypanosomicidal effects. These results provided useful information about the structural characteristics for developing new aryloxy-quinones with higher potency against the protozoan parasite T. cruzi.

Cathepsin L Inhibitors with Activity against the Liver Fluke Identified From a Focus Library of Quinoxaline 1,4-di-N-Oxide Derivatives.

Infections caused by Fasciola species are widely distributed in cattle and sheep causing significant economic losses, and are emerging as human zoonosis with increasing reports of human cases, especially in children in endemic areas. The current treatment is chemotherapeutic, triclabendazole being the drug of preference since it is active against all parasite stages. Due to the emergence of resistance in several countries, the discovery of new chemical entities with fasciolicidal activity is urgently needed. In our continuous search for new fasciolicide compounds, we identified and characterized six quinoxaline 1,4-di-N-oxide derivatives from our in-house library. We selected them from a screening of novel inhibitors against FhCL1 and FhCL3 proteases, two essential enzymes secreted by juvenile and adult flukes. We report compounds C7, C17, C18, C19, C23, and C24 with an IC50 of less than 10 µM in at least one cathepsin. We studied their binding kinetics in vitro and their enzyme-ligand interactions in silico by molecular docking and molecular dynamic (MD) simulations. These compounds readily kill newly excysted juveniles in vitro and have low cytotoxicity in a Hep-G2 cell line and bovine spermatozoa. Our findings are valuable for the development of new chemotherapeutic approaches against fascioliasis, and other pathologies involving cysteine proteases.

Discovery of Potent EGFR Inhibitors through the Incorporation of a 3D-Aromatic-Boron-Rich-Cluster into the 4-Anilinoquinazoline Scaffold: Potential Drugs for Glioma Treatment.

New 1,7-closo-carboranylanilinoquinazoline hybrids have been identified as EGFR inhibitors, one of them with higher affinity than the parent compound erlotinib. The comparative docking analysis with compounds bearing bioisoster-substructures, demonstrated the relevance of the 3D aromatic-boron-rich moiety for interacting into the EGFR ATP binding region. The capability to accumulate in glioma cells, the ability to cross the blood-brain barrier and the stability on simulated biological conditions, render these molecules as lead compounds for further structural modifications to obtain dual action drugs to treat glioblastoma.

Synthesis of hydrophilic HYNIC-[1,2,4,5]tetrazine conjugates and their use in antibody pretargeting with 99mTc.

Pretargeted imaging, based on the highly reactive process between [1,2,4,5]tetrazines with trans-cyclooctene (TCO), appears as an attractive strategy to overcome disadvantages associated with traditional radioimmunoconjugates. To be successful, the radiolabeled component should react in vivo with the conjugated antibody and the non reactive excess clear fast from the organism. Herein, we explore the in vivo effects of hydrophilic linker incorporation into [1,2,4,5]tetrazine systems bearing a 6-hydrazinonicotinyl (HYNIC) moiety for technetium-99m coordination. Incorporation of a polypeptide chain containing hydrophilic aminoacids, resulted in a derivative with renal clearance. Pretargeted bevacizumab imaging was used as proof of concept.

Looking for combination of benznidazole and Trypanosoma cruzi-triosephosphate isomerase inhibitors for Chagas disease treatment.

BACKGROUND: The current chemotherapy for Chagas disease is based on monopharmacology with low efficacy and drug tolerance. Polypharmacology is one of the strategies to overcome these limitations. OBJECTIVES: Study the anti-Trypanosoma cruzi activity of associations of benznidazole (Bnz) with three new synthetic T. cruzi-triosephosphate isomerase inhibitors, 2, 3, and 4, in order to potentiate their actions. METHODS: The in vitro effect of the drug combinations were determined constructing the corresponding isobolograms. In vivo activities were assessed using an acute murine model of Chagas disease evaluating parasitaemias, mortalities and IgG anti-T. cruzi antibodies. FINDINGS: The effect of Bnz combined with each of these compounds, on the growth of epimastigotes, indicated an additive action or a synergic action, when combining it with 2 or 3, respectively, and an antagonic action when combining it with 4. In vivo studies, for the two chosen combinations, 2 or 3 plus one fifth equivalent of Bnz, showed that Bnz can also potentiate the in vivo therapeutic effects. For both combinations a decrease in the number of trypomastigote and lower levels of anti-T. cruzi IgG-antibodies were detected, as well clear protection against death. MAIN CONCLUSIONS: These results suggest the studied combinations could be used in the treatment of Chagas disease.

Small-Molecule Kinase-Inhibitors-Loaded Boron Cluster as Hybrid Agents for Glioma-Cell-Targeting Therapy.

The reported new anilinoquinazoline-icosahedral borane hybrids have been evaluated as glioma targeting for potential use in cancer therapy. Their anti-glioma activity depends on hybrids' lipophilicity; the most powerful compound against glioma cells, a 1,7-closo-derivative, displayed at least 3.3 times higher activity than the parent drug erlotinib. According to the cytotoxic effects on normal glia cells, the hybrids were selective for epidermal growth factor receptor (EGFR)-overexpressed tumor cells. These boron carriers could be used to enrich glioma cancer cells with boron for cancer therapy.

Development of new PTK7-targeting aptamer-fluorescent and -radiolabelled probes for evaluation as molecular imaging agents: Lymphoma and melanoma in vivo proof of concept.

Aptamers are single-stranded oligonucleotides that recognize molecular targets with high affinity and specificity. Aptamer that selectively bind to the protein tyrosine kinase-7 (PTK7) receptor, overexpressed on many cancers, has been labelled as probes for molecular imaging of cancer. Two new PTK7-targeting aptamer probes were developed by coupling frameworks from the fluorescent dye AlexaFluor647 or the 6-hydrazinonicotinamide (HYNIC) chelator-labelled to 99mTc. The derivatizations via a 5'-aminohexyl terminal linker were done at room temperature and under mild buffer conditions. Physicochemical and biological controls for both imaging agents were performed verifying the integrity of the aptamer-conjugates by HPLC. Recognition of melanoma (B16F1) and lymphoma (A20) mouse cell lines by the aptamer was studied using cell binding, flow cytometry and confocal microscopy. Finally, in vivo imaging studies in tumour-bearing mice were performed. The new probes were able to bind to melanoma and lymphoma cell lines in vitro, the in vivo imaging in tumour-bearing mice showed different uptake behaviours showing for the fluorescent conjugate good uptake by B cell lymphoma while the radiolabelled conjugate did not display tumour uptake due to its high extravascular distribution, and both showed rapid clearance properties in tumour-bearing mice.

Multi-Anti-Parasitic Activity of Arylidene Ketones and Thiazolidene Hydrazines against Trypanosoma cruzi and Leishmania spp.

A series of fifty arylideneketones and thiazolidenehydrazines was evaluated against Leishmania infantum and Leishmania braziliensis. Furthermore, new simplified thiazolidenehydrazine derivatives were evaluated against Trypanosoma cruzi. The cytotoxicity of the active compounds on non-infected fibroblasts or macrophages was established in vitro to evaluate the selectivity of their anti-parasitic effects. Seven thiazolidenehydrazine derivatives and ten arylideneketones had good activity against the three parasites. The IC50 values for T. cruzi and Leishmania spp. ranged from 90 nM-25 µM. Eight compounds had multi-trypanocidal activity against T. cruzi and Leishmania spp. (the etiological agents of cutaneous and visceral forms). The selectivity of these active compounds was better than the three reference drugs: benznidazole, glucantime and miltefosine. They also had low toxicity when tested in vivo on zebrafish. Trying to understand the mechanism of action of these compounds, two possible molecular targets were investigated: triosephosphate isomerase and cruzipain. We also used a molecular stripping approach to elucidate the minimal structural requirements for their anti-T. cruzi activity.

Synthesis and biological evaluation of quinoxaline di-N-oxide derivatives with in vitro trypanocidal activity.

We report the synthesis and in vitro activity against Trypanosoma cruzi epimastigotes of 15 novel quinoxaline derivatives. Ten of the derivatives presented IC50 values lower than the reference drugs Nfx and Bzn; four of them standed out with IC50 values lower than 1.5 µM. Moreover, unspecific cytotoxicity and genotoxicity studies are also reported. Compound 14 showed a SI higher than 24, whereas compound 10 was the only one that was negative in the genotoxicity screening.

In vivo phase II-enzymes inducers, as potential chemopreventive agents, based on the chalcone and furoxan skeletons.

Cancer chemoprevention involves prevention/delay/reverse of the carcinogenic process through administration of cancer chemopreventive agents (CCA). Compounds which are able to induce detoxification-enzymes, especially monofunctional phase II enzymes, have become in excellent approaches for new CCA. Herein, we report the synthesis of new furoxanyl chalcone-like hybrid compounds as CCA. In vitro studies showed that phenylfuroxanyl derivatives 6 and 9 displayed the best activities being 9 the greatest monofunctional-inducer. Additionally, compounds were non-mutagenic against TA98 Salmonella typhimurium strain (Ames test) and could be used in the prevention of the progression of pre-malignant lesions for their cytotoxic activity against tumoral cells. In vivo proof of concept showed increment on phase II-enzymes activities in liver, colon and mammary gland having derivative 9 the best induction profiles. We probed Nrf2 nuclear translocation is operative for both compounds allowing to exert protective effects via expression of downstream phase-II enzymes.