General alkyl fluoride functionalization via short-lived carbocation-organozincate ion pairs.

Fluorinated organic compounds are frequently used across the chemical and life sciences. Although a large, structurally diverse pool of alkyl fluorides is nowadays available, synthetic applications trail behind the widely accepted utility of other halides. We envisioned that C(sp2)-C(sp3) cross-coupling reactions of alkyl fluorides with fluorophilic organozinc compounds should be possible through a heterolytic mechanism that involves short-lived ion pairs and uses the stability of the Zn-F bond as the thermodynamic driving force. This would be mechanistically different from previously reported radical reactions and overcome long-standing limitations of organometallic cross-coupling methodology, including competing ß-hydride elimination, homodimerization and hydrodefluorination. Here, we show a practical Csp3-F bond functionalization method that expands the currently restricted synthetic space of unactivated primary, secondary and tertiary C(sp3)-F bonds but also uses benzylic, propargylic and acyl fluorides. Many functional groups and sterically demanding substrates are tolerated, which allows practical carbon-carbon bond formation and late-stage functionalization.

Design, synthesis and biological evaluation of Nucleosidic CD99 inhibitors that selectively reduce Ewing sarcoma viability.

Ewing Sarcoma (ES) is a cancer of bone and soft tissues affecting mostly children and young adults. Aggressive progression and poor prognosis of this malignancy call for novel and targeted treatments. CD99 is a transmembrane protein that is abundantly expressed on ES cells and is a diagnostic marker for the disease. ES cells are selectively sensitive to CD99 inhibition compared to most normal cells and other tumors. Therefore, CD99 is a good molecular target for ES treatment. Clofarabine and cladribine are two FDA approved drugs that are administered for their inhibitory acts on DNA synthesis to treat relapsed or refractory acute lymphoblastic and myeloid leukemia. They have also been shown to directly bind to CD99 and inhibit ES growth through a distinct mechanism. In the current study, we designed, synthesized and tested new ES specific derivatives of both drugs that would continue to target CD99 but with expected reduction in cellular membrane permeability and rendered unsuitable for inhibiting DNA synthesis. By using commercially available clofarabine and cladribine purine nucleoside analogs, we modified the primary alcohol moiety at the deoxyribose C-5' terminal site to suppress phosphorylation and thus inhibition of subsequent DNA synthesis pathways. In addition, we incorporated a variety of polar groups in the ribose and purine rings to reduce membrane permeability and investigated the effects of configurational changes in the sugar moiety. Among 26 new derivatives, we identified two compounds, BK50164 and BK60106, that cause cell death specifically in ES primarily due to inhibition of CD99 but not via inhibition of DNA synthesis. These findings provide a road map for the future development selective CD99 inhibitors for targeted treatment of ES.

Selective Csp3-F Bond Functionalization with Lithium Iodide.

A highly efficient method for C-F bond functionalization of a broad variety of activated and unactivated aliphatic substrates with inexpensive lithium iodide is presented. Primary, secondary, tertiary, benzylic, propargylic and α-functionalized alkyl fluorides react in chlorinated or aromatic solvents at room temperature or upon heating to the corresponding iodides which are isolated in 91-99% yield. The reaction is selective for aliphatic monofluorides and can be coupled with in situ nucleophilic iodide replacements to install carbon-carbon, carbon-nitrogen and carbon-sulfur bonds with high yields. Alkyl difluorides, trifluorides, even in activated benzylic positions, are inert under the same conditions and aryl fluoride bonds are also tolerated.

Organocuprate Cross-Coupling Reactions with Alkyl Fluorides.

Cross-coupling of alkyl fluorides and organocuprates is accomplished via aluminum halide mediated C-F bond activation and subsequent Csp2-Csp3 and Csp3-Csp3 bond formation. Relatively mild conditions allow for smooth activation of notoriously challenging primary and secondary alkyl fluorides while competing alkyl chain rearrangement, HF elimination, and homocoupling reactions are effectively controlled. The utility and functional group tolerance are demonstrated with 23 examples and a variety of coupling products obtained in up to 88% yield.

Inhibition of Ubiquitin-Specific Protease-13 Improves Behavioral Performance in Alpha-Synuclein Expressing Mice.

Ubiquitin-Specific Protease-13 (USP13) promotes protein de-ubiquitination. USP13 levels are upregulated in post-mortem Parkinson's disease, whereas USP13 knockdown via shRNA reduces alpha-synuclein levels in animal models. We studied the role of USP13 in knockout mice expressing lentiviral human alpha-synuclein and investigated the impact of a small molecule inhibitor of USP13, BK50118-C, on alpha-synuclein pathology and animal behavior. Alpha-synuclein was expressed unilaterally in substantia nigra (SN) of USP13 deficient mice that were treated with a daily intraperitoneal injection of 100 mg/kg BK50118-C or DMSO for four consecutive weeks, and behavioral and functional assays were performed. Wild-type USP13+/+ mice expressing lentiviral human alpha-synuclein showed motor and behavioral defects that were not seen in partially (USP13+/-) or completely (USP13-/-) deficient USP13 mice. BK50118-C displayed a wide and favorable therapeutic dose range in vivo. Treatment with BK50118-C significantly reduced ubiquitinated alpha-synuclein, increased dopamine levels, and improved motor and behavioral symptoms in wild-type (USP13+/+), but not USP13 deficient, mice. These data suggest that USP13 is critical to the neuropathology of alpha-synuclein, whereas a novel small molecule inhibitor of USP13 is a potential therapeutic agent of alpha-synucleinopathies.

Chemodivergent Csp3─F bond functionalization and cross-electrophile alkyl-alkyl coupling with alkyl fluorides.

The widespread use of fluorinated organic compounds in the health, agrochemical, and materials sciences is sustained by a steadily growing pool of commercially available fine chemicals. The synthetic utility of the increasingly ubiquitous Csp3─F bond, however, remains to be fully exploited, which is often a difficult task because of its paramount stability and chemical inertness. Here, we demonstrate chemodivergent activation of monofluoroalkyl compounds toward either nucleophilic or electrophilic intermediates. This is accomplished under conditions that are compatible with several reaction types and many functional groups, which drastically widens the current scope of organofluorine chemistry and sets the stage for carbon-carbon and carbon-heteroatom bond formations, stereoselective construction of bisoxindole alkaloid scaffolds via in situ Umpolung, and cross-electrophilic coupling methodology. The selective generation of either nucleophilic or electrophilic species and the possibility of doing so simultaneously or, alternatively, switching molecular polarity enable previously unidentified synthetic opportunities that recognize alkyl fluorides as chemodivergent building blocks.

The Potential of 2-Substituted Quinolines as Antileishmanial Drug Candidates.

There is a need for new, cost-effective drugs to treat leishmaniasis. A strategy based on traditional medicine practiced in Bolivia led to the discovery of the 2-substituted quinoline series as a source of molecules with antileishmanial activity and low toxicity. This review documents the development of the series from the first isolated natural compounds through several hundred synthetized molecules to an optimized compound exhibiting an in vitro IC50 value of 0.2 µM against Leishmania donovani, and a selectivity index value of 187, together with in vivo activity on the L. donovani/hamster model. Attempts to establish structure-activity relationships are described, as well as studies that have attempted to determine the mechanism of action. For the latter, it appears that molecules of this series act on multiple targets, possibly including the immune system, which could explain the observed lack of drug resistance after in vitro drug pressure. We also show how nanotechnology strategies could valorize these drugs through adapted formulations and how a mechanistic targeting approach could generate new compounds with increased activity.

Palladium and Nickel Catalyzed Suzuki Cross-Coupling with Alkyl Fluorides.

Suzuki cross-coupling of benzylic and unactivated aliphatic fluorides with aryl- and alkenylboronic acids has been achieved via mechanistically distinct Pd and Ni catalyzed pathways that outperform competing protodeboronation, ß-hydride elimination, and homocoupling processes. The utility is demonstrated with more than 20 examples including heterocyclic structures, 1,1-disubstituted and trans-1,2-disubstituted alkenes, and by the incorporation of acetonitrile into functionalized (hetero)arenes.

Novel Ubiquitin Specific Protease-13 Inhibitors Alleviate Neurodegenerative Pathology.

Ubiquitin Specific Protease-13 (USP13) promotes protein de-ubiquitination and is poorly understood in neurodegeneration. USP13 is upregulated in Alzheimer's disease (AD) and Parkinson's disease (PD), and USP13 knockdown via shRNA reduces neurotoxic proteins and increases proteasome activity in models of neurodegeneration. We synthesized novel analogues of spautin-1 which is a non-specific USP13 inhibitor but unable to penetrate the brain. Our synthesized small molecule compounds are able to enter the brain, more potently inhibit USP13, and significantly reduce alpha-synuclein levels in vivo and in vitro. USP13 inhibition in transgenic mutant alpha-synuclein (A53T) mice increased the ubiquitination of alpha-synuclein and reduced its protein levels. The data suggest that novel USP13 inhibitors improve neurodegenerative pathology via antagonism of de-ubiquitination, thus alleviating neurotoxic protein burden in neurodegenerative diseases.

Discoidin Domain Receptor 1 is a therapeutic target for neurodegenerative diseases.

The role of Discoidin Domain Receptors (DDRs) is poorly understood in neurodegeneration. DDRs are upregulated in Alzheimer's and Parkinson's disease (PD), and DDRs knockdown reduces neurotoxic protein levels. Here we show that potent and preferential DDR1 inhibitors reduce neurotoxic protein levels in vitro and in vivo. Partial or complete deletion or inhibition of DDR1 in a mouse model challenged with α-synuclein increases autophagy and reduces inflammation and neurotoxic proteins. Significant changes of cerebrospinal fluid microRNAs that control inflammation, neuronal injury, autophagy and vesicular transport genes are observed in PD with and without dementia and Lewy body dementia, but these changes are attenuated or reversed after treatment with the DDR1 inhibitor, nilotinib. Collectively, these data demonstrate that DDR1 regulates autophagy and reduces neurotoxic proteins and inflammation and is a therapeutic target in neurodegeneration.

Quantitative Chirality and Concentration Sensing of Alcohols, Diols, Hydroxy Acids, Amines and Amino Alcohols using Chlorophosphite Sensors in a Relay Assay.

Analytical methods that allow simultaneous determination of the concentration and enantiomeric composition of small sample amounts and are also compatible with high-throughput multi-well plate technology have received increasing attention in recent years. We now introduce a new class of broadly useful small-molecule probes and a relay sensing strategy that together accomplish these tasks with five classes of compounds including the challenging group of mono-alcohols-a scope that stands out among previously reported UV, fluorescence, and CD assays. Several chlorophosphite probes and aniline indicators have been evaluated and used for on-the-fly CD/UV sensing following a continuous workflow. The wide application range of the readily available sensors is highlighted with almost 30 alcohols, diols, hydroxy acids, amines and amino alcohols, and the accuracy of the stereochemical analysis is showcased with samples covering a wide range of concentrations and enantiomeric ratios.

Catalytic Asymmetric Allylic Amination with Isatins, Sulfonamides, Imides, Amines, and N-Heterocycles.

A generally useful palladium-catalyzed method for the asymmetric allylic amination with a large variety of isatins, sulfonamides, imides, amines, and N-heterocycles is introduced. A single protocol with a readily available catalyst accomplishes this reaction at room temperature with high yields and enantioselectivities often exceeding 90%, which is demonstrated with 31 examples.

Streamlined Asymmetric Reaction Development: A Case Study with Isatins.

Asymmetric reaction development within a day or two has been a dream of synthetic chemists for several decades. We now show that such a task is feasible with a highly efficient streamlined screening strategy using the asymmetric allylation of isatins with a chiral boron complex as a case study. Our high-throughput screening (HTS) method is based on fast optical UV/CD analysis of minute amounts of crude reaction mixtures (≈3 mg scale) and it obviates product isolation and the general need for reference compounds which greatly reduces preliminary work and analysis time. The setup, reaction screening, analysis and data processing for 54 asymmetric allylations of nine different isatins in six different solvents was handled by a single operator in less than 20 work hours. One could easily extend this HTS strategy to hundreds of reactions in roughly the same time frame and further reduce the labor with commercially available automated high-throughput experimentation equipment. The effectiveness of this asymmetric reaction development strategy is confirmed with the upscale synthesis of two representative 3-allyl-3-hydroxyisatins in 98-99 % yield and with 91-94 % ee under optimized conditions.

Click chemistry enables quantitative chiroptical sensing of chiral compounds in protic media and complex mixtures.

Click reactions have become powerful synthetic tools with unique applications in the health and materials sciences. Despite the progress with optical sensors that exploit the principles of dynamic covalent chemistry, metal coordination or supramolecular assemblies, quantitative analysis of complex mixtures remains challenging. Herein, we report the use of a readily available coumarin conjugate acceptor for chiroptical click chirality sensing of the absolute configuration, concentration and enantiomeric excess of several compound classes. This method has several attractive features, including wide scope, fast substrate fixation without by-product formation or complicate equilibria often encountered in reversible substrate binding, excellent solvent compatibility, and tolerance of air and water. The ruggedness and practicality of this approach are demonstrated by comprehensive analysis of nonracemic monoamine samples and crude asymmetric imine hydrogenation mixtures without work-up. Click chemosensing addresses increasingly important time efficiency, cost, labor and chemical sustainability aspects and streamlines asymmetric reaction development at the mg scale.

In-vitro and in-vivo antileishmanial activity of inexpensive Amphotericin B formulations: Heated Amphotericin B and Amphotericin B-loaded microemulsion.

Amphotericin B (AmB) is effective against visceral leishmaniasis (VL), but the renal toxicity of the conventional form, mixed micelles with deoxycholate (M-AmB), is often dose-limiting, while the less toxic lipid-based formulations such as AmBisome® are very expensive. Two different strategies to improve the therapeutic index of AmB with inexpensive ingredients were evaluated on this work: (i) the heat treatment of the commercial formulation (H-AmB) and (ii) the preparation of an AmB-loaded microemulsion (ME-AmB). M-AmB was heated to 70 °C for 20 min. The resulting product was characterized by UV spectrophotometry and circular dichroism, showing super-aggregates formation. ME-AmB was prepared from phosphate buffer pH 7.4, Tween 80®, Lipoid S100® and Mygliol 812® with AmB at 5 mg/mL. The droplet size, measured by dynamic light scattering, was about 40 nm and transmission electron microscopy confirmed a spherical shape. Rheological analysis showed low viscosity and Newtonian behavior. All the formulations were active in vitro and in vivo against Leishmania donovani (LV9). A selectivity index (CC50 on RAW/IC50 on LV9) higher than 10 was observed for ME-AmB, H-AmB and AmBisome®. Furthermore, no important in vivo toxicity was observed for all the samples. The in-vivo efficacy of the formulations after IV administration was evaluated in Balb/C mice infected with LV9 (three doses of 1 mg/kg AmB) and no significant difference was observed between H-AmB, M-AmB, ME-AmB and AmBisome®. In conclusion, these two inexpensive alternative formulations for AmB showing good efficacy and selectivity for Leishmania donovani merit further investigation.

Organocatalytic Stereoselective Synthesis of Fluorinated 3,3'-Linked Bisoxindoles.

A highly diastereoselective organocatalytic method that produces 3-fluoro-3'-hydroxy-3,3'-bisoxindoles and the corresponding 3-fluoro-3'-amino derivatives having two adjacent chirality centers from fluorooxindoles and isatins in high yields is described. The reaction occurs in protic solvents at room temperature, it can be upscaled without compromising yield and stereoselectivity, and chromatographic product purification is not required.

Organocatalytic Decarboxylative Cyanomethylation of Difluoromethyl and Trifluoromethyl Ketones.

An efficient organocatalytic method for the synthesis of difluoromethyl and trifluoromethyl substituted ß-hydroxynitriles is introduced. The decarboxylative cyanomethylation of fluorinated ketones with readily available cyanoacetic acid gives a variety of tertiary alcohols in high yields and without concomitant water elimination. The reaction occurs in the presence of catalytic amounts of triethylamine, can be upscaled and applied to chlorofluoromethyl ketones and difluoromethyl ketimines.

Stereoselective Synthesis of 3,3'-Bisindolines by Organocatalytic Michael Additions of Fluorooxindole Enolates to Isatylidene Malononitriles in Aqueous Solution.

A highly diastereoselective organocatalytic reaction for the synthesis of fluorinated 3,3'-bisindolines exhibiting adjacent tetrasubstituted carbon stereocenters is described. A broad variety of heterochiral bisindolines was prepared in 91-99% yield using 3-fluorooxindoles and isatylidene malononitriles in the presence of catalytic amounts of triethylamine in water or aqueous solution. The reaction can be upscaled without compromising yield and diastereoselectivity and the general usefulness of this method was demonstrated with various Michael acceptors and extended to aldol and Mannich reactions.

Catalytic Enantioselective and Diastereoselective Allylic Alkylation with Fluoroenolates: Efficient Access to C3-Fluorinated and All-Carbon Quaternary Oxindoles.

Synthetically versatile 3,3-disubstituted fluorooxindoles exhibiting vicinal chirality centers were obtained in high yields and with excellent enantio-, diastereo-, and regioselectivity through catalytic asymmetric fluoroenolate alkylation with allylic acetates. The reaction proceeds under mild conditions and can be scaled up without compromising the asymmetric induction. The unique synthetic usefulness of the products is highlighted by the incorporation of additional functionalities and the formation of 3-fluorinated oxindoles exhibiting an array of four adjacent centers of chirality. A new C-F bond functionalization path that provides unprecedented possibilities for the stereoselective generation of a chiral quaternary carbon center in the alkaloid scaffold is introduced.