Antascomicin B stabilizes FKBP51-Akt1 complexes as a molecular glue.

Antascomicin B is a natural product that similarly to the macrolides FK506 and Rapamycin binds to the FK506-binding protein 12 (FKBP12). FK506 and Rapamycin act as molecular glues by inducing ternary complexes between FKBPs and additional target proteins. Whether Antascomicin B can induce ternary complexes is unknown. Here we show that Antascomicin B binds tightly to larger human FKBP homologs. The cocrystal structure of FKBP51 in complex with Antascomicin B revealed that large parts of Antascomicin B are solvent-exposed and available to engage additional proteins. Cellular studies demonstrated that Antascomicin B enhances the interaction between human FKBP51 and human Akt. Our studies show that molecules with molecular glue-like properties are more prominent in nature than previously thought. We predict the existence of additional 'orphan' molecular glues that evolved to induce ternary protein complexes but where the relevant ternary complex partners are unknown.

Continuous flow synthesis enabling reaction discovery.

This article defines the role that continuous flow chemistry can have in new reaction discovery, thereby creating molecular assembly opportunities beyond our current capabilities. Most notably the focus is based upon photochemical, electrochemical and temperature sensitive processes where continuous flow methods and machine assisted processing can have significant impact on chemical reactivity patterns. These flow chemical platforms are ideally placed to exploit future innovation in data acquisition, feed-back and control through artificial intelligence (AI) and machine learning (ML) techniques.

Nitro-sulfinate Reductive Coupling to Access (Hetero)aryl Sulfonamides.

A method to assemble (hetero)aryl sulfonamides via the reductive coupling of aryl sulfinates and nitroarenes is reported. Various reducing conditions with sodium bisulfite and with or without tin(II) chloride in DMSO were developed using an ultrasound bath to improve reaction homogeneity and mixing. A range of (hetero)aryl sulfonamides bearing a selection of functional groups were prepared, and the mechanism of the transformation was investigated. These investigations have led us to propose the formation of nitrosoarene intermediates, which were established via an independent molecular coupling strategy.

Photoredox-Catalyzed Preparation of Sulfones Using Bis-Piperidine Sulfur Dioxide - An Underutilized Reagent for SO2 Transfer.

Sulfonyl groups are widely observed in biologically relevant molecules and consequently, SO2 capture is an increasingly attractive method to prepare these sulfonyl-containing compounds given the range of SO2 -surrogates now available as alternatives to using the neat gas. This, along with the advent of photoredox catalysis, has enabled mild radical capture of SO2 to emerge as an effective route to sulfonyl compounds. Here we report a photoredox-catalyzed cross-electrophile sulfonylation of aryl and alkyl bromides making use of a previously under-used amine-SO2 surrogate; bis(piperidine) sulfur dioxide (PIPSO). A broad selection of alkyl and aryl bromides were photocatalytically converted to their corresponding sulfinates and then trapped with various electrophiles in a one-pot multistep procedure to prepare sulfones and sulfonamides.

The Evolution of Flow Chemistry: An Opinion on Factors Driving Innovation.

This article seeks to provide an overview of the environmental factors within the pharmaceutical industry that have contributed to the emergence of flow chemistry over the past two decades. It highlights some of the challenges facing the industry and describes how they are being overcome by the exponential trajectory of scientific progress in the area. We identify current trends and offer a speculative glimpse into the future of drug development and manufacturing with some examples of progress being made at CARBOGEN AMCIS.

p38γ and p38δ modulate innate immune response by regulating MEF2D activation.

Evidence implicating p38γ and p38δ (p38γ/p38δ) in inflammation are mainly based on experiments using Mapk12/Mapk13-deficient (p38γ/δKO) mice, which show low levels of TPL2, the kinase upstream of MKK1-ERK1/2 in myeloid cells. This could obscure p38γ/p38δ roles, since TPL2 is essential for regulating inflammation. Here, we generated a Mapk12D171A/D171A/Mapk13-/- (p38γ/δKIKO) mouse, expressing kinase-inactive p38γ and lacking p38δ. This mouse exhibited normal TPL2 levels, making it an excellent tool to elucidate specific p38γ/p38δ functions. p38γ/δKIKO mice showed a reduced inflammatory response and less susceptibility to lipopolysaccharide (LPS)-induced septic shock and Candida albicans infection than wild-type (WT) mice. Gene expression analyses in LPS-activated wild-type and p38γ/δKIKO macrophages revealed that p38γ/p38δ-regulated numerous genes implicated in innate immune response. Additionally, phospho-proteomic analyses and in vitro kinase assays showed that the transcription factor myocyte enhancer factor-2D (MEF2D) was phosphorylated at Ser444 via p38γ/p38δ. Mutation of MEF2D Ser444 to the non-phosphorylatable residue Ala increased its transcriptional activity and the expression of Nos2 and Il1b mRNA. These results suggest that p38γ/p38δ govern innate immune responses by regulating MEF2D phosphorylation and transcriptional activity.

Multicomponent Direct Assembly of N-Heterospirocycles Facilitated by Visible-Light-Driven Photocatalysis.

N-heterospirocycles are interesting structural units found in both natural products and medicinal compounds but have relatively few reliable methods for their synthesis. Here, we enlist the photocatalytic generation of N-centered radicals to construct ß-spirocyclic pyrrolidines from N-allylsulfonamides and alkenes. A variety of ß-spirocyclic pyrrolidines have been constructed, including drug derivatives, in moderate to very good yields. Further derivatization of the products has also been demonstrated as has a viable scale-up procedure, making use of flow chemistry techniques.

TPL-2 Inhibits IFN-ß Expression via an ERK1/2-TCF-FOS Axis in TLR4-Stimulated Macrophages.

TPL-2 kinase plays an important role in innate immunity, activating ERK1/2 MAPKs in myeloid cells following TLR stimulation. We investigated how TPL-2 controls transcription in TLR4-stimulated mouse macrophages. TPL-2 activation of ERK1/2 regulated expression of genes encoding transcription factors, cytokines, chemokines, and signaling regulators. Bioinformatics analysis of gene clusters most rapidly induced by TPL-2 suggested that their transcription was mediated by the ternary complex factor (TCF) and FOS transcription factor families. Consistently, TPL-2 induced ERK1/2 phosphorylation of the ELK1 TCF and the expression of TCF target genes. Furthermore, transcriptomic analysis of TCF-deficient macrophages demonstrated that TCFs mediate approximately half of the transcriptional output of TPL-2 signaling, partially via induced expression of secondary transcription factors. TPL-2 signaling and TCFs were required for maximal TLR4-induced FOS expression. Comparative analysis of the transcriptome of TLR4-stimulated Fos -/- macrophages indicated that TPL-2 regulated a significant fraction of genes by controlling FOS expression levels. A key function of this ERK1/2-TCF-FOS pathway was to mediate TPL-2 suppression of type I IFN signaling, which is essential for host resistance against intracellular bacterial infection.

Photoredox-Catalyzed Dehydrogenative Csp3-Csp2 Cross-Coupling of Alkylarenes to Aldehydes in Flow.

Executing photoredox reactions in flow offers solutions to frequently encountered issues regarding reproducibility, reaction time, and scale-up. Here, we report the transfer of a photoredox-catalyzed benzylic coupling of alkylarenes to aldehydes to a flow chemistry setting leading to improvements in terms of higher concentration, shorter residence times, better yields, ease of catalyst preparation, and enhanced substrate scope. Its applicability has been demonstrated by a multi-gram-scale reaction using high-power light-emitting diodes (LEDs), late-stage functionalization of selected active pharmaceutical ingredients (APIs), and also a photocatalyst recycling method.

CLICK-enabled analogues reveal pregnenolone interactomes in cancer and immune cells.

Pregnenolone (P5) promotes prostate cancer cell growth, and de novo synthesis of intratumoural P5 is a potential cause of development of castration resistance. Immune cells can also synthesize P5 de novo. Despite its biological importance, little is known about P5's mode of actions, which appears to be context dependent and pleiotropic. A comprehensive proteome-wide spectrum of P5-binding proteins that are involved in its trafficking and functionality remains unknown. Here, we describe an approach that integrates chemical biology for probe synthesis with chemoproteomics to map P5-protein interactions in live prostate cancer cells and murine CD8+ T cells. We subsequently identified P5-binding proteins potentially involved in P5-trafficking and in P5's non-genomic action that may drive the promotion of castrate-resistance prostate cancer and regulate CD8+ T cell function. We envisage that this methodology could be employed for other steroids to map their interactomes directly in a broad range of living cells, tissues, and organisms.

Endothelial Tpl2 regulates vascular barrier function via JNK-mediated degradation of claudin-5 promoting neuroinflammation or tumor metastasis.

Increased vascular permeability and leakage are hallmarks of several pathologies and determine disease progression and severity by facilitating inflammatory/metastatic cell infiltration. Using tissue-specific genetic ablation in endothelial cells, we have investigated in vivo the role of Tumor progression locus 2 (Tpl2), a mitogen-activated protein kinase kinase kinase (MAP3K) member with pleiotropic effects in inflammation and cancer. In response to proinflammatory stimuli, endothelial Tpl2 deletion alters tight junction claudin-5 protein expression through inhibition of JNK signaling and lysosomal degradation activation, resulting in reduced vascular permeability and immune cell infiltration. This results in significantly attenuated disease scores in experimental autoimmune encephalomyelitis and fewer tumor nodules in a hematogenic lung cancer metastasis model. Accordingly, pharmacologic inhibition of Tpl2 or small interfering RNA (siRNA)-mediated Tpl2 knockdown recapitulates our findings and reduces lung metastatic tumor invasions. These results establish an endothelial-specific role for Tpl2 and highlight the therapeutic potential of blocking the endothelial-specific Tpl2 pathway in chronic inflammatory and metastatic diseases.

TPL-2 kinase induces phagosome acidification to promote macrophage killing of bacteria.

Tumour progression locus 2 (TPL-2) kinase mediates Toll-like receptor (TLR) activation of ERK1/2 and p38α MAP kinases in myeloid cells to modulate expression of key cytokines in innate immunity. This study identified a novel MAP kinase-independent regulatory function for TPL-2 in phagosome maturation, an essential process for killing of phagocytosed microbes. TPL-2 catalytic activity was demonstrated to induce phagosome acidification and proteolysis in primary mouse and human macrophages following uptake of latex beads. Quantitative proteomics revealed that blocking TPL-2 catalytic activity significantly altered the protein composition of phagosomes, particularly reducing the abundance of V-ATPase proton pump subunits. Furthermore, TPL-2 stimulated the phosphorylation of DMXL1, a regulator of V-ATPases, to induce V-ATPase assembly and phagosome acidification. Consistent with these results, TPL-2 catalytic activity was required for phagosome acidification and the efficient killing of Staphylococcus aureus and Citrobacter rodentium following phagocytic uptake by macrophages. TPL-2 therefore controls innate immune responses of macrophages to bacteria via V-ATPase induction of phagosome maturation.

Integrated Batch and Continuous Flow Process for the Synthesis of Goniothalamin.

An integrated batch and continuous flow process has been developed for the gram-scale synthesis of goniothalamin. The synthetic route hinges upon a telescoped continuous flow Grignard addition followed by an acylation reaction capable of delivering a racemic goniothalamin precursor (16) (20.9 g prepared over 3 h), with a productivity of 7 g·h-1. An asymmetric Brown allylation protocol was also evaluated under continuous flow conditions. This approach employing (-)-Ipc2B(allyl) provided an (S)-goniothalamin intermediate in 98% yield and 91.5% enantiomeric excess (ee) with a productivity of 1.8 g·h-1. For the final step, a ring-closing metathesis reaction was explored under several conditions in both batch and flow regimes. In a batch operation, the Grubbs second-generation was shown to be effective and highly selective for the desired ring closure product over those arising from other modes of reactivity, and the reaction was complete in 1.5 h. In a flow operation, reactivity and selectivity were attenuated relative to the batch mode; however, after further optimization, the residence time could be reduced to 16 min with good selectivity and good yield of the target product. A tube-in-tube reactor was investigated for in-situ ethylene removal to favor ring-closing over cross-metathesis, in this context. These results provide further evidence of the utility of flow chemistry for organometallic processing and reaction telescoping. Using the developed integrated batch and flow methods, a total of 7.75 g of goniothalamin (1) was synthesized.

In silico rationalisation of selectivity and reactivity in Pd-catalysed C-H activation reactions.

A computational approach has been developed to automatically generate and analyse the structures of the intermediates of palladium-catalysed carbon-hydrogen (C-H) activation reactions as well as to predict the final products. Implemented as a high-performance computing cluster tool, it has been shown to correctly choose the mechanism and rationalise regioselectivity of chosen examples from open literature reports. The developed methodology is capable of predicting reactivity of various substrates by differentiation between two major mechanisms - proton abstraction and electrophilic aromatic substitution. An attempt has been made to predict new C-H activation reactions. This methodology can also be used for the automated reaction planning, as well as a starting point for microkinetic modelling.

CARD14E138A signalling in keratinocytes induces TNF-dependent skin and systemic inflammation.

To investigate how the CARD14E138A psoriasis-associated mutation induces skin inflammation, a knock-in mouse strain was generated that allows tamoxifen-induced expression of the homologous Card14E138A mutation from the endogenous mouse Card14 locus. Heterozygous expression of CARD14E138A rapidly induced skin acanthosis, immune cell infiltration and expression of psoriasis-associated pro-inflammatory genes. Homozygous expression of CARD14E138A induced more extensive skin inflammation and a severe systemic disease involving infiltration of myeloid cells in multiple organs, temperature reduction, weight loss and organ failure. This severe phenotype resembled acute exacerbations of generalised pustular psoriasis (GPP), a rare form of psoriasis that can be caused by CARD14 mutations in patients. CARD14E138A-induced skin inflammation and systemic disease were independent of adaptive immune cells, ameliorated by blocking TNF and induced by CARD14E138A signalling only in keratinocytes. These results suggest that anti-inflammatory therapies specifically targeting keratinocytes, rather than systemic biologicals, might be effective for GPP treatment early in disease progression.

Photoredox Generation of Sulfonyl Radicals and Coupling with Electron Deficient Olefins.

Various sulfone tetrazoles were activated via iridium photoredox catalysis in the presence of DMAP to give dialkyl sulfones. The presumed sulfone radical intermediates were trapped by a range of electron-deficient olefins in generally good to excellent yields.

A Practical Method for Continuous Production of sp3-Rich Compounds from (Hetero)Aryl Halides and Redox-Active Esters.

A practically useful coupling reaction between aromatic halides and redox-active esters was realized by nickel catalysis through the use of a packed zinc bed column in continuous flow. Multiple reuse of the column showed a negligible decrease in efficiency, affording high space/time yields. A wide range of substrates, including a number of heteroaryl halides and polyfunctional materials were coupled in generally good yields. Longer-time and larger-scale experiments further demonstrates the robustness of the system.

A New World for Chemical Synthesis?

This perspective seeks to provide an overarching vision of the current state of chemical synthesis methodology using machinery as enabling tools. It highlights current capabilities and limitations in this highly digitallyconnected world and suggests areas where new opportunities may arise in the future by going well beyond our present levels of innovation and automation. There is a new need for improved downstream processing tools, advanced reactor design, computational predictive algorithms and integration of robotic systems to maximise the human resource to facilitate a new era in the assembly of our functional materials.

Continuous Pd-Catalyzed Carbonylative Cyclization Using Iron Pentacarbonyl as a CO Source.

This work discloses a continuous flow carbonylation reaction using iron pentacarbonyl as source of CO. The described transformation using this surrogate was designed for use in commonly accessible flow equipment. Optimized conditions were applied to a scalable synthesis of the natural compound isolated from perianal glandular pheromone secretion of the African civet cat. In addition, a flow Pd-catalyzed carbonylation of aryl halides is successfully reported.