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Identification and analysis of small molecule bioactivity in target-agnostic cellular assays and monitoring changes in phenotype followed by identification of the biological target are a powerful approach for the identification of novel bioactive chemical matter in particular when the monitored phenotype is disease-related and physiologically relevant. Profiling methods that enable the unbiased analysis of compound-perturbed states can suggest mechanisms of action or even targets for bioactive small molecules and may yield novel insights into biology. Here we report the enantioselective synthesis of natural-product-inspired 8-oxotetrahydroprotoberberines and the identification of Picoberin, a low picomolar inhibitor of Hedgehog (Hh)-induced osteoblast differentiation. Global transcriptome and proteome profiling revealed the aryl hydrocarbon receptor (AhR) as the molecular target of this compound and identified a cross talk between Hh and AhR signaling during osteoblast differentiation.

Phenotypic drug discovery (PDD) continues to fuel the research and development pipelines with first-in-class therapeutic modalities, but success rates critically depend on the quality of the underlying model system. Here, we employed a stem cell-based approach for the target-agnostic, yet pathway-centric discovery of small-molecule cytokine signaling activators to act as morphogens during development and regeneration. Unbiased screening identified triazolo[1,5

We describe the synthesis and biological evaluation of a new natural product‐inspired compound class obtained by combining the conceptually complementary pseudo‐natural product (pseudo‐NP) design strategy and a formal adaptation of the complexity‐to‐diversity ring distortion approach. Fragment‐sized α‐methylene‐sesquiterpene lactones, whose scaffolds can formally be viewed as related to each other or are obtained by ring distortion, were combined with alkaloid‐derived pyrrolidine fragments by means of highly selective stereocomplementary 1,3‐dipolar cycloaddition reactions. The resulting pseudo‐sesquiterpenoid alkaloids were found to be both chemically and biologically diverse, and their biological performance distinctly depends on both the structure of the sesquiterpene lactone‐derived scaffolds and the stereochemistry of the pyrrolidine fragment. Biological investigation of the compound collection led to the discovery of a novel chemotype inhibiting Hedgehog‐dependent osteoblast differentiation, The combination of the conceptually complementary formal ring distortion‐ and pseudo‐natural product strategies yields novel natural product‐inspired bioactive pseudo‐sesquiterpenoid alkaloids.

Pseudo-natural products (pseudo-NPs) are de novo combinations of natural product (NP) fragments that define novel bioactive chemotypes. For their discovery, new design principles are being sought. Previously, pseudo-NPs were synthesized by the combination of fragments originating from biosynthetically unrelated NPs to guarantee structural novelty and novel bioactivity. We report the combination of fragments from biosynthetically related NPs in novel arrangements to yield a novel chemotype with activity not shared by the guiding fragments. We describe the synthesis of the polyketide pseudo-NP grismonone and identify it as a structurally novel and potent inhibitor of Hedgehog signaling. The insight that the de novo combination of fragments derived from biosynthetically related NPs may also yield new biologically relevant compound classes with unexpected bioactivity may be considered a chemical extension or diversion of existing biosynthetic pathways and greatly expands the opportunities for exploration of biologically relevant chemical space by means of the pseudo-NP principle.

For the discovery of novel chemical matter generally endowed with bioactivity, strategies may be particularly efficient that combine previous insight about biological relevance, e.g., natural product (NP) structure, with methods that enable efficient coverage of chemical space, such as fragment-based design. We describe the de novo combination of different 5-membered NP-derived N-heteroatom fragments to structurally unprecedented “pseudo-natural products” in an efficient complexity-generating and enantioselective one-pot synthesis sequence. The pseudo-NPs inherit characteristic elements of NP structure but occupy areas of chemical space not covered by NP-derived chemotypes, and may have novel biological targets. Investigation of the pseudo-NPs in unbiased phenotypic assays and target identification led to the discovery of the first small-molecule ligand of the RHO GDP-dissociation inhibitor 1 (RHOGDI1), termed Rhonin. Rhonin inhibits the binding of the RHOGDI1 chaperone to GDP-bound RHO GTPases and alters the subcellular localization of RHO GTPases.

An enantioselective hetero-Diels–Alder reaction of alkylidene­ oxindoles and 2-aza-3-silyloxy-1,3-butadienes, catalyzed by divalent transition metal complexes with N,N′-dioxide ligands offered an efficient access to natural-product-based 3,3′-piperidinoyl spiroox­indole class of small molecules. exo-Cycloadducts formed via stereospecific cycloaddition with Z-olefin displayed potent activity in modulation of hedgehog pathway.

RhIII‐catalyzed C−H functionalization reaction yielding isoindolinones from aryl hydroxamates and ortho‐substituted styrenes is reported. The reaction proceeds smoothly under mild conditions at room temperature, and tolerates a range of functional groups. Experimental and computational investigations support that the high regioselectivity observed for these substrates results from the presence of an ortho‐substituent embedded in the styrene. The resulting isoindolinones are valuable building blocks for the synthesis of bioactive compounds. They provide easy access to the natural‐product‐like compounds, isoindolobenzazepines, in a one‐pot two‐step reaction. Selected isoindolinones inhibited Hedgehog (Hh)‐dependent differentiation of multipotent murine mesenchymal progenitor stem cells into osteoblasts., Isoindolinones: The first RhIII‐catalyzed C−H functionalization toward the synthesis of isoindolinones with styrenes is presented. The use of styrenes with a coordinating side arm together with OBoc‐aryl hydroxamates completely switched the regioselectivity. Mechanistic investigations supported the β‐hydride elimination pathway. This method gave an access to NPs scaffold (isoindolobenzazepines). Some of the synthesized derivatives inhibited Hedgehog (Hh)‐dependent differentiation of multipotent murine mesenchymal progenitor stem cells into osteoblasts.

The first synthetic access to stable and acyclic 1,3-butadienes with two electron-withdrawing carbonyl groups and their potential to deliver new molecular scaffolds through intriguing endo-selective Diels-Alder cycloadditions are presented. The bicyclic scaffolds produced through the cycloaddition chemistry of electron-deficient dienes afforded potent Hedgehog signaling pathway inhibitors.

Reported is the identification of the furo[3,2-b]pyridine core as a novel scaffold for potent and highly selective inhibitors of cdc-like kinases (CLKs) and efficient modulators of the Hedgehog signaling pathway. Initially, a diverse target compound set was prepared by synthetic sequences based on chemoselective metal-mediated couplings, including assembly of the furo[3,2-b]pyridine scaffold by copper-mediated oxidative cyclization. Optimization of the subseries containing 3,5-disubstituted furo[3,2-b]pyridines afforded potent, cell-active, and highly selective inhibitors of CLKs. Profiling of the kinase-inactive subset of 3,5,7-trisubstituted furo[3,2-b]pyridines revealed sub-micromolar modulators of the Hedgehog pathway.

Herein, we report the first enantioselective annulation of α-arylidene pyrazolones through a formal C(sp3 )-H activation under mild conditions enabled by highly variable RhIII -Cpx catalysts. The method has a wide substrate scope and proceeds with good to excellent yields and enantioselectivities. Its synthetic utility was demonstrated by the late-stage functionalization of drugs and natural products as well as the preparation of enantioenriched [3]dendralenes. Preliminary biological investigations also identified the spiropyrazolones as a novel class of Hedgehog pathway inhibitors.

The Hedgehog (Hh) signaling pathway is crucial for vertebrate embryonic development, tissue homeostasis and regeneration. Hh signaling is upregulated in basal cell carcinoma and medulloblastoma and Hh pathway inhibitors targeting the Smoothened (SMO) protein are in clinical use. However, the signaling cascade is incompletely understood and novel druggable proteins in the pathway are in high demand. We describe the discovery of the Hh‐pathway modulator Pipinib by means of cell‐based screening. Target identification and validation revealed that Pipinib selectively inhibits phosphatidylinositol 4‐kinase IIIβ (PI4KB) and suppresses GLI‐mediated transcription and Hh target gene expression by impairing SMO translocation to the cilium. Therefore, inhibition of PI4KB and, consequently, reduction in phosphatidyl‐4‐phosphate levels may be considered an alternative approach to inhibit SMO function and thus, Hedgehog signaling., Cell‐based screening identifies a thieno[3,2]pyrimidine derivative as an inhibitor of Hedgehog signaling by selective inhibition of phosphatidylinositol 4‐kinase IIIβ (PI4KB), reduction of PI4P levels and prevention of Hedgehog‐mediated ciliary localization of Smoothened. Thus, PI4KB may be a relevant target protein for inhibition of Hedgehog signaling.

Herein, we report the first enantioselective annulation of α-arylidene pyrazolones through a formal C(sp

Axially chiral 4-arylisoquinolones are endowed with pronounced bioactivity, and methods for their efficient synthesis have gained widespread attention. However, enantioselective synthesis of axially chiral 4-arylisoquinolones by means of C-H activation has not been reported to date. Described here is a rhodium (III)-catalyzed C-H bond activation and annulation for the atroposelective synthesis of axially chiral 4-arylisoquinolones. The method employs chiral cyclopentadienyl ligands embodying a piperidine ring as backbone and yields the atropisomers with good to excellent yields and enantioselectivity. Biological relevance of the 4-arylisoquinolones was demonstrated by their investigation in different cellular assays, leading to the discovery of novel non-SMO (SMO= smoothened) binding Hedgehog pathway inhibitors.