In Vitro Angiogenesis Inhibition and Endothelial Cell Growth and Morphology.

A co-culture assay with human umbilical vein endothelial cells (HUVECs) and normal human dermal fibroblasts (NHDFs) was used to study whether selected angiogenesis inhibitors were able to inhibit differentiation and network formation of HUVECs in vitro. The effect of the inhibitors was determined by the morphology and the calculated percentage area covered by HUVECs. Neutralizing VEGF with avastin and polyclonal goat anti-VEGF antibody and inhibiting VEGFR2 with sorafenib and vatalanib resulted in the formation of HUVEC clusters of variable sizes as a result of inhibited EC differentiation. Furthermore, numerous inhibitors of the VEGF signaling pathways were tested for their effect on the growth and differentiation of HUVECs. The effects of these inhibitors did not reveal a cluster morphology, either individually or when combined to block VEGFR2 downstream pathways. Only the addition of N-methyl-p-bromolevamisole revealed a similar morphology as when targeting VEGF and VEGFR2, meaning it may have an inhibitory influence directly on VEGFR signaling. Additionally, several nuclear receptor ligands and miscellaneous compounds that might affect EC growth and differentiation were tested, but only dexamethasone gave rise to cluster formation similarly to VEGF-neutralizing compounds. These results point to a link between angiogenesis, HUVEC differentiation and glucocorticoid receptor activation.

Chemical Editing of Macrocyclic Natural Products and Kinetic Profiling Reveal Slow, Tight-Binding Histone Deacetylase Inhibitors with Picomolar Affinities.

Histone deacetylases (HDACs) are validated targets for treatment of certain cancer types and play numerous regulatory roles in biology, ranging from epigenetics to metabolism. Small molecules are highly important as tool compounds for probing these mechanisms as well as for the development of new medicines. Therefore, detailed mechanistic information and precise characterization of the chemical probes used to investigate the effects of HDAC enzymes are vital. We interrogated Nature's arsenal of macrocyclic nonribosomal peptide HDAC inhibitors by chemical synthesis and evaluation of more than 30 natural products and analogues. This furnished surprising trends in binding affinities for the various macrocycles, which were then exploited for the design of highly potent class I and IIb HDAC inhibitors. Furthermore, thorough kinetic investigation revealed unexpected inhibitory mechanisms of important tool compounds as well as the approved drug Istodax (romidepsin). This work provides novel inhibitors with varying potencies, selectivity profiles, and mechanisms of inhibition and, importantly, affords insight into known tool compounds that will improve the interpretation of their effects in biology and medicine.

Synthesis of oxacyclic scaffolds via dual ruthenium hydride/Brønsted acid-catalyzed isomerization/cyclization of allylic ethers.

A ruthenium hydride/Brønsted acid-catalyzed tandem sequence is reported for the synthesis of 1,3,4,9-tetrahydropyrano[3,4-b]indoles (THPIs) and related oxacyclic scaffolds. The process was designed on the premise that readily available allylic ethers would undergo sequential isomerization, first to enol ethers (Ru catalysis), then to oxocarbenium ions (Brønsted acid catalysis) amenable to endo cyclization with tethered nucleophiles. This methodology provides not only an attractive alternative to the traditional oxa-Pictet-Spengler reaction for the synthesis of THPIs, but also convenient access to THPI congeners and other important oxacycles such as acetals.

Ruthenium hydride/Brønsted acid-catalyzed tandem isomerization/N-acyliminium cyclization sequence for the synthesis of tetrahydro-ß-carbolines.

This paper describes an efficient tandem sequence for the synthesis of 1,2,3,4-tetrahydro-ß-carbolines (THBCs) relying on a ruthenium hydride/Brønsted acid-catalyzed isomerization of allylic amides to N-acyliminium ion intermediates which are trapped by a tethered indole nucleophile. The methodology provides not only a convenient "aldehyde-free" alternative to the classical Pictet-Spengler reaction but also attractive possibilities for total synthesis, including rapid generation of molecular complexity and formation of quaternary stereogenic centers. TBHCs can also be accessed by harnessing the Suzuki cross-coupling reaction to the isomerization/N-acyliminium cyclization sequence. Finally, diastereo- and enantioselective versions of the title reaction have been examined using substrate control (with dr >15: 1) and asymmetric catalysis (ee up to 57%), respectively.

Diversity-Oriented A3-Macrocyclization for Studying Influences of Ring-Size and Shape of Cyclic Peptides: CD36 Receptor Modulators.

Cyclic peptide diversity has been broadened by elaborating the A3-macrocyclization to include various di-amino carboxylate components with different Nε-amine substituents. Triple-bond reduction provided new cyclic peptide macrocycles with Z-olefin and completely saturated structures. Moreover, cyclic azasulfurylpeptides were prepared by exchanging the propargylglycine (Pra) component for an amino sulfamide surrogate. Examination of such diversity-oriented methods on potent cyclic azapeptide modulators of the cluster of differentiation 36 receptor (CD36) identified the importance of the triple bond as well as the Nε-allyl lysine and azaPra residues for high CD36 binding affinity. Cyclic azapeptides which engaged CD36 effectively reduced pro-inflammatory nitric oxide and downstream cytokine and chemokine production in macrophages stimulated with a Toll-like receptor-2 agonist. Studying the triple bond and amine components in the multiple-component A3-macrocyclization has given a diverse array of macrocycles and pertinent information to guide the development of ideal CD36 modulators with biomedical potential for curbing macrophage-driven inflammation.

Dynamic Chirality in the Mechanism of Action of Allosteric CD36 Modulators of Macrophage-Driven Inflammation.

Dynamic chirality influences numerous processes in nature from protein folding to catalysis. Azapeptides are peptidomimetics possessing semicarbazide residues that can interconvert between sp2 and sp3 hybridization, resulting in stereodynamic interconversions of pseudo-R and -S-configurations by means of a planar intermediate. Cyclic azapeptides have shown unprecedented binding affinity to the cluster of differentiation 36 receptor (CD36) and ability to mitigate macrophage-driven inflammation by modulation of the toll-like receptor 2/6 pathway. A novel approach to synthesize cyclic peptides via A3-macrocyclization has been used to make R- and S-configuration controls to study the relevance of semicarbazide hybridization for modulator activity. Nuclear magnetic resonance spectroscopy analysis of potent cyclic azapeptide CD36 modulators (e.g., 1 and 2) and related cyclic peptides demonstrated that binding affinity correlated with conformational rigidity, and a hybridization preference for sp2 > S- > R-sp3 semicarbazide nitrogen configuration was evaluated. Evidence of the active conformation and the relevance for dynamic chirality serve as insights for creating cyclic (aza)peptide CD36 modulators to curb inflammation.

Catalytic Enantioselective Synthesis of Tetrahydocarbazoles and Exocyclic Pictet-Spengler-Type Reactions.

A synthetic strategy for the synthesis of chiral tetrahydrocarbazoles (THCAs) has been developed. The strategy relies on two types of 6-exo-trig cyclization of 3-substituted indole substrates. Enantioselective domino Friedel-Crafts-type reactions leading to THCAs can be catalyzed by chiral phosphoric acid derivatives (with up to >99% ee), and the first examples of exocyclic Pictet-Spengler reactions to form THCAs are reported.