Phenotypic screen identifies calcineurin-sparing FK506 analogs as BMP potentiators for treatment of acute kidney injury.

Acute kidney injury (AKI) is a life-threatening disease with no known curative or preventive therapies. Data from multiple animal models and human studies have linked dysregulation of bone morphogenetic protein (BMP) signaling to AKI. Small molecules that potentiate endogenous BMP signaling should have a beneficial effect in AKI. We performed a high-throughput phenotypic screen and identified a series of FK506 analogs that act as potent BMP potentiators by sequestering FKBP12 from BMP type I receptors. We further showed that calcineurin inhibition was not required for this activity. We identified a calcineurin-sparing FK506 analog oxtFK through late-stage functionalization and structure-guided design. OxtFK demonstrated an improved safety profile in vivo relative to FK506. OxtFK stimulated BMP signaling in vitro and in vivo and protected the kidneys in an AKI mouse model, making it a promising candidate for future development as a first-in-class therapeutic for diseases with dysregulated BMP signaling.

A Stem Cell-Based Screening Platform Identifies Compounds that Desensitize Motor Neurons to Endoplasmic Reticulum Stress.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease selectively targeting motor neurons in the brain and spinal cord. The reasons for differential motor neuron susceptibility remain elusive. We developed a stem cell-based motor neuron assay to study cell-autonomous mechanisms causing motor neuron degeneration, with implications for ALS. A small-molecule screen identified cyclopiazonic acid (CPA) as a stressor to which stem cell-derived motor neurons were more sensitive than interneurons. CPA induced endoplasmic reticulum stress and the unfolded protein response. Furthermore, CPA resulted in an accelerated degeneration of motor neurons expressing human superoxide dismutase 1 (hSOD1) carrying the ALS-causing G93A mutation, compared to motor neurons expressing wild-type hSOD1. A secondary screen identified compounds that alleviated CPA-mediated motor neuron degeneration: three kinase inhibitors and tauroursodeoxycholic acid (TUDCA), a bile acid derivative. The neuroprotective effects of these compounds were validated in human stem cell-derived motor neurons carrying a mutated SOD1 allele (hSOD1A4V). Moreover, we found that the administration of TUDCA in an hSOD1G93A mouse model of ALS reduced muscle denervation. Jointly, these results provide insights into the mechanisms contributing to the preferential susceptibility of ALS motor neurons, and they demonstrate the utility of stem cell-derived motor neurons for the discovery of new neuroprotective compounds.

Design of Small Molecules That Compete with Nucleotide Binding to an Engineered Oncogenic KRAS Allele.

RAS mutations are found in 30% of all human cancers, with KRAS the most frequently mutated among the three RAS isoforms (KRAS, NRAS, and HRAS). However, directly targeting oncogenic KRAS with small molecules in the nucleotide-binding site has been difficult because of the high affinity of KRAS for GDP and GTP. We designed an engineered allele of KRAS and a covalent inhibitor that competes for GTP and GDP. This ligand-receptor combination demonstrates that the high affinity of GTP and GDP for RAS proteins can be overcome with a covalent inhibitor and a suitably engineered binding site. The covalent inhibitor irreversibly modifies the protein at the engineered nucleotide-binding site and is able to compete with GDP and GTP. This provides a new tool for studying KRAS function and suggests strategies for targeting the nucleotide-binding site of oncogenic RAS proteins.

Incorporation of metabolically stable ketones into a small molecule probe to increase potency and water solubility.

Introducing a reactive carbonyl to a scaffold that does not otherwise have an electrophilic functionality to create a reversible covalent inhibitor is a potentially useful strategy for enhancing compound potency. However, aldehydes are metabolically unstable, which precludes the use of this strategy for compounds to be tested in animal models or in human clinical studies. To overcome this limitation, we designed ketone-based functionalities capable of forming reversible covalent adducts, while displaying high metabolic stability, and imparting improved water solubility to their pendant scaffold. We tested this strategy on the ferroptosis inducer and experimental therapeutic erastin, and observed substantial increases in compound potency. In particular, a new carbonyl erastin analog, termed IKE, displayed improved potency, solubility and metabolic stability, thus representing an ideal candidate for future in vivo cancer therapeutic applications.

Rearrangements of N-acyl isothioureas. Alternate access to acylguanidines from cyanamides.

We report a tin-free one-pot radical approach to the synthesis of N-acyl isothioureas and acylguanidines from N-acyl cyanamides. Photoactivated reduction of aromatic disulfides in the presence of Hünig's base results in hydrothiolation of the cyanamide moiety, followed by spontaneous 1,3-migration of the acyl group. Onward reaction of the isothioureas obtained with amines led to the corresponding N-acylguanidines, where the acyl group is attached to the nitrogen atom formerly at the cyano-end of the starting material.

Expeditious synthesis of phenanthridines from benzylamines via dual palladium catalysis.

A method for the synthesis of phenanthridines from benzylamines and aryl iodides which uses a dual palladium-catalyzed process is developed. The domino sequence ends via an intramolecular amination and an oxidative dehydrogenation. No protecting group or prefunctionalization of the amine is required, and the process uses dioxygen as the terminal oxidant.

Radical migration of substituents of aryl groups on quinazolinones derived from N-acyl cyanamides.

A newly designed radical cascade involving N-acyl cyanamides is reported. It builds on aromatic homolytic substitutions as intermediate events and leads to complex heteroaromatic structures via an unprecedented radical migration of a substituent on aryl groups of quinazolinones (hydrogen or alkyl). Mechanistic considerations are detailed, which allowed us to devise fine control over the domino processes. The latter could be predictably stopped at several stages, depending on the reaction conditions. Finally, a surgical introduction of a trifluoromethyl substituent on a quinazolinone was achieved via the reported migration.

Intramolecular homolytic substitution of sulfinates and sulfinamides.

A general and efficient method for the synthesis of cyclic sulfinates and sulfinamides based on intramolecular homolytic substitution (S(H)i) at the sulfur atom by aryl or alkyl radicals is described. Both alkyl and benzofused compounds can be accessed directly from easily prepared acyclic precursors. Enantiomerically enriched sulfur-based heterocycles were formed through an S(H)i process with inversion of configuration at the sulfur atom. Cyclization of prochiral radicals proceeded with varying stereochemical outcomes, depending on the size of the incoming radical. 2-Pyridyl and 2-quinolyl radicals led to biaryl compounds, which result from attack onto the ortho position of the arylsulfinate rather than a thiophilic substitution.