Salt-inducible kinases (SIKs) are promising therapeutic targets for modulating cytokine responses during innate immune activation. The study of SIK inhibition in animal models of disease has been limited by the lack of selective small-molecule probes suitable for modulating SIK function in vivo. We used the pan-SIK inhibitor HG-9-91-01 as a starting point to develop improved analogs, yielding a novel probe 5 (YKL-05-099) that displays increased selectivity for SIKs versus other kinases and enhanced pharmacokinetic properties. Well-tolerated doses of YKL-05-099 achieve free serum concentrations above its IC50 for SIK2 inhibition for > 16 hours and reduce phosphorylation of a known SIK substrate in vivo. While in vivo active doses of YKL-05-099 recapitulate the effects of SIK inhibition on inflammatory cytokine responses, they did not induce metabolic abnormalities observed in Sik2 knockout mice. These results identify YKL-05-099 as a useful probe to investigate SIK function in vivo, and further support the development of SIK inhibitors for treatment of inflammatory disorders.

An important objective of a cell is to accurately replicate its genetic material and evenly divide along with its subcellular components into two identical daughter cells. To do so, a cell reduces or halts growth, transcription, cap-dependent translation, and undergoes dramatic changes in cellular structure and organization. These changes include chromosome condensation, nuclear envelope breakdown, disassembly of the endoplasmic reticulum and Golgi apparatus, reorganization of the actin cortex, and the formation of the mitotic spindle. Deregulation and errors in these processes can produce nonidentical daughter cells with aberrant chromosome numbers, a state known as aneuploidy and a hallmark of human cancer and the origin of many birth defects (1-5). Therefore, it is imperative that mitosis proceeds in a highly accurate and controlled manner. This is achieved by a sophisticated network of proteins that engage in a multitude of protein-protein interactions regulated by post-translational modifications, including dynamic protein phosphorylation by protein kinases and phosphatases (summarized in (6 -8)).One of the master regulators of mitosis that is conserved from yeast to human is the cyclin-dependent kinase 1 (Cdk1) 1 (9). Cdk1 expression is constant across the cell cycle and the regulation of its activity relies on its association with cyclin A and B, as well as on post-translational modifications including phosphorylation. Specifically, mRNA and protein abundance of cyclin A and B oscillate during the cell cycle due to temporally regulated transcription, translation, and degradation cycles that restrict Cdk1 activity from S-phase to mitosis

Immune checkpoint blockade (ICB) has substantially improved the prognosis of patients with cancer, but the majority experiences limited benefit, supporting the need for new therapeutic approaches. Up-regulation of sialic acid–containing glycans, termed hypersialylation, is a common feature of cancer-associated glycosylation, driving disease progression and immune escape through the engagement of Siglec receptors on tumor-infiltrating immune cells. Here, we show that tumor sialylation correlates with distinct immune states and reduced survival in human cancers. The targeted removal of Siglec ligands in the tumor microenvironment, using an antibody-sialidase conjugate, enhanced antitumor immunity and halted tumor progression in several murine models. Using single-cell RNA sequencing, we revealed that desialylation repolarized tumor-associated macrophages (TAMs). We also identified Siglec-E as the main receptor for hypersialylation on TAMs. Last, we found that genetic and therapeutic desialylation, as well as loss of Siglec-E, enhanced the efficacy of ICB. Thus, therapeutic desialylation represents an immunotherapeutic approach to reshape macrophage phenotypes and augment the adaptive antitumor immune response.

Advances in human genetics have dramatically expanded our understanding of complex heritable diseases. Genome-wide association studies have identified an allelic series of variants associated with increased risk of or protection from inflammatory bowel disease (IBD). The predisposing variant of CARD9 is associated with increased NF-κB-mediated cytokine production. Conversely, the protective variant lacks a functional C-terminal domain and is unable to recruit the E3 ubiquitin ligase TRIM62. Here, we used biochemical insights into CARD9 variant proteins to create a blueprint for IBD therapeutics and recapitulated the mechanism of the CARD9 protective variant using small molecules. We developed a multiplexed bead-based technology to screen compounds for disruption of the CARD9-TRIM62 interaction. We identified compounds that directly and selectively bind CARD9, disrupt TRIM62 recruitment, inhibit TRIM62-mediated ubiquitinylation of CARD9, and demonstrate cellular activity and selectivity in CARD9-dependent pathways. Taken together, small molecules targeting CARD9 illustrate a path toward improved IBD therapeutics.

No abstract