Structure-Based Design and Characterization of the Highly Potent and Selective Covalent Inhibitors Targeting the Lysine Methyltransferases G9a/GLP.

Protein lysine methyltransferases G9a and GLP, which catalyze mono- and di-methylation of histone H3K9 and nonhistone proteins, play important roles in diverse cellular processes. Overexpression or dysregulation of G9a and GLP has been identified in various types of cancer. Here, we report the discovery of a highly potent and selective covalent inhibitor 27 of G9a/GLP via the structure-based drug design approach following structure-activity relationship exploration and cellular potency optimization. Mass spectrometry assays and washout experiments confirmed its covalent inhibition mechanism. Compound 27 displayed improved potency in inhibiting the proliferation and colony formation of PANC-1 and MDA-MB-231 cell lines and exhibited enhanced potency in reducing the levels of H3K9me2 in cells compared to noncovalent inhibitor 26. In vivo, 27 showed significant antitumor efficacy in the PANC-1 xenograft model with good safety. These results clearly indicate that 27 is a highly potent and selective covalent inhibitor of G9a/GLP.

Reprogramming of TAMs via the STAT3/CD47-SIRPα axis promotes acquired resistance to EGFR-TKIs in lung cancer.

Cross-talk between the tumor microenvironment (TME) and cancer cells plays an important role in acquired drug resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs). The role of tumor-associated macrophages (TAMs), the major component of the TME, in acquired resistance remains unclear. In this study, M2-like reprogramming of TAMs and reduced phagocytosis by macrophages were observed in gefitinib-resistant lung cancer cells and tumor xenografts. CD47 was upregulated in TKI-resistant lung cancer cells, and M2 macrophage polarization and cancer cell escape from macrophage phagocytosis were enhanced. Culture medium from TKI-resistant cells led to metabolic reprogramming of TAMs. STAT3 was associated with CD47 expression in TKI-resistant lung cancer cells. Genetic and pharmacological inhibition of STAT3 enhanced the phagocytic activity of TAMs and alleviated the acquired resistance to EGFR-TKIs via inhibiting the CD47-SIRPα signaling axis and M2 polarization in the co-culture system. Moreover, STAT3 transcriptionally regulated CD47 expression by binding to consensus DNA response elements in the intron of the CD47 gene. Furthermore, the combination of gefitinib with a STAT3 inhibitor and an anti-CD47 monoclonal antibody alleviated the acquired resistance to gefitinib in vitro and in vivo. Collectively, our study reveals the role of TAM reprogramming and the CD47-SIRPα axis in acquired EGFR-TKI resistance and provides a novel therapeutic strategy to overcome the acquired resistance to EGFR-TKIs in lung cancer.

Discovery of a New-Generation S-Adenosylmethionine-Noncompetitive Covalent Inhibitor Targeting the Lysine Methyltransferase Enhancer of Zeste Homologue 2.

The first-generation enhancer of zeste homologue 2 (EZH2) inhibitors suffer from several limitations, such as high dosage, cofactor S-adenosylmethionine (SAM) competition, and acquired drug resistance. Development of covalent EZH2 inhibitors that are noncompetitive with cofactor SAM offers an opportunity to overcome these disadvantages. The structure-based design of compound 16 (BBDDL2059) as a highly potent and selective covalent inhibitor of EZH2 is presented in this context. 16 inhibits EZH2 enzymatic activity at sub-nanomolar concentrations and achieves low nanomolar potencies in cell growth inhibition. The kinetic assay revealed that 16 is noncompetitive with the cofactor SAM, providing the basis for its superior activity over noncovalent and positive controls by reducing competition with cofactor SAM and offering a preliminary proof for its covalent inhibition nature. Mass spectrometric analysis and washout experiments firmly establish its covalent inhibition mechanism. This study demonstrates that covalent inhibition of EZH2 can offer a new opportunity for the development of promising new-generation drug candidates.

Structural optimization of Imidazo[1, 2-a]pyridine derivatives for the treatment of gastric cancer via STAT3 signaling pathway.

STAT3 is a promising therapeutic target for the treatment of gastric cancer, which is one of the most common solid tumors worldwide. In the previous works, we discovered a series of novel STAT3 inhibitors bearing an imidazo[1,2-a] pyridine scaffold. In order to improve the metabolic stability of these compounds, herein we performed a systematic structural optimization leading to a bioactive inhibitor 42, which demonstrated significant effects on inhibiting the growth, migration and invasion of human gastric cancer cells lines (AGS and MGC-803). Meanwhile, it was able to block the phosphorylation and dimerization of STAT3 at low micromolar concentration. Furthermore, compound 42 obviously suppressed tumor growth in MGC-803 derived xenograft mouse model, suggesting that it deserves further exploration as a promising anti-cancer agent for advanced gastric cancer.

Boronic Acid: A Novel Pharmacophore Targeting Src Homology 2 (SH2) Domain of STAT3.

SH2 domains have been recognized as promising targets for various human diseases. However, targeting SH2 domains with phosphopeptides or small-molecule inhibitors derived from bioisosteres of the phosphate group is still challenging. Identifying novel bioisosteres of the phosphate group to achieve favorable in vivo potency is urgently needed. Here, we report the feasibility of targeting the STAT3-SH2 domain with a boronic acid group and the identification of a highly potent inhibitor compound 7 by replacing the carboxylic acid of compound 4 with a boronic acid. Compound 7 shows higher binding affinity, better cellular potency, more favorable PK profiles, and higher in vivo antitumor activity than 4. The stronger anticancer effect of 7 partially stems from its covalent binding mode with the SH2 domain, verified by the washout experiments. The relatively high level of sequence conservation among SH2 domains makes the results presented here of general significance.

Inhibition of STAT3-ferroptosis negative regulatory axis suppresses tumor growth and alleviates chemoresistance in gastric cancer.

Chemotherapy is still one of the principal treatments for gastric cancer, but the clinical application of 5-FU is limited by drug resistance. Here, we demonstrate that ferroptosis triggered by STAT3 inhibition may provide a novel opportunity to explore a new effective therapeutic strategy for gastric cancer and chemotherapy resistance. We find that ferroptosis negative regulation (FNR) signatures are closely correlated with the progression and chemoresistance of gastric cancer. FNR associated genes (GPX4, SLC7A11, and FTH1) and STAT3 are upregulated in 5-FU resistant cells and xenografts. Further evidence demonstrates that STAT3 binds to consensus DNA response elements in the promoters of the FNR associated genes (GPX4, SLC7A11, and FTH1) and regulates their expression, thereby establishing a negative STAT3-ferroptosis regulatory axis in gastric cancer. Genetic inhibition of STAT3 activity triggers ferroptosis through lipid peroxidation and Fe2+ accumulation in gastric cancer cells. We further develop a potent and selective STAT3 inhibitor, W1131, which demonstrates significant anti-tumor effects in gastric cancer cell xenograft model, organoids model, and patient-derived xenografts (PDX) model partly by inducing ferroptosis, thus providing a new candidate compound for advanced gastric cancer. Moreover, targeting the STAT3-ferroptosis circuit promotes ferroptosis and restores sensitivity to chemotherapy. Our finding reveals that STAT3 acts as a key negative regulator of ferroptosis in gastric cancer through a multi-pronged mechanism and provides a new therapeutic strategy for advanced gastric cancer and chemotherapy resistance.

Discovery of Marine Natural Products as Promising Antibiotics against Pseudomonas aeruginosa.

Pseudomonas aeruginosa, one of the most intractable Gram-negative bacteria, has become a public health threat due to its outer polysaccharide layer, efflux transporter system, and high level of biofilm formation, all of which contribute to multi-drug resistance. Even though it is a pathogen of the highest concern, the status of the antibiotic development pipeline is unsatisfactory. In this review, we summarize marine natural products (MNPs) isolated from marine plants, animals, and microorganisms which possess unique structures and promising antibiotic activities against P. aeruginosa. In the last decade, nearly 80 such MNPs, ranging from polyketides to alkaloids, peptides, and terpenoids, have been discovered. Representative compounds exhibited impressive in vitro anti-P. aeruginosa activities with MIC values in the single-digit nanomolar range and in vivo efficacy in infectious mouse models. For some of the compounds, the preliminary structure-activity-relationship (SAR) and anti-bacterial mechanisms of selected compounds were introduced. Compounds that can disrupt biofilm formation or membrane integrity displayed potent inhibition of multi-resistant clinical P. aeruginosa isolates and could be considered as lead compounds for future development. Challenges on how to translate hits into useful candidates for clinical development are also proposed and discussed.

Dodelates A-E: Five dimeric eudesmane sesquiterpenoids from Dobinea delavayi.

Dodelates A-E (1-5), five angeloylated eudesmane sesquiterpenoid dimers were isolated from the roots of Dobinea delavayi. Their structures were elucidated by extensive spectroscopic data and single-crystal X-ray diffraction analyses. A possible biosynthetic pathway of these five compounds was proposed. Compounds 1 and 3 exhibited moderate antimalarial activities against Plasmodium yoelii BY265RFP.