Aim Hepatic ischemia reperfusion injury (HIRI) is a leading cause of mortality post liver transplantation, hypovolemic shock and trauma. In this study, we tested, on molecular bases, the possible protective role of two different derivatives of 2-oxindole in a preclinical model of HIRI in rats.
METHODS: HIRI was operated in male Wistar albino rats and prophylactic treatment with oxindole-curcumin (Coxi) or oxindole-vanillin (Voxi) was carried out before the operation. The biochemical and histopathological investigations, in addition to the mechanistic characterizations of the effect of the tested drugs were performed.
RESULTS: HIRI was assured with elevated liver enzymes and marked changes in histopathological features, inflammatory response and oxidative stress. Pretreatment with Coxi and Voxi improved the hepatic histopathological alterations, reduced the elevated serum liver enzymes level and hepatic Malondialdehyde (MDA) content, increased the hepatic Superoxide Dismutase (SOD) activity and reduced Glutathione (GSH) content, downregulated the expression of TNF-α, IL-6, Nod-Like Receptor p3 (NLRP3), Cleaved caspase1, Cleaved caspase 3 proteins, alongside the expression level of IL-1β, ICAM-1, VCAM-1 and BAX genes, attenuated NF-кB p-P65 Ser536 and Myeloperoxidase (MPO)-positive neutrophils, and activated the PI3K/AKT pathway.
CONCLUSIONS: Coxi and Voxi have promising hepatoprotective activity against HIRI in rats through ameliorating the biochemical and histopathological alterations, attenuating inflammatory and oxidative stress status by modulating the inflammatory TNF-α/ICAM-1, the pyroptosis NLRP3/Caspase-1, and the antioxidant PI3K/AKT pathways.

Respiratory syncytial virus (RSV) is a major cause of hospitalization in infants, the elderly, and immune-compromised patients. While a half-life extended monoclonal antibody and 2 vaccines have recently been approved for infants and the elderly, respectively, options to prevent disease in immune-compromised patients are still needed. Here, we describe spiro-azetidine oxindoles as small molecule RSV entry inhibitors displaying favorable potency, developability attributes, and long-acting PK when injected as an aqueous suspension, suggesting their potential to prevent complications following RSV infection over a period of 3 to 6 months with 1 or 2 long-acting intramuscular (IM) or subcutaneous (SC) injections in these immune-compromised patients.

BACKGROUND: IA-0130 is a derivative of 3-(1,3-diarylallylidene)oxindoles, which is a selective estrogen receptor modulator (SERM). A previous study demonstrated that SERM exhibits anti-inflammatory effects on colitis by promoting the anti-inflammatory phenotype of monocytes in murine colitis. However, the therapeutic effects of oxindole on colitis remain unknown. Therefore, we evaluated the efficacy of IA-0130 on dextran sulfate sodium (DSS)-induced mouse colitis.
METHODS: The DSS-induced colitis mouse model was established by administration of 2.5% DSS for 5 days. Mice were orally administered with IA-0130 (0.01 mg/kg or 0.1 mg/kg) or cyclosporin A (CsA; 30 mg/kg). Body weight, disease activity index score and colon length of mice were calculated and histological features of mouse colonic tissues were analyzed using hematoxylin and eosin staining. The expression of inflammatory cytokines and tight junction (TJ) proteins were analyzed using quantitative real-time PCR and enzyme-linked immunosorbent assay. The expression of interleukin-6 (IL-6) signaling molecules in colonic tissues were investigated using Western blotting and immunohistochemistry (IHC).
RESULTS: IA-0130 (0.1 mg/kg) and CsA (30 mg/kg) prevented colitis symptom, including weight loss, bleeding, colon shortening, and expression of pro-inflammatory cytokines in colon tissues. IA-0130 treatment regulated the mouse intestinal barrier permeability and inhibited abnormal TJ protein expression. IA-0130 down-regulated IL-6 expression and prevented the phosphorylation of signaling molecules in colonic tissues.
CONCLUSIONS: This study demonstrated that IA-0130 suppressed colitis progression by inhibiting the gp130 signaling pathway and expression of pro-inflammatory cytokines, and maintaining TJ integrity.

OBJECTIVE: This study aims to disclose how loss of fucosyltransferase 2 (Fut2) impacts intestinal inflammation through cGAS-STING pathway that is closely associated with gut microbiota, and which microbial metabolite improves colitis in Fut2 deficiency.
METHODS: Chronic colitis was induced in intestinal epithelial Fut2 knock out mice (Fut2△IEC), whose intestinal inflammation and activity of cGAS-STING pathway were evaluated. 16S rRNA sequencing and metabolomics were performed using intestinal samples. 2-oxindole was used to treat RAW264.7 cells and Fut2△IEC mice with colitis (Fut2△IEC-DSS) to investigate the effect of 2-oxindole on cGAS-STING response and intestinal inflammation.
RESULTS: Fut2 loss exacerbated chronic colitis in mice, manifested by declined body weight, reduced colon length, increased disease activity index (DAI) and more colon injury in Fut2△IEC-DSS mice compared with WT-DSS (wild type mice with colitis). Lack of Fut2 promoted activation of cGAS-STING pathway. Fut2 deficiency had a primary impact on colonic microbiota, as shown by alteration of microbial diversity and structure, as well as decreased Lactobacillus. Metabolic structure and tryptophan metabolism in colonic luminal microbiota were also influenced by Fut2 loss. Fut2 deficiency also led to decreased levels of aryl hydrocarbon receptor (AHR) and its ligand 2-oxindole derived from tryptophan metabolism. 2-oxindole compromised cGAS-STING response through activating AHR in macrophages, and protected against intestinal inflammation and overactive cGAS-STING pathway in Fut2△IEC-DSS mice.
CONCLUSIONS: Fut2 deficiency promotes cGAS-STING pathway through suppressing 2-oxindole-AHR axis, ultimately facilitating the susceptibility to chronic colitis.

Cyclin-dependent kinase 2 (CDK2) regulates cell cycle checkpoints in the synthesis and mitosis phases and plays a pivotal role in cancerous cell proliferation. The activation of CDK2, influenced by various protein signaling pathways, initiates the phosphorylation process. Due to its crucial role in carcinogenesis, CDK2 is a druggable hotspot target to suppress cancer cell proliferation. In this context, several studies have identified spirooxindoles as an effective class of CDK2 inhibitors. In the present study, three spirooxindoles (SOI1, SOI2, and SOI3) were studied to understand their inhibitory mechanism against CDK2 through a structure-based approach. Molecular docking and molecular dynamics (MD) simulations were performed to explore their interactions with CDK2 at the molecular level. The calculated binding free energy for the spirooxindole-based CDK2 inhibitors aligned well with experimental results regarding CDK2 inhibition. Energy decomposition (ED) analysis identified key binding residues, including I10, G11, T14, R36, F82, K89, L134, P155, T158, Y159, and T160, in the CDK2 active site and T-loop phosphorylation. Molecular mechanics (MM) energy was identified as the primary contributor to stabilizing inhibitor binding in the CDK2 protein structure. Furthermore, the analysis of binding affinity revealed that the inhibitor SOI1 binds more strongly to CDK2 compared to the other inhibitors under investigation. It demonstrated a robust interaction with the crucial residue T160 in the T-loop phosphorylation site, responsible for kinase activation. These insights into the inhibitory mechanism are anticipated to contribute to the development of potential CDK2 inhibitors using the spirooxindole scaffold.

Kratom is a natural psychoactive product known primarily in Southeast Asia, including Thailand, Malaysia, etc. It is also known as krathom, kakuam, ithang, thom (Thailand), biak-biak, ketum (Malaysia) and mambog (Philippines) and is sometimes used as an opium substitute. It is stimulant at doses of 1-5 g, analgesic at doses of 5-15 g and euphoric and sedative at doses of >15 g. Mitragynine is the most abundant indole compound in kratom (Mitragyna speciosa) and is metabolized in humans to 7-hydroxymitragynine, the more active metabolite. Adverse effects include seizures, nausea, vomiting, diarrhea, tachycardia, restlessness, tremors, hallucinations and death. There are few studies on the analytical method for the detection of mitragynine and 7-hydroxymitragynine in hair. Therefore, this study proposes a liquid chromatography-tandem mass spectrometry (LC-MS-MS) method for the analysis of kratom in hair. Hair samples were first weighed to ∼10 mg and washed with methanol. Then the washed hair samples were cut into pieces and incubated in methanol with stirring and heating (16 h/38℃). Extracts were then analyzed by LC-MS-MS. This method was validated by determining the limit of detection (LOD), limit of quantification, linearity, intra- and inter-day accuracy and precision, recovery and matrix effects. The intra- and inter-day precision (CV%) and accuracy (bias%) were within ±20%, which was considered acceptable. Using this newly developed LC-MS-MS method, the simultaneous detection of mitragynine and 7-hydroxymitragynine in six authentic hair samples was achieved to provide the direct evidence of kratom use in the past. Mitragynine concentrations ranged from 16.0 to 2,067 pg/mg (mean 905.3 pg/mg), and 7-hydroxymitragynine concentrations ranged from 0.34 to 15 pg/mg (mean 7.4 pg/mg) in six authentic hair samples from kratom abusers. This may be due to the higher sensitivity of the LOD in this study, with values of 0.05 pg/mg for mitragynine and 0.2 pg/mg for 7-hydroxymitragynine in hair.

Alkenyl oxindoles have been characterized as autophagosome-tethering compounds (ATTECs), which can target mutant huntingtin protein (mHTT) for lysosomal degradation. In order to expand the application of alkenyl oxindoles for targeted protein degradation, we designed and synthesized a series of heterobifunctional compounds by conjugating different alkenyl oxindoles with bromodomain-containing protein 4 (BRD4) inhibitor JQ1. Through structure-activity relationship study, we successfully developed JQ1-alkenyl oxindole conjugates that potently degrade BRD4. Unexpectedly, we found that these molecules degrade BRD4 through the ubiquitin-proteasome system, rather than the autophagy-lysosomal pathway. Using pooled CRISPR interference (CRISPRi) screening, we revealed that JQ1-alkenyl oxindole conjugates recruit the E3 ubiquitin ligase complex CRL4DCAF11 for substrate degradation. Furthermore, we validated the most potent heterobifunctional molecule HL435 as a promising drug-like lead compound to exert antitumor activity both in vitro and in a mouse xenograft tumor model. Our research provides new employable proteolysis targeting chimera (PROTAC) moieties for targeted protein degradation, providing new possibilities for drug discovery.

Environment-benign, multicomponent synthetic methodologies are vital in modern pharmaceutical research and facilitates multi-targeted drug development via synergistic approach. Herein, we reported green and efficient synthesis of pyrano[2,3-c]pyrazole fused spirooxindole linked 1,2,3-triazoles using a tea waste supported copper catalyst (TWCu). The synthetic approach involves a one-pot, five-component reaction using N-propargylated isatin, hydrazine hydrate, ethyl acetoacetate, malononitrile/ethyl cyanoacetate and aryl azides as model substrates. Mechanistically, the reaction was found to proceed via in situ pyrazolone formation followed by Knoevenagel condensation, azide alkyne cycloaddition and Michael\'s addition reactions. The molecules were developed using structure-based drug design. The primary goal is to identifying anti-oxidant molecules with potential ability to modulate α-amylase and DPP4 (dipeptidyl-peptidase 4) activity. The anti-oxidant analysis, as determined via DPPH, suggested that the synthesized compounds, A6 and A10 possessed excellent anti-oxidant potential compared to butylated hydroxytoluene (BHT). In contrast, compounds A3, A5, A8, A9, A13, A15, and A18 were found to possess comparable anti-oxidant potential. Among these, A3 and A13 possessed potential α-amylase inhibitory activity compared to the acarbose, and A3 further emerged as dual inhibitors of both DPP4 and α-amylase with anti-oxidant potential. The relationship of functionalities on their anti-oxidant and enzymatic inhibition was explored in context to their SAR that was further corroborated using in silico techniques and enzyme kinetics.

The oxindole scaffold has been the center of several kinase drug discovery programs, some of which have led to approved medicines. A series of two oxindole matched pairs from the literature were identified where TLK2 was potently inhibited as an off-target kinase. The oxindole has long been considered a promiscuous kinase inhibitor template, but across these four specific literature oxindoles TLK2 activity was consistent, while the kinome profile was radically different ranging from narrow to broad spectrum kinome coverage. We synthesized a large series of analogues, utilizing quantitative structure-activity relationship (QSAR) analysis, water mapping of the kinase ATP binding sites, kinome profiling, and small-molecule x-ray structural analysis to optimize TLK2 inhibition and kinome selectivity. This resulted in the identification of several narrow spectrum, sub-family selective, chemical tool compounds including 128 (UNC-CA2-103) that could enable elucidation of TLK2 biology.

Background: A dual COX/5-LOX strategy was adopted to develop new oxindole derivatives with superior anti-inflammatory activity. Methods: Three series of oxindoles - esters 4a-p, 6a-l and imines 7a-o - were synthesized and evaluated for their anti-inflammatory and analgesic activities. Molecular docking and predicted pharmacokinetic parameters were done for the most active compounds. A new LC-MS/MS method was developed and validated for the quantification of 4h in rat plasma. Results: Compounds 4h, 6d, 6f, 6j and 7m revealed % edema inhibition up to 100.00%; also, 4l and 7j showed 100.00% writhing protection. Compound 4h showed dual inhibitory activity with IC50 = 0.0533 and 0.4195 μM for COX-2 and 5-LOX, respectively. Molecular docking rationalized the obtained biological activity. The pharmacokinetic parameters of 4h from rat plasma were obtained.
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