Hypoglycemia, as a complication of type 2 diabetes (T2D), causes increased morbidity and mortality but the physiological response underlying hypoglycemia has not been fully elucidated. Small noncoding microRNA (miRNA) have multiple downstream biological effects. This pilot exploratory study was undertaken to determine if induced miRNA changes would persist and contribute to effects seen 24 h post-hypoglycemia. A parallel, prospective study design was employed, involving T2D (n = 23) and control (n = 23) subjects. The subjects underwent insulin-induced hypoglycemia (2 mmol/L; 36 mg/dL); blood samples were drawn at baseline, upon the induction of hypoglycemia, and 4 h and 24 h post-hypoglycemia, with a quantitative polymerase chain reaction analysis of miRNA undertaken. The baseline miRNAs did not differ. In the controls, 15 miRNAs were downregulated and one was upregulated (FDR < 0.05) from the induction of hypoglycemia to 4 h later while, in T2D, only four miRNAs were altered (downregulated), and these were common to both cohorts (miR-191-5p; miR-143-3p; let-7b-5p; let-7g-5p), correlated with elevated glucagon levels, and all were associated with energy balance. From the induction of hypoglycemia to 24 h, 14 miRNAs were downregulated and 5 were upregulated (FDR < 0.05) in the controls; 7 miRNAs were downregulated and 7 upregulated (FDR < 0.05) in T2D; a total of 6 miRNAs were common between cohorts, 5 were downregulated (miR-93-5p, let-7b-5p, miR-191-5p, miR-185-5p, and miR-652-3p), and 1 was upregulated (miR-369-3p). An ingenuity pathway analysis indicated that many of the altered miRNAs were associated with metabolic and coagulation pathways; however, of the inflammatory proteins expressed, only miR-143-3p at 24 h correlated positively with tumor necrosis factor-α (TNFa; p < 0.05 and r = 0.46) and negatively with toll-like receptor-4 (TLR4; p < 0.05 and r = 0.43). The MiRNA levels altered by hypoglycemia reflected changes in counter-regulatory glucagon and differed between cohorts, and their expression at 24 h suggests miRNAs may potentiate and prolong the physiological response. Trial registration: ClinicalTrials.gov NCT03102801.

Background: Colorectal cancer (CRC) is a complex and increasingly prevalent malignancy with significant challenges in its treatment and prognosis. This study aims to explore the role of the SLC4A4 transporter as a biomarker in CRC progression and its potential as a therapeutic target, particularly in relation to tumor acidity and immune response. Methods: The study utilized computational approaches, including receptor-based virtual screening and high-throughput docking, to identify potential SLC4A4 inhibitors. A model of the human SLC4A4 structure was generated based on CryoEM data (PDB ID 6CAA), and drug candidates from the DrugBank database were evaluated using two computational tools (DrugRep and CB-DOCK2). Results: The study identified the compound (5R)-N-[(1r)-3-(4-hydroxyphenyl)butanoyl]-2-decanamide (DB07991) as the best ligand, demonstrating favorable binding affinity and stability. Molecular dynamics simulations revealed strong protein–ligand interactions with consistent RMSD (~0.25 nm), RMSF (~0.5 nm), compact Rg (4.0–3.9 nm), and stable SASA profiles, indicating that the SLC4A4 structure remains stable upon ligand binding. Conclusions: The findings suggest that DB07991 is a promising drug candidate for further investigation as a therapeutic agent against CRC, particularly for targeting SLC4A4. This study highlights the potential of computational drug repositioning in identifying effective treatments for colorectal cancer.

Introduction: Colorectal cancer is a highly complex disease that continues to rise in prevalence, posing significant difficulties in its management and treatment outcomes. Reduced expression of the SLC4A4 gene has been identified as a critical factor in driving tumor development and poor clinical prognosis in CRC. This reduction disrupts several key cellular mechanisms, including cellular proliferation, programmed cell death, and metastasis, largely by altering pH regulation within the cells. Given its influence on tumor behavior and patient survival, SLC4A4 expression levels could serve as a reliable prognostic marker in colorectal cancer. Methods: The CryoEM structure of SLC4A4, retrieved from the Protein Data Bank (PDB), was refined using SwissModel and subjected to virtual drug screening via the DrugRep web server. This screening employed a database comprising FDA-approved drugs. Results: Nilotinib emerged as the most potent inhibitor following further validation through redocking using CBDOCK2, ranking it highest among the top three results from DrugRep. The docking analysis yielded a score of −11.2 kcal/mol for Nilotinib. A 50-nanosecond molecular dynamics simulation further validated these findings, revealing that Nilotinib formed robust interactions with the SLC4A4 protein. The simulation yielded consistent results, with a root mean square deviation of around 0.30 nm, a root mean square fluctuation of near 0.5 nm, a compact radius of gyration between 3.8 and 4.0 nm, and stable solvent-accessible surface area profiles. These findings confirmed that the drug-protein complex maintained structural stability throughout the simulation. Discussion: The computational findings suggest that Nilotinib binds effectively and stably to SLC4A4, indicating its potential to modulate the function of this protein in CRC. Its established safety profile as an FDA-approved drug supports the feasibility of drug repurposing. These results also reinforce the utility of integrating structure-based drug screening with molecular dynamics simulations to identify novel therapeutic agents for cancer. Conclusion: Nilotinib holds significant potential as a therapeutic agent for colorectal cancer and warrants further experimental investigation to validate its effectiveness.