Mahasarakham University

This investigation compares in vitro release, ex vivo release and permeation, and in vivo ocular pharmacokinetics to render biologically informed evaluations of ophthalmic semi-solid drug products containing dexamethasone (hydrophobic) and tobramycin (hydrophilic). Both drugs were formulated with three petrolatum matrices (IGI® 320 A, IGI® 386, or Spectrum®) with distinct rheological character and benchmarked against the reference listed drug, Tobradex®. Temperature-sweep rheology revealed that IGI® 386 most closely reproduced the viscoelastic profile of the reference product. USP Apparatus I release testing with surfactant-free medium provided maximal discrimination for dexamethasone (Tobradex® > IGI® 320 A > IGI® 386 > Spectrum®), and rank-order release rates correlated strongly with ex vivo corneal permeation and in vivo corneal exposure. In contrast, tobramycin required a polysorbate-containing medium to resolve formulation differences in vitro, yet those differences did not persist ex vivo or in vivo, consistent with its rapid dissolution and diffusion, which attenuate matrix effects. The data demonstrate that drug solubility dictates the choice of biorelevant release conditions in petrolatum-based ophthalmic ointments: surfactant-free media capture formulation-dependent release for hydrophobic actives, whereas hydrophilic actives may yield artifactual discrimination when surfactant is present. However, formulations indistinguishable in vitro were typically similar in their in vivo ocular pharmacokinetics. By integrating tiered models, the framework enhances understanding of critical quality attributes, supports regulatory decision-making, and may help reduce reliance on animal studies, thereby expediting the development of therapeutically equivalent generic ophthalmic ointments.

Phosphatidylinositol-4,5-bisphosphate 3-kinase alpha (PI3Kα) is a central signaling enzyme driving cell proliferation and growth in cancers including breast cancer. Selective inhibition of PI3Kα isoform has become a promising therapeutic approach. In this work, 2000 in-house natural compounds were virtually screened against the ATP-binding site of PI3Kα. Of these, 618 compounds were predicted to have acceptable drug-likeness, pharmacokinetic, and toxicity properties based on in silico ADMET screening. Docking analysis highlighted four candidates forming stable hydrogen bonds with key residues V851, S854, and Q859 in the PI3Kα binding pocket. Molecular dynamics simulations were then used to assess their structural features and dynamic stability. Hit 2 was found to form strong hydrogen bonds with E849 and V851 of the PI3Kα protein. MM/GBSA-based binding free energy analysis supported that Hit 2 possessed the most favorable binding affinity to PI3Kα among the identified candidates. In vitro cytotoxicity assays were then performed in MCF-7 and MDA-MB-231 breast cancer cell lines, with alpelisib as a reference compound. Hit 2 reduced cell viability in both cell lines, but its effect was particularly pronounced in MDA-MB-231 cells, a model of triple-negative breast cancer (TNBC). These results suggest that Hit 2 represents a promising natural scaffold for further design and development in breast cancer therapy, with particular relevance for aggressive TNBC.

Anisakis simplex, a parasitic nematode transmitted through the consumption of raw or undercooked seafood, is known to cause gastric anisakidosis, often accompanied by acute inflammatory and allergic responses. However, the underlying immunological mechanisms remain poorly understood. In this study, we established a murine model by orally infecting BALB/c mice with live Anisakis larvae to elucidate the host immune response. Repeated infection with live Anisakis larvae, but not with homogenates, induced severe gastric inflammation, accompanied by a marked elevation of total IgE, Anisakis-specific IgE, Ani s 1-specific IgE, and interleukin (IL)-5 in the serum. Active cutaneous anaphylaxis confirmed the presence of functional Anisakis-specific IgE capable of mediating immediate hypersensitivity. Flow cytometry of gastric lamina propria cells showed increased infiltration of IL-5 producing group 2 innate lymphoid cells (ILC2s), eosinophils, IL-4 producing T helper (Th) cells and B cells in mice with repeated infection. Furthermore, immunohistochemistry revealed widespread localization of IL-33-positive cells in the gastric mucosa, consistent with the observed increased levels of truncated IL-33 and total IL-33. These findings suggest that Anisakis infection induces a type 2 immune response through IL-33-driven activation of ILC2s followed by Th2 development, thereby promoting eosinophilia and antigen-specific IgE. This model enables new insights of Anisakis-associated allergic inflammation and may serve as future therapeutic studies.