Inappropriate expression of histone deacetylase (HDAC-6) and deregulation of the phosphatidylinositol 3-kinase (PI3K) signalling pathway are common aberrations observed in cancers. LASSBio-2208, has been previously described as a dual inhibitor in the nanomolar range of HDAC-6 and PI3Kα and is three times more potent in inhibiting HDAC-6. In this paper we described the cytotoxic and antiproliferative potency of LASSBio-2208 on different tumour cell lines, its possible synergism effect in association with PI3K and HDAC-6 inhibitors, and its drug metabolism and pharmacokinetics (DMPK) in vitro profile. Our studies have demonstrated that LASSBio-2208 has moderate cytotoxic potency on breast cancer cell line MCF-7 (IC₅₀ = 23 µM), human leukaemia cell line CCRF-CEM (IC₅₀ = 8.54 µM) and T lymphoblast cell line MOLT-4 (IC₅₀ = 7.15 µM), with no cytotoxic effect on human peripheral blood mononuclear cells (hPBMC). In addition, it has a good antiproliferative effect on MCF-7 cells (IC₅₀ = 5.44 µM), low absorption by parallel artificial membrane permeability—gastrointestinal tract (PAMPA—GIT) and low permeation by parallel artificial membrane permeability—blood–brain barrier (BBB) (PAMPA—BBB), exhibiting high metabolic stability in rat plasma. Moreover, LASSBio-2208 exhibited synergism when combined with getadolisib and tubastatin A, using the concentrations corresponding to their CC₅₀ values on MOLT-4 and CCRF-CEM cells.

Rho-associated coiled-coil containing kinases (ROCK1 and ROCK2) are central regulators of actin cytoskeleton organization and cell contractility under physiological conditions. Dysregulation of ROCK signaling contributes to aberrant cell migration, invasion, and tissue remodeling, positioning these kinases as attractive therapeutic targets in cancer and fibrotic diseases. In this work, we report the discovery of N-acylhydrazone (NAH) derivatives as potent and selective ROCK inhibitors, integrating structure-based virtual screening (SBVS), de novo design, and biological evaluation. Initial hit identification from the LASSBio Chemical Library revealed three inhibitors (LASSBio-1828 (1), LASSBio-1829 (2), and LASSBio-1919 (3)), which guided the rational design of a virtual library of 321 NAH analogues. Docking-based prioritization, synthesis, and SAR exploration yielded compounds with low nanomolar potency, among which LASSBio-2360 (12), LASSBio-2380 (17), and LASSBio-2382 (18) exhibited dual ROCK1/2 inhibition (IC₅₀ values in the 1–15 nM range), while LASSBio-2389 (21) showed remarkable ROCK2 selectivity (IC₅₀ = 0.051 μM; 21-fold vs ROCK1 - IC₅₀ = 1.143 μM) and minimal inhibition of other related kinases at 500 nM. Molecular dynamics simulations demonstrated that 21 stabilizes the DFG-out conformation of ROCK2, providing a structural rationale for isoform selectivity. In vitro studies using MDA-MB-231 triple-negative breast cancer cells confirmed that compounds 12, 17, 18, and 21 inhibit migration more effectively than fasudil and comparably to belumosudil. Altogether, this work identifies NAH as a privileged scaffold for ROCK inhibition, delineates the molecular determinants of ROCK2 selectivity, and highlights new chemical leads for the development of antimetastatic and antifibrotic agents targeting the Rho/ROCK pathway.

Background: Nrf2 plays a key role in regulating the antioxidant response against oxidative stress. Therefore, it is imperative to examine the advantages of Nrf2 activation by new small molecules capable of inhibiting the Nrf2-Keap1 protein interaction that do not present electrophilic sites, since electrophilic compounds have intrinsic toxicity. The bixin pigment has been used as a form of treatment and prevention of several pathological conditions in animal models since it was described as an Nrf2 activator without electrophilic sites. This study aims to synthetize a soluble derivate KBx (potassium bixinate) and evaluate its ability to activate Nrf2/ARE in a model of exposure to cigarette smoke extract (CSE; in vitro) and intranasal LPS administration (in vivo). Methods: In the in vivo study, C57BL/6 mice were pretreated with 200 mg/kg of KBx (gavage) during 5 consecutive days and then challenged with 60 µg of LPS i.n. for 16 h. Bronchoalveolar lavage was collected to examine cytokines dosage. In the in vitro study, RAW 264.7 macrophages were exposed to CSE and post-treated with KBx to evaluate their ability to revert the redox imbalance caused by the stressor. Results: KBx was characterized using mass spectrometry (433.1778 m/z). KC levels were increased in the LPS group (p = 0.021), and KBx inhibited this (p = 0.001). IL-10 levels were decreased (p = 0.055) in the LPS group that was prevented when pretreated with KBx (p = 0.037). The in vitro study showed KBx to be a more potent derivate of bixin through its ability to intercept ROS formation with three-fold more potency, and it showed an anti-inflammatory propriety by reducing the nuclear translocation of p65 (p < 0.001). Conclusions: In conclusion, these data suggest that KBx was able to activate the Nrf2/ARE pathway and intercept ROS formation induced by CSE and LPS in both in vivo and in vitro studies.

Inappropriate expression of histone deacetylase (HDAC6) and deregulation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway are common aberrations observed in cancers. LASSBio-2208, was previously described as a dual inhibitor in the nanomolar range of HDAC6 and PI3Kα, being three times more potent in inhibiting HDAC-6. In this paper we described the cytotoxic and antiproliferative potency of LASSBio-2208 on different tumour cell lines, its possible synergism effect in association with PI3K and HDAC-6 inhibitors, and its DMPK in vitro profile. Our studies have demonstrated that LASSBio-2208 has moderate cytotoxic potency on MCF-7 (IC50 = 23 µM), CCRF-CEM (IC50 = 8.54 µM) and MOLT-4 (IC50 = 7.15 µM), with no cytotoxic effect on hPBMC. In addition, it has a good antiproliferative effect on MCF-7 cells (IC50 = 5.44 µM), low absorption by PAMPA-GIT and low permeation through the blood-brain barrier by PAMPA-BBB, exhibiting high metabolic stability in rat plasma. Moreover, LASSBio-2208 exhibited synergism when combined with getadolisib and tubastatin A, using the concentration corresponding to their CC50 values on MOLT-4 and CCRF-CEM cells.

The PI3K/AKT/mTOR pathway is frequently dysregulated in cancer, contributing to tumor progression, drug resistance, and poor prognosis. Dual PI3K/mTOR inhibitors such as gedatolisib have shown clinical promise, but they still face challenges, including low solubility, poor metabolic stability, and limited activity against resistant tumor phenotypes. Here, we report a proof-of-concept study exploring structural modifications on compound 5f, a simplified gedatolisib analog, to generate a novel small subseries of morpholino-triazine derivatives (9a–f). The goal was to improve molecular interactions within the affinity site of PI3K, investigate the impact on isoform selectivity, and evaluate pharmacological properties relevant to early optimization. Among these, compound 9a (LASSBio-2337) emerged as a dual pan-PI3K/mTOR inhibitor (IC₅₀: 0.3–5.8 μM), showing cytotoxic effects in leukemia cell lines (CC₅₀: 4.37–9.44 μM), including those with multidrug resistance (Lucena, MDR phenotype), while sparing nontumor hPBMCs. Although aqueous insoluble, 9a displayed moderate PAMPA-GIT permeability and low metabolic stability in rat liver microsomes, underscoring its potential as a lead for further optimization. This integrated study provides structural, mechanistic, and pharmacokinetic insights to guide next-generation PI3K/mTOR inhibitor design.