From EGFR kinase inhibitors to anti-inflammatory drugs: Optimization and biological evaluation of (4-(phenylamino)quinazolinyl)-phenylthiourea derivatives as novel NF-κB inhibitors.

The transcription factor NF-κB is a pivotal mediator of chronic inflammatory and autoimmune diseases. Based on our previously published dual EGFR/NF-κB inhibitors, we designed and synthesized new thiourea quinazoline derivatives that retained only the NF-κB inhibitory activity. Several congeners displayed a strong suppression of NF-κB activity in a reporter gene assay, yet low cytotoxicity, and were further evaluated in differentiated macrophage-like THP-1 cells. The compounds exhibited a strong inhibition of IL-6 and, less potently, of TNFα release, which was accompanied by a selective induction of macrophage cell death. The mode of action was investigated with a selected inhibitor, 18, revealing that the translocation of p65/RelA to the nucleus but not its release from the IκB complex was inhibited. Eventually, 18 was identified as the first small molecule inhibitor affecting only the phosphorylation of p65-Ser468 but not of Ser536, which may be causally related to the retention of NF-κB in the cytoplasm. Altogether, our novel NF-κB inhibitors seem applicable for the suppression of cytokine release and the additional selective depletion of activated macrophages in various inflammatory diseases.

Synthesis, biological evaluation and molecular docking studies of novel 1,2,3-triazole-quinazolines as antiproliferative agents displaying ERK inhibitory activity.

ERK1/2 inhibitors have attracted special attention concerning the ability of circumventing cases of innate or log-term acquired resistance to RAF and MEK kinase inhibitors. Based on the 4-aminoquinazoline pharmacophore of kinases, herein we describe the synthesis of 4-aminoquinazoline derivatives bearing a 1,2,3-triazole stable core to bridge different aromatic and heterocyclic rings using copper-catalysed azide-alkyne cycloaddition reaction (CuAAC) as a Click Chemistry strategy. The initial screening of twelve derivatives in tumoral cells (CAL-27, HN13, HGC-27, and BT-20) revealed that the most active in BT-20 cells (25a, IC50 24.6 µM and a SI of 3.25) contains a more polar side chain (sulfone). Furthermore, compound 25a promoted a significant release of lactate dehydrogenase (LDH), suggesting the induction of cell death by necrosis. In addition, this compound induced G0/G1 stalling in BT-20 cells, which was accompanied by a decrease in the S phase. Western blot analysis of the levels of p-STAT3, p-ERK, PARP, p53 and cleaved caspase-3 revealed p-ERK1/2 and p-STA3 were drastically decreased in BT-20 cells under 25a incubation, suggesting the involvement of these two kinases in the mechanisms underlying 25a-induced cell cycle arrest, besides loss of proliferation and viability of the breast cancer cell. Molecular docking simulations using the ERK-ulixertinib crystallographic complex showed compound 25a could potentially compete with ATP for binding to ERK in a slightly higher affinity than the reference ERK1/2 inhibitor. Further in silico analyses showed comparable toxicity and pharmacokinetic profiles for compound 25a in relation to ulixertinib.

Design, synthesis, and biological evaluation of some novel 4-aminoquinazolines as Pan-PI3K inhibitors.

A series of 4-aminoquinazolines derivatives containing hydrophilic group were designed and identified as potent Pan-PI3K inhibitors in this study. The results of antiproliferative assays in vitro showed that this series of compounds had strong inhibition of tumor growth, especially compound 7b for MCF-7 cells but weak inhibition to normal cells. PI3K kinase assay showed that 7b had high activity for three PI3K isoforms with the IC50 values of picomole. The western blot assay indicated that 7b could decrease the phospho-Akt (S473) in a dose-dependent manner. Further experiments showed that 7b could induce apoptosis in MCF-7 cells. Four key hydrogen bonding interactions were found in the docking of 7b with PI3K kinase. All these results suggested that 7b is a potent PI3K inhibitor and could be considered as a potential candidate for the development of anticancer agents.

Discovery of substituted 4-aminoquinazolines as selective Toll-like receptor 4 ligands.

The Toll-like receptors (TLRs) are critical components of the innate immune system that regulate immune recognition in part through NF-κB activation. A human cell-based high throughput screen (HTS) revealed substituted 4-aminoquinazolines to be small molecular weight activators of NF-κB. The most potent hit compound predominantly stimulated through the human TLR4/MD2 complex, and had less activity with the mouse TLR4/MD2. There was no activity with other TLRs and the TLR4 activation was MD-2 dependent and CD14 independent. Synthetic modifications of the quinazoline scaffold at the 2 and 4 positions revealed trends in structure-activity relationships with respect to TLR dependent production of the NF-κB associated cytokine IL-8 in human peripheral blood mononuclear cells, as well as IL-6 in mouse antigen presenting cells. Furthermore, the hit compound in this series also activated the interferon signaling pathway resulting in type I interferon production. Substitution at the O-phenyl moiety with groups such as bromine, chlorine and methyl resulted in enhanced immunological activity. Computational studies indicated that the 4-aminoquinazoline compounds bind primarily to human MD-2 in the TLR4/MD-2 complex. These small molecules, which preferentially stimulate human rather than mouse innate immune cells, may be useful as adjuvants or immunotherapeutic agents.

Discovery of new sulfonamide-tethered 2-aryl-4-anilinoquinazolines as the first-in-class dual carbonic anhydrase and EGFR inhibitors.

In today's medical field, there is a growing trend of exploiting a single small molecule to target two different molecular targets concurrently. This approach is proving to be highly effective in fighting against cancer. The 4-anilinoquinazoline scaffold, known for its potential in cancer therapy and its effectiveness as a leading class of tyrosine kinase inhibitors, was employed to develop a novel series of anilinoquinazoline-sulfonamides (AQSs) (8a-d, 9a-f, and 10a-d) as dual inhibitors of the tumor-associated carbonic anhydrases (CA) IX/XII and EGFR. 2-(3-Methoxyphenyl)quinazoline bearing p-sulfanilamide 10b elicited superior hCA IX and XII inhibition in the low nanomolar range (KIs = 38.4 and 8.9 nM, respectively). Also, 10b shined as a potent and selective EGFR inhibitor, boasting an impressive IC50 value of 51.2 ±â€¯0.97 nM, surpassing the reference EGFR inhibitor Erlotinib (IC50 = 80 ±â€¯2.0 nM). Compound 10b exhibited broadest-spectrum antiproliferative activity against the NCI-tumor panel with a mean GI% value of 68 %. Of special interest, 10b demonstrated potent growth inhibition (GI% ≥ 80-97 %) toward cell lines reported to express high levels of EGFR belonging to renal, colon, breast, and lung cancers. Compound 10b's molecular docking in the CA IX/XII and EGFR active sites revealed binding modes that justify its potent enzyme inhibitory effects. Additionally, molecular dynamic simulations demonstrated strong and stable interactions of 10b with the binding sites of these targets.

Development of (4-Phenylamino)quinazoline Alkylthiourea Derivatives as Novel NF-κB Inhibitors.

For many inflammatory diseases, new effective drugs with fewer side effects are needed. While it appears promising to target the activation of the central pro-inflammatory transcription factor NF-κB, many previously discovered agents suffered from cytotoxicity. In this study, new alkylthiourea quinazoline derivatives were developed that selectively inhibit the activation of NF-κB in macrophage-like THP-1 cells while showing low general cytotoxicity. One of the best compounds, 19, strongly inhibited the production of IL-6 (IC50 = 0.84 µM) and, less potently, of TNFα (IC50 = 4.0 µM); in comparison, the reference compound, caffeic acid phenethyl ester (CAPE), showed IC50s of 1.1 and 11.4 µM, respectively. Interestingly, 19 was found to block the translocation of the NF-κB dimer to the nucleus, although its release from the IκB complex was unaffected. Furthermore, 19 suppressed the phosphorylation of NF-κB-p65 at Ser468 but not at Ser536; however, 19 did not inhibit any kinase involved in NF-κB activation. The only partial suppression of p65 phosphorylation might be associated with fewer side effects. Since several compounds selectively induced cell death in activated macrophage-like THP-1 cells, they might be particularly effective in various inflammatory diseases that are exacerbated by excess activated macrophages, such as arteriosclerosis and autoimmune diseases.

Design, synthesis and biological evaluation of novel 4-aminoquinazolines as dual target inhibitors of EGFR-PI3Kα.

The overexpression of EGFR correlates with rapidly progressive disease, resistance to chemotherapy and poor prognosis. In certain human cancers, PI3K works synergistically with EGFR to promote proliferation, survival, invasion and metastasis. Development of dual-target drugs against EGFR and PI3K has therapeutic advantage and was an attractive approach against tumors. In this work, based on the molecular docking and previous studies, a series of 4-aminoquinazolines derivatives containing 6-sulfonamide substituted pyridyl group were rationally designed and identified as potent EGFR and PI3K dual inhibitors. The cytotoxicity experiment results showed that this series of compounds could effectively inhibit cell growth. The kinase assay demonstrated that 6c and 6i had high inhibition for EGFR and selectivity for PI3Kα distinguished from other isoforms. Further experiments showed that 6c could induce cell cycle arrest in G1 phase and apoptosis in BT549 cells. The western blot assay indicated that 6c inhibited the proliferation of BT549 cell through EGFR and PI3Kα/Akt signaling pathway. Our study suggested that compound 6c was a potential dual inhibitors of EGFR and PI3Kα.