So-Deok Lee

Structure–Activity Relationships and Optimization of 3,5-Dichloropyridine Derivatives As Novel P2X7 Receptor Antagonists

Screening of a library of chemical compounds showed that the dichloropyridine-based analogue 9 was a novel P2X(7) receptor antagonist. To optimize its activity, we assessed the structure-activity relationships (SAR) of 9, focusing on the hydrazide linker, the dichloropyridine skeleton, and the hydrophobic acyl (R(2)) group. We found that the hydrazide linker and the 3,5-disubstituted chlorides in the pyridine skeleton were critical for P2X(7) antagonistic activity and that the presence of hydrophobic polycycloalkyl groups at the R(2) position optimized antagonistic activity. In the EtBr uptake assay in hP2X(7)-expressing HEK293 cells, the optimized antagonists, 51 and 52, had IC(50) values of 4.9 and 13 nM, respectively. The antagonistic effects of 51 and 52 were paralleled by their ability to inhibit the release of the pro-inflammatory cytokine, IL-1β, by LPS/IFN-γ/BzATP stimulation of THP-1 cells (IC(50) = 1.3 and 9.2 nM, respectively). In addition, 52 strongly inhibited iNOS/COX-2 expression and NO production in THP-1 cells, further indicating that this compound blocks inflammatory signaling and suggesting that the dichloropyridine analogues may be useful in developing P2X(7) receptor targeted anti-inflammatory agents.

Discovery of Novel 2,5-Dioxoimidazolidine-Based P2X7 Receptor Antagonists as Constrained Analogues of KN62

Novel 2,5-dioxoimidazolidine-based conformationally constrained analogues of KN62 (1) were developed as P2X7 receptor (P2X7R) antagonists using a rigidification strategy of the tyrosine backbone of 1. SAR analysis of the 2,5-dioxoimidazolidine scaffold indicated that piperidine substitution at the N3 position and no substitution at N1 position were preferable. Further optimization of the substituents at the piperidine nitrogen and the spacer around the skeleton resulted in several superior antagonists to 1, including 1-adamantanecarbonyl analogue 21i (IC50 = 23 nM in ethidium uptake assay; IC50 = 14 nM in IL-1β ELISA assay) and (3-CF3-4-Cl)benzoyl analogue (-)-21w (54 nM in ethidium uptake assay; 9 nM in IL-1β ELISA assay), which was more potent than the corresponding (+) isomer. Compound 21w displayed potent inhibitory activity in an ex vivo model of LTP-induced pain signaling in the spinal cord and significant anti-inflammatory activity in in vivo models of carrageenan-induced paw edema and type II collagen-induced joint arthritis.

Potent Suppressive Effects of 1-Piperidinylimidazole Based Novel P2X7 Receptor Antagonists on Cancer Cell Migration and Invasion.

The P2X7 receptor (P2X7R) has been reported as a key mediator in inflammatory processes and cancer invasion/metastasis. In this study, we report the discovery of novel P2X7R antagonists and their functional activities as potential antimetastatic agents. Modifications of the hydantoin core-skeleton and the side chain substituents of the P2X7R antagonist 7 were performed. The structure-activity relationships (SAR) and optimization demonstrated the importance of the sulfonyl group at the R1 position and the substituted position and overall size of R2 for P2X7R antagonism. The optimized novel analogues displayed potent P2X7 receptor antagonism (IC50 = 0.11-112 nM) along with significant suppressive effects on IL-1β release (IC50 = 0.32-210 nM). Moreover, representative antagonists (12g, 13k, and 17d) with imidazole and uracil core skeletons significantly inhibited the invasion of MDA-MB-231 triple negative breast cancer cells and cancer cell migration in a zebrafish xenograft model, suggesting the potential therapeutic application of these novel P2X7 antagonists to block metastatic cancer.

Solid-phase synthesis of quinolinone library.

Quinolinones have various biological activities, including antibacterial, anticancer, and antiviral properties. The 3-substituted amide quinolin-2(1H)-ones not only show antibacterial activity, but also act as immunomodulators, 5-HT4 receptor agonists, cannabinoid receptor inverse agonists, and AchE and, BuchE inhibitors. To investigate the potent biological activity of 3-substituted amide quinolin-2(1H)-ones, a large number of 3,5-amide substituted-2-oxoquinolinones were prepared by parallel solid-phase synthesis. The compound 5-amino-1-(4-methoxybenzyl)-2-oxo-1,2-dihydroquinoline-3-carboxylic acid was loaded onto 4-formyl-3,5-dimethoxyphenoxy (PL-FDMP) resin by reductive amination with high efficiency. Various building blocks were attached to the 3 and 5 positions to yield 3,5-disubstituted-2-oxoquinolinones with high purity and good yield. The ability some of these compound to inhibit the release of IL-1β, a cytokine involved in the immune response was measured, and they showed about 50% inhibition at 10 μM.

Nanoparticle-encapsulated P2X7 receptor antagonist in a pH-sensitive polymer as a potential local drug delivery system to acidic inflammatory environments

We have developed nanoparticles of anti-inflammatory P2X7 receptor antagonist encapsulated in a pH-sensitive polymer, poly(tetrahydropyran-2-yl methacrylate) (poly(THPMA)), as a potential local drug delivery system to target to acidic inflammatory environments, in which P2X7 receptors are implicated in the pathology of inflammation via the activation of immune cells. The nanoparticles were prepared using single emulsion methods, also their size and shape were confirmed by microscopy and spectroscopy, etc. The profiles of the pH-dependent degradation, release of antagonist and biological activities were investigated. The nanoparticles that encapsulated the 3,5-dichloropyridine derivative (2) with poly(THPMA), were observed to be more slowly cleaved than the blank nanoparticles. Moreover, the free P2X7 receptor antagonists potently inhibited the receptor activation, whereas the nanoparticles of the 3,5-dichloropyridine derivative (2) encapsulated poly(THPMA) exhibited much lower P2X7 antagonistic activity through sustained encapsulation. Thus, the nanoparticles of the 3,5-dichloropyridine derivative (2) encapsulated poly(THPMA) may be utilized to develop a pH-sensitive local drug delivery system for controlled release of anti-inflammatory therapeutics in acidic physiological environments.

Escape from adamantane: Scaffold optimization of novel P2X7 antagonists featuring complex polycycles

The adamantane scaffold, despite being widely used in medicinal chemistry, is not devoid of problems. In recent years we have developed new polycyclic scaffolds as surrogates of the adamantane group with encouraging results in multiple targets. As an adamantane scaffold is a common structural feature in several P2X7 receptor antagonists, herein we report the synthesis and pharmacological evaluation of multiple replacement options of adamantane that maintain a good activity profile. Molecular modeling studies support the binding of the compounds to a site close to the central pore, rather than to the ATP-binding site and shed light on the structural requirements for novel P2X7 antagonists.

Structure-activity relationship studies of pyrimidine-2,4-dione derivatives as potent P2X7 receptor antagonists.

As an optimization strategy, the flexible structure of KN-62, a known P2X7 receptor antagonist, was converted into conformationally constrained derivatives using pyrimidine-2,4-dione as the core skeleton. Various modifications at the 4-position of the piperazine moiety of the new lead compound were performed to improve P2X7 receptor antagonistic activities, which were evaluated in HEK293 cells stably expressing the human P2X7 receptor (EtBr uptake assay) and in THP-1 cells (IL-1β ELISA assay). According to the results, polycycloalkyl acyl or di-halogenated benzoyl substituents were much more favorable than the original phenyl group of KN-62. Among these compounds, the trifluoromethyl-chloro benzoyl derivative 18 m and adamantyl carbonyl derivatives 19 g-19 i and 19k showed potent antagonistic effects, with IC50 values ranging from 10 to 30 nM. In addition, the in vitro adsorption, distribution, metabolism, excretion, and toxicity (ADMET) profile of 18 m was determined to be in acceptable ranges in terms of metabolic stability and cytotoxicity. These results suggest that pyrimidine-2,4-dione derivatives may be promising novel P2X7 receptor antagonists for the development of anti-inflammatory drugs.

Towards a Novel Class of Multitarget-Directed Ligands: Dual P2X7-NMDA Receptor Antagonists

Multi-target-directed ligands (MTDLs) offer new hope for the treatment of multifactorial complex diseases such as Alzheimer’s Disease (AD). Herein, we present compounds aimed at targeting the NMDA and the P2X7 receptors, which embody a different approach to AD therapy. On one hand, we are seeking to delay neurodegeneration targeting the glutamatergic NMDA receptors; on the other hand, we also aim to reduce neuroinflammation, targeting P2X7 receptors. Although the NMDA receptor is a widely recognized therapeutic target in treating AD, the P2X7 receptor remains largely unexplored for this purpose; therefore, the dual inhibitor presented herein—which is open to further optimization—represents the first member of a new class of MTDLs.

Discovery of Potent Antiallodynic Agents for Neuropathic Pain Targeting P2X3 Receptors

Antagonism of the P2X3 receptor is one of the potential therapeutic strategies for the management of neuropathic pain because P2X3 receptors are predominantly localized on small to medium diameter C- and Aδ-fiber primary afferent neurons, which are related to the pain-sensing system. In this study, 5-hydroxy pyridine derivatives were designed, synthesized, and evaluated for their in vitro biological activities by two-electrode voltage clamp assay at hP2X3 receptors. Among the novel hP2X3 receptor antagonists, intrathecal treatment of compound 29 showed parallel efficacy with pregabalin (calcium channel modulator) and higher efficacy than AF353 (P2X3 receptor antagonist) in the evaluation of its antiallodynic effects in spinal nerve ligation rats. However, because compound 29 was inactive by intraperitoneal administration in neuropathic pain animal models due to low cell permeability, the corresponding methyl ester analogue, 28, which could be converted to compound 29 in vivo, was investigated as a prodrug concept. Intravenous injection of compound 28 resulted in potent antiallodynic effects, with ED50 values of 2.62 and 2.93 mg/kg in spinal nerve ligation and chemotherapy-induced peripheral neuropathy rats, respectively, indicating that new drug development targeting the P2X3 receptor could be promising for neuropathic pain, a disease with high unmet medical needs.

Discovery and structure-activity relationship studies of quinolinone derivatives as potent IL-2 suppressive agents.

The quinolinone skeleton has been utilized to develop various mechanism-based immune modulators. However, the effects of quinolinone derivatives on the release of T cell-associated interleukin-2 (IL-2) have not been established. In this study, a series of novel quinolinone derivatives was synthesized, and their immunosuppressive activity was evaluated by measuring suppression of IL-2 release from activated Jurkat T cells. Optimizing the three side chains around the quinolinone skeleton revealed the most active compound: 11l. This compound exhibits potent inhibitory activity toward IL-2 release in both 12-o-tetradecanoylphorbol-13-acetate (PMA)/A23187 (ionomycin) (IC50=80±10nM) and anti-CD3/CD28-stimulated Jurkat T cells (83% inhibition at 10μM) without cytotoxic effects. Further investigation into the underlying mechanism of 11l indicated the suppression of NF-κB and nuclear factor of activated T cells (NFAT) promoter activities in Jurkat T cells.