ECDD-S16 targets vacuolar ATPase: A potential inhibitor compound for pyroptosis-induced inflammation.

BACKGROUND: Cleistanthin A (CA), extracted from Phyllanthus taxodiifolius Beille, was previously reported as a potential V-ATPase inhibitor relevant to cancer cell survival. In the present study, ECDD-S16, a derivative of cleistanthin A, was investigated and found to interfere with pyroptosis induction via V-ATPase inhibition. OBJECTIVE: This study examined the ability of ECDD-S16 to inhibit endolysosome acidification leading to the attenuation of pyroptosis in Raw264.7 macrophages activated by both surface and endosomal TLR ligands. METHODS: To elucidate the activity of ECDD-S16 on pyroptosis-induced inflammation, Raw264.7 cells were pretreated with the compound before stimulation with surface and endosomal TLR ligands. The release of lactate dehydrogenase (LDH) was determined by LDH assay. Additionally, the production of cytokines and the expression of pyroptosis markers were examined by ELISA and immunoblotting. Moreover, molecular docking was performed to demonstrate the binding of ECDD-S16 to the vacuolar (V-)ATPase. RESULTS: This study showed that ECDD-S16 could inhibit pyroptosis in Raw264.7 cells activated with surface and endosomal TLR ligands. The attenuation of pyroptosis by ECDD-S16 was due to the impairment of endosome acidification, which also led to decreased Reactive Oxygen Species (ROS) production. Furthermore, molecular docking also showed the possibility of inhibiting endosome acidification by the binding of ECDD-S16 to the vacuolar (V-)ATPase in the region of V0. CONCLUSION: Our findings indicate the potential of ECDD-S16 for inhibiting pyroptosis and prove that vacuolar H+ ATPase is essential for pyroptosis induced by TLR ligands.

Rational Design and Lead Optimisation of Potent Antimalarial Quinazolinediones and Their Cytotoxicity against MCF-7.

Quinazolinedione is one of the most outstanding heterocycles in medicinal chemistry thanks to its wide ranges of biological activities including antimalarial, anticancer, and anti-inflammatory. TCMDC-125133 containing a quinazolinedione pharmacophore displays promising antimalarial activity and low toxicity, as described in the GlaxoSmithKline (GSK) report. Herein, the design and synthesis of novel quinazolinedione derivatives is described on the basis of our previous work on the synthesis of TCMDC-125133, where low-cost chemicals and greener alternatives were used when possible. The initial SAR study focused on the replacement of the valine linker moiety; according to the in silico prediction using SwissADME, concise four-step syntheses toward compounds 4-10 were developed. The in-house synthesized compounds 4-10 were assayed for antimalarial activity against P. falciparum 3D7, and the result revealed that only the compound 2 containing a valine linker was tolerated. Another round of lead optimization focused on the replacement of the m-anisidine moiety in compound 2. A library of 12 derivatives was prepared, and the antimalarial assay showed that potent antimalarial activity could be maintained by replacing the methoxy group in the meta position of the phenyl side chain with a fluorine or chlorine atom (21: IC50 = 36 ± 5 nM, 24: IC50 = 22 ± 5 nM). Further lead optimization is underway to enhance the antimalarial activity of this class of compound. The compounds included in the study possess little to no antiproliferative activity against MCF-7 cells.

Phyllanthus taxodiifolius Beille Disrupted N-cadherin, Vimentin, Paxillin and Actin Stress Fibers in Glioblastoma.

OBJECTIVE: Glioblastoma is the most aggressive and lethal brain tumor in adults with highly invasive properties. In this present study, we explored the effects of Phyllanthus taxodiifolius Beille extract on molecules known to be hallmarks of aggressive glioblastoma including N-cadherin and vimentin, mesenchymal markers, as well as paxillin, a major adaptor protein that regulates the linking of focal adhesions to the actin cytoskeleton. METHODS: P. taxodiifolius were air-dried, powdered and percolated with methanol, filtered, concentrated and lyophilized to yield a crude methanol extract. C6 glioblastoma cell line was used in this study. The expression of N-cadherin and vimentin, as well as the activation of paxillin was determined using Western blot analysis. The effect of the extract on focal adhesions and actin cytoskeleton were investigated using immunofluorescence staining and confocal imaging. RESULTS: In the presence of 40 µg/ml Phyllanthus taxodiifolius Beille extract, the expression of N-cadherin and vimentin were significantly decreased (p<0.001 and p<0.05, respectively). Activation of paxillin was also diminished as indicated by a reduction of phosphorylated-paxillin (p<0.01). Consequently, actin stress fibers in glioblastoma cells were abolished as evidenced by the decrease in focal adhesion (p<0.001) and stress fibers numbers (p<0.001). CONCLUSION: Our study demonstrates for the first time that P. taxodiifolius interferes with multiple key molecules related to pathological hallmarks of glioblastoma. These molecules are involved with cell contacts, focal adhesions, and the formation and stabilization of actin stress fibers, which are required for glioblastoma metastatic behavior. These results provide further evidence supporting the potential of P. taxodiifolius and its bioactive compounds as anti-cancer agents.

Pharmacological Effects of Panduratin A on Renal Cyst Development in In Vitro and In Vivo Models of Polycystic Kidney Disease.

Renal cyst expansion in polycystic kidney disease (PKD) involves abnormalities in both cyst-lining-cell proliferation and fluid accumulation. Suppression of these processes may retard the progression of PKD. Evidence suggests that the activation of 5' AMP-activated protein kinase (AMPK) inhibits cystic fibrosis transmembrane conductance regulator (CFTR)-mediated chloride secretion, leading to reduced progression of PKD. Here we investigated the pharmacological effects of panduratin A, a bioactive compound known as an AMPK activator, on CFTR-mediated chloride secretion and renal cyst development using in vitro and animal models of PKD. We demonstrated that AMPK was activated in immortalized normal renal cells and autosomal dominant polycystic kidney disease (ADPKD) cells following treatment with panduratin A. Treatment with panduratin A reduced the number of renal cyst colonies corresponding with a decrease in cell proliferation and phosphorylated p70/S6K, a downstream target of mTOR signaling. Additionally, panduratin A slowed cyst expansion via inhibition of the protein expression and transport function of CFTR. In heterozygous Han:Sprague-Dawley (Cy/+) rats, an animal model of PKD, intraperitoneal administration of panduratin A (25 mg/kg BW) for 5 weeks significantly decreased the kidney weight per body weight ratios and the cystic index. Panduratin A also reduced collagen deposition in renal tissue. Intraperitoneal administration of panduratin A caused abdominal bleeding and reduced body weight. However, 25 mg/kg BW of panduratin A via oral administration in the PCK rats, another non-orthologous PKD model, showed a significant decrease in the cystic index without severe adverse effects, indicating that the route of administration is critical in preventing adverse effects while still slowing disease progression. These findings reveal that panduratin A might hold therapeutic properties for the treatment of PKD.

Pinostrobin inhibits renal CFTR-mediated Cl- secretion and retards cyst growth in cell-derived cyst and polycystic kidney disease rats.

Cystic fibrosis transmembrane conductance regulator (CFTR) plays crucial role in renal cyst expansion via increase in fluid accumulation. Inhibition of CFTR has been proposed to retard cyst development and enlargement in polycystic kidney disease (PKD). Pinostrobin, a bioactive natural flavonoid, possesses several pharmacological effects. The present study investigated pharmacological effects of pinostrobin on CFTR-mediated Cl- secretion and renal cyst expansion in in vitro and in vivo models. Pinostrobin (10 and 50 µM) reduced number of MDCK cell-derived cyst colonies and inhibited cyst expansion via inhibition of cell proliferation and CFTR-mediated Cl- secretion. The inhibitory effect of pinostrobin was not due to the decrease in cell viability and activity of Na+-K+-ATPase. We also investigated the natural analogue pinocembrin as well as the synthetic analogue pinostrobin oxime. Both pinocembrin and pinostrobin oxime did not reduce CFTR-mediated Cl- secretion. In PKD rats, oral administration of pinostrobin (40 mg/kg/day) exhibited a decreasing in cystic area compared to vehicle-treated rats. Pinostrobin treatment inhibited renal expression of CFTR protein in PKD rats. Our findings highlighted the potential therapeutic application of pinostrobin in PKD.

Panduratin A Derivative Protects against Cisplatin-Induced Apoptosis of Renal Proximal Tubular Cells and Kidney Injury in Mice.

BACKGROUND: Panduratin A is a bioactive cyclohexanyl chalcone exhibiting several pharmacological activities, such as anti-inflammatory, anti-oxidative, and anti-cancer activities. Recently, the nephroprotective effect of panduratin A in cisplatin (CDDP) treatment was revealed. The present study examined the potential of certain compounds derived from panduratin A to protect against CDDP-induced nephrotoxicity. METHODS: Three derivatives of panduratin A (DD-217, DD-218, and DD-219) were semi-synthesized from panduratin A. We investigated the effects and corresponding mechanisms of the derivatives of panduratin A for preventing nephrotoxicity of CDDP in both immortalized human renal proximal tubular cells (RPTEC/TERT1 cells) and mice. RESULTS: Treating the cell with 10 µM panduratin A significantly reduced the viability of RPTEC/TERT1 cells compared to control (panduratin A: 72% ± 4.85%). Interestingly, DD-217, DD-218, and DD-219 at the same concentration did not significantly affect cell viability (92% ± 8.44%, 90% ± 7.50%, and 87 ± 5.2%, respectively). Among those derivatives, DD-218 exhibited the most protective effect against CDDP-induced renal proximal tubular cell apoptosis (control: 57% ± 1.23%; DD-218: 19% ± 10.14%; DD-219: 33% ± 14.06%). The cytoprotective effect of DD-218 was mediated via decreases in CDDP-induced mitochondria dysfunction, intracellular reactive oxygen species (ROS) generation, activation of ERK1/2, and cleaved-caspase 3 and 7. In addition, DD-218 attenuated CDDP-induced nephrotoxicity by a decrease in renal injury and improved in renal dysfunction in C57BL/6 mice. Importantly, DD-218 did not attenuate the anti-cancer efficacy of CDDP in non-small-cell lung cancer cells or colon cancer cells. CONCLUSIONS: This finding suggests that DD-218, a derivative of panduratin A, holds promise as an adjuvant therapy in patients receiving CDDP.

Protective Effect of Panduratin A on Cisplatin-Induced Apoptosis of Human Renal Proximal Tubular Cells and Acute Kidney Injury in Mice.

BACKGROUND: Cisplatin is an effective chemotherapy but its main side effect, acute kidney injury, limits its use. Panduratin A, a bioactive compound extracted from Boesenbergia rotunda, shows several biological activities such as anti-oxidative effects. The present study investigated the nephroprotective effect of panduratin A on cisplatin-induced renal injury. METHODS: We investigated the effect of panduratin A on the toxicity of cisplatin in both mice and human renal cell cultures using RPTEC/TERT1 cells. RESULTS: The results demonstrated that panduratin A ameliorates cisplatin-induced renal toxicity in both mice and RPTEC/TERT1 cells by reducing apoptosis. Mice treated with a single intraperitoneal (i.p.) injection of cisplatin (20 mg/kg body weight (BW)) exhibited renal tubule injury and impaired kidney function as shown by histological examination and increased serum creatinine. Co-administration of panduratin A (50 mg/kg BW) orally improved kidney function and ameliorated renal tubule injury of cisplatin by inhibiting activation of extracellular signal-regulated kinase (ERK)1/2 and caspase 3. In human renal proximal tubular cells, cisplatin induced cell apoptosis by activating pro-apoptotic proteins (ERK1/2 and caspase 3), and reducing the anti-apoptotic protein (Bcl-2). These effects were significantly ameliorated by co-treatment with panduratin A. Interestingly, panduratin A did not alter intracellular accumulation of cisplatin. It did not alter the anti-cancer efficacy of cisplatin in either human colon or non-small cell lung cancer cell lines. CONCLUSIONS: The present study highlights panduratin A has a potential protective effect on cisplatin's nephrotoxicity.

High-content screening of Thai medicinal plants reveals Boesenbergia rotunda extract and its component Panduratin A as anti-SARS-CoV-2 agents.

Since December 2019, the emergence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has caused severe pneumonia, a disease named COVID-19, that became pandemic and created an acute threat to public health. The effective therapeutics are in urgent need. Here, we developed a high-content screening for the antiviral candidates using fluorescence-based SARS-CoV-2 nucleoprotein detection in Vero E6 cells coupled with plaque reduction assay. Among 122 Thai natural products, we found that Boesenbergia rotunda extract and its phytochemical compound, panduratin A, exhibited the potent anti-SARS-CoV-2 activity. Treatment with B. rotunda extract and panduratin A after viral infection drastically suppressed SARS-CoV-2 infectivity in Vero E6 cells with IC50 of 3.62 µg/mL (CC50 = 28.06 µg/mL) and 0.81 µΜ (CC50 = 14.71 µM), respectively. Also, the treatment of panduratin A at the pre-entry phase inhibited SARS-CoV-2 infection with IC50 of 5.30 µM (CC50 = 43.47 µM). Our study demonstrated, for the first time, that panduratin A exerts the inhibitory effect against SARS-CoV-2 infection at both pre-entry and post-infection phases. Apart from Vero E6 cells, treatment with this compound was able to suppress viral infectivity in human airway epithelial cells. This result confirmed the potential of panduratin A as the anti-SARS-CoV-2 agent in the major target cells in human. Since B. rotunda is a culinary herb generally grown in China and Southeast Asia, its extract and the purified panduratin A may serve as the promising candidates for therapeutic purposes with economic advantage during COVID-19 situation.

Phyllanthus taxodiifolius Beille suppresses microtubule dynamics and restricts glioblastoma aggressiveness.

Glioblastoma is the most common and the most malignant form of brain tumor. This devastating tumor results in death within a year after diagnosis. Although the tumor mass can be surgically removed, glioma cells invade other areas in the brain leading to tumor recurrence and poor prognosis. Therefore, new agents that can overcome cancer cell invasion are urgently required. Phyllanthus taxodiifolius Beille (P. taxodiifolius), has been reported to have potent anti-cancer activities. However, its effects on glioblastoma cells and its underlying mechanisms have never been revealed. Here we investigated the effect and underlying mechanisms of P. taxodiifolius extract on aggressive properties of the glioblastoma, including adhesion, migration, and invasion. P. taxodiifolius extract disrupted adhesion, delayed migration and interfered with the invasion of glioblastoma cells. In addition, the extract suppressed microtubule dynamics as shown by live imaging of a microtubule plus tip protein and decreased focal adhesion by decreasing focal adhesion kinase activity. Our study is the first evidence showing that P. taxodiifolius extract suppresses invasive properties of glioblastoma cells by disrupting microtubule structure and interfering with microtubule dynamics, suggesting the possibility to further develop P. taxodiifolius and its bioactive compounds as an anti-cancer drug targeting microtubules in glioblastoma.