New quinoline/chalcone hybrids as anti-cancer agents: Design,synthesis, and evaluations of cytotoxicity and PI3K inhibitory activity

A series of quinoline-chalcone hybrids was designed as potential anti-cancer agents, synthesized and evaluated. Different cytotoxic assays revealed that compounds experienced promising activity. Compounds 9i and 9j were the most potent against all the cell lines tested with IC₅₀ = 1.91–5.29 µM against A549 and K-562 cells. Mechanistically, 9i and 9j induced G₂/M cell cycle arrest and apoptosis in both A549 and K562 cells. Moreover, all PI3K isoforms were inhibited non selectively with IC₅₀s of 52–473 nM when tested against the two mentioned compounds with 9i being most potent against PI3K-γ (IC₅₀ = 52 nM). Docking of 9i and 9j showed a possible formation of H-bonding with essential valine residues in the active site of PI3K-γ isoform. Meanwhile, Western blotting analysis revealed that 9i and 9j inhibited the phosphorylation of PI3K, Akt, mTOR, as well as GSK-3β in both A549 and K562 cells, suggesting the correlation of blocking PI3K/Akt/mTOR pathway with the above antitumor activities. Together, our findings support the antitumor potential of quinoline-chalcone derivatives for NSCLC and CML by inhibiting the PI3K/Akt/mTOR pathway.     

Sesquiterpenoids from the roots of Inula helenium inhibit acute myelogenous leukemia progenitor cells

One new eudesmane sesquiterpenoid, 11β-hydroxy-13-chloro-eudesm-5-en-12, 8-olide (1), was isolated from the roots of Inula helenium together with nine eudesmanolides (2–10) and one germacranolide (11). Their structures were elucidated on the basis of detailed spectroscopic analyses. All isolates were evaluated for their antiproliferative activities against human leukemia stem-like cell line KG1a. Compound 10 exhibited the most potent effect with the IC₅₀ value of 3.36 ± 0.18 μM. A further investigation revealed that compound 10 could significantly induce apoptosis of KG1a cells. Additionally, compound 10 had an obvious effect on the levels of apoptosis-related proteins (Bcl-2, Bax, cytochrome c, caspase 9 and caspase 3), indicating that the antiproliferative effect of compound 10 on KG1a cells might be mediated through a mitochondria-dependent apoptotic pathway.     

Synthesis,anticancer effect and molecular modeling of new thiazolylpyrazolyl coumarin derivatives targeting VEGFR-2 kinase and inducing cell cycle arrest and apoptosis

New thiazolylpyrazolyl coumarin derivatives were synthesized and tested for their anticancer potential in vitro against five different human cell lines, including breast MCF-7, lung A549, prostate PC3, liver HepG2 and normal melanocyte HFB4. Breast carcinoma revealed higher sensitivity towards compounds 7a, 8c, 9b, 9c and 9d with IC₅₀ values ranging from 5.41 to 10.75 μM in comparison to the reference drug doxorubicin (IC₅₀ = 6.73 μM). In addition, no noticeable toxicity was exhibited towards normal cells HFB4. Moreover, in vitro studies of the VEGFR-2 inhibition in human breast cancer MCF-7 cell line for the promising cytotoxic compounds showed that compounds 7a, 8c, 9b, 9c and 9d were potent inhibitors at low micromolar concentrations (IC₅₀ = 0.034–0.582 μM) compared to the reference drug, sorafenib (IC₅₀ = 0.019 μM). Several theoretical and experimental studies were done to reveal the molecular mechanisms that control breast carcinoma metastasis. The mechanistic effectiveness in cell cycle progression, apoptotic induction and gene regulation were assessed for the promising compound 9d due to its remarkable cytotoxic activity against MCF-7 and significant VEGFR-2 inhibition. Flow cytometeric analysis showed that compound 9d induced cell growth cessation at G2/M phase and increased the percentage of cells at pre-G1 phase that stimulates the apoptotic death of MCF-7 cells. Furthermore, real time PCR assay illustrated that compound 9d up regulated p53 gene expression and elevated Bax/Bcl-2 ratio which confirmed the mechanistic pathway of compound 9d. Moreover, the apoptotic induction of breast cancer cells MCF-7 was enhanced effectively through activation of caspases-7 and 9 by compound 9d. On the other hand, a set of in silico methods such as molecular docking, molecular dynamics simulation, QSAR analysis as well as ADMET analysis was performed in order to study the protein-ligand interactions and the relationship between the physicochemical properties and the inhibitory activity of the promising compounds 7a, 8c and 9d. Based on the aforementioned findings, compound 9d could be considered as effective apoptosis modulator and promising lead for future development of new anti-breast cancer agents.     

Cytotoxic diterpenoids as potential anticancer agents from the twigs of Casearia kurzii

A search for bioactive natural products as anticancer lead compounds has led to the isolation of five new clerodane diterpenoids (1–5) from the twigs of Casearia kurzii. Their structures were elucidated by extensive analysis of their NMR, IR, and HRESIMS data, and the absolute configurations were determined by experimental and calculated electronic circular dichroism (ECD) data analysis. The isolates were biologically evaluated and showed cytotoxic activities toward human lung cancer cells (A549), human cervical cancer cells (HeLa), and human hepatocellular carcinoma cells (HepG2). The most active compound (5) with an IC₅₀ value of 5.3 μM against HeLa cells, was found to induce apoptosis and arrest the HeLa cell cycle at G0/G1 stage to exert cytotoxic effects.     

Design,synthesis and in vitro apoptotic mechanism of novel pyrrolopyrimidine derivatives

In this work we described the synthesis and evaluation of cytotoxic and apoptotic activity of novel pyrrolopyrimidine derivatives against A549, PC3 and MCF-7 cells. Among the synthesized compounds, 6b, 8a, 9a and 7a, 8b displayed the significant cytotoxic activities against A549 and PC3 cells with IC₅₀ value of 0.35, 1.48, 1.56 and 1.04, 1.89 µM, respectively. It was found that A549 cells were more sensitive to synthesized compounds than PC3 and MCF-7 cells. In order to evaluate the mechanism of cytotoxic activity in A549, compounds 6b, 8a and 9a were selected for further studies. Annexin V binding assay and western blot analysis results revealed that 6b, 8a and 9a induced apoptosis in A549 cells by intrinsic apoptotic pathway through the activation pro-apoptotic proteins such as Bim, Bax, Bak, Puma and deactivation of anti-apoptotic proteins including Bcl-2, Mcl-1 and Bcl-XL accompanied by the activation of caspase-3, caspase-9 and cleavage of PARP. Also, compounds 6b, 8a and 9a triggered apoptosis in HCT116 wt cells via activation of caspase-3 and caspase-9, but not in HCT116 Bax/Bak KO cells, indicating resistance to 6b, 8a and 9a treatment.     

Discovery of certain benzyl/phenethyl thiazolidinone-indole hybrids as potential anti-proliferative agents: Synthesis,molecular modeling and tubulin polymerization inhibition study

A series of certain benzyl/phenethyl thiazolidinone-indole hybrids were synthesized for the study of anti-proliferative activity against A549, NCI-H460 (lung cancer), MDA-MB-231 (breast cancer), HCT-29 and HCT-15 (colon cancer) cell lines by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). We found that compound G37 displayed highest cytotoxicity with IC₅₀ value of 0.92 ± 0.12 µM towards HCT-15 cancer cell line among all the synthesized compounds. Moreover, compound G37 was also tested on normal human lung epithelial cells (L132) and was found to be safe in contrast to HCT-15 cells. The lead compound G37 showed significant G2/M phase arrest in HCT-15 cells. Additionally, compound G37 significantly inhibited tubulin polymerization with IC₅₀ value of 2.92 ± 0.23 µM. Mechanistic studies such as acridine orange/ethidium bromide (AO/EB) dual staining, DAPI nuclear staining, annexinV/propidium iodide dual staining, clonogenic growth inhibition assays inferred that compound G37 induced apoptotic cell death in HCT-15 cells. Moreover, loss of mitochondrial membrane potential with elevated intracellular ROS levels was observed by compound G37. These compounds bind at the active pocket of the α/β-tubulin with higher number of stable hydrogen bonds, hydrophobic and arene-cation interactions confirmed by molecular modeling studies.     

Synthesis and biological evaluation of new bisindole-imidazopyridine hybrids as apoptosis inducers

A series of diindolylmethanes (5a-t) were designed, synthesized, and examined for their cytotoxicity against four human cancer cell lines like prostate (DU-145), lung (A549), breast (MCF-7) and cervical cancer (HeLa). These results revealed that among all the hybrids, two (5k and 5r) were identified and exhibited significant cytotoxic effect against A549 cancer cells with IC₅₀ values of 1.65 ± 0.3 and 1.80 ± 0.8 µM respectively. To investigate the reasons for the cytotoxic activity, the conventional biological assays were carried out with 5k and 5r on the A549 cancer cells. Both hybrids led to the arrest of A549 cell lines at the G2/M phase of the cell cycle and strongly induced apoptosis. Further the apoptotic effects of 5k and 5r were confirmed by ROS, annexin-V FITC, and mitochondrial membrane potential. Moreover, structure–activity relationships were elucidated with various substitutions on these hybrids.     

Deficiency of mouse mast cell protease 4 mitigates cardiac dysfunctions in mice after myocardium infarction

Mouse mast cell protease-4 (mMCP4) is a chymase that has been implicated in cardiovascular diseases, including myocardial infarction (MI). This study tested a direct role of mMCP4 in mouse post-MI cardiac dysfunction and myocardial remodeling. Immunoblot and immunofluorescent double staining demonstrated mMCP4 expression in cardiomyocytes from the infarct zone from mouse heart at 28 day post-MI. At this time point, mMCP4-deficient Mcpt4^?/? mice showed no difference in survival from wild-type (WT) control mice, yet demonstrated smaller infarct size, improved cardiac functions, reduced macrophage content but increased T-cell accumulation in the infarct region compared with those of WT littermates. mMCP4-deficiency also reduced cardiomyocyte apoptosis and expression of TGF-β1, p-Smad2, and p-Smad3 in the infarct region, but did not affect collagen deposition or α-smooth muscle actin expression in the same area. Gelatin gel zymography and immunoblot analysis revealed reduced activities of matrix metalloproteinases and expression of cysteinyl cathepsins in the myocardium, macrophages, and T cells from Mcpt4^?/? mice. Immunoblot analysis also found reduced p-Smad2 and p-Smad3 in the myocardium from Mcpt4^?/? mice, yet fibroblasts from Mcpt4^?/? mice showed comparable levels of p-Smad2 and p-Smad3 to those of WT fibroblasts. Flow cytometry, immunoblot analysis, and immunofluorescent staining demonstrated that mMCP4-deficiency reduced the expression of proapoptotic cathepsins in cardiomyocytes and protected cardiomyocytes from H₂O₂-induced apoptosis. This study established a role of mMCP4 in mouse post-MI dysfunction by regulating myocardial protease expression and cardiomyocyte death without significant impact on myocardial fibrosis or survival post-MI in mice.     

Apoptosis: A mammalian cell bioprocessing perspective

Apoptosis is a form of programmed and controlled cell death that accounts for the majority of cellular death in bioprocesses. Cell death affects culture longevity and product quality; it is instigated by several stresses experienced by the cells within a bioreactor. Understanding the factors that cause apoptosis as well as developing strategies that can protect cells is crucial for robust bioprocess development. This review aims to a) address apoptosis from a bioprocess perspective; b) describe the significant apoptotic mechanisms linking them to the most relevant stresses encountered in bioreactors; c) discuss the design of operating conditions in order to avoid cell death; d) focus on industrially relevant cell lines; and e) present anti-apoptosis strategies including cell engineering and model-based optimization of bioprocesses. In addition, the importance of apoptosis in quality-by-design bioprocess development from clone screening to production scale are highlighted.     

Downregulation of TIMP2 attenuates sepsis-induced AKI through the NF-κb pathway

Acute kidney injury (AKI) is a frequent complication of sepsis and contributes to increased morbidity and mortality. Urinary tissue inhibitor of metalloproteinases-2 (TIMP2) has been recently recognized as an early biomarker to predict AKI in critically ill patients. However, the biological functions of TIMP2 remain largely unknown. In this study, we investigated the role of TIMP2 in mediating inflammation and tubular cell apoptosis in AKI. In kidney tissue taken from mice exposed to cecal ligation and puncture (CLP) and in human kidney 2 (HK-2) cells exposed to lipopolysaccharide (LPS) in culture, TIMP2 expression was significantly upregulated. The expression of TIMP2 in the kidney tissue correlated with the severity of AKI in vivo. In cultured HK-2 cells, LPS challenge markedly induced cytokine release, and recombinant cytokines promoted TIMP2 expression and apoptosis. However, TIMP2 silencing ameliorated LPS-induced cytokine release, apoptosis, and cell injury. We further found that the effects of downregulation of TIMP2 on a suppression of release of inflammatory cytokines were mediated by p-P65. Stable, kidney-specific TIMP2 knockdown mice were transduced by injecting the TIMP2 knockdown lentiviral vector into kidney parenchyma. TIMP2 silencing ameliorated CLP-induced proinflammatory cytokines, kidney dysfunction as measured by serum creatinine level, and histopathological changes. Downregulation of TIMP2 showed renoprotective effects on endotoxin-induced AKI, which was associated with the anti-inflammatory activity through inhibition of the nuclear factor (NF)-κB pathway. Collectively, our results indicate that TIMP2 plays an important role in mediating sepsis-induced AKI through regulation of NF-κB. These findings reveal the pathogenic role of TIMP2 in AKI and suggest a novel target for the treatment of AKI.