Human Renal Carcinoma A498 Cells Research Articles

Characterization and apoptotic effect of copper nanoparticles biosynthesized from Ziziphus zizyphus leaf on human renal cell carcinoma A498 cells

Biological formulation of copper nanoparticles (Cu-Nps) in cancer treatment has augmented due to its beneficial and curative effects. Ziziphus zizyphus (ZZ) is a species of medicinal plant traditionally used for various ailments. Hence, this work deals with formulation of copper nanoparticles (Cu-NP’s) from Z. zizyphus leaf extracts (Cu-ZZ NP’s) and examine its effect on human renal carcinoma A498 cells. The characterization of the formulated Cu-ZZ NPs was done by spectroscopic and microscopic methods. The anti-cancer tests (MTT, ROS, MMP, Hoechst staining, comet, Real‐time PCR, and western blot) were used to evaluate the biological properties of Cu-ZZ NPs on the growth and proliferation of A498 cells. The cytotoxic activity of Cu-ZZ NPs resulted in potent toxicity of A498 cells. Further, Cu-ZZ NP’s triggered an intracellular ROS generation through mitochondrial pathways in the A498 cells, revealing a noteworthy cytotoxicity activity. Also, fluorescent microscopic studies using Hoechst staining, and comet analysis confirmed the occurrence of apoptosis in Cu-ZZ NPs treated A498 cells. Following, quantitative RT-PCR presented a negative regulation of Bcl-2 expression and positive regulation of Bid, Bax, and caspases (3 and 9) gene expression in Cu-ZZ NP’s treated A498 cells. To further justify, inhibition of mTOR/PI3K/AKT protein expression by Cu-ZZ NPs in A498 cells, lead to an inference that Cu-ZZ NPs induce apoptosis in A498 cells by inhibiting the mTOR, PI3K/Akt pathway.

YC‐1 induces apoptosis of human renal carcinoma A498 cells in vitro and in vivo through activation of the JNK pathway

The aim of this study was to elucidate the mechanism of YC-1{3-(5'-hydroxy methyl-2'-furyl)-1-benzylindazole}-induced human renal carcinoma cells apoptosis and to evaluate the potency of YC-1 in models of tumour growth in mice. YC-1-mediated apoptosis was assessed by analysis of MTT, SRB, DAPI staining and flow cytometry analysis. Knockdown of JNK protein was achieved by transient transfection using siRNA. The mechanisms of action of YC-1 on different signalling pathways involved were studied using western blot. Fas clustering was analysed by confocal microscopy and in vivo efficacy was examined in a A498 xenograft model. YC-1 displayed cytotoxicity in renal carcinoma cells at 10(-7)-10(-8) M. Increased condensation of chromatin was observed and an increase in the cell population in subG1 phase. Moreover, YC-1 triggered mitochondria-mediated and caspase-dependent pathways. YC-1 significantly induced Fas ligand expression, but did not modify either the protein levels of death receptors or ligands. In addition, Fas clustering in cells responsive to YC-1 was observed, suggesting involvement of a Fas-mediated pathway. Furthermore, YC-1 markedly induced phosphorylation of JNK and a JNK inhibitor, SP600125, and siRNA JNK1/2 significantly reversed YC-1-induced cytotoxicity and protein expression. We suggest that YC-1 induced JNK phosphorylation, the upregulation of FasL and Fas receptor clustering to promote the activation of caspases 8 and 3, resulting in apoptosis. Finally, we demonstrated the antitumour effect of YC-1 in vivo. These data suggest that YC-1 is a good candidate for development as an anticancer drug.

Enhanced antitumor activity of combinations of free and HPMA copolymer-bound drugs

The synergism in anticancer effect toward human renal carcinoma A498 cells by binary combinations of free and N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-bound anticancer drugs, SOS thiophene (SOS), doxorubicin (DOX), and mesochlorin e 6 monoethylenediamine (Mce 6), was evaluated. The combination index (CI) analysis was used to quantify the synergism, antagonism, and additive effects. Both free drugs and HPMA copolymer conjugates, when used as single agents or in combination, exhibited cytotoxic activities against A498 cells, as determined using a modified MTT assay. As single agents, SOS and P-GFLG-SOS (HPMA copolymer conjugates containing SOS bound via glycylphenylalanylleucylglycine [GFLG] spacer) were significantly more effective than the other agents evaluated. The synergistic effects ranked in the order SOS + DOX > P-GFLG-DOX + P-GFLG-Mce 6 ≈ DOX + Mce 6 > P-GFLG-SOS + P-GFLG-DOX ≈ SOS + Mce 6 > P-GFLG-SOS + P-GFLG-Mce 6. The combination of SOS + DOX proved to be synergistic over all cell growth inhibition levels. All other combinations exhibited synergism in a wide range of drug effect levels. The SOS + Mce 6 and P-GFLG-SOS + P-GFLG-Mce 6 combinations displayed synergism up to drug affected fraction ( f a) values of about 0.8 and reached slight antagonism and nearly additivity at f a = 0.95, respectively. However, all other combinations were synergistic up to f a < 0.9 and were additive at higher f a values. The observations that most combinations produced synergistic effects will be important for clinical translation.

D-501036, a novel selenophene-based triheterocycle derivative, exhibits potentin vitroandin vivoantitumoral activity which involves DNA damage and ataxia telangiectasia–mutated nuclear protein kinase activation

D-501036 [2,5-bis(5-hydroxymethyl-2-selenienyl)-3-hydroxymethyl-N-methylpyrrole] is herein identified as a novel antineoplastic agent with a broad spectrum of antitumoral activity against several human cancer cells and an IC(50) value in the nanomolar range. The IC(50) values for D-501036 in the renal proximal tubule, normal bronchial epithelial, and fibroblast cells were >10 mumol/L. D-501036 exhibited no cross-resistance with vincristine- and paclitaxel-resistant cell lines, whereas a low level of resistance toward the etoposide-resistant KB variant was observed. Cell cycle analysis established that D-501036 treatment resulted in a dose-dependent accumulation in S phase with concomitant loss of both the G(0)-G(1) and G(2)-M phase in both Hep 3B and A-498 cells. Pulsed-field gel electrophoresis showed D-501036-induced, concentration-dependent DNA breaks in both Hep 3B and A-498 cells. These breaks did not involve interference with either topoisomerase-I and topoisomerase-II function or DNA binding. Rapid reactive oxygen species production and formation of Se-DNA adducts were evident following exposure of cells to D-501036, indicating that D-501036-mediated DNA breaks were attributable to the induction of reactive oxygen species and DNA adduct formation. Moreover, D-501036-induced DNA damage activated ataxia telangiectasia-mutated nuclear protein kinase, leading to hyperphosphorylation of Chk1, Chk2, and p53, decreased expression of CDC25A, and up-regulation of p21(WAF1) in both p53-proficient and p53-deficient cells. Collectively, the results indicate that D-501036-induced cell death was associated with DNA damage-mediated induction of ataxia telangiectasia-mutated activation, and p53-dependent and -independent apoptosis pathways. Notably, D-501036 shows potent activity against the growth of xenograft tumors of human renal carcinoma A-498 cells. Thus, D-501036 is a promising anticancer compound that has strong potential for the management of human cancers.