Induction of apoptosis in human retinoblastoma cells by topoisomerase inhibitors

PURPOSE:To examine the apoptotic effect induced in human retinoblastoma Y79 cells by camptothecin, etoposide, and amsacrine, to examine the effect of these drugs on the expression of many apoptosis-related modulators, and to test the antiapoptotic effect exerted by insulin-like growth factor-I (IGF-I). METHODS:Morphologic features of apoptosis were demonstrated using acridine orange- ethidium bromide staining and electron microscopy. DNA fragmentation was determined by means of an in situ cell detection procedure (TdT-dUTP terminal nick-end labeling [TUNEL]) or by electrophoresis on agarose gels and was quantified by enzyme-linked immunosorbent assay. The expression of apoptosis-related mod…

The apoptotic effects and synergistic interaction of sodium butyrate and MG132 in human retinoblastoma Y79 cells

This study deals with the apoptotic effect exerted on human retinoblastoma Y79 cells by both sodium butyrate and an inhibitor of 26S proteasome [z-Leu-Leu-Leu-CHO (MG132)] and their synergistic effect. Exposure to sodium butyrate (1-4 mM) induced an accumulation of cells in the G2-M phase that was already visible after 24 h of treatment, when morphological and biochemical signs of apoptosis appeared only in a small number of cells (5-10%). Thereafter, the apoptotic effects increased progressively with slow kinetics, reaching a maximum after 72 h of exposure, when they concerned a large fraction of cells (>75% with 4 mM sodium butyrate). Sodium butyrate stimulated the conversion of procaspas…

Increased cyclin E level in retinoblastoma cells during programmed cell death

Identification and synthesis of insulin in human retinoblastoma Y-79 cells

Non-canonical roles of caspase-8 in MDA-MB-231 breast cancer cell line

Caspase-8 (casp-8) is well known as an initiator caspase involved in cell death signalling, although its activity in many cancer cell types seems to work under non-apoptotic conditions. Moreover, in several types of cancer, casp-8 is only rarely mutated and often its expression is very elevated. Since cancer cell growth also depends on evasion of apoptosis, the upregulation of casp-8 in tumours may suggest one or more non-apoptotic roles (1). Here we report our recent studies carried out in MDA-MB-231 cells, derived from clinically aggressive forms of Triple-Negative Breast Cancer, where we have assessed the non-canonical roles of casp-8. Firstly, we evaluated casp-8 mRNA and protein levels…

A‐1210477 sensitizes TRAIL-induced apoptosis in MDA-MB-231 Triple Negative Breast Cancer cells.

Triple negative breast cancer (TNBC) is a form of BC characterized by high aggressiveness, therapy resistance, short time to relapse, poor prognosis. The presence of Cancer Stem Cells (CSCs) could be responsible for TNBC resistance to therapy, recurrence and metastasis, and might explain the difficult of its eradication. Mcl-1 is one of the key regulators of CSCs self-renewal and its expression can limit the efficacy of antitumorigenic agents as TRAIL, a selective anticancer agent but with limited effects against some cancer cell lines. Here we investigated the expression profiles of Mcl-1 in TNBC tissue and cell lines. We also evaluated the effect of A-1210477, a selective Mcl-1 inhibitor,…

Let-7d-5p miRNA shows oncogenic functions in triple negative breast cancer

Triple negative breast cancer (TNBC) is a highly aggressive subtype of BC which lack of targeted therapies and is associated with poor prognosis. The presence of Cancer Stem Cells (CSCs) could be responsible for TNBC resistance to therapy, recurrence and metastasis, and might explain the difficult of its eradication. MiRNAs -a class of small non-coding RNAs- can modulate gene expression and their dysregulation may cause cancer formation. The let-7 family is dysregulated in various cancers and often its roles are unclear and of difficult interpretation. For example, let-7d can be over- or down-expressed and can act as tumor suppressor or oncogene. Here, we evaluated the expression profiles o…

Regulatory properties of nucleoside phosphotransferase from mucosa of chicken intestine

Nucleoside phosphotransferase from chicken intestinal mucosa is an associated multisubunit protein which can dissociate into components of lower molecular weight. The associated and the dissociated forms have the same substrate specificity but the first (A) shows a higher V(max) and a lower S(0.5) value than the second (B), whichever phosphate donor or nucleoside acceptor was employed. Moreover with form A the interaction coefficient in the Hill plots; as measured with a phosphate donor, varied: they always showed a higher result (about 2) than with form B (about 1). A brief preincubation at 37°C of form A modified the values both of the inactivation constant and of the enzyme kinetic param…

Apoptosis meets proteasome, an invaluable therapeutic target of anticancer drugs.

Purification of nucleoside phosphotransferase from mucosa of chicken intestine

The authors describe the purification procedure and some properties of a nucleoside phosphotransferase obtained from chicken intestinal mucosa, the tissue of which in preliminary studies showed the highest specific activity. Sepharose 6B chromatography and electrophoresis on polyacrylamide gel subdivided the enzyme into many forms which represent various levels of an associated multisubunit protein derived by assembly of a component at lower molecular weight. Nucleotide protectors regulate the equilibrium among these different forms, favoring the production and the stabilization of supramolecular complexes of nucleoside phosphotransferase. Similar results were obtained with enzyme purified …

Inhibitory effect of phosphate on nucleoside phosphotransferase from chicken intestinal mucosa

Nucleoside phosphotransferase purified from chicken intestinal mucosa is a multisubunit protein of high molecular weight regulated by many nucleotides. The adsorption of a nucleotide effector to regulatory sites favors the conversion of the enzyme to a stable form and induces a modification of the substrate site, increasing its affinity for substrate donors. We studied the effects exerted by orthophosphate on the reaction and on the stability of the enzyme. P(i) inhibits competitively nucleoside phosphotransferase activity, whichever nucleoside monophosphate is employed as donor. Nucleotide effector (d-TDP) removes entirely the inhibitory effect exerted by different concentrations of P(i). …