Biomedical subjects

Hamid Morjani

Publications and source records attributed to Hamid Morjani.

15 recordsLinked to original sources

PubMed2006

Telomestatin-induced telomere uncapping is modulated by POT1 through G-overhang extension in HT1080 human tumor cells.

Telomestatin is a potent G-quadruplex ligand that interacts with the 3' telomeric overhang, leading to its degradation, and induces a delayed senescence and apoptosis of cancer cells. POT1 and TRF2 were recently identified as specific telomere-binding proteins involved in telomere capping and t-loop maintenance and whose interaction with telomeres is modulated by telomestatin. We show here that the treatment of HT1080 human tumor cells by telomestatin induces a rapid decrease of the telomeric G-overhang and of the double-stranded telomeric repeats. Telomestatin treatment also provokes a strong decrease of POT1 and TRF2 from their telomere sites, suggesting that the ligand triggers the uncapping of the telomere ends. The effect of the ligand is associated with an increase of the gamma-H2AX foci, one part of them colocalizing at telomeres, thus indicating the occurrence of a DNA damage response at the telomere, but also the presence of additional DNA targets for telomestatin. Interestingly, the expression of GFP-POT1 in HT1080 cells increases both telomere and G-overhang length. As compared with HT1080 cells, HT1080GFP-POT1 cells presented a resistance to telomestatin treatment characterized by a protection to the telomestatin-induced growth inhibition and the G-overhang shortening. This protection is related to the initial G-overhang length rather than to its degradation rate and is overcome by increased telomestatin concentration. Altogether these results suggest that telomestatin induced a telomere dysfunction in which G-overhang length and POT1 level are important factors but also suggest the presence of additional DNA sites of action for the ligand.

Base Sequence↗

PubMed2005

Interleukin-1beta-induced apoptosis through adenylyl cyclase and ERK1/2 inhibition in primary cultured thyroid cells.

The programmed cell death plays a crucial role in the regulation of numerous physiological and pathological phenomena. In this study, we show that interleukin-1 beta (IL-1beta) induces an early production of endogenous ceramides via N-sphingomyelinase (N-Smase) as well as an inhibition of adenylyl cyclase activity in pig thyroid cells. This effect is followed by a down-regulation of the extracellular signal-regulated protein kinase (ERK1/2) phosphorylation, an activation of caspase-3, and ends by setting up the programmed cell death. The permeable exogenous C(2)-ceramide reproduces IL-1beta effects on: (i) inhibition of adenylyl cyclase activity, (ii) down-regulation of ERK1/2 phosphorylation, (iii) activation of caspase-3, and (iv) apoptosis in pig thyroid cells. Cell treatment with a PKA inhibitor down-regulates ERK1/2 phosphorylation. Furthermore, inhibition of ERK1/2 signaling pathway by U-0126 enhances caspase-3 activity and sets up programmed cell death. Both IL-1beta and exogenous C(2)-ceramide effects are reproduced by U-0126 so illustrating the implication of ERK1/2 down-regulation in both caspase-3 activation and apoptosis induction. Our study shows for the first time that endogenous ceramides are important second messengers in IL-1beta-induced apoptosis in pig thyroid cells through inhibition of adenylyl cyclase and ERK1/2 activities.

Adenylyl Cyclase Inhibitors↗

PubMed2005

MRP3, BCRP, and P-glycoprotein activities are prognostic factors in adult acute myeloid leukemia.

PURPOSE: P-Glycoprotein (Pgp) is associated with poor outcome in acute myeloid leukemia (AML). We have investigated other ATP-binding cassette proteins such as BCRP, MRP1, MRP2, MRP3, and MRP5 for their potential implication in chemoresistance. EXPERIMENTAL DESIGN AND RESULTS: Eighty five AML patient samples were analyzed in this study. First, MRP3 function was higher in patients which had a high level of leukocytes (P = 0.01), a M5 FAB subtype (P = 0.04), and an intermediate or poor cytogenesis (P = 0.05). BCRP activity was not correlated with clinical or biological variables, but high Pgp activity was correlated with the following variables: CD34 expression (P = 0.002), FAB subtype (P = 0.002), intermediate or poor cytogenesis (P = 0.02), and elderly patients (P = 0.03). Second, Pgp, MRP3, and BCRP activities were correlated with complete remission (P = 0.02, P = 0.04, and P = 0.04, respectively), disease-free survival (P = 0.02, P = 0.03, and P = 0.25, respectively), and overall survival (P = 0.04, P = 0.04, and P = 0.05, respectively) in multivariate analysis. The patient samples expressing one or none of these Pgp, MRP3, or BCRP functional proteins have a better prognosis than the patients expressing two or three of these functional proteins (complete remission, P = 0.02; disease-free survival, P = 0.01; overall survival, P < 0.001). CONCLUSIONS: BCRP and MRP3 may also be involved in chemoresistance in AML, especially MRP3 in patients with M5 FAB. Additional modulation of BCRP or MRP3 to Pgp modulation may be necessary in some patients in order to improve the treatment outcome.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2005

Overexpression of Bcl-2 is associated with apoptotic resistance to the G-quadruplex ligand 12459 but is not sufficient to confer resistance to long-term senescence.

The triazine derivative 12459 is a potent G-quadruplex interacting agent that inhibits telomerase activity. This agent induces time- and dose-dependent telomere shortening, senescence-like growth arrest and apoptosis in the human A549 tumour cell line. We show here that 12459 induces a delayed apoptosis that activates the mitochondrial pathway. A549 cell lines selected for resistance to 12459 and previously characterized for an altered hTERT expression also showed Bcl-2 overexpression. Transfection of Bcl-2 into A549 cells induced a resistance to the short-term apoptotic effect triggered by 12459, suggesting that Bcl-2 is an important determinant for the activity of 12459. In sharp contrast, the Bcl-2 overexpression was not sufficient to confer resistance to the senescence-like growth arrest induced by prolonged treatment with 12459. We also show that 12459 provokes a rapid degradation of the telomeric G-overhang in conditions that paralleled the apoptosis induction. In contrast, the G-overhang degradation was not observed when apoptosis was induced by camptothecin. Bcl-2 overexpression did not modify the G-overhang degradation, suggesting that this event is an early process uncoupled from the final apoptotic pathway.

Antineoplastic Agents↗

PubMed2005

Immunosuppressors and reversion of multidrug-resistance.

Drug resistance is the major reason for failure of cancer therapy. When one drug elicits a response in tumour cells resulting in resistance to a large variety of chemically unrelated drugs, this is called multidrug-resistance (MDR). ATP-binding cassette (ABC) transporters contribute to drug resistance via ATP-dependent drug efflux. P-glycoprotein (Pgp) encoded by MDR1 gene, confers resistance to certain anticancer agents. The development of agents able to modulate MDR mediated by Pgp and ABC transporters remained a major goal for the past 10 years. Immunosuppressors, cyclosporin A (CSA) in particular, were shown to modulate Pgp activity in laboratory models and entered very early into clinical trials for reversal of MDR. The proof of reversing activity of CSA was found in phase II studies with myeloma and acute leukaemia. In phase III studies, the results were less convincing regarding the response rate, progression-free survival and overall survival were detected in advanced refractory myeloma. The non-immunosuppressive derivative PSC833 was then extensively studied. This compound shows 10-fold higher potency in reversal of MDR mediated by Pgp. Results from clinical trials with this modulator are still emerging and the notable finding was the need to reduce the dose of anticancer agent used in combination with it. Other effects of CSA and PSC833 on MDR have been described. These two molecules have been shown to have an action on the metabolism of ceramide which stands as second messenger of anticancer agents-induced apoptosis. PSC833 stimulates de novo ceramide synthesis and enhances cell death induced by anticancer agents, such as camptothecins and anthracyclines. In addition, ceramide glycosylation and storage in some cell lines have been described to play a crucial role in resistance to anticancer drugs. CSA is able to inhibit ceramide glucosylation and modulate MDR phenotype. The emergence of other modulators with several ABC protein targets like VX710 are of clinical interest in malignancies expressing several efflux pumps.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2005

From molecular characteristics to cellular events in apoptosis-resistant HL-60 cells.

The ability to induce apoptosis in tumor cells is critical to elicit a positive response to cytotoxic chemo-therapy. In this study, we investigated the effect of the topoisomerase I inhibitors camptothecin and SN-38, known to cause an unusual form of DNA damage, on apoptotic pathways using the leukemic cell line HL-60 and its vincristine-resistant variant HL-60 VCR. Both camptothecin and SN-38 induced high levels of apoptosis in sensitive cells when compared to the multidrug-resistant ones. Interestingly, a higher BCL-2/BAX ratio was observed in HL-60 VCR at the basal state and during treatments. Moreover, these cells which did not exhibit Bcr-abl translocation or bcrp efflux pump, overexpressed topoisomerase I protein. The data provide evidence that BCL-2 protein could protect HL-60 VCR from mitochondrial membrane depolarization and block ROS production in these cells. Finally, our results suggest that dysregulation of proteins associated with DNA replication and apoptotic process could contribute to the multidrug-resistance phenotype.

Antineoplastic Agents, Phytogenic↗

PubMed2005

[Regulation of telomeres length: making the telomeres accessible?].

Under a normal state, the extremities of chromosomes, telomeres, are protected against undesired fusion events. Alterations of the telomere structure are associated with genetic instability, while erosion of the telomeric repeats, occurring at each cell division, provides a mechanism controlling the long-term proliferation of somatic cells. Although the structure and composition of the human telomerase enzyme are now well characterized, the protein partners regulating the stability and conformation of its DNA substrate, the telomeric end, are much less known. A functional link has been recently evidenced between proteins that bind the double-stranded telomere repeats and those recruited at the 3' G-rich telomeric overhang. This review presents an update on these telomeric factors controlling telomere lengthening and discuss the actual models proposed for its regulation.

Cell Division↗

PubMed2003

Resistance to the short term antiproliferative activity of the G-quadruplex ligand 12459 is associated with telomerase overexpression and telomere capping alteration.

Ligands that stabilize the telomeric G-rich single-stranded DNA overhang into G-quadruplex can be considered as potential antitumor agents that block telomere replication. Ligand 12459, a potent G-quadruplex ligand that belongs to the triazine series, has been previously shown to induce both telomere shortening and apoptosis in the human A549 cell line as a function of its concentration and time exposure. We show here that A549 clones obtained after mutagenesis and selected for resistance to the short term effect of ligand 12459 frequently displayed hTERT transcript overexpression (2-6-fold). Overexpression of hTERT was also characterized in two resistant clones (JFD10 and JFD18) as an increase in telomerase activity, leading to an increase in telomere length. An increased frequency of anaphase bridges was also detected in JFD10 and JFD18, suggesting an alteration of telomere capping functions. Transfection of either hTERT or DN-hTERT cDNAs into A549 cells did not confer resistance or hypersensitivity to the short term effect of ligand 12459, indicating that telomerase expression is not the main determinant of the antiproliferative effect of ligand 12459. In contrast, transfection of DN-hTERT cDNA into resistant JFD18 cells restored sensitivity to apoptotic concentrations of ligand 12459, suggesting that telomerase does participate in the resistance to this G-quadruplex ligand. This work provides evidence that telomerase activity is not the main target for the 12459 G-quadruplex ligand but that hTERT functions contribute to the resistance phenotype to this class of agents.

Cell Division↗

PubMed2003

Resistance to senescence induction and telomere shortening by a G-quadruplex ligand inhibitor of telomerase.

The molecular mechanisms induced by G-quadruplex ligands to trigger senescence in mammalian cells are still unknown, although the critical role of telomerase is highly suspected. JFA2 cells selected for resistance to senescence induced by the G-quadruplex ligand 12459 presented an overexpression of hTERT transcript that correlated to a functional increase in telomerase activity and telomere length. Consistently, treatment with 12459 failed to trigger senescence and telomere shortening in JFA2 cells. Resistant cells also presented cross-resistance for senescence induction to telomestatin, another G-quadruplex ligand from a different series, but not to other anticancer agents, indicating the selectivity of the resistance mechanism. We, thus, provide evidence that telomerase activity and telomere length are key cellular determinants of the resistance to G-quadruplex ligands.

Cell Line, Tumor↗

PubMed2003

Enhanced cytotoxicity and nuclear accumulation of doxorubicin-loaded nanospheres in human breast cancer MCF7 cells expressing MRP1.

Previous studies have demonstrated that doxorubicin (DOX) encapsulated in polyisohexylcyano-acrylate nanospheres (NS-DOX) circumvented the resistance of breast cancer cells overexpressing P-glycoprotein (Pgp). Another protein is involved in multidrug resistance phenotype, the multidrug resistance associated protein (MRP1). We report that NS-DOX overcomes multidrug resistance in breast cancer cells overexpressing MRP1. Taking into account that anthracyclines are conjugated to or co-transported with glutathione by MRP1, these data suggest that probably due to ion pair formation (NS-DOX), MRP1 could not transport the anthracycline. Pgp is probably able to transport the ion pair drug complex and the mechanisms of drug resistance reversion in Pgp expressing cells need to be further elucidated.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2002

Reversal of multidrug resistance-associated protein-mediated daunorubicin resistance by camptothecin.

The multidrug-resistance (MR) status of camptothecin (CPT) was investigated in colon adenocarcinoma HT29 cells, leukemia K562, and breast carcinoma MCF7 cells expressing P-glycoprotein (Pgp) and/or MR-associated protein (MRP1). The concentration that induced 50% growth inhibition (IC(50)) against CPT was 0.14 and 0.20 microM in parental K562/WT and MCF7/WT cells, respectively. The drug resistant subline KH30 and MCF7/VP cells, which both overexpress MRP1, presented IC(50) values of 0.63 and 3.10 microM, respectively. The resulting resistance indexes were 3.80 and 12.50, respectively. However, in KH300 cells, a cell line that preferentially overexpresses Pgp, the IC(50) of CPT was 0.08 microM and thus did not exhibit resistance against CPT. In MCF7/DoX cells, preferentially overexpressing Pgp, but also a significant level of MRP1, the IC(50) of CPT was 0.64 microM and thus presented a resistance index of 3.26 against CPT. The cytotoxic effect of CPT was modulated in cells expressing MRP1 (MCF7/VP, HT29 cells) by the specific MRP1 modulators, probenecid and MK571. These results led us to consider CPT as a substrate for MRP1 and a potential modulator of MRP1 activity. To test this hypothesis, we examined the ability of nontoxic concentrations of CPT to sensitize MRP1-overexpressing cells to daunorubicin (DNR). In MCF7/VP and KH30 cells, nontoxic concentrations of CPT were able to enhance cytotoxicity of DNR and its nuclear accumulation. Sequential and simultaneous associations of CPT (100 nM) and DNR provided complete reversal of resistance, thus showing a synergistic effect in KH30 cells. However, simultaneous association (with 10 or 20 nM CPT) had an additive effect in MCF7/VP. These data suggest that CPT could be proposed as a candidate for the reversal of the MRP1 phenotype at clinically achievable concentrations.

Antibiotics, Antineoplastic↗

PubMed2002

Free radical production and labile iron pool decrease triggered by subtoxic concentration of aclarubicin in human leukemia cell lines.

Aclarubicin (ACLA), which belongs to the antracycline class of antineoplasic agents, has been demonstrated as a differentiating agent for a broad range of human solid tumors and leukemia. By using dihydroethidium as a fluorescent probe, we show the ability of subtoxic (i.e. differentiating) concentration of ACLA to generate reactive oxygen species in both K562 and HL-60 leukemia cell lines. Besides, we have used a calcein-based spectrofluorimetric assay to determine the influence of ACLA treatment on the cellular labile iron pool (LIP). In both cell lines, the LIP level was markedly decreased in the presence of ACLA. Nevertheless, whereas ACLA-induced differentiation was obviously ROS-dependent, the LIP decrease was rather ROS-independent.

Aclarubicin↗

PubMed2002

Homocamptothecin-daunorubicin association overcomes multidrug-resistance in breast cancer MCF7 cells.

The multidrug-resistance (MDR) status of a novel camptothecin analogue, homocamptothecin (hCPT), was investigated in human colon adenocarcinoma HT29 cells, myelogenous leukemia K562 cells and breast carcinoma MCF7 cells. The cytotoxicity of hCPT was not sensitive to the MDR status in K562 cell lines. However, its cytotoxicity was altered by MRP1, but not Pgp, in naturally MRP1-expressing HT29 cells, and etoposide- and doxorubicin-resistant MCF7/VP and MCF7/DOX cells, respectively. These cells were sensitized to hCPT in presence of MK571, probenecid but not verapamil. These results led to consider hCPT as a substrate for MRP1 and a potential modulator of MRP1 activity. The relationship between the cytotoxic effect of anthracyclines and their nuclear localization had been previously demonstrated. We show that MRPI mediated the daunorubicin (DNR) efflux in MCF7/VP and MCF7/DOX cells. The combination of sub-toxic doses of hCPT with DNR resulted in the potentiation of DNR activity, well-correlated with an increase in its nuclear accumulation in MCF7/VP cells. Simultaneous pattern was shown to provide higher cytotoxic response than sequential one. In agreement, hCPT increased also the DNR nuclear accumulation in low MRP1-expressing MCF7/DOX cells. However, the enhancement of cytotoxicity in the DNR-hCPT combination was poorly correlated with the nuclear concentration of DNR in MCF7/DOX cells. In addition to the increase in DNR accumulation, the potentiation of DNR activity by hCPT in MCF7/DOX cells implied a synergistic mechanism between both drugs. These data suggest that the present topoisomerase I/II inhibitors combination may be of clinical interest to overcome MDR phenotype in DNR-treated breast cancer patients.

Adenocarcinoma↗

PubMed2002

Impact of endogenous nitric oxide on microglial cell energy metabolism and labile iron pool.

Microglial activation is common in several neurodegenerative disorders. In the present study, we used the murine BV-2 microglial cell line stimulated with gamma-interferon and lipopolysaccharide to gain new insights into the effects of endogenously produced NO on mitochondrial respiratory capacity, iron regulatory protein activity, and redox-active iron level. Using polarographic measurement of respiration of both intact and digitonin-permeabilized cells, and spectrophotometric determination of individual respiratory chain complex activity, we showed that in addition to the reversible inhibition of cytochrome-c oxidase, long-term endogenous NO production reduced complex-I and complex-II activities in an irreversible manner. As a consequence, the cellular ATP level was decreased in NO-producing cells, whereas ATPase activity was unaffected. We show that NO up-regulates RNA-binding of iron regulatory protein 1 in microglial cells, and strongly reduces the labile iron pool. Together these results point to a contribution of NO derived from inflammatory microglia to the misregulation of energy-producing reactions and iron metabolism, often associated with the pathogenesis of neurodegenerative disorders.

Aconitate Hydratase↗

PubMed2002

Ceramide involvement in homocamptothecin- and camptothecin-induced cytotoxicity and apoptosis in colon HT29 cells.

Topoisomerase I inhibitors of the camptothecin (CPT) family have emerged as potent clinical chemotherapeutic agents in first-line treatment of solid colorectal cancer and in second-line for 5-fluorouracil resistant patients. CPT and homocamptothecin (hCPT), derivative with enhanced lactone stability, induced growth inhibition in HT29 cells via p53-independent apoptosis. hCPT- and CPT-induced apoptosis was dependent on caspase-3 but not caspase-1. We report here substantial evidence that ceramide, resulting from de novo pathway or catabolism modulation, acted as a second messenger of these antitumor drugs in HT29 cells and leads to the activation of caspase-3. In addition, hCPT and CPT may favor ceramide signaling by disturbing sites of synthesis (Golgi) and trafficking of glucosylceramide from Golgi to lipid droplets. This work contributes to the understanding of the mechanism of action of CPTs, and suggests that inhibitors of glycosylation or activators of de novo metabolism could be of clinical interest in enhancing the effects of CPTs.

Antineoplastic Agents, Phytogenic↗