Biomedical subjects

S Soddu

Publications and source records attributed to S Soddu.

At least 37 records · Page 2Linked to original sources

PubMed1998

p53 expression in B-cell chronic lymphocytic leukemia: a marker of disease progression and poor prognosis.

We have analyzed by immunocytochemistry (ICC) the frequency of p53 protein expression in 181 cases of B-cell chronic lymphocytic leukemia (CLL) followed at a single institution to assess the relationship between p53 and the clinical and morphological features of the disease, as well as the possible involvement of this protein in the pathogenesis of the more aggressive forms of CLL. The overall frequency of p53 protein positivity in CLL was 15% (27 of 181 cases). There were no significant differences in age, sex, absolute lymphocyte count, or lymphocyte doubling time between p53-positive and -negative patients. By contrast, p53-positive patients had a significantly higher percentage of prolymphocytes (P = .002) and a significantly lower percentage of residual CD3-positive T lymphocytes (P = .0001). No correlation was found between the percentage of p53-positive cells and the percentage of cells in cycle assessed by the monoclonal antibody Ki-67. When the percentage of p53 positivity was correlated with the clinical stage of the disease, the proportion of p53-positive cases increased significantly from Binet's stage A (8 of 108; 7.4%), to stage B (12 of 49; 24.4%) and C (7 of 24; 29.2%) (P = .002). p53 positivity correlated also with the phase of the disease, showing a low expression at diagnosis (8 of 112; 7.1%) and a significantly higher expression in patients studied during the course of the disease (7 of 35; 20%) and, to a further extent, with disease progression (12 of 34; 35.3%) (P = .0001). The association of p53 protein expression with mutations in the gene was confirmed by direct sequence of the entire cDNA in 15 of the 17 ICC positive cases tested (88%). A significantly shorter treatment-free interval from diagnosis (P = .003) and a poorer response to therapy (P = .007) was observed in p53-positive compared with p53-negative patients. Overall survival from the time of diagnosis, as well as from the time of p53 protein analysis, was significantly shorter in patients with p53 protein expression (P = .03 and .0001, respectively). Moreover, in multivariate analysis, p53 expression and stage C were independently associated with a short survival. The results of this study indicate that in CLL the expression of the p53 protein, analyzed by a simple and reliable immunocytochemical method, is strongly associated with p53 gene mutations, a morphological variant (CLL with >10% prolymphocytes), advanced clinical stage, progressive disease, poor response to therapy, and short survival.

Adult↗

PubMed1998

Genomic instability associated with myotonic dystrophy does not involve p53 expression and activity.

We tested the hypothesis that the instability of the trinucleotide CTG at the myotonic dystrophy (DM) locus could be an intrinsic DNA damage recognisable by the p53 cell-cycle checkpoint system. p53 mRNA and protein levels were assayed in muscle biopsies and fibroblast cell lines of DM patients and unaffected controls. No differences in mRNA and protein levels were found between patients and controls, regardless of their expansion size. However, in the cells treated with adryamicin, p53 protein levels were comparable in DM and control cells. We conclude that the CTG trinucleotide expansion within the myotonin gene does not activate the p53 surveillance system, at least in adult tissues. The escape of trinucleotide expansion from the p53-mediated DNA repair system could explain some of the biological characteristics of genome instability.

Adult↗

PubMed1998

Wt-p53 action in human leukaemia cell lines corresponding to different stages of differentiation.

Recent studies support the potential application of the wt-p53 gene in cancer therapy. Expression of exogenous wt-p53 suppresses a variety of leukaemia phenotypes by acting on cell survival, proliferation and/or differentiation. As for tumour gene therapy, the final fate of the neoplastic cells is one of the most relevant points. We examined the effects of exogenous wt-p53 gene expression in several leukaemia cell lines to identify p53-responsive leukaemia. The temperature-sensitive p53Val135 mutant or the human wt-p53 cDNA was transduced in leukaemia cell lines representative of different acute leukaemia FAB subtypes, including M1 (KG1), M2 (HL-60), M3 (NB4), M5 (U937) and M6 (HEL 92.1.7), as well as blast crisis of chronic myelogenous leukaemia (BC-CML: K562, BV173) showing diverse differentiation features. By morphological, molecular and biochemical analyses, we have shown that exogenous wt-p53 gene expression induces apoptosis only in cells corresponding to M1, M2 and M3 of the FAB classification and in BC-CML showing morphological and cytochemical features of undifferentiated blast cells. In contrast, it promotes differentiation in the others. Interestingly, cell responsiveness was independent of the vector used and the status of the endogenous p53 gene.

Apoptosis↗

PubMed1998

Wild-type p53-mediated down-modulation of interleukin 15 and interleukin 15 receptors in human rhabdomyosarcoma cells.

We recently reported that rhabdomyosarcoma cell lines express and secrete interleukin 15 (IL-15), a tightly regulated cytokine with IL-2-like activity. To test whether the p53-impaired function that is frequently found in this tumour type could play a role in the IL-15 production, wild-type p53 gene was transduced in the human rhabdomyosarcoma cell line RD (which harbours a mutated p53 gene), and its effect on proliferation and expression of IL-15 was studied. Arrest of proliferation was induced by wild-type p53; increased proportions of G1-arrested cells and of apoptotic cells were observed. A marked down-modulation of IL-15 expression, at both the mRNA and protein level, was found in p53-transduced cells. Because a direct effect of IL-15 on normal muscle cells has been reported, the presence of IL-15 membrane receptors was studied by cytofluorometric analysis. Rhabdomyosarcoma cells showed IL-15 membrane receptors, which are down-modulated by wild-type p53 transfected gene. In conclusion, wild-type p53 transduction in human rhabdomyosarcoma cells induces the down-modulation of both IL-15 production and IL-15 receptor expression.

Cell Division↗

PubMed1998

p53: prospects for cancer gene therapy.

Initially described as an oncogene, at the end of the 1980s, the wild-type p53 gene (TP53) was later shown to be capable of suppressing the proliferation of transformed cells. In the following years, an increasing number of studies demonstrated that intact p53 function is essential for the maintenance of the non-tumorigenic phenotype of cells. Indeed, functional inactivation of the p53 protein is one of the most common alterations observed in human cancers. More recently, it has been shown that inactivation of the TP53 tumor suppressor gene may lead to radioresistant and chemoresistant tumors, possibly by the induction of apoptosis-resistant phenotypes. Finally, a large number of in vitro and in vivo transduction experiments have demonstrated that exogenous TP53 overexpression can suppress the transformed phenotype of many cell types by inducing growth arrest, apoptosis or cell differentiation. All of these findings have rendered p53 a potentially helpful target for the therapy of many types of human cancers. In this review we shall discuss the different approaches to p53-related cancer therapy that have been proposed on the basis of this large number of experimental studies, and we shall try to dissect the biological questions that are still open and need to be clarified to improve p53-based therapy.

Animals↗

PubMed1997

Oncogenes belonging to the CSF-1 transduction pathway direct p53 tumor suppressor effects to monocytic differentiation in 32D cells.

Expression of exogenous wt-p53 in different tumor cell lines can induce growth arrest, apoptosis, or differentiation. Several experimental works have highlighted the relevance of cellular context in the determination of p53-mediated final outcomes. We recently observed that these diverse wt-p53 effects can also be induced by overexpressing wt-p53 in a single cell type-the 32D myeloid progenitors-transformed with different activated oncogenes. Here we show that 32D cells transformed with two different oncogenes, v-src or c-fms [S301,F969], both belonging to the CSF-1 transduction pathway, respond to exogenous wt-p53 expression with the same final outcome-monocytic differentiation. This result is particularly significant since 32D cells do not spontaneously express the CSF-1 receptor, whereas they undergo granulocytic differentiation upon G-CSF stimulation. These data strongly support the idea that wt-p53 suppressing effects result from interactions between p53 activity and the signaling pathways activated in different transformed cells.

Cell Differentiation↗

PubMed1997

p53 re-expression inhibits proliferation and restores differentiation of human thyroid anaplastic carcinoma cells.

Alterations of the tumor suppressor gene p53 are uncommon in differentiated thyroid neoplasia but are detected at high frequency in anaplastic thyroid carcinoma suggesting that impaired p53 function may contribute to the undifferentiated and highly aggressive phenotype of these tumors. Effects of wild type p53 (wt-p53) re-expression were investigated in a human anaplastic thyroid carcinoma cell line (ARO) expressing a mutated p53. ARO cells were stably transfected with the temperature-sensitive p53 Val135 gene (ts-p53) which exhibits wild type-like activity at 32 degrees C. Exogenous wt-p53 function in ARO-tsp53 clones was assessed by evaluating its transcriptional activity on a CAT reporter vector containing p53 binding sites. At 32 degrees C, a significant reduction in the proliferation rate (approximately or equal to 50%) was observed, with accumulation of cells in the G0/G1 phase of the cell cycle. This effect was accompanied by induction of the expression of the growth inhibitor p21/Waf1 gene. At 32 degrees C, ARO-tsp53 clones also showed a marked impairment of their tumorigenic potential. Furthermore, transfected clones re-acquired the ability to respond to thyrotropin (TSH) stimulation showing an increased expression of thyroid-specific genes (thyroglobulin, thyroperoxidase and TSH receptor). In conclusion, re-expression of wt-p53 activity in ARO cells, inhibits cell proliferation and restores responsiveness to physiological stimuli.

Carcinoma↗

PubMed1997

Expression of exogenous wt-p53 does not affect normal hematopoiesis: implications for bone marrow purging.

Some gene therapy approaches for cancer treatment attempt to transduce onco-suppressor genes into tumor cells. A central problem of this strategy is the targeting of tumor cells to avoid damage to normal ones. It has been noticed that transduction of wt-p53 into a large number of cancer cells induces tumor suppression. In contrast, some observations suggest that introduction of exogenous wt-p53 into nontransformed cells does not impair proliferation. If normal bone marrow (BM) cells are not affected by wt-p53 transduction, BM purging from p53-responding leukemic cells might be achieved in vitro by delivering the wild-type onco-suppressor to all marrow cells. We undertook a series of experiments to assess whether transduction of wt-p53 into normal hematopoietic cells is harmful. Two different wt-p53-recombinant retroviruses were used to infect primary, murine BM cells. Expression of exogenous wt-p53 in these cells did not affect in vitro colony formation, and did not induce any observable effects on morphology and differentiation. In contrast, the same viruses suppressed the tumor phenotype of v-src-transformed 32D cells. These results might open the way to gene therapy approaches to leukemias with the p53 gene without the need to target specifically and uniquely the tumor cells, sparing the normal ones.

Bone Marrow Cells↗

PubMed1997

P53 role in DNA repair and tumorigenesis.

The p53 oncosuppressor is the gene which has been found to be most frequently altered in human cancers. How the p53 gene contributes to tumor induction and progression is presently being investigated by numerous researchers whose specialties range from basic science to clinics. This is a brief overview of the hypotheses on p53 role in tumorigenesis and the possible use of p53 analyses in tumor prognosis and therapy.

DNA Repair↗

PubMed1996

The beta 4 integrin subunit is expressed in mouse fibroblasts and modulated by transforming growth factor-beta 1.

Integrin beta 4 subunit is present in association with alpha 6 chain on both normal and transformed epithelial cells. Recently alpha 6 beta 4 heterodimer was found on the endothelium of medium-sized blood vessels and on immature thymocytes. In this report we show, by Northern blotting, indirect immunofluorescence, immunoprecipitation, and Western blotting, that beta 4 subunit is expressed also on cells of mesenchymal origin such as fibroblasts, myoblasts, and myotubes. Increased expression of alpha 6 beta 4 has been related to the aggressive metastatic phenotype of human and murine carcinomas. The transforming growth factor beta 1 (TGF-beta 1) has been found to modulate the expression of several integrins and intracellular matrix proteins, as well as to stimulate cell invasion and metastatic potential. To evaluate whether alpha 6 beta 4 expression is modulated by TGF-beta 1, we transfected 3T3 fibroblasts with an expression vector carrying the human TGF-beta 1 cDNA driven by the SV40 early promoter. We observed by indirect immunofluorescence a modification in the subcellular distribution of beta 4 subunit, which acquires a perinuclear localization. This finding suggests this integrin subunit correlates with the cytoskeletal reorganization induced by TGF-beta 1.

3T3 Cells↗

PubMed1996

Lonidamine induces apoptosis in drug-resistant cells independently of the p53 gene.

Lonidamine, a dichlorinated derivative of indazole-3-carboxylic acid, was shown to play a significant role in reversing or overcoming multidrug resistance. Here, we show that exposure to 50 microg/ml of lonidamine induces apoptosis in adriamycin and nitrosourea-resistant cells (MCF-7 ADR(r) human breast cancer cell line, and LB9 glioblastoma multiform cell line), as demonstrated by sub-G1 peaks in DNA content histograms, condensation of nuclear chromatin, and internucleosomal DNA fragmentation. Moreover, we find that apoptosis is preceded by accumulation of the cells in the G0/G1 phase of the cell cycle. Interestingly, lonidamine fails to activate the apoptotic program in the corresponding sensitive parental cell lines (ADR-sensitive MCF-7 WT, and nitrosourea-sensitive LI cells) even after long exposure times. The evaluation of bcl-2 protein expression suggests that this different effect of lonidamine treatment in drug-resistant and -sensitive cell lines might not simply be due to dissimilar expression levels of bcl-2 protein. To determine whether the lonidamine-induced apoptosis is mediated by p53 protein, we used cells lacking endogenous p53 and overexpressing either wild-type p53 or dominant-negative p53 mutant. We find that apoptosis by lonidamine is independent of the p53 gene.

Antineoplastic Agents↗

PubMed1996

Interference with p53 protein inhibits hematopoietic and muscle differentiation.

The involvement of p53 protein in cell differentiation has been recently suggested by some observations made with tumor cells and the correlation found between differentiation and increased levels of p53. However, the effect of p53 on differentiation is in apparent contrast with the normal development of p53-null mice. To test directly whether p53 has a function in cell differentiation, we interfered with the endogenous wt-p53 protein of nontransformed cells of two different murine histotypes: 32D myeloid progenitors, and C2C12 myoblasts. A drastic inhibition of terminal differentiation into granulocytes or myotubes, respectively, was observed upon expression of dominant-negative p53 proteins. This inhibition did not alter the cell cycle withdrawal typical of terminal differentiation, nor p21(WAF1/CIP1) upregulation, indicating that interference with endogenous p53 directly affects cell differentiation, independently of the p53 activity on the cell cycle. We also found that the endogenous wt-p53 protein of C2C12 cells becomes transcriptionally active during myogenesis, and this activity is inhibited by p53 dominant-negative expression. Moreover, we found that p53 DNA-binding and transcriptional activities are both required to induce differentiation in p53-negative K562 cells. Taken together, these data strongly indicate that p53 is a regulator of cell differentiation and it exerts this role, at least in part, through its transcriptional activity.

Animals↗

PubMed1996

Wild-type p53 induces diverse effects in 32D cells expressing different oncogenes.

Expression of exogenous wild-type (wt) p53 in different leukemia cell lines can induce growth arrest, apoptotic cell death, or cell differentiation. The hematopoietic cell lines that have been used so far to study wt p53 functions have in common the characteristic of not expressing endogenous p53. However, the mechanisms involved in the transformation of these cells are different, and the cells are at different stages of tumor progression. It can be postulated that each type of neoplastic cell offers a particular environment in which p53 might generate different effects. To test this hypothesis, we introduced individual oncogenes into untransformed, interleukin-3 (IL-3)-dependent myeloid precursor 32D cells to have a single transforming agent at a time. The effects induced by wt p53 overexpression were subsequently evaluated in each oncogene-expressing 32D derivative. We found that in not fully transformed, v-ras-expressing 32D cells, as already shown for the parental 32D cells, overexpression of the wt p53 gene caused no phenotypic changes and no reduction of the proliferative rate as long as the cells were maintained in their normal culture conditions (presence of IL-3 and serum). An accelerated rate of apoptosis was observed after IL-3 withdrawal. In contrast, in transformed, IL-3-independent 32D cells, wt p53 overexpression induced different effects. The v-abl-transformed cells manifested a reduction in growth rate, while the v-src-transformed cells underwent monocytic differentiation. These results show that the phenotype effects of wt p53 action(s) can vary as a function of the cellular environment.

Animals↗

PubMed1996

Terminally differentiated skeletal myotubes are not confined to G0 but can enter G1 upon growth factor stimulation.

Terminally differentiated cells are specialized cells unable to proliferate that constitute most of the mammalian body. Despite their abundance, little information exists on the characteristics of cell cycle control in these cells and the molecular mechanisms that prevent their proliferation. They are generally believed to be irreversibly restricted to the G0 state. In this report, we define some features of a paradigmatic terminally differentiated system, the skeletal muscle, by studying its responses to various mitogenic stimuli. We show that forced expression of a number of cell cycle-regulatory genes, including erbB-2, v-ras, v-myc, B-myb, ld-1, and E2F-1, alone or in combinations, cannot induce terminally differentiated skeletal muscle cells (myotubes) to synthesize DNA. However, serum-stimulated myotubes display a typical immediate-early response, including the up-regulation of c-fos, c-jun, c-myc, and ld-1. They also elevate the expression of cyclin D1 after 4 hours of serum treatment. All these events take place in myotubes in a way that is indistinguishable from that of quiescent, undifferentiated myoblasts reactivated by serum. Moreover, pretreatment with serum shortens the time required by E1A to induce DNA synthesis, confirming that myotubes can partially traverse G1. Serum growth factors do not activate late-G1 genes in myotubes, suggesting that the block that prevents terminally differentiated cells from proliferating acts in mid-G1. Our results show that terminally differentiated cells are not confined to G0 but can partially reenter G1 in response to growth factors; they contribute to a much-needed definition of terminal differentiation. The important differences in the control of the cell cycle between terminally differentiated and senescent cells are discussed.

Adenovirus E1A Proteins↗

PubMed1995

The effects of end point overdispersions on the validity of single-dose tumorigenicity assays.

Overdispersion can be found in tumor incidence and tumor latency end point values obtained by the conventional assays that are being used to assess the tumorigenicity of neoplastic cells growing in tissue culture. Failure to account for such wide variations in end point data can lead to incorrect assessments of the neoplastic cell tumorigenic phenotype and misinterpretations of data relating genetic functions to tumor-forming capacity. This problem suggests the need for more detailed analyses of the relationships that exist between tumor cell dose and the parameters being used to measure tumorigenicity.

Animals↗

PubMed1995

Adenovirus infection induces reentry into the cell cycle of terminally differentiated skeletal muscle cells.

Different cell types in the body of higher animals undergo terminal differentiation. In such a process, cells acquire specialized functions and irreversibly lose their ability to divide, therefore entering the postmitotic state. Terminally differentiated cells do not proliferate in response to growth factors or following the expression of activated, retroviral oncogenes. In this paper we demonstrate that adenovirus infection is an efficient and convenient means to induce terminally differentiated cells to reenter the cell cycle. These findings constitute a first step toward defining the molecular determinants of the irreversible withdrawal from the cell cycle of terminally differentiated cells. They may also open the way to therapeutic applications.

Adenoviruses, Human↗

PubMed1995

Wild-type p53 modulates apoptosis of normal, IL-3 deprived, hematopoietic cells.

Apoptotic cell death is an active process which regulates the maintenance of the hematopoietic homeostasis. It has been reported that wild-type p53 (wt-p53) protein induces apoptosis in leukemia cells. To assess whether p53 is involved in the apoptotic process of normal hematopoietic cells, we introduced the temperature-sensitive p53Val135 mutant into the murine myeloid precursor cell line 32Dcl3. These are diploid, non-tumorigenic cells whose survival and proliferation are dependent upon growth factor supply (IL-3 and serum). Overexpression of wt-p53 protein does not affect morphology and proliferation of 32D cells as long as they are maintained in the presence of IL-3. However, after IL-3 withdrawal, wt-p53 overexpression significantly accelerates apoptosis. This phenomenon is IL-3 specific since no differences in death rates induced by serum starvation are found between parental cells and p53-transfectants. When the latter experiments are carried out at 37 degrees C with p53 protein in mutant conformation, an extended survival of 32D cells is observed after IL-3 deprivation, but not after serum withdrawal. Taken together, these results show that wt-p53 actively mediates the apoptosis due to the absence of specific growth factors, such as IL-3, suggesting that p53 might be involved in the response of myeloid precursors to environmental cytokines for the maintenance of the hematopoietic homeostasis.

Animals↗

PubMed1995

Mitotic cycle reactivation in terminally differentiated cells by adenovirus infection.

Different cell types (e.g., neurons, skeletal and heart myocytes, adipocytes, keratinocytes) undergo terminal differentiation, in which acquisition of specialized functions entails definitive withdrawal from the cell cycle. Such cells are distinct from quiescent (reversibly growth-arrested) cells, such as contact-inhibited fibroblasts. Terminally differentiated cells can not be induced to proliferate by means of growth factor stimulation or transduction of cellular oncogenes. An important first step toward defining the molecular basis for such unresponsiveness is to find a practical means to overcome the proliferative block. Furthermore, determining whether terminally differentiated, postmitotic cells still retain a potential competence for proliferation that can be reactivated would have important theoretical and practical implications. To address these questions, we exploited the properties of adenoviruses. These viruses can infect postmitotic cells and express E1A, a powerful activator of proliferation in reversibly growth-arrested cells. We infected terminally differentiated skeletal muscle cells and adipocytes with human adenovirus type 5 or 12, obtaining full reentry into the cell cycle, including DNA synthesis, mitosis, cytokinesis, and extended proliferation. Similar results were obtained with established cell lines and primary cells belonging to several species, from quail to humans. Genetic analysis indicated that the smaller splice product of E1A, E1A 12S, is sufficient to induce cell cycle reactivation in otherwise permanently nonmitotic cells. These results demonstrate that terminally differentiated cells retain proliferative potential and establish adenovirus as a convenient and powerful means to force such cells to reenter the cell cycle.

Adenovirus E1A Proteins↗