Showing papers by "Carlo M. Croce published in 2002"

Frequent deletions and down-regulation of micro- RNA genes miR15 and miR16 at 13q14 in chronic lymphocytic leukemia

Abstract: Micro-RNAs (miR genes) are a large family of highly conserved noncoding genes thought to be involved in temporal and tissue-specific gene regulation MiRs are transcribed as short hairpin precursors (≈70 nt) and are processed into active 21- to 22-nt RNAs by Dicer, a ribonuclease that recognizes target mRNAs via base-pairing interactions Here we show that miR15 and miR16 are located at chromosome 13q14, a region deleted in more than half of B cell chronic lymphocytic leukemias (B-CLL) Detailed deletion and expression analysis shows that miR15 and miR16 are located within a 30-kb region of loss in CLL, and that both genes are deleted or down-regulated in the majority (≈68%) of CLL cases

Akt induces enhanced myocardial contractility and cell size in vivo in transgenic mice.

Abstract: The serine-threonine kinase Akt seems to be central in mediating stimuli from different classes of receptors. In fact, both IGF-1 and IL6-like cytokines induce hypertrophic and antiapoptotic signals in cardiomyocytes through PI3K-dependent Akt activation. More recently, it was shown that Akt is involved also in the hypertrophic and antiapoptotic effects of β-adrenergic stimulation. Thus, to determine the effects of Akt on cardiac function in vivo, we generated a model of cardiac-specific Akt overexpression in mice. Transgenic mice were generated by using the E40K, constitutively active mutant of Akt linked to the rat α-myosin heavy chain promoter. The effects of cardiac-selective Akt overexpression were studied by echocardiography, cardiac catheterization, histological and biochemical techniques. We found that Akt overexpression produced cardiac hypertrophy at the molecular and histological levels, with a significant increase in cardiomyocyte cell size and concentric LV hypertrophy. Akt-transgenic mice also showed a remarkable increase in cardiac contractility compared with wild-type controls as demonstrated by the analysis of left ventricular (dP/dtmax) in an invasive hemodynamic study, although with graded dobutamine infusion, the maximum response was not different from that in controls. Diastolic function, evaluated by left ventricular dP/dtmin, was not affected at rest but was impaired during graded dobutamine infusion. Isoproterenol-induced cAMP levels, β-adrenergic receptor (β-AR) density, and β-AR affinity were not altered compared with control mice. Moreover, studies on signaling pathway activation from myocardial extracts demonstrated that glycogen synthase kinase3-β is phosphorylated, whereas p42/44 mitogen-activated protein kinases is not, indicating that Akt induces hypertrophy in vivo by activating the glycogen synthase kinase3-β/GATA 4 pathway. In summary, our results not only demonstrate that Akt regulates cardiomyocyte cell size in vivo, but, importantly, show that Akt modulates cardiac contractility in vivo without directly affecting β-AR signaling capacity.

Overexpression of the HMGA2 gene in transgenic mice leads to the onset of pituitary adenomas.

Abstract: Overexpression of the HMGA2 gene is a common feature of neoplastic cells both in experimental and human models. Intragenic and extragenic HMGA2 rearrangements responsible for HMGA2 gene overexpression have been frequently detected in human benign tumours of mesenchymal origin. To better understand the role of HMGA2 overexpression in human tumorigenesis, we have generated transgenic mice carrying the HMGA2 gene under the transcriptional control of the cytomegalovirus promoter. High expression of the transgene was demonstrated in all the mouse tissues analysed, whereas no expression of the endogenous HMGA2 gene was detected in the same tissues from wild-type mice. In this study, two independent lines of transgenic mice have been generated. By 6 months of age, 85% of female animals of both transgenic lines developed pituitary adenomas secreting prolactin and growth hormone. The transgenic males developed the same phenotype with a lower penetrance (40%) and a longer latency period (about 18 months). Therefore, these data demonstrate that the overexpression of HMGA2 leads to the onset of mixed growth hormone/prolactin cell pituitary adenomas. These transgenic mice may represent an important tool for the study of this kind of neoplasia.

Genetic alterations of the tumor suppressor gene WWOX in esophageal squamous cell carcinoma.

Abstract: The WWOX (WW domain containing oxidoreductase) gene was recently identified as a candidate tumor suppressor gene at 16q23.3-24.1, a chromosome region that spans the common fragile site FRA16D. To evaluate the potential role of the WWOX gene in esophageal squamous cell carcinomas, we examined 36 tumors for genetic alterations of the WWOX gene. Loss of heterozygosity (LOH) at the WWOX locus was observed in 14 (39%) tumors. A tumor-specific missense mutation was found in one tumor, and LOH analysis had shown that the other allele was missing. Furthermore, we detected aberrant WWOX gene transcripts with absence of exons 6-8 in two tumors, and complete absence of transcript in one tumor. These results indicate that alteration and inactivation of the WWOX gene may play a role in esophageal squamous cell carcinogenesis.

Restoration of fragile histidine triad (FHIT) expression induces apoptosis and suppresses tumorigenicity in lung and cervical cancer cell lines.

Abstract: Loss of expression of the Fhit protein is often associated with the development of many human epithelial cancers, including lung and cervical carcinomas. Restoration of Fhit expression in cell lines derived from these tumors has however yielded conflicting results, prompting the need for careful evaluation of the oncosuppressive potential of FHIT. In the present study, we have investigated the effect of Fhit reintroduction in seven lung cancer and three cervical cancer cell lines. To achieve efficient gene transfer and high levels of transgene expression, we have used an adenoviral vector to transduce the FHIT gene. The induction of apoptosis was evaluated by using the terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling assay and propidium iodide staining. Activation of caspases was detected by using Western blot analysis, and tumorigenic potential of transduced cells in the nude mouse was also assessed. Restoration of Fhit expression induced apoptosis in all Fhit-negative cell lines, with Calu-1, H460, and A549 being the most susceptible among the lung cancer cell lines and SiHa cells among cervical carcinomas. Activation of caspase-8 was always associated with Fhit-mediated apoptosis, and in vivo tumorigenicity was either abolished by FHIT gene transfer (in H460 and SK-Mes cells) or strongly suppressed (in A549 and SiHa cells). Our data demonstrate oncosuppressive properties and strong proapoptotic activity of the Fhit protein in lung and cervical cancer cell lines and strengthens the hypothesis of its possible use as a therapeutic tool.

Association of SV40 with human tumors

Abstract: In 1994, PCR and protein studies suggested that SV40 DNA sequences and proteins were present in 29/48 (60%) USA human mesothelioma samples. Sequence analysis confirmed that the sequences were homologous to SV40. One year later, SV40 was also found in 5/9 human mesotheliomas, and in 1996 SV40 was also reported to be present in 1/3 of the tumor specimens examined. These reports, in combination with an earlier study in 1992 which had detected SV40 in human brain tumors, raised concerns that SV40 was associated with certain types of human tumors, specifically mesothelioma, bone, and brain tumors. These findings raised concerns, because these tumor types are the same malignancies that had been observed in animals injected with SV40. However, a study in 1996 and a presentation made at the International Mesothelioma Interest Group, IMIG in 1997 failed to detect SV40 in mesotheliomas, suggesting the possibility that laboratory artifacts, such as PCR contamination, had caused the previous positive findings. In 1997, the FDA, the NIH, and the CDC organized an international conference in Bethesda to review the literature and to address the possibility that SV40 was present in, and was possibly the cause of, some human tumors. The results of that conference were reported the same year in a meeting review in Oncogene by Carbone and colleagues. Briefly, the consensus was that before accepting the possibility that SV40 was present in human tumors, a multi-laboratory study needed to be conducted. It was recommended that a blinded multi-laboratory study be directed by an independent scientist not previously associated with the controversial reports of SV40 in human specimens. It was also recommended that this study include laboratories that had reported positive findings as well as laboratories that had failed to detect SV40 in human specimens. Since 1997, about 30 independent reports have been published on this topic, including the multi-laboratory study. Evidence in favor and against a possible association of SV40 with human cancer was reviewed at an international consensus meeting at the University of Chicago on 20, 21 April 2001, entitled "Malignant Mesothelioma: Therapeutic Options and the Role of SV40, 2001". The main focus was the association of SV40 with mesothelioma and other human tumors. At the end of the meeting, a panel discussion, which included independent experts who had not published on this topic, critically reviewed the evidence presented at the meeting. The results of the meeting and of the final panel discussion are outlined below.

FHIT: from gene discovery to cancer treatment and prevention.

Abstract: Chromosomal abnormalities, including homozygous deletions and loss of heterozygosity, are among the most common features of human tumours. The short arm of human chromosome 3, particularly the region 3p14.2, is a major site of such rearrangements. The 3p14.2 region spans the most active common fragile site of the human genome, encompassing a familial-kidney-cancer-associated breakpoint and a papilloma virus integration site. 6 years ago, the FHIT gene was identified in this region. Subsequent studies have shown that FHIT is commonly the target of chromosomal aberrations involving the long arm of human chromosome 3 and is thereby inactivated in most of the common human malignant diseases, including cancers of the lung, oesophagus, stomach, breast, and kidney. During the past 5 years, evidence has accumulated in support of a tumour-suppressor function for FHIT. In this review, we describe the recent findings in the molecular biology of FHIT with particular focus on the opportunities for treatment and prevention of cancer that have emerged.

The High Mobility Group A2 Gene Is Amplified and Overexpressed in Human Prolactinomas

Abstract: Trisomy of chromosome 12 is a nonrandom chromosomal change in pituitary adenomas, particularly prolactinomas. This and the finding that prolactin-secreting pituitary adenomas develop in transgenic mice overexpressing the wild-type HMGA2 gene (which maps to 12q14-15) prompted us to investigate HMGA2 rearrangements and expression in human prolactinomas. By dual-color interphase fluorescence in situ hybridization analysis using HMGA2-specific PACs and BACs, we found that the HMGA2 locus was amplified in seven of the eight prolactinoma samples examined. The cytogenetic manifestations of elevated HMGA2 concentrations ranged from simple trisomy to tetrasomy 12 and der(12) chromosomes to marker chromosomes bearing 12q14-15-derived regions. Reverse transcription-PCR, Western blot and immunohistochemical analysis showed HMGA2 overexpression in a number of prolactinomas bearing rearrangement of regions 12q14-15. These data suggest a critical role of the HMGA2 overexpression in the generation of prolactin-secreting pituitary adenomas in humans.

TCL1 participates in early embryonic development and is overexpressed in human seminomas.

Abstract: Overexpression of the TCL1 oncogene has been shown to play a causative role in T cell leukemias of humans and mice. The characterization of Tcl1-deficient mice in these studies indicates an important developmental role for Tcl1 in early embryogenesis. In wild-type embryos, Tcl1 is abundant in the first three mitotic cycles, during which it shuttles between nuclei and the embryo cortical regions in a cell-cycle-dependent fashion. The absence of this protein in early embryogenesis results in reduced fertility of female mice. The present studies elucidate the mechanism responsible for the reduced female fertility through analysis of the oogenesis stages and early embryo development in Tcl1-deficient mice. Even though Tcl1−/− females display normal oogenesis and rates of oocyte maturation/ovulation and fertilization, the lack of maternally derived Tcl1 impairs the embryo's ability to undergo normal cleavage and develop to the morula stage, especially under in vitro culture conditions. Beyond this crisis point, differentiative traits of zygotic genome activation and embryo compaction can take place normally. In contrast with this unanticipated role in early embryogenesis, we observed an overexpression of TCL1 in human seminomas. This finding suggests that TCL1 dysregulation could contribute to the development of this germinal cell cancer as well as lymphoid malignancies.

TNF-α signal transduction in rat neonatal cardiac myocytes: definition of pathways generating from the TNF-α receptor

Abstract: Cardiomyocyte hypertrophy and apoptosis have been implicated in the loss of contractile function during heart failure (HF). Moreover, patients with HF have been shown to exhibit increased levels of tumor necrosis factor α (TNF-α) in the myocardium. However, the multiple signal transduction pathways generating from the TNF-α receptor in cardiomyocytes and leading preferentially to apoptosis or hypertrophy are still unknown. Here we demonstrate in neonatal rat cardiomyocytes that 1) TNF-α induces phosphorylation of AKT, activation of NF-κB, and the phosphorylation of JUN kinase; 2) blocking AKT activity prevents NF-κB activation, suggesting a role for AKT in regulating NF-κB function; 3) AKT and JUN are both critical for the hypertrophic effects of TNF-α, since dominant-negative mutants of these genes are capable of inhibiting TNF-α-induced ANF-promoter up-regulation and increase in cardiomyocyte cell size, and 4) blocking NF-κB, AKT, or JUN alone or in combination does not sensitize cardiomyocytes to the p...

FEZ1/LZTS1 is down-regulated in high-grade bladder cancer, and its restoration suppresses tumorigenicity in transitional cell carcinoma cells.

Abstract: FEZ1/LZTS1 is a tumor suppressor gene that maps to chromosome 8p22, a chromosomal region frequently deleted in many human malignancies, including transitional cell carcinoma (TCC) of the urinary bladder. FEZ1/LZTS1 alterations have been reported in esophageal, breast, prostate, and gastric carcinomas. Fez1 expression was studied in five TCC-derived cancer cell lines by Western blot analysis and in 60 primary TCCs of the urinary bladder by immunohistochemistry. Fez1 protein was absent or reduced in four of five cell lines and in 37 of 60 primary TCC examined. We also restored Fez1 protein expression in human SW780 TCC-derived cells lacking endogenous Fez1 protein to study the effects of Fez1 expression on cell proliferation, cell kinetics, and tumorigenicity in BALB/c nude mice. In vitro transduction of SW780 Fez1-negative cell, with Ad-FEZ1, inhibited cell growth, altered cell cycle progression, and suppressed subcutaneous tumor growth in nude mice. These results suggest that FEZ1/LZTS1 gene plays a role in the development of TCC of the urinary bladder by acting as a bona fide tumor suppressor gene both in vitro and in vivo.

FHIT as tumor suppressor: mechanisms and therapeutic opportunities.

Abstract: Chromosomal abnormalities including homozygous deletions and loss of heterozygosity at 3p l4.2 are commonly observed in most human tumors, including lung, breast and kidney cancers. This region also contains the most common human fragile site FRA3B, a familial kidney cancer-associated translocation breakpoint and papilloma virus integration sites. The FHIT gene is a tumor suppressor, which is frequently inactivated by mentioned genomic alterations at 3pl4.2. In the last few years considerable amount of data describing inactivation of FHIT in a variety of human malignancies and demonstrating the tumor suppressor potential of Fhit has accumulated. However, these have not yet led to major advances in uncovering the precise molecular mechanism of Fhit action. This review focuses on the most recent progress in understanding of Fhit function as a tumor suppressor and opportunities for gene cancer therapy with Fhit.

Isolation and characterization of a novel gene, hRFI, preferentially expressed in esophageal cancer.

Abstract: Isolation and characterization of a novel gene, hRFI, preferentially expressed in esophageal cancer

Adenoviral transduction of fragile histidine triad (fhit) into cancer cells

Abstract: Inactivation of FHIT, a tumor suppressor gene, and loss of Fhit protein expression is observed in many human cancers. The present invention provides a method for the treatment of cancers. Specifically, the invention provides a method for inducing apoptosis in cancer cells via introduction of a tumor suppressor gene into the cells. The present invention relates to the delivery of the FHIT gene by way of an adenoviral vector, or other DNA transport system, into the cancer cells of an individual. This invention has particular application to the treatment of esophageal cancer and other tumors that carry alterations of the FHIT gene.

Crystal structures of Tcl1 family oncoproteins and their conserved surface features.

Abstract: Members of the TCL1 family of oncogenes are abnormally expressed in mature T-cell leukemias and B-cell lymphomas. The proteins are involved in the coactivation of protein kinase B (Akt/PKB), a key intracellular kinase. The sequences and crystal structures of three Tcl1 proteins were analyzed in order to understand their interactions with Akt/PKB and the implications for lymphocyte malignancies. Tcl1 proteins are approximately 15 kD and share 25-80% amino acid sequence identity. The tertiary structures of mouse Tcl1, human Tcl1, and Mtcp1 are very similar. Analysis of the structures revealed conserved semi-planar surfaces that have characteristics of surfaces involved in protein-protein interactions. The Tcl1 proteins show differences in surface charge distribution and oligomeric state suggesting that they do not interact in the same way with Akt/PKB and other cellular protein(s).

Crystal Structure of Murine Tcl1 Oncoprotein and Conserved Surface Features of the Molecules of the Tcl1 Family.

Abstract: INTRODUCTION. The structures of members of the Tcl1 oncoprotein family are being studied in order to understand their roles in lymphocyte biology and their development of lymphocytic diseases[1-3]. These ~15 kD proteins share 25–80% sequence identity between the members of the family. No sequence similarity was found with other human genes suggesting a unique cellular role(s). Family members share an uncommon tertiary structure of an eight-stranded beta barrel. Recently, the crystal structure of murine Tcl1 was determined[3]. The three structures were analyzed to reveal conserved features indicative of a potential binding site for an interacting protein.

Altered Expression of FHIT in Esophageal Neoplasm

Abstract: The FHIT (fragile histidine triad) gene located at chromosome region 3p14 has been reported to be deleted in a variety of common tumors including esophageal carcinoma

Control of apoptosis by using small molecule regulators of Bcl-2 family proteins

Abstract: The potent biological activity of CPM-1285 suggests that it may represent a promising lead for the development of new anticancer agents. The cell permeable Bcl-2 inhibitor can also be used as a chemical probe to study the in vivo mechanism and signaling pathway of the Bcl-2 family. Unlike other peptides that are active only in vitro or in the cell-free system, the cell-permeable peptide approach described here provides a new tool to analyze the function of the Bcl-2 family in living cells and animals.

Therapie par le gene fhit empechant le developpement des tumeurs chez des souris deficientes en fhit

Abstract: Il a ete observe dans une large fraction de cellules, qui sont des cellules premalignes ou malignes, une inactivation de FHIT, un gene suppresseur de tumeur. La presente invention porte sur une therapie genique dans laquelle le gene FHIT est administre, in vivo ou ex vivo, a un patient afin d'inhiber le developpement tumoral. Les procedes de therapie genique de cette invention peuvent etre utilises dans le traitement des troubles impliquant une surproliferation de cellules. Ils sont egalement utiles pour proteger un patient predispose a developper un cancer.