[Show abstract][Hide abstract] ABSTRACT: Signal Transducer and Activator of Transcription factors (STATs) are proteins able to translocate into the nucleus, bind DNA and activate gene transcription. STATs proteins play a crucial role in cell proliferation, apoptosis and differentiation. The prevalent view is that STATs proteins are able to form dimers and bind DNA only upon phosphorylation of specific tyrosine residues in the Trans-Activation-domain. However, this paradigm has been questioned recently by the observation of dimers of unphosphorylated STATs (USTATs) by X-ray, FRET and site-directed mutagenesis. A more complex picture of the dimerization process and of the role of the dimers is, thus, emerging. Here we present an integrated modeling study of STAT3, a member of the STAT family of utmost importance in cancer development and therapy, in which we combine available experimental data with several computational methodologies such as homology modeling, protein-protein docking and molecular dynamics, to build reliable atomistic models of USTAT3 dimers. The models generated with integrative approach presented here, were, then, validated by performing computational alanine scanning for all the residues in the protein-protein interface. These results confirmed the experimental observation of the importance of some of these residues (in particular Leu78 and Asp19) in the USTAT3 dimerization process. Given the growing importance of USTAT3 dimers in several cellular pathways, our models provide an important tool to study the effects of pathological mutations at molecular/atomistic level, and in the rational design of new inhibitors of dimerization.
[Show abstract][Hide abstract] ABSTRACT: Enhancement of cellular senescence in tumours triggers a stable cell growth arrest and activation of an antitumour immune response that can be exploited for cancer therapy. Currently, there are only a limited number of targeted therapies that act by increasing senescence in cancers, but the majority of them are not selective and also target healthy cells. Here we developed a chemogenomic screening to identify compounds that enhance senescence in PTEN-deficient cells without affecting normal cells. By using this approach, we identified casein kinase 2 (CK2) as a pro-senescent target. Mechanistically, we show that Pten loss increases CK2 levels by activating STAT3. CK2 upregulation in Pten null tumours affects the stability of Pml, an essential regulator of senescence. However, CK2 inhibition stabilizes Pml levels enhancing senescence in Pten null tumours. Taken together, our screening strategy has identified a novel STAT3-CK2-PML network that can be targeted for pro-senescence therapy for cancer.
[Show abstract][Hide abstract] ABSTRACT: MicroRNAs (miRNAs) originate from stem-loop-containing precursors (pre-miRNAs, pri-miRNAs) and mature by means of the Drosha and Dicer endonucleases and their associated factors. The let-7 miRNAs have prominent roles in developmental differentiation and in regulating cell proliferation. In cancer, the tumor suppressor function of let-7 is abrogated by overexpression of Lin28, one of several RNA-binding proteins that regulate let-7 biogenesis by interacting with conserved motifs in let-7 precursors close to the Dicer cleavage site. Using in vitro assays, we have identified a binding site for short modified oligoribonucleotides ('looptomirs') overlapping that of Lin28 in pre-let-7a-2. These looptomirs selectively antagonize the docking of Lin28, but still permit processing of pre-let-7a-2 by Dicer. Looptomirs restored synthesis of mature let-7 and inhibited growth and clonogenic potential in Lin28 overexpressing hepatocarcinoma cells, thereby demonstrating a promising new means to rescue defective miRNA biogenesis in Lin28-dependent cancers.
Nucleic Acids Research 11/2014; 43(2). DOI:10.1093/nar/gku1090 · 9.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Prosenescence therapy has recently emerged as a novel therapeutic approach for treating cancer. However, this concept is challenged by conflicting evidence showing that the senescence-associated secretory phenotype (SASP) of senescent tumor cells can have pro-as well as antitumorigenic effects. Herein, we report that, in Pten-null senescent tumors, activation of the Jak2/Stat3 pathway establishes an immunosuppressive tumor microenvironment that contributes to tumor growth and chemoresistance. Activation of the Jak2/Stat3 pathway in Pten-null tumors is sustained by the downregulation of the protein tyrosine phosphatase PTPN11/SHP2, providing evidence for the existence of a novel PTEN/SHP2 axis. Importantly, treatment with docetaxel in combination with a JAK2 inhibitor reprograms the SASP and improves the efficacy of docetaxel-induced senescence by triggering a strong antitumor immune response in Pten-null tumors. Altogether, these data demonstrate that immune surveillance of senescent tumor cells can be suppressed in specific genetic backgrounds but also evoked by pharmacological treatments.
[Show abstract][Hide abstract] ABSTRACT: Several studies link disease progression, recurrence and treatment failures to the cancer stem-like cell (CSC) subpopulation within the heterogeneous tumor cell population. Myc is a transcription factor having a central function in stem cell biology and in human cancers. Hence, Myc represents an attractive target to develop CSC-specific therapies. Recent findings suggest that Myc transcription can be silenced using an RNAi-based strategy that targets noncoding promoter-associated RNA (paRNA) overlapping the transcription start site. In this study, we investigated the effects of silencing Myc transcription on prostate CSC in cell culture and xenograft models of human prostate cancer. Treatment with an effective promoter-targeting siRNA reduced the fraction of CSCs leading to reduced self-renewal, tumor-initiating and metastatic capability. Combined analysis of stem-like cells and senescence markers indicated that Myc silencing triggered a phenotypic shift and senescence in the CSC subpopulation. Notably, systemic delivery of the promoter-targeting siRNA in the xenograft model produced a striking suppression in the development of prostate tumors. Our results support a pivotal role for Myc in CSC maintenance and show that Myc targeting via RNAi-based transcriptional silencing can trigger CSC senescence and loss of their tumor-initiating capability. More generally, our findings demonstrate the efficacy of RNAi-based transcriptional strategies and the potential to target regulatory noncoding paRNAs for therapeutic applications.
Cancer Research 09/2013; 73(22). DOI:10.1158/0008-5472.CAN-13-0615 · 9.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: RNAi-based nucleic acid molecules have attracted considerable attention as compelling therapeutics providing safe and competent delivery systems are available. Dendrimers are emerging as appealing nanocarriers for nucleic acid delivery thanks to their unique well-defined architecture and the resulting cooperativity and multivalency confined within a nanostructure. The present review offers a brief overview of the structurally flexible triethanolamine-core poly(amidoamine) (PAMAM) dendrimers developed in our group as nanovectors for the delivery of RNAi therapeutics. Their excellent activity for delivering different RNAi therapeutics in various disease models in vitro and in vivo will be highlighted here.
[Show abstract][Hide abstract] ABSTRACT: Testin (TES) is a putative tumour-suppressor gene downregulated in various types of cancers. Survivin is a nodal protein involved in multiple signalling pathways, tumour maintenance and inhibition of apoptosis. Previous studies indicate that TES and survivin can functionally interact and modulate cell death and proliferation in breast cancer cells. The aim of the present study was to investigate the expression and prognostic relevance of TES and survivin in breast cancer subtypes examining a large cohort of breast cancer patients. We determined the expression of TES and survivin by immunohistochemistry (IHC) in tissue samples from 242 breast cancer patients diagnosed between 1981 and 2009. The expression of these proteins was compared with clinical and pathological data. There was a significant association of nuclear survivin overexpression and TES downregulation with triple-negative tumours [P=0.009; univariate odds ratio (OR), 3.20; 95% CI, 1.34-7.66] (P=0.018; multivariate OR, 2.90; 95% CI, 1.20‑6.97). A further significant correlation was observed between TES downregulation and the luminal B subtype (P=0.019, univariate OR: 2.90; 95% CI, 1.19‑7.06) (P=0.032, multivariate OR, 2.67; 95% CI, 1.09-6.65), independent of survivin expression. Our results demonstrated a statistically significant association between TES downregulation and highly aggressive breast tumour subtypes, such as triple-negative and luminal B tumours, along with the prognostic relevance of nuclear expression of survivin. To our knowledge, this is the first demonstration of such an association.
[Show abstract][Hide abstract] ABSTRACT: Chromosomal translocations leading to deregulated expression of ETS transcription factors are frequent in prostate tumors. Here, we report a novel mechanism leading to oncogenic activation of the ETS factor ESE1/ELF3 in prostate tumors. ESE1/ELF3 was overexpressed in human primary and metastatic tumors. It mediated transforming phenotypes in vitro and in vivo and induced an inflammatory transcriptome with changes in relevant oncogenic pathways. ESE1/ELF3 was induced by IL-1β through NF-κB and was a crucial mediator of the phenotypic and transcriptional changes induced by IL-1β in prostate cancer cells. This linkage was mediated by interaction of ESE1/ELF3 with the NF-κB subunits p65 and p50, acting by enhancing their nuclear translocation and transcriptional activity and by inducing p50 transcription. Supporting these findings, gene expression profiling revealed an enrichment of NF-κB effector functions in prostate cancer cells or tumors expressing high levels of ESE1/ELF3. We observed concordant upregulation of ESE1/ELF3 and NF-κB in human prostate tumors that was associated with adverse prognosis. Collectively, our results define an important new mechanistic link between inflammatory signaling and the progression of prostate cancer.
Cancer Research 05/2013; 73(14). DOI:10.1158/0008-5472.CAN-12-4537 · 9.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Peroxisome proliferator-activated receptor β/δ (PPARβ/δ) is a nuclear receptor involved in regulation of lipid and glucose metabolism, wound healing and inflammation. PPARβ/δ has been associated also with cancer. Here we investigated the expression of PPARβ/δ and components of the prostaglandin biosynthetic pathway in non-small cell lung cancer (NSCLC). We found increased expression of PPARβ/δ, Cox-2, cPLA(2), PGES and VEGF in human NSCLC compared to normal lung. In NSCLC cell lines PPARβ/δ activation increased proliferation and survival, while PPARβ/δ knock-down reduced viability and increased apoptosis. PPARβ/δ agonists induced Cox-2 and VEGF transcription, suggesting the existence of feed-forward loops promoting cell survival, inflammation and angiogenesis. These effects were seen only in high PPARβ/δ expressing cells, while low expressing cells were less or not affected. The effects were also abolished by PPARβ/δ knock-down or incubation with a PPARβ/δ antagonist. Induction of VEGF was due to both binding of PPARβ/δ to the VEGF promoter and PI3K activation through a non-genomic mechanism. We found that PPARβ/δ interacted with the PI3K regulatory subunit p85α leading to PI3K activation and Akt phosphorylation. Collectively, these data indicate that PPARβ/δ might be a central element in lung carcinogenesis controlling multiple pathways and representing a potential target for NSCLC treatment.
PLoS ONE 09/2012; 7(9):e46009. DOI:10.1371/journal.pone.0046009 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pegylated liposomal doxorubicin (PLD) is an established treatment for relapsed ovarian cancer. Preclinical and clinical evidences in other tumor types suggest that the proteasome inhibitor bortezomib can act synergistically with PLD.
Patients with relapsed ovarian cancer (N = 58), previously treated with platinum (100%) and taxane (95%), received bortezomib, 1.3 mg/m intravenous (days 1, 4, 8, and 11), and PLD, 30 mg/m intravenous (day 1), every 3 weeks. Tumor responses were assessed using Response Evaluation Criteria In Solid Tumors and Gynecologic Cancer Intergroup criteria. An optimal 2-stage design was implemented. Gene expression profiling in peripheral blood was characterized before and during treatment in 10 platinum-sensitive patients enrolled in stage 2 of the study.
Median number of bortezomib-PLD cycles was 3.5. Of 38 patients in the platinum-sensitive group, 9 responses were observed (median duration, 4.8 months). The platinum-resistant group was closed at stage 1 owing to lack of response. Toxicity was moderate and mainly consisted of hematologic, gastrointestinal, and mucositis events. Of the total 58 patients, peripheral neuropathy was reported in 9 patients (none were grade 3). Transcription profiling identified the prevalence of genes associated with ribonucleoprotein complexes, RNA processing, and protein translation. The gene expression changes were more robust in patients who responded or had stable disease compared with patients who had progressive disease.
The combination of bortezomib and PLD was well tolerated, but the antitumor activity is insufficient to warrant further investigation in ovarian cancer.
International Journal of Gynecological Cancer 06/2012; 22(5):792-800. DOI:10.1097/IGC.0b013e318251051a · 1.95 Impact Factor