Reduced expression of secretogranin VGF in laryngeal squamous cell carcinoma.

Laryngeal cancer accounts for one-third of all head and neck tumors, with squamous cell carcinoma (SCC) being the most predominant type, followed by neuroendocrine tumors. Chromogranins, are commonly used as biomarkers for neuroendocrine tumors, including laryngeal cancer. It has been reported that secretogranin VGF, a member of the chromogranin family, can be also used as a significant biomarker for neuroendocrine tumors. However, the expression and role of VGF in laryngeal carcinomas have not been previously investigated. Therefore, the present study aimed to determine the expression levels of VGF in laryngeal SCC (LSCC). The present study collected tumor tissues, as well as serum samples, from a cohort of 15 patients with LSCC. The results of reverse transcription-quantitative PCR, western blot analysis and immunofluorescence assays showed that the selective VGF precursor was downregulated in patients with LSCC. Notably, in tumor tissue, the immunoreactivity for VGF was found in vimentin-positive cells, probably corresponding to T lymphocytes. The current preliminary study suggested that the reduced expression levels of VGF observed in tumor tissue and at the systemic level could sustain LSCC phenotype. Overall, VGF could be a potential biomarker for detecting neoplastic lesions with a higher risk of tumor invasiveness, even in non-neuroendocrine tumors.

Blocking prokineticin receptors attenuates synovitis and joint destruction in collagen-induced arthritis.

Rheumatoid arthritis (RA) is a chronic inflammatory disease mediated by an interdependent network of proinflammatory molecules such as chemokines. Prokineticin 2 (PK2) is a chemokine-like peptide that modulates nociceptive threshold and immuno-inflammatory processes via two G-protein-linked receptors, prokineticin receptor 1 and 2 (PKR1 and PKR2). In the present study, we investigated the effects of the prokineticin receptor antagonist PC1 on arthritic pain and the inflammatory response in type II collagen-induced arthritis (CIA) in mice. We demonstrated that PC1, administered subcutaneously from day 25 to day 35 after CIA, improved clinical signs of arthritis such as paw edema, pain, and impaired locomotor activity. In CIA mice, PC1 was also able to lower plasma malondialdehyde (MDA) levels, suggesting a role in reducing oxidative damage, as well as joint expression levels of PK2, PKRs, TNFα, IL-1ß, CD4, CD8, and NF-kB. These results suggest that blocking PKRs may be a successful strategy to control arthritic pain and pathology development. KEY MESSAGES: PK2/PKRs expression levels strongly increase in the synovium of RA mice. PC1 treatment shows anti-arthritic activity and reduces arthritis-induced pain. PC1 treatment significantly lowers synovial PK2/PKRs levels. PC1 treatment lowers plasma MDA levels and synovial levels of TNFα and IL -1ß PC1 treatment is a viable therapeutic option for RA.

Prokineticin 2 in cancer-related inflammation.

Inflammation, which triggers the release of a variety of growth factors, cytokines, and chemokines, is a critical component of tumor progression. Prokineticin 2 belongs to a new family of chemokines bound to two G-protein-coupled receptors called prokineticin receptor 1 and 2 that exert various tissue-specific biological functions. Under pathological conditions, prokineticin 2 can induce the proliferation, migration, and angiogenesis of endothelial cells, suggesting that this molecule plays a role in tumor growth, angiogenesis, and metastasis. The aim of this review is to provide a complete compendium of the involvement of prokineticin 2 in some cancers and to evaluate its role not only in the tumor microenvironment as an angiogenic factor and a mediator of immune cell migration, but also in modulating tumor growth and spread as a suppressor of tumor cell apoptosis, and as a trigger of their proliferation and movements required for metastasis. The involvement of prokineticin 2 in tumor pain and resistance responses is also described, and finally, the potential role of prokineticin 2 as a novel prognostic tumor biomarker is highlighted.

Pilot Investigation on p75ICD Expression in Laryngeal Squamous Cell Carcinoma.

We investigated the p75 Neurotrophin Receptor (p75NTR) expression and cleavage product p75NTR Intracellular Domain (p75ICD) as potential oncogenic and metastatic markers in human Laryngeal Squamous Cell Carcinoma (LSCC). p75NTR is highly expressed in Cancer Stem Cells (CSCs) of the laryngeal epithelia and it has been proposed as a marker for stemness, cell migration, and chemo-resistance in different squamous carcinomas. To investigate the clinical significance of p75NTR cleavage products in solid tumors, full-length and cleaved p75NTR expression was analyzed in laryngeal primary tumors from different-stage LSCC patients, diagnosed at the Policlinico Umberto I Hospital. Molecular and histological techniques were used to detect the expressions of p75NTR and p75ICD, and ATP Binding Cassette Subfamily G Member 2 (ABCG2), a CSC marker. We found regulated p75NTR cleavage during squamous epithelial tumor progression and tissue invasion. Our preliminary investigation suggests p75ICD expression and localization as possible features of tumorigenesis and metastaticity. Its co-localization with ABCG2 in squamous cells in the parenchyma invaded by the tumor formation allows us to hypothesize p75NTR and p75ICD roles in tumor invasion and CSC spreading in LSCC patients. These data might represent a starting point for a comprehensive analysis of p75NTR cleavage and of its clinical relevance as a potential molecular LSCC signature, possibly helping diagnosis, and improving prognosis and personalized therapy.

Transcriptional Profiles of Cell Fate Transitions Reveal Early Drivers of Neuronal Apoptosis and Survival.

Neuronal apoptosis and survival are regulated at the transcriptional level. To identify key genes and upstream regulators primarily responsible for these processes, we overlayed the temporal transcriptome of cerebellar granule neurons following induction of apoptosis and their rescue by three different neurotrophic factors. We identified a core set of 175 genes showing opposite expression trends at the intersection of apoptosis and survival. Their functional annotations and expression signatures significantly correlated to neurological, psychiatric and oncological disorders. Transcription regulatory network analysis revealed the action of nine upstream transcription factors, converging pro-apoptosis and pro-survival-inducing signals in a highly interconnected functionally and temporally ordered manner. Five of these transcription factors are potential drug targets. Transcriptome-based computational drug repurposing produced a list of drug candidates that may revert the apoptotic core set signature. Besides elucidating early drivers of neuronal apoptosis and survival, our systems biology-based perspective paves the way to innovative pharmacology focused on upstream targets and regulatory networks.

Mediterranean Diet, Brain and Muscle: Olive Polyphenols and Resveratrol Protection in Neurodegenerative and Neuromuscular Disorders.

The Mediterranean diet is worldwide recognized as a good prototype of nutrition due to the conspicuous intake of olive oil, nuts, red wine, legumes, fruit, and vegetables, all fundamental elements rich in antioxidant substances and polyphenols. Polyphenols are a wide range of phytochemicals and/or synthetic chemical compounds with proven beneficial properties for human health. In the present review, we critically summarize the wellcharacterized antioxidant and anti-inflammatory properties of polyphenols contained in the olives and extra virgin olive oil and of resveratrol, a non-flavonoid phenolic compound. We discuss the potential use of these polyphenols as pharmaceutical formulations for the treatment of human diseases. We also show the emerging importance of their consumption in the prevention and management of crucial neurodegenerative conditions (alcohol-related brain disorders and aging) and in neuromuscular disorders (Spinal Muscular Atrophy and Amyotrophic Lateral Sclerosis and Duchenne Muscular Dystrophy), where oxidative stress plays a predominant role.

Systemic Amyloidosis: a Contemporary Overview.

Amyloidosis constitutes a large spectrum of diseases characterized by an extracellular deposition of a fibrillar aggregate, generating insoluble and toxic amasses that may be deposited in tissues in bundles with an abnormal cross-ß-sheet conformation, known as amyloid. Amyloid may lead to a cell damage and an impairment of organ function. Several different proteins are recognized as able to produce amyloid fibrils with a different tissue tropism related to the molecular structure. The deposition of amyloid may occur as a consequence of the presence of an abnormal protein, caused by high plasma levels of a normal protein, or as a result of the aging process along with some environmental factors. Although amyloidosis is rare, amyloid deposits play a role in several conditions as degenerative diseases. Thus, the development of antiamyloid curative treatments may be a rational approach to treat neurodegenerative conditions like Alzheimer's disease in the future. Nowadays, novel treatment options are currently refined through controlled trials, as new drug targets and different therapeutic approaches have been identified and validated through modern advances in basic research. Fibril formation stabilizers, proteasome inhibitors, and immunotherapy revealed promising results in improving the outcomes of patients with systemic amyloidosis, and these novel algorithms will be effectively combined with current treatments based on chemotherapeutic regimens. The aim of this review is to provide an update on diagnosis and treatment for systemic amyloidosis.

Utrophin up-regulation by artificial transcription factors induces muscle rescue and impacts the neuromuscular junction in mdx mice.

Up-regulation of the dystrophin-related gene utrophin represents a promising therapeutic strategy for the treatment of Duchenne Muscular Dystrophy (DMD). In order to re-program the utrophin expression level in muscle, we engineered artificial zinc finger transcription factors (ZF-ATFs) that target the utrophin 'A' promoter. We have previously shown that the ZF-ATF "Jazz", either by transgenic manipulation or by systemic adeno-associated viral delivery, induces significant rescue of muscle function in dystrophic "mdx" mice. We present the full characterization of an upgraded version of Jazz gene named "JZif1" designed to minimize any possible host immune response. JZif1 was engineered on the Zif268 gene-backbone using selective amino acid substitutions to address JZif1 to the utrophin 'A' promoter. Here, we show that JZif1 induces remarkable amelioration of the pathological phenotype in mdx mice. To investigate the molecular mechanisms underlying Jazz and JZif1 induced muscle functional rescue, we focused on utrophin related pathways. Coherently with utrophin subcellular localization and role in neuromuscular junction (NMJ) plasticity, we found that our ZF-ATFs positively impact the NMJ. We report on ZF-ATF effects on post-synaptic membranes in myogenic cell line, as well as in wild type and mdx mice. These results candidate our ZF-ATFs as novel therapeutic molecules for DMD treatment.

Bromelain Degrades Aß1-42 Monomers and Soluble Aggregates: An In Vitro Study in Cerebrospinal Fluid of Alzheimer's Disease Patients.

BACKGROUND: Therapeutic approaches targeting amyloid ß42 (Aß42) oligomers may represent a promising neuroprotective strategy for the prevention and treatment of Alzheimer's disease (AD). OBJECTIVE: In this study we evaluated the ability of bromelain, a plant cysteine protease derived from pineapple stems, to interact with synthetic Aß42 monomers and oligomers. We also examined the ability of bromelain to interfere in vitro with synthetic Aß42 aggregates in the cerebrospinal fluid (CSF) of Alzheimer's disease as well as of control patients affected by other neurological diseases. METHOD: Both synthetic monomers and aggregates of Aß42 were incubated in CSF with varying concentrations of bromelain. The effects of digestion were evaluated by Western Blot analysis using the specific monoclonal antibody 4G8 to identify the patterns of residual content of Aß42. We further used rat primary cortical culture neurons (CN) to examine the cytotoxic action of this natural compound. RESULTS: We found that bromelain successfully degraded Aß42 monomers and low and high molecular weight oligomers. Indeed, when bromelain preparations of 3 and 6 mU were added to the CSF, the residual amount of Aß42 monomers and oligomers were significantly reduced when compared to the same standard Aß42 preparations incubated in CSF without bromelain. Moreover, bromelain incubations of 0.1, 0.5, and 1 mU/ml were not toxic to CN, as compared to vehicle treated cells. CONCLUSION: Overall, these results represent an important insight into the action of bromelain on Aß42 oligomers, suggesting its potential use in the therapy of AD.

The Intersection of NGF/TrkA Signaling and Amyloid Precursor Protein Processing in Alzheimer's Disease Neuropathology.

Dysfunction of nerve growth factor (NGF) and its high-affinity Tropomyosin receptor kinase A (TrkA) receptor has been suggested to contribute to the selective degeneration of basal forebrain cholinergic neurons (BFCN) associated with the progressive cognitive decline in Alzheimer's disease (AD). The aim of this review is to describe our progress in elucidating the molecular mechanisms underlying the dynamic interplay between NGF/TrkA signaling and amyloid precursor protein (APP) metabolism within the context of AD neuropathology. This is mainly based on the finding that TrkA receptor binding to APP depends on a minimal stretch of ~20 amino acids located in the juxtamembrane/extracellular domain of APP that carries the α- and ß-secretase cleavage sites. Here, we provide evidence that: (i) NGF could be one of the "routing" proteins responsible for modulating the metabolism of APP from amyloidogenic towards non-amyloidogenic processing via binding to the TrkA receptor; (ii) the loss of NGF/TrkA signaling could be linked to sporadic AD contributing to the classical hallmarks of the neuropathology, such as synaptic loss, ß-amyloid peptide (Aß) deposition and tau abnormalities. These findings will hopefully help to design therapeutic strategies for AD treatment aimed at preserving cholinergic function and anti-amyloidogenic activity of the physiological NGF/TrkA pathway in the septo-hippocampal system.

Drug target identification at the crossroad of neuronal apoptosis and survival.

INTRODUCTION: Inappropriate activation of apoptosis may contribute to neurodegeneration, a multifaceted process that results in various chronic disorders, including Alzheimer's and Parkinson's diseases. Several in vitro and in vivo studies demonstrated that neuronal apoptosis is a multi-pathway cell-death program that requires RNA synthesis. Thus, transcriptionally activated genes whose products induce cell death can be triggered by different stimuli and antagonized by neurotrophic factors. Systems biology is now unveiling the series of intracellular signaling pathways and key drug targets at the intersection of neuronal apoptosis and survival. Areas covered: This review introduces a genomic approach that can be used to elucidate the systems biology of neuronal apoptosis and survival, and to rationally select drug targets, no longer oriented to emulate the action of growth factors at the membrane receptor level, but rather to modulate their downstream signals. Expert opinion: The advent of genomics is offering an unprecedented opportunity to explore how the delicate balance between apoptosis and survival-inducing signals triggers a transcriptional program. Characterization of this program can be useful to identify potential pharmacological targets for existing drugs. Such knowledge might pave the way towards an innovative pharmacology.

Bv8/prokineticin 2 is involved in Aß-induced neurotoxicity.

Bv8/Prokineticin 2 (PROK2) is a bioactive peptide initially discovered as a regulator of gastrointestinal motility. Among multiple biological roles demonstrated for PROK2, it was recently established that PROK2 is an insult-inducible endangering mediator for cerebral damage. Aim of the present study was to evaluate the PROK2 and its receptors' potential involvement in amyloid beta (Aß) neurotoxicity, a hallmark of Alzheimer's disease (AD) and various forms of traumatic brain injury (TBI). Analyzing primary cortical cultures (CNs) and cortex and hippocampus from Aß treated rats, we found that PROK2 and its receptors PKR1 and PKR2 mRNA are up-regulated by Aß, suggesting their potential involvement in AD. Hence we evaluated if impairing the prokineticin system activation might have protective effect against neuronal death induced by Aß. We found that a PKR antagonist concentration-dependently protects CNs against Aß(1-42)-induced neurotoxicity, by reducing the Aß-induced PROK2 neuronal up-regulation. Moreover, the antagonist completely rescued LTP impairment in hippocampal slices from 6 month-old Tg2576 AD mice without affecting basal synaptic transmission and paired pulse-facilitation paradigms. These results indicate that PROK2 plays a role in cerebral amyloidosis and that PROK2 antagonists may represent a new approach for ameliorating the defining pathology of AD.

The VGF-derived peptide TLQP-62 modulates insulin secretion and glucose homeostasis.

Insulin secretion control is critical for glucose homeostasis. Paracrine and autocrine molecules secreted by cells of the islet of Langerhans, as well as by intramural and autonomic neurons, control the release of different hormones that modulate insulin secretion. In pancreatic islets, the abundant presence of the granin protein VGF (nonacronymic; unrelated to VEGF) suggests that some of its proteolytically derived peptides could modulate hormone release. Thus, in the present study, we screened several VGF-derived peptides for their ability to induce insulin secretion, and we identified the VGF C-terminal peptide TLQP-62 as the most effective fragment. TLQP-62 induced a potent increase in basal insulin secretion as well as in glucose-stimulated insulin secretion in several insulinoma cell lines. We found that this peptide stimulated insulin release via increased intracellular calcium mobilization and fast expression of the insulin 1 gene. Moreover, the peripheral injection of TLQP-62 in mice improved glucose tolerance. Together, the present findings suggest that TLQP-62, acting as an endocrine, paracrine, or autocrine factor, can be considered a new, strong insulinotropic peptide that can be targeted for innovative antidiabetic drug discovery programs.

Igf1 and Pacap rescue cerebellar granule neurons from apoptosis via a common transcriptional program.

A shift of the delicate balance between apoptosis and survival-inducing signals determines the fate of neurons during the development of the central nervous system and its homeostasis throughout adulthood. Both pathways, promoting or protecting from apoptosis, trigger a transcriptional program. We conducted whole-genome expression profiling to decipher the transcriptional regulatory elements controlling the apoptotic/survival switch in cerebellar granule neurons following the induction of apoptosis by serum and potassium deprivation or their rescue by either insulin-like growth factor-1 (Igf1) or pituitary adenylyl cyclase-activating polypeptide (Pacap). Although depending on different upstream signaling pathways, the survival effects of Igf1 and Pacap converged into common transcriptional cascades, thus suggesting the existence of a general transcriptional program underlying neuronal survival.

Novel adeno-associated viral vector delivering the utrophin gene regulator jazz counteracts dystrophic pathology in mdx mice.

Over-expression of the dystrophin-related gene utrophin represents a promising therapeutic strategy for Duchenne muscular dystrophy (DMD). The strategy is based on the ability of utrophin to functionally replace defective dystrophin. We developed the artificial zinc finger transcription factor "Jazz" that up-regulates both the human and mouse utrophin promoter. We observed a significant recovery of muscle strength in dystrophic Jazz-transgenic mdx mice. Here we demonstrate the efficacy of an experimental gene therapy based on the systemic delivery of Jazz gene in mdx mice by adeno-associated virus (AAV). AAV serotype 8 was chosen on the basis of its high affinity for skeletal muscle. Muscle-specific expression of the therapeutic Jazz gene was enhanced by adding the muscle α-actin promoter to the AAV vector (mAAV). Injection of mAAV8-Jazz viral preparations into mdx mice resulted in muscle-specific Jazz expression coupled with up-regulation of the utrophin gene. We show a significant recovery from the dystrophic phenotype in mAAV8-Jazz-treated mdx mice. Histological and physiological analysis revealed a reduction of fiber necrosis and inflammatory cell infiltration associated with functional recovery in muscle contractile force. The combination of ZF-ATF technology with the AAV delivery can open a new avenue to obtain a therapeutic strategy for treatment of DMD.

Bindarit, inhibitor of CCL2 synthesis, protects neurons against amyloid-ß-induced toxicity.

One of the hallmarks of Alzheimer's disease (AD), the most common age-related neurodegenerative pathology, is the abnormal extracellular deposition of neurotoxic amyloid-ß (Aß) peptides that accumulate in senile plaques. Aß aggregates are toxic to neurons and are thought to contribute to neuronal loss. Evidence indicates that inflammation is involved in the pathophysiology of AD, and activation of glial cells by a variety of factors, including Aß, appears to be a central event. Among molecules produced during inflammation associated with neuronal death, CCL2, also known as monocyte chemotactic protein-1 (MCP-1), seems to be particularly important. Indeed, CCL2 levels are higher in the cerebrospinal fluid of patients with AD than in controls. In the present study, we demonstrated the protective effect of bindarit (which inhibits CCL2 synthesis) against both Aß25-35 and Aß1-42-induced toxicity in primary mixed neural cultures. Bindarit (30-500 µM) reversed cell death induced by Aß in a dose-dependent manner and reduced the transcription and release of CCL2 by astrocytes after Aß treatment, as revealed by qRT-PCR, ELISA, and immunofluorescence staining. Astroglial activation and CCL2 release was induced by ATP released by damaged neurons through interaction with P2X7 receptors present on astrocyte surface. CCL2, interacting with its cognate receptor CCR2, present on neuron surface, strongly contributes to the toxic activity of Aß. Bindarit was able to disconnect this neuro-glial interaction. Our results demonstrate the ability of bindarit to inhibit Aß-induced neuronal death and suggest the potential role of CCL2 inhibitors in the treatment of neuroinflammatory/neurodegenerative diseases.

Systemic administration of substance P recovers beta amyloid-induced cognitive deficits in rat: involvement of Kv potassium channels.

Reduced levels of Substance P (SP), an endogenous neuropeptide endowed with neuroprotective and anti-apoptotic properties, have been found in brain and spinal fluid of Alzheimer's disease (AD) patients. Potassium (K(+)) channel dysfunction is implicated in AD development and the amyloid-ß (Aß)-induced up-regulation of voltage-gated potassium channel subunits could be considered a significant step in Aß brain toxicity. The aim of this study was to evaluate whether SP could reduce, in vivo, Aß-induced overexpression of Kv subunits. Rats were intracerebroventricularly infused with amyloid-ß 25-35 (Aß25-35, 20 µg) peptide. SP (50 µg/Kg, i.p.) was daily administered, for 7 days starting from the day of the surgery. Here we demonstrate that the Aß infused rats showed impairment in cognitive performances in the Morris water maze task 4 weeks after Aß25-35 infusion and that this impairing effect was prevented by SP administration. Kv1.4, Kv2.1 and Kv4.2 subunit levels were quantified in hippocampus and in cerebral cortex by Western blot analysis and immunofluorescence. Interestingly, SP reduced Kv1.4 levels overexpressed by Aß, both in hippocampus and cerebral cortex. Our findings provide in vivo evidence for a neuroprotective activity of systemic administration of SP in a rat model of AD and suggest a possible mechanism underlying this effect.

Neuropeptide TLQP-21, a VGF internal fragment, modulates hormonal gene expression and secretion in GH3 cell line.

In the present study we demonstrated that TLQP-21, a biologically active peptide derived from the processing of the larger pro-VGF granin, plays a role in mammotrophic cell differentiation. We used an established in vitro model, the GH3 cell line, which upon treatment with epidermal growth factor develops a mammotrophic phenotype consisting of induction of prolactin expression and secretion, and inhibition of growth hormone. Here we determined for the first time that during mammotrophic differentiation, epidermal growth factor also induces Vgf gene expression and increases VGF protein precursor processing and peptide secretion. After this initial observation we set out to determine the specific role of the VGF encoded TLQP-21 peptide on this model. TLQP-21 induced a trophic effect on GH3 cells and increased prolactin expression and its own gene transcription without affecting growth hormone expression. TLQP-21 was also able to induce a significant rise of cytoplasmic calcium, as measured by Fura2AM, due to the release from a thapsigargin-sensitive store. TLQP-21-dependent rise in cytoplasmic calcium was, at least in part, dependent on the activation of phospholipase followed by phosphorylation of PKC and ERK. Taken together, the present results demonstrate that TLQP-21 contributes to differentiation of the GH3 cell line toward a mammotrophic phenotype and suggest that it may exert a neuroendocrine role in vivo on lactotroph cells in the pituitary gland.

Gene expression changes in medical workers exposed to radiation.

The use of nuclear resources for medical purposes causes considerable concern about occupational exposure. Nevertheless, little information is available regarding the effects of low-dose irradiations protracted over time. We used oligomicroarrays to identify the genes that are transcriptionally regulated by persistent exposure to extremely low doses of ionizing radiation in 28 exposed professionals (mean cumulative effective dose +/- SD, 19 +/- 38 mSv) compared with a matched sample of nonexposed subjects. We identified 256 modulated genes from peripheral blood mononuclear cells profiles, and the main biological processes we found were DNA packaging and mitochondrial electron transport NADH to ubiquinone. Next we investigated whether a different pattern existed when only 22 exposed subjects with accumulated doses >2.5 mSv, a threshold corresponding to the natural background radiation in Italy per year, and mean equal to 25 +/- 41 mSv were used. In addition to DNA packaging and NADH dehydrogenase function, the analysis of the higher-exposed subgroup revealed a significant modulation of ion homeostasis and programmed cell death as well. The changes in gene expression that we found suggest different mechanisms from those involved in high-dose studies that may help to define new biomarkers of radiation exposure for accumulated doses below 25 mSv.

Gene expression time-series analysis of camptothecin effects in U87-MG and DBTRG-05 glioblastoma cell lines.

BACKGROUND: The clinical efficacy of camptothecin (CPT), a drug specifically targeting topoisomerase I (TopoI), is under evaluation for the treatment of malignant gliomas. Due to the high unresponsiveness of these tumours to chemotherapy, it would be very important to study the signalling network that drives camptothecin outcome in this type of cancer cells. To address this issue, we had previously compared the expression profile of human U87-MG glioblastoma cells with that of a CPT-resistant counterpart, giving evidence that the development of a robust inflammatory response was the main transcriptional effect associated with CPT resistance. Here we report time-related changes and cell line specific patterns of gene expression after CPT treatment by using two p53 wild-type glioblastoma cell lines, U87-MG and DBTRG-05, with different sensitivities to TopoI inhibition. RESULTS: First, we demonstrated that CPT treatment brings the two cell lines to completely different outcomes: accelerated senescence in U87-MG and apoptosis in DBTRG-05 cells. Then, to understand the different susceptibility to CPT, we used oligo-microarray to identify the genes whose expression was regulated during a time-course treatment, ranging from 2 h to 72 h. The statistical analysis of microarray data by MAANOVA (MicroArray ANalysis Of VAriance) showed much less modulated genes in apoptotic DBTRG-05 cells (155) with respect to the senescent U87-MG cells (3168), where the number of down-regulated genes largely exceeded that of the up-regulated ones (80% vs. 20%). Despite this great difference, the two data-sets showed a large overlapping (60% circa) mainly due to the expression of early stress responsive genes. The use of High-Throughput GoMINER and EASE tools, for functional analysis of significantly enriched GO terms, highlighted common cellular processes and showed that U87-MG and DBTRG-05 cells shared many GO terms, which are related to the down-regulation of cell cycle and mitosis and to the up-regulation of cell growth inhibition and DNA damage.Furthermore, the down-regulation of MYC and DP1 genes, which act as key transcription factors in cell growth control, together with the inhibition of BUB1, BUB3 and MAD2 mRNAs, which are known to be involved in the spindle checkpoint pathway, were specifically associated with the execution of senescence in U87-MG cells and addressed as critical factors that could drive the choice between different CPT-inducible effectors programs. In U87-MG cells we also found inflammation response and IL1-beta induction, as late transcriptional effects of Topo I treatment but these changes were only partially involved in the senescence development, as shown by IL1-beta gene silencing. CONCLUSION: By comparing the transcription profile of two glioblastoma cell lines treated with camptothecin, we were able to identify the common cellular pathways activated upon Topo I inhibition. Moreover, our results helped in identifying some key genes whose expression seemed to be associated with the execution of senescence or apoptosis in U87-MG and DBTRG-05 cells, respectively.