Pierpaolo Peruzzi

MicroRNAs in cancer: biomarkers, functions and therapy

The emergence of microRNAs has been one of the defining developments in cancer biology over the past decade, and the explosion of knowledge in this area has brought forward new diagnostic and therapeutic opportunities. The importance of microRNAs in cancer has been underlined by the identification of alterations in microRNA target binding sites and the microRNA processing machinery in tumor cells. Clinical trials utilizing microRNA profiling for patient prognosis and clinical response are now underway, and the first microRNA mimic entered the clinic for cancer therapy in 2013. In this article we review the potential applications of microRNAs for the clinical assessment of patient outcome in cancer, as well as in cancer monitoring and therapy.

Extracellular Vesicles Modulate the Glioblastoma Microenvironment via a Tumor Suppression Signaling Network Directed by miR-1

Extracellular vesicles have emerged as important mediators of intercellular communication in cancer, including by conveying tumor-promoting microRNAs between cells, but their regulation is poorly understood. In this study, we report the findings of a comparative microRNA profiling and functional analysis in human glioblastoma that identifies miR-1 as an orchestrator of extracellular vesicle function and glioblastoma growth and invasion. Ectopic expression of miR-1 in glioblastoma cells blocked in vivo growth, neovascularization, and invasiveness. These effects were associated with a role for miR-1 in intercellular communication in the microenvironment mediated by extracellular vesicles released by cancer stem-like glioblastoma cells. An extracellular vesicle-dependent phenotype defined by glioblastoma invasion, neurosphere growth, and endothelial tube formation was mitigated by loading miR-1 into glioblastoma-derived extracellular vesicles. Protein cargo in extracellular vesicles was characterized to learn how miR-1 directed extracellular vesicle function. The mRNA encoding Annexin A2 (ANXA2), one of the most abundant proteins in glioblastoma-derived extracellular vesicles, was found to be a direct target of miR-1 control. In addition, extracellular vesicle-derived miR-1 along with other ANXA2 extracellular vesicle networking partners targeted multiple pro-oncogenic signals in cells within the glioblastoma microenvironment. Together, our results showed how extracellular vesicle signaling promotes the malignant character of glioblastoma and how ectopic expression of miR-1 can mitigate this character, with possible implications for how to develop a unique miRNA-based therapy for glioblastoma management.

MicroRNA-128 coordinately targets Polycomb Repressor Complexes in glioma stem cells

BACKGROUND The Polycomb Repressor Complex (PRC) is an epigenetic regulator of transcription whose action is mediated by 2 protein complexes, PRC1 and PRC2. PRC is oncogenic in glioblastoma, where it is involved in cancer stem cell maintenance and radioresistance. METHODS We used a set of glioblastoma patient samples, glioma stem cells, and neural stem cells from a mouse model of glioblastoma. We characterized gene/protein expression and cellular phenotypes by quantitative PCR/Western blotting and clonogenic, cell-cycle, and DNA damage assays. We performed overexpression/knockdown studies by lentiviral infection and microRNA/small interfering RNA oligonucleotide transfection. RESULTS We show that microRNA-128 (miR-128) directly targets mRNA of SUZ12, a key component of PRC2, in addition to BMI1, a component of PRC1 that we previously showed as a target as well. This blocks the partially redundant functions of PRC1/PRC2, thereby significantly reducing PRC activity and its associated histone modifications. MiR-128 and SUZ12/BMI1 show opposite expression in human glioblastomas versus normal brain and in glioma stemlike versus neural stem cells. Furthermore, miR-128 renders glioma stemlike cells less radioresistant by preventing the radiation-induced expression of both PRC components. Finally, miR-128 expression is significantly reduced in neural stem cells from the brain of young, presymptomatic mice in our mouse model of glioblastoma. This suggests that loss of miR-128 expression in brain is an early event in gliomagenesis. Moreover, knockdown of miR-128 expression in nonmalignant mouse and human neural stem cells led to elevated expression of PRC components and increased clonogenicity. CONCLUSIONS MiR-128 is an important suppressor of PRC activity, and its absence is an early event in gliomagenesis.

Genetic strategies for brain tumor therapy

Gene therapy is a potentially useful approach in the treatment of human brain tumors, which are notoriously refractory to conventional approaches. Most human clinical trials to date have been unsuccessful in terms of improving patient outcome. Recent improvements in viral vectors, the development of stem cell technology, and increased understanding of the mechanism of action of therapeutic transgenes provide hope that the next generation of gene therapeutics may show increased efficacy in treatment of this devastating disease.

Design and application of oncolytic HSV vectors for glioblastoma therapy.

Glioblastoma multiforme is one of the most common human brain tumors. The tumor is generally highly infiltrative, making it extremely difficult to treat by surgical resection or radiotherapy. This feature contributes to recurrence and a very poor prognosis. Few anticancer drugs have been shown to alter rapid tumor growth and none are ultimately effective. Oncolytic vectors have been employed as a treatment alternative based on the ability to tailor virus replication to tumor cells. The human neurotropic herpes simplex virus (HSV) is especially attractive for development of oncolytic vectors (oHSV) because this virus is highly infectious, replicates rapidly and can be readily modified to achieve vector attenuation in normal brain tissue. Tumor specificity can be achieved by deleting viral genes that are only required for virus replication in normal cells and permit mutant virus replication selectively in tumor cells. The anti-tumor activity of oHSV can be enhanced by arming the vector with genes that either activate chemotherapeutic drugs within the tumor tissue or promote anti-tumor immunity. In this review, we describe current designs of oHSV and the experience thus far with their potential utility for glioblastoma therapy. In addition, we discuss the impediments to vector effectiveness and describe our view of future developments in vector improvement.

Magnetic resonance imaging versus ultrasonography for the in utero evaluation of central nervous system anomalies

OBJECT The use of fetal MR imaging for the in utero evaluation of pathological conditions of the CNS is widely accepted as an adjunct to fetal ultrasonography studies. Magnetic resonance imaging is thought to characterize CNS anomalies better, and to provide a more exact diagnosis and accurate prognosis. The purpose of this study was to determine the role of and indications for fetal MR imaging in evaluating fetuses with different CNS abnormalities that were seen initially on prenatal sonograms. METHODS Over a 3-year period, fetuses with prior sonographic evidence of CNS abnormalities who consequently received prenatal MR imaging at Columbus Nationwide Childrens Hospital within 2 weeks of the fetal ultrasonography study were included in this study. For each patient, radiological reports from both studies were reviewed, analyzed, and compared with the findings at postnatal imaging or physical examination. Results of the 2 modalities were then compared in terms of diagnostic accuracy. RESULTS Twenty-six fetuses were included in this study on the basis of an in utero sonogram showing a CNS anomaly. Their gestational age ranged from 17 to 35 weeks, with a mean of 25 weeks at the time of fetal ultrasonography. Hydrocephalus was identified in 16 fetuses, 6 had evidence of a spinal dysraphic defect, 2 had holoprosencephaly, 1 had an encephalocele, and 1 had multiple body abnormalities requiring detailed CNS evaluation. Twenty-five of the fetuses were correctly evaluated as having abnormal CNS findings on both fetal ultrasonography and fetal MR imaging. Fetal ultrasonography provided a correct prenatal diagnosis in 20 cases, whereas fetal MR imaging was correct in 22 cases. There were 9 cumulative false-positive results for fetal ultrasonography and 7 for fetal MR imaging, whereas for false-negative results there were a total of 34 and 19, respectively. CONCLUSIONS Fetal MR imaging is more sensitive in detecting fetal CNS abnormalities, but its ability to provide a correct prenatal diagnosis is only marginally superior to fetal ultrasonography. Moreover, fetal MR imaging is not exempt from misdiagnosis, and still shows a significantly high rate of false-negative results. Particularly for spinal dysraphic defects, fetal MR imaging does not seem to add important diagnostic or prognostic details when compared with fetal ultrasonography.

Physiological transgene regulation and functional complementation of a neurological disease gene deficiency in neurons.

The microtubule-associated protein tau (MAPT) and alpha-synuclein (SNCA) genes play central roles in neurodegenerative disorders. Mutations in each gene cause familial disease, whereas common genetic variation at both loci contributes to susceptibility to sporadic neurodegenerative disease. Here, we demonstrate exquisite gene regulation of the human MAPT and SNCA transgene loci and functional complementation in neuronal cell cultures and organotypic brain slices using the herpes simplex virus type 1 (HSV-1) amplicon-based infectious bacterial artificial chromosome (iBAC) vector to express complete loci >100 kb. Cell cultures transduced by iBAC vectors carrying a 143 kb MAPT or 135 kb SNCA locus expressed the human loci similar to the endogenous gene. We focused on analysis of the iBAC-MAPT vector carrying the complete MAPT locus. On transduction into neuronal cultures, multiple MAPT transcripts were expressed from iBAC-MAPT under strict developmental and cell type-specific control. In primary neurons from Mapt(-/-) mice, the iBAC-MAPT vector expressed the human tau protein, as detected by enzyme-linked immunosorbent assay and immunocytochemistry, and restored sensitivity of Mapt(-/-) neurons to Abeta peptide treatment in dissociated neuronal cultures and in organotypic slice cultures. The faithful retention of gene expression and phenotype complementation by the system provides a novel method to analyze neurological disease genes.

Cancer immunotherapy: A vaccine from plant virus proteins.

Cowpea mosaic virus nanoparticles can induce the immune system to clear metastatic cancers.

Intraoperative MRI (ioMRI) in the Setting of Awake Craniotomies for Supratentorial Glioma Resection

Both awake craniotomy under conscious sedation and use of intraoperative MRI can increase the efficiency and safety of glioma resections. In contrast to craniotomies under general anesthesia, neurosurgery under conscious sedation requires several changes to the routine operative setup when performed in the ioMRI environment. This work reports our experience with awake craniotomies under conscious sedation using ioMRI. Seven patients underwent awake-craniotomies for resection of supratentorial gliomas using ioMRI at the Ohio State University Medical Center and James Cancer Hospital by a single surgeon. ioMRI can be safely employed in patients who are undergoing craniotomies under conscious sedation. Particularly important is the evaluation by the anesthesiologist whether the patient is a good candidate to sustain a likely longer than average procedure in a setting where his active cooperation is not only required, but also the essential aspect of this procedure.

Reversal of neurological deficit after chemotherapy in BCL-6-positive neurolymphomatosis. Case report.

Neurolymphomatosis, the infiltration of the peripheral nervous system (PNS) by malignant lymphatic cells, is a rare condition whose prognosis and treatment are not fully characterized. The authors report the case of a 69-year-old, previously healthy man who had a 1-month history of progressive pain in his right arm and associated weakness of several muscles of the right upper extremity when they first examined him. Initial MR imaging of the right brachial plexus showed no abnormalities, but over 3 months, symptoms gradually progressed to almost complete plegia of his right upper extremity. Subsequent MR imaging of his right brachial plexus showed an enhancing mass of the posterior cord of the plexus that encroached on the other cords. Positron emission tomography confirmed the presence of a hypermetabolic lesion in the right axillary region and also detected an asymptomatic hot spot in the gastric wall. Biopsy of the gastric lesion demonstrated a CD20+, diffuse large B-cell lymphoma that was immunohistochemically positive for BCL-6 and negative for p16. The patient underwent 6 cycles of dose-adjusted etoposide-vincristine-doxorubicin-cyclophosphamide-prednisone (EPOCH) and rituximab, intermixed with 3 cycles of high-dose intravenous and intrathecal methotrexate, and followed by 6 monthly doses of rituximab for consolidation. Follow-up MR imaging and PET of the plexus showed complete radiological response after 3 months of treatment, as demonstrated by normalization of brachial plexus caliber, contrast enhancement, and metabolic activity. Twenty-eight months after symptom onset and 20 months after beginning therapy, the patient was disease-free, had recovered most upper extremity neurological function, and had only minimal remaining weakness of the right wrist and finger extension.