MicroRNA-Attenuated Clone of Virulent Semliki Forest Virus Overcomes Antiviral Type I Interferon in Resistant Mouse CT-2A Glioma.

UNLABELLED: Glioblastoma is a terminal disease with no effective treatment currently available. Among the new therapy candidates are oncolytic viruses capable of selectively replicating in cancer cells, causing tumor lysis and inducing adaptive immune responses against the tumor. However, tumor antiviral responses, primarily mediated by type I interferon (IFN-I), remain a key problem that severely restricts viral replication and oncolysis. We show here that the Semliki Forest virus (SFV) strain SFV4, which causes lethal encephalitis in mice, is able to infect and replicate independent of the IFN-I defense in mouse glioblastoma cells and cell lines originating from primary human glioblastoma patient samples. The ability to tolerate IFN-I was retained in SFV4-miRT124 cells, a derivative cell line of strain SFV4 with a restricted capacity to replicate in neurons due to insertion of target sites for neuronal microRNA 124. The IFN-I tolerance was associated with the viral nsp3-nsp4 gene region and distinct from the genetic loci responsible for SFV neurovirulence. In contrast to the naturally attenuated strain SFV A7(74) and its derivatives, SFV4-miRT124 displayed increased oncolytic potency in CT-2A murine astrocytoma cells and in the human glioblastoma cell lines pretreated with IFN-I. Following a single intraperitoneal injection of SFV4-miRT124 into C57BL/6 mice bearing CT-2A orthotopic gliomas, the virus homed to the brain and was amplified in the tumor, resulting in significant tumor growth inhibition and improved survival. IMPORTANCE: Although progress has been made in development of replicative oncolytic viruses, information regarding their overall therapeutic potency in a clinical setting is still lacking. This could be at least partially dependent on the IFN-I sensitivity of the viruses used. Here, we show that the conditionally replicating SFV4-miRT124 virus shares the IFN-I tolerance of the pathogenic wild-type SFV, thereby allowing efficient targeting of a glioma that is refractory to naturally attenuated therapy vector strains sensitive to IFN-I. This is the first evidence of orthotopic syngeneic mouse glioma eradication following peripheral alphavirus administration. Our findings indicate a clear benefit in harnessing the wild-type virus replicative potency in development of next-generation oncolytic alphaviruses.

Rate and Risk Factors for Shunt Revision in Pediatric Patients with Hydrocephalus-A Population-Based Study.

BACKGROUND: Ventriculoperitoneal shunt (VPS) is a common treatment for patients with hydrocephalus (HC). VPS is associated with complications that may lead to shunt revisions. We studied the surgical outcome of pediatric patients with HC in a population-based setting. METHODS: The medical charts and imaging findings of 80 patients ≤16 years old who required VPS secondary to HC were studied. RESULTS: Mean age at time of initial shunt placement was 3.2 years (SD 4.5) and mean follow-up time was 3.3 years (SD 2.9); 57% of patients were male. Half of patients underwent shunt revision with mean time to first revision of 8 months. Patients ≤6 months old had a higher shunt revision rate compared with patients >6 months old (P < 0.001). The most common causes of HC requiring VPS were tumors (27.5%), congenital defects (22.5%), and intraventricular hemorrhage (19%). Revision rates in the intraventricular hemorrhage and congenital defects groups were 67% (P = 0.017) and 72% (P = 0.016) compared with 32% in the tumor group. Programmable valves (56%) were more common than nonprogrammable valves, but there was no significant difference in shunt survival (P = 0.632). The mean biparietal measurement change between preoperative and postoperative images was +0.9 mm in the no revision group and +6.6 mm in the revision group (P = 0.003). CONCLUSIONS: Half of patients with shunts required revision. Age ≤6 months and intraventricular hemorrhage and congenital defects etiologies of HC were associated with increased risk for shunt revision. Most revisions were done during the first year after the initial VPS.

A 42-year-old woman with subacute reversible dementia: A cautionary tale.

A 42-year-old woman presented with a 6-month history of diffuse headache of moderate intensity and gradual onset of generalized weakness, imbalance, apathy, memory decline, hypophonia, dysphagia, constipation and urinary incontinence. Clinical examination revealed several elements of a frontal lobe dysfunction including apathy with motor impersistence, presence of primitive reflexes, generalized hyperreflexia with bilateral Hoffman sign and ankle clonus. The biological workup was unremarkable and a brain computed tomography scan identified a giant olfactory groove meningioma. A prompt neurosurgical intervention helped to reverse the symptoms. This case illustrates the benefits of actively looking for treatable conditions in young patients presenting with acute or subacute dementia and emphasizes the pivotal role of early brain imaging.

Immunohistochemical Characterization and Sensitivity to Human Adenovirus Serotypes 3, 5, and 11p of New Cell Lines Derived from Human Diffuse Grade II to IV Gliomas.

BACKGROUND: Oncolytic adenoviruses show promise in targeting gliomas because they do not replicate in normal brain cells. However, clinical responses occur only in a subset of patients. One explanation could be the heterogenic expression level of virus receptors. Another contributing factor could be variable activity of tumor antiviral defenses in different glioma subtypes. METHODS: We established a collection of primary low-passage cell lines from different glioma subtypes (3 glioblastomas, 3 oligoastrocytomas, and 2 oligodendrogliomas) and assessed them for receptor expression and sensitivity to human adenovirus (HAd) serotypes 3, 5, and 11p. To gauge the impact of antiviral defenses, we also compared the infectivity of the oncolytic adenoviruses in interferon (IFN)-pretreated cells with IFN-sensitive Semliki Forest virus (SFV). RESULTS: Immunostaining revealed generally low expression of HAd5 receptor CAR in both primary tumors and derived cell lines. HAd11p receptor CD46 levels were maintained at moderate levels in both primary tumor samples and derived cell lines. HAd3 receptor DSG-2 was reduced in the cell lines compared to the tumors. Yet, at equal multiplicities of infection, the oncolytic potency of HAd5 in vitro in tumor-derived cells was comparable to HAd11p, whereas HAd3 lysed fewer cells than either of the other two HAd serotypes in 72 hours. IFN blocked replication of SFV, while HAds were rather unaffected. CONCLUSIONS: Adenovirus receptor levels on glioma-derived cell lines did not correlate with infection efficacy and may not be a relevant indicator of clinical oncolytic potency. Adenovirus receptor analysis should be preferentially performed on biopsies obtained perioperatively.

Prevention of NMDA-induced death of cortical neurons by inhibition of protein kinase Czeta.

Excitotoxicity through stimulation of N-methyl-d-aspartate (NMDA) receptors contributes to neuronal death in brain injuries, including stroke. Several lines of evidence suggest a role for protein kinase C (PKC) isoforms in NMDA excitotoxicity. We have used specific peptide inhibitors of classical PKCs (alpha, beta, and gamma), novel PKCs delta and epsilon, and an atypical PKCzeta in order to delineate which subspecies are involved in NMDA-induced cell death. Neuronal cell cultures were prepared from 15-day-old mouse embryos and plated onto the astrocytic monolayer. After 2 weeks in vitro the neurons were exposed to 100 micro m NMDA for 5 min, and 24 h later the cell viability was examined by measuring the lactate dehydrogenase release and bis-benzimide staining. While inhibitors directed to classical (alpha, beta, and gamma) or novel PKCs (delta or epsilon) had no effect, the PKCzeta inhibitor completely prevented the NMDA-induced necrotic neuronal death. Confocal microscopy confirmed that NMDA induced PKCzeta translocation, which was blocked by the PKCzeta inhibitor. The NMDA-induced changes in intracellular free Ca2+ were not affected by the peptides. In situ hybridization experiments demonstrated that PKCzeta mRNA is induced in the cortex after focal brain ischemia. Altogether, the results indicate that PKCzeta activation is a downstream signal in NMDA-induced death of cortical neurons.