Arto Immonen

Thymidine kinase gene therapy for human malignant glioma, using replication-deficient retroviruses or adenoviruses.

Herpes simplex virus thymidine kinase (HSV tk) gene therapy combined with ganciclovir (GCV) medication is a potential new method for the treatment of malignant glioma. We have used both retrovirus-packaging cells (PA317/tk) and adenoviruses (Adv/tk) for gene therapy for malignant glioma. Retrovirus-packaging cells were used for eight tumors in seven patients and adenoviruses were used for seven tumors in seven patients. As a control group, seven tumors in seven patients were transduced with lacZ marker gene 4-5 days before tumor resection. Safety and efficacy of the gene therapy were studied with clinical evaluation, blood and urine samples, MRI follow-up, and survival of the patients. Four patients with adenovirus injections had a significant increase in anti-adenovirus antibodies and two of them had a short-term fever reaction. Frequency of epileptic seizures increased in two patients. No other adverse events possibly related to gene therapy were detected. In the retrovirus group, all treated gliomas showed progression by MRI at the 3-month time point, whereas three of the seven patients treated with Adv/tk remained stable (p < 0.05). Mean survival times for retrovirus, adenovirus, and control groups were 7.4, 15.0, and 8. 3 months, respectively. The difference in the survival times between the adenovirus and retrovirus groups was significant (p < 0.012). It is concluded that HSV tk gene therapy is safe and well tolerated. On the basis of these results further trials are justified, especially with adenovirus vectors.

Long term outcome of temporal lobe epilepsy surgery: analyses of 140 consecutive patients

Objective: To analyse the long term results of temporal lobe epilepsy surgery in a national epilepsy surgery centre for adults, and to evaluate preoperative factors predicting a good postoperative outcome on long term follow up. Methods: Longitudinal follow up of 140 consecutive adult patients operated on for drug resistant temporal lobe epilepsy. Results: 46% of patients with unilateral temporal lobe epilepsy became seizure-free, 10% had only postoperative auras, and 15% had rare seizures on follow up for (mean (SD)) 5.4 (2.6) years, range 0.25 to 10.5 years. The best outcome was after introduction of a standardised magnetic resonance (MR) imaging protocol (1993–99): in unilateral temporal lobe epilepsy, 52% of patients became seizure-free, 7% had only postoperative auras, and 17% had rare seizures (median follow up 3.8 years, range 0.25 to 6.5 years); in palliative cases (incomplete removal of focus), a reduction in seizures of at least 80% was achieved in 71% of cases (median follow up 3.1 years, range 1.1 to 6.8 years). Most seizure relapses (86%) occurred within one year of the operation, and outcome at one year did not differ from the long term outcome. Unilateral hippocampal atrophy with or without temporal cortical atrophy on qualitative MR imaging (p < 0.001, odds ratio (OR) 5.2, 95% confidence interval (CI) 2.0 to 13.7), other unitemporal structural lesions on qualitative MR imaging (p ≤ 0.001, OR 6.9, 95% CI 2.2 to 21.5), onset of epilepsy before the age of five years (p < 0.05, OR 2.9, 95% CI 1.2 to 7.2), and focal seizures with ictal impairment of consciousness and focal ictal EEG as a predominant seizure type (p < 0.05, OR 3.4, 95% CI 1.2 to 9.1) predicted Engel I–II outcome. Hippocampal volume reduction of at least 1 SD from the mean of controls on the side of the seizure onset (p < 0.05, OR 3.1, 95% CI 1.1 to 9.2) also predicted Engel I–II outcome. Conclusions: Outcome at one year postoperatively is highly predictive of long term outcome after temporal lobe epilepsy surgery. Unitemporal MR imaging abnormalities, early onset of epilepsy, and seizure type predominance are factors associated with good postoperative outcome.

Long-term epilepsy surgery outcomes in patients with MRI-negative temporal lobe epilepsy

Purpose:  The outcome of surgery in patients with temporal lobe epilepsy (TLE) and normal high‐resolution magnetic resonance imaging (MRI) has been significantly worse than in patients with unilateral hippocampal damage upon MRI. The purpose of this study was to determine the long‐term outcomes of consecutive true MRI‐negative TLE patients who all underwent standardized preoperative evaluation with intracranial electroencephalography (EEG) electrodes.

Cystatin C expression is associated with granule cell dispersion in epilepsy

Human temporal lobe epilepsy (TLE) is associated with cellular alterations (eg, hilar cell death, neurogenesis, and granule cell dispersion) in the dentate gyrus but their underlying molecular mechanism are not known. We previously demonstrated increased expression of cystatin C, a protease inhibitor linked to both neurodegeneration and neurogenesis, during epileptogenesis in the rat hippocampus. Here, we investigated cystatin C expression in the dentate gyrus in chronic epilepsy and its association with neuronal loss and neurogenesis. In both rats with epilepsy and human patients with TLE, cystatin C expression was increased in glial cells in the molecular layer of the dentate gyrus, being most prominent in cases with granule cell dispersion. In patients with TLE, high cystatin C expression associated with greater numbers of polysialylated neural cell adhesion molecule–positive newborn cells in the molecular layer, although the overall number was decreased, indicating that the newborn cells migrate to abnormal locations in the epileptic dentate gyrus. These data thus demonstrate that cystatin C expression is altered during the chronic phase of epilepsy and suggest that cystatin C plays a role in network reorganization in the epileptic dentate gyrus, especially in granule cell dispersion and guidance of migrating newborn granule cells. Ann Neurol 2005;58:211–223

Non-invasive preoperative localization of primary motor cortex in epilepsy surgery by navigated transcranial magnetic stimulation

BACKGROUND Navigated transcranial magnetic stimulation (nTMS) is a non-invasive method to localize the primary motor cortex (M1). OBJECTIVE/HYPOTHESIS To assess the safety and feasibility of nTMS as a non-invasive preoperative mode of functional localization of M1 in epilepsy surgery candidates with intractable focal epilepsy due to lesions in the vicinity of M1. METHODS We mapped the muscle representation areas of M1 with nTMS in 10 patients (age 2 to 55 years) with intractable epilepsy. The lesions were focal cortical dysplasia (n=6), ganglioglioma (n=2) polymicrogyria (n=1) or dysembryoblastic neuroepithelial tumour (n=1). The optimal stimulation sites and motor threshold (MT) of the distal hand or leg muscles were determined in both hemispheres. Cortical areas were mapped with stimulation intensities 100-120% of the MT to localize functional M1. Patients were on their stabile antiepileptic medication, and EEG was continuously monitored. The clinical benefit obtained with the preoperative nTMS mapping in the surgical decision making was scored as (1) essential, (2) beneficial, or (3) not beneficial, depending mainly on the difference between the functional and the presumed anatomic M1. RESULTS The M1 was successfully assessed in all but the 2 youngest patients (aged 2 and 5 years), in whom nTMS was unable to elicit motor responses. nTMS was regarded as essential or beneficial in the localization of M1 in relation to the lesions in 6 out of 10 cases. The optimal motor representation areas were mainly located symmetrically on the precentral gyrus, and corresponded to the presumed location of M1 in MRI. No clinical or EEG evidence of acute epileptogenic adverse effects were observed during the localization procedure. None of the operated patients developed post-operative motor deficits. CONCLUSIONS nTMS is a safe and feasible clinical tool for the non-invasive preoperative localization of motor cortex in patients with intractable epilepsy due to focal lesions adjacent or within the presumed M1 in MRI.

Epilepsy after aneurysmal subarachnoid hemorrhage A population-based, long-term follow-up study

Objective: The aim was to elucidate the incidence and risk factors of epilepsy after subarachnoid hemorrhage (SAH) from saccular intracranial aneurysm (sIA) in a population-based cohort. Methods: The Kuopio sIA Database (www.uef.fi/ns) includes all unruptured and ruptured sIA cases admitted to the Kuopio University Hospital from its defined catchment population in Eastern Finland. The use of prescribed medicines, including reimbursable antiepileptic drugs, has been entered from the Finnish national registries. The cumulative incidence and independent risk factors of epilepsy and death were analyzed in 876 patients with sIA-SAH admitted from 1995 to 2007. The competing risks analysis was used to correctly estimate the probability of epilepsy, because epilepsy and death after sIA-SAH may share risk factors. Results: The follow-up ended at death (n = 200) or December 31, 2008; median follow-up time was 76 months. Epilepsy was diagnosed in 113 patients in a median of 8 months after sIA-SAH. Cumulative incidence of epilepsy after sIA-SAH was 8% at 1 year and 12% at 5 years. Thirty-three percent of patients with intracerebral hemorrhage (ICH) >15 cm3 developed epilepsy. In the 876 patients with sIA-SAH, the independent risk factors for epilepsy were ICH >15 cm3, Hunt and Hess grade III–V, and acute seizures. Conclusions: Cumulative incidence of epilepsy is 12% at 5 years. Epilepsy and 12-month mortality after sIA-SAH share poor Hunt and Hess grading as an independent risk factor. Epilepsy in the 2-week survivors of sIA-SAH is predicted by signs of primary injury in the brain tissue, most notably ICH.OBJECTIVE The aim was to elucidate the incidence and risk factors of epilepsy after subarachnoid hemorrhage (SAH) from saccular intracranial aneurysm (sIA) in a population-based cohort. METHODS The Kuopio sIA Database (www.uef.fi/ns) includes all unruptured and ruptured sIA cases admitted to the Kuopio University Hospital from its defined catchment population in Eastern Finland. The use of prescribed medicines, including reimbursable antiepileptic drugs, has been entered from the Finnish national registries. The cumulative incidence and independent risk factors of epilepsy and death were analyzed in 876 patients with sIA-SAH admitted from 1995 to 2007. The competing risks analysis was used to correctly estimate the probability of epilepsy, because epilepsy and death after sIA-SAH may share risk factors. RESULTS The follow-up ended at death (n = 200) or December 31, 2008; median follow-up time was 76 months. Epilepsy was diagnosed in 113 patients in a median of 8 months after sIA-SAH. Cumulative incidence of epilepsy after sIA-SAH was 8% at 1 year and 12% at 5 years. Thirty-three percent of patients with intracerebral hemorrhage (ICH) >15 cm(3) developed epilepsy. In the 876 patients with sIA-SAH, the independent risk factors for epilepsy were ICH >15 cm(3), Hunt and Hess grade III-V, and acute seizures. CONCLUSIONS Cumulative incidence of epilepsy is 12% at 5 years. Epilepsy and 12-month mortality after sIA-SAH share poor Hunt and Hess grading as an independent risk factor. Epilepsy in the 2-week survivors of sIA-SAH is predicted by signs of primary injury in the brain tissue, most notably ICH.

Temporal anteroinferior encephalocele: An underrecognized etiology of temporal lobe epilepsy?

Objective: To report the increasing frequency with which temporal anteroinferior encephalocele is a cause of adult temporal lobe epilepsy, to illustrate the clinical and imaging characteristics of this condition, and to report its surgical treatment in a series of 23 adult patients. Methods: Epilepsy patients diagnosed with temporal anteroinferior encephalocele from January 2006 to December 2013 in a national epilepsy reference center were included in this noninterventional study. Results: Twenty-three epilepsy patients (14 female, mean age 43.8 years) were diagnosed with temporal anteroinferior encephalocele in our institute. Thirteen patients had ≥2 encephaloceles; 7 cases presented bilaterally. The estimated frequency of this condition was 0.3% among MRI examinations performed due to newly diagnosed epilepsy (n = 6) and 1.9% among drug-resistant patients referred to our center (n = 17). Nine patients with local encephalocele disconnection (n = 4) or anterior temporal lobectomy and amygdalohippocampectomy (n = 5) have become seizure-free (Engel 1) for a mean 2.8 years (range 3 months–6.2 years) of follow-up. Three patients with local encephalocele disconnection were almost seizure-free or exhibited worthwhile improvement. Histologically, all 12 surgical patients had gliosis at the base of the encephalocele; some had cortical laminar disorganization (n = 5) or mild hippocampal degeneration (n = 1). Conclusions: The possibility of a temporal encephalocele should be considered when interpreting MRI examinations of patients with medically intractable focal epilepsy. These patients can significantly benefit from unitemporal epilepsy surgery, even in cases with bilateral encephaloceles.

Safety Profile of Plasmid/Liposomes and Virus Vectors in Clinical Gene Therapy

Despite of more than 500 gene therapy trials worldwide very little systematic safety information is available from gene therapy. Safety information was collected from 146 consecutive patients who participated in three randomized, controlled phase II gene therapy trials in cardiovascular diseases and malignant glioma using adenoviruses, plasmid/liposomes and retrovirus packaging cells. Total follow-up time of the patients was 78794 days which equals 1.5 years per patient. The main outcome measures were serious adverse events, other adverse events and changes in general laboratory parameters. Except fever and increases in CRP values plasmid/liposomes were safe and well tolerated. The incidence of serious adverse events in adenovirus-treated patients was 0.9 and 4.0/10000 patient days in cardiovascular and malignant glioma trials as compared to 0.5 and 2.1 in randomized control patients, respectively. Transient fever, leukopenia and increases in CRP and liver enzymes were detected in virus-treated patients. No deaths from side effects or no new cancers were associated with gene therapy. It is concluded that gene therapy, like any other therapy, is associated with side effects which depend on the administered vector, dose, and route of delivery and properties of the transgene. However, given the limitations of this study and length of the follow-up, the safety profile of gene therapy seems to be acceptable for the treatment of severe human diseases.

3-D reconstructed magnetic resonance imaging in localization of subdural EEG electrodes: Case illustration

We report an illustrative case of presurgical evaluation for epilepsy surgery, where the three-dimensional reconstructed magnetic resonance imaging played a pivotal role in determining the exact location of the subdural strip electrodes in temporomesial area. The tip of one the frontal strip electrodes was actually recording the temporopolar ictal activity. This contributed conclusively to the decision for surgical treatment and to the excellent outcome.

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

ABSTRACT 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.