Intracranial stimulation for children with epilepsy

OBJECTIVES: To evaluate the efficacy of intracranial stimulation to treat refractory epilepsy in children. METHODS: This is a retrospective analysis of a pilot study on all 8 children who had intracranial electrical stimulation for the investigation and treatment of refractory epilepsy at King's College Hospital between 2014 and 2015. Five children (one with temporal lobe epilepsy and four with frontal lobe epilepsy) had subacute cortical stimulation (SCS) for a period of 20-161 h during intracranial video-telemetry. Efficacy of stimulation was evaluated by counting interictal discharges and seizures. Two children had thalamic deep brain stimulation (DBS) of the centromedian nucleus (one with idiopathic generalized epilepsy, one with presumed symptomatic generalized epilepsy), and one child on the anterior nucleus (right fronto-temporal epilepsy). The incidence of interictal discharges was evaluated visually and quantified automatically. RESULTS: Among the three children with DBS, two had >60% improvement in seizure frequency and severity and one had no improvement. Among the five children with SCS, four showed improvement in seizure frequency (>50%) and one chid did not show improvement. Procedures were well tolerated by children. CONCLUSION: Cortical and thalamic stimulation appear to be effective and well tolerated in children with refractory epilepsy. SCS can be used to identify the focus and predict the effects of resective surgery or chronic cortical stimulation. Further larger studies are necessary

Antibodies against the mono-methylated arginine-glycine repeat (MMA-RG) of the Epstein-Barr virus nuclear antigen 2 (EBNA2) identify potential cellular proteins targeted in viral transformation.

The Epstein-Barr virus is a human herpes virus with oncogenic potential. The virus-encoded nuclear antigen 2 (EBNA2) is a key mediator of viral tumorigenesis. EBNA2 features an arginine-glycine (RG) repeat at amino acids (aa)339-354 that is essential for the transformation of lymphocytes and contains symmetrically (SDMA) and asymmetrically (ADMA) di-methylated arginine residues. The SDMA-modified EBNA2 binds the survival motor neuron protein (SMN), thus mimicking SMD3, a cellular SDMA-containing protein that interacts with SMN. Accordingly, a monoclonal antibody (mAb) specific for the SDMA-modified RG repeat of EBNA2 also binds to SMD3. With the novel mAb 19D4 we now show that EBNA2 contains mono-methylated arginine (MMA) residues within the RG repeat. Using 19D4, we immune-precipitated and analysed by mass spectrometry cellular proteins in EBV-transformed B-cells that feature MMA motifs that are similar to the one in EBNA2. Among the cellular proteins identified, we confirmed by immunoprecipitation and/or Western blot analyses Aly/REF, Coilin, DDX5, FXR1, HNRNPK, LSM4, MRE11, NRIP, nucleolin, PRPF8, RBM26, SMD1 (SNRDP1) and THRAP3 proteins that are either known to contain MMA residues or feature RG repeat sequences that probably serve as methylation substrates. The identified proteins are involved in splicing, tumorigenesis, transcriptional activation, DNA stability and RNA processing or export. Furthermore, we found that several proteins involved in energy metabolism are associated with MMA-modified proteins. Interestingly, the viral EBNA1 protein that features methylated RG repeat motifs also reacted with the antibodies. Our results indicate that the region between aa 34-52 of EBNA1 contains ADMA or SDMA residues, while the region between aa 328-377 mainly contains MMA residues

Epstein-Barr Virus Infection of Cell Lines Derived from Diffuse Large B-Cell Lymphomas Alters MicroRNA Loading of the Ago2 Complex

Diffuse large B-cell lymphoma (DLBCL) is an aggressive lymphoid tumor which is occasionally Epstein-Barr virus (EBV)-positive and is further subtyped as activated B-cell (ABC) and germinal center B-cell (GCB) DLBCL, which has implications for prognosis and treatment. We performed Ago2-RNA immunoprecipitation followed by high throughput RNA sequencing (Ago2-RIP-Seq) to capture functionally active miRNAs in EBV-negative ABC-DLBCL and GC-DLBCL cell lines and their EBV-infected counterparts. In parallel, total miRNomes of these cells were sequenced to capture the cellular miRNA profile for comparison with the functionally active profile. Selected miRNAs with differential abundance were validated using RT-qPCR and Northern Blot. We found 6 miRNAs with differential abundance (2 upregulated and 4 downregulated miRNAs) between EBV-neg. and pos. ABC-DLBCL, and 12 miRNAs with differential abundance (3 upregulated and 9 downregulated miRNAs) between EBV-neg and -pos GC-DLBCL. Eight and twelve miRNAs were confirmed using RT-qPCR in ABC-DLBCL and GC-DLBCL, respectively. Selected miRNs were analyzed in additional type I/II vs. type III EBV latency DLBCL cell lines. Furthermore, up regulation of miR-221-3p and down regulation of let-7c-5p in ABC-DLBCL and up regulation of miR-363-3p and down regulation of 423-5p in GC-DLBCL was verified using RIP-Northern blot. Our comprehensive sequence analysis of the DLBCL miRNomes identified sets of deregulated miRNAs in the Ago2-RIP-seq. Our Ago2-IP-seq miRNomes profile could be considered as an important data set for detection of deregulated functionally active miRNAs in DLBCL and could possibly lead to identification of miRNAs as biomarkers for classification of DLBCL or even as targets for personalized targeted treatment