Intraparenchymal convection enhanced delivery of AAV in sheep to treat Mucopolysaccharidosis IIIC.

BACKGROUND: Mucopolysaccharidosis IIIC (MPSIIIC) is one of four Sanfilippo diseases sharing clinical symptoms of severe cognitive decline and shortened lifespan. The missing enzyme, heparan sulfate acetyl-CoA: α-glucosaminide-N-acetyltransferase (HGSNAT), is bound to the lysosomal membrane, therefore cannot cross the blood-brain barrier or diffuse between cells. We previously demonstrated disease correction in MPSIIIC mice using an Adeno-Associated Vector (AAV) delivering HGSNAT via intraparenchymal brain injections using an AAV2 derived AAV-truetype (AAV-TT) serotype with improved distribution over AAV9. METHODS: Here, intraparenchymal AAV was delivered in sheep using catheters or Hamilton syringes, placed using Brainlab cranial navigation for convection enhanced delivery, to reduce proximal vector expression and improve spread. RESULTS: Hamilton syringes gave improved AAV-GFP distribution, despite lower vector doses and titres. AAV-TT-GFP displayed moderately better transduction compared to AAV9-GFP but both serotypes almost exclusively transduced neurons. Functional HGSNAT enzyme was detected in 24-37% of a 140g gyrencephalic sheep brain using AAV9-HGSNAT with three injections in one hemisphere. CONCLUSIONS: Despite variabilities in volume and titre, catheter design may be critical for efficient brain delivery. These data help inform a clinical trial for MPSIIIC.

Awake brain surgery for autistic patients: Is it possible?

Background: Awake neurosurgery is currently the mainstay for eloquent brain lesions. Opting for an awake operation is affected by a number of patient-related factors. We present a case of a patient with autistic spectrum disorder (ASD) that was successfully operated for a brain tumor through awake craniotomy. To the best of our knowledge, this is the first reported case in the literature. Case Description: A 42-year-old patient, with known ASD since his childhood, underwent awake craniotomy for a left supplementary motor area tumor. Detailed preoperative preparation of the patient was done to identify special requirements and establish a good patient-team relationship. Intraoperatively, continuous language and motor testing were performed. Conversation and music were the main distractors used. Throughout the operation, the patient remained calm and cooperative, even during a focal seizure. Mapping allowed for >80% resection of the tumor. Postoperatively, the patient recovered without any deficits. Conclusion: This case shows that with growing experience and meticulous preparation, the limits of awake craniotomy can be expanded to include more patients that were previously considered unfit.

A risk assessment strategy to re-introduce elective neurosurgery patients during COVID-19.

OBJECTIVES: To demonstrate the utilisation of a risk assessment protocol designed to prioritise elective neurosurgical patients against the risks of COVID-19. This tool can be applied to all other surgical specialties. DESIGN: Prospective case series of 166 patients. SETTING: Single-centre tertiary neurosurgical department. PARTICIPANTS: All patients awaiting an elective neurosurgical procedure were included in this study. All emergency or life-threatening neurosurgical pathologies affecting patients were excluded. MAIN OUTCOME MEASURES: The risk assessment tool identified patients with progressive neurology and stratified need for surgery against risk of harm during the COVID-19 pandemic. RESULTS: Using our risk stratification tool, 6.6% patients required expedited surgery and a further 11.4% patients were removed completely from the waiting list. The majority of patients 47%, required surgery within 3 months. CONCLUSIONS: This simple tool encourages surgical departments to establish contact with patients during COVID-19. The clinician acquires up-to-date information regarding patient symptomatology and subsequently determines surgical priority, a timescale required for surgery and overall uses of NHS resources efficiently. We recommend the use of this tool for all neurosurgical departments, with a wider application to other surgical specialties during the ongoing pressures of elective backlogs secondary to the persistent COVID-19 pandemic.

Pre-clinical Safety and Efficacy of Lentiviral Vector-Mediated Ex Vivo Stem Cell Gene Therapy for the Treatment of Mucopolysaccharidosis IIIA.

Hematopoietic stem cell gene therapy is a promising therapeutic strategy for the treatment of neurological disorders, since transplanted gene-corrected cells can traffic to the brain, bypassing the blood-brain barrier, to deliver therapeutic protein to the CNS. We have developed this approach for the treatment of Mucopolysaccharidosis type IIIA (MPSIIIA), a devastating lysosomal storage disease that causes progressive cognitive decline, leading to death in early adulthood. In a previous pre-clinical proof-of-concept study, we demonstrated neurological correction of MPSIIIA utilizing hematopoietic stem cell gene therapy via a lentiviral vector encoding the SGSH gene. Prior to moving to clinical trial, we have undertaken further studies to evaluate the efficiency of gene transfer into human cells and also safety studies of biodistribution and genotoxicity. Here, we have optimized hCD34+ cell transduction with clinical grade SGSH vector to provide improved pharmacodynamics and cell viability and validated effective scale-up and cryopreservation to generate an investigational medicinal product. Utilizing a humanized NSG mouse model, we demonstrate effective engraftment and biodistribution, with no vector shedding or transmission to germline cells. SGSH vector genotoxicity assessment demonstrated low transformation potential, comparable to other lentiviral vectors in the clinic. This data establishes pre-clinical safety and efficacy of HSCGT for MPSIIIA.

A novel adeno-associated virus capsid with enhanced neurotropism corrects a lysosomal transmembrane enzyme deficiency.

Recombinant adeno-associated viruses (AAVs) are popular in vivo gene transfer vehicles. However, vector doses needed to achieve therapeutic effect are high and some target tissues in the central nervous system remain difficult to transduce. Gene therapy trials using AAV for the treatment of neurological disorders have seldom led to demonstrated clinical efficacy. Important contributing factors are low transduction rates and inefficient distribution of the vector. To overcome these hurdles, a variety of capsid engineering methods have been utilized to generate capsids with improved transduction properties. Here we describe an alternative approach to capsid engineering, which draws on the natural evolution of the virus and aims to yield capsids that are better suited to infect human tissues. We generated an AAV capsid to include amino acids that are conserved among natural AAV2 isolates and tested its biodistribution properties in mice and rats. Intriguingly, this novel variant, AAV-TT, demonstrates strong neurotropism in rodents and displays significantly improved distribution throughout the central nervous system as compared to AAV2. Additionally, sub-retinal injections in mice revealed markedly enhanced transduction of photoreceptor cells when compared to AAV2. Importantly, AAV-TT exceeds the distribution abilities of benchmark neurotropic serotypes AAV9 and AAVrh10 in the central nervous system of mice, and is the only virus, when administered at low dose, that is able to correct the neurological phenotype in a mouse model of mucopolysaccharidosis IIIC, a transmembrane enzyme lysosomal storage disease, which requires delivery to every cell for biochemical correction. These data represent unprecedented correction of a lysosomal transmembrane enzyme deficiency in mice and suggest that AAV-TT-based gene therapies may be suitable for treatment of human neurological diseases such as mucopolysaccharidosis IIIC, which is characterized by global neuropathology.

High content screening of patient-derived cell lines highlights the potential of non-standard chemotherapeutic agents for the treatment of glioblastoma.

BACKGROUND: Glioblastoma (GBM) is the most common primary brain malignancy in adults, yet survival outcomes remain poor. First line treatment is well established, however disease invariably recurs and improving prognosis is challenging. With the aim of personalizing therapy at recurrence, we have established a high content screening (HCS) platform to analyze the sensitivity profile of seven patient-derived cancer stem cell lines to 83 FDA-approved chemotherapy drugs, with and without irradiation. METHODS: Seven cancer stem cell lines were derived from patients with GBM and, along with the established cell line U87-MG, each patient-derived line was cultured in tandem in serum-free conditions as adherent monolayers and three-dimensional neurospheres. Chemotherapeutics were screened at multiple concentrations and cells double-stained to observe their effect on both cell death and proliferation. Sensitivity was classified using high-throughput algorithmic image analysis. RESULTS: Cell line specific drug responses were observed across the seven patient-derived cell lines. Few agents were seen to have radio-sensitizing effects, yet some drug classes showed a marked difference in efficacy between monolayers and neurospheres. In vivo validation of six drugs suggested that cell death readout in a three-dimensional culture scenario is a more physiologically relevant screening model and could be used effectively to assess the chemosensitivity of patient-derived GBM lines. CONCLUSION: The study puts forward a number of non-standard chemotherapeutics that could be useful in the treatment of recurrent GBM, namely mitoxantrone, bortezomib and actinomycin D, whilst demonstrating the potential of HCS to be used for personalized treatment based on the chemosensitivity profile of patient tumor cells.

Escherichia coli positive infratentorial subdural empyema secondary to mastoiditis and underlying cholesteatoma.

Infratentorial subdural empyema is a neurosurgical emergency that is associated with an alarmingly high morbidity and mortality if appropriate management is delayed. It is an important differential to consider when confronted with a patient with a reduced Glasgow Coma Scale, focal neurology and symptoms of raised intracranial pressure in the presence of a head and neck infection. It is also important that the primary team managing these patients is aware of the many pathogens that may be involved, including Escherichia coli. Early recognition, prompt diagnosis, timely involvement of the appropriate multidisciplinary teams, including neurosurgery, otorhinolaryngology, radiology and microbiology should be sought, and urgent intervention are imperative in avoiding a fatal outcome. This article presents a case of E coli-positive infratentorial subdural empyema secondary to mastoiditis due to underlying cholesteatoma, and a review of the pertinent literature.

Day case tonsillectomy for the treatment of obstructive sleep apnoea syndrome in children: Alder Hey experience.

BACKGROUND: The aim of this study was to assess the safety of paediatric adeno-tonsillectomy for obstructive sleep apnoea syndrome (OSAS) performed as a day case procedure. METHODOLOGY: Postoperative airway related complications were prospectively evaluated in children undergoing adeno-tonsillectomy for OSAS over a 12-month period in a tertiary care centre. All data was analysed and descriptive statistics performed. RESULTS: The overall incidence of postoperative airway related complications were 21 (10.4%) in 202 consecutively operated children. Eleven were in children with other medical co-morbidities (n=37) of which 8 occurred after 6h. Amongst those without any medical illnesses (n=165), two (1.2%) developed OSAS related complication after 6h of surgery only one of which needed medical/nursing intervention. The incidence of postoperative desaturations after 6h was 21.6% in those with co-morbidities. CONCLUSION: It is safe to perform day case adeno-tonsillectomy for OSAS in children without any medical co-morbidities while those with other medical illnesses should be planned as in-patient procedures.