Comparison of different promoters to improve AAV vector-mediated gene therapy for neuronopathic Gaucher disease.

Gaucher Disease (GD) is an inherited metabolic disorder caused by mutations in the GBA1 gene. It can manifest with severe neurodegeneration and visceral pathology. The most acute neuronopathic form (nGD), for which there are no curative therapeutic options, is characterised by devastating neuropathology and death during infancy. In this study, we investigated the therapeutic benefit of systemically delivered AAV9 vectors expressing the human GBA1 gene at two different doses comparing a neuronal-selective promoter with ubiquitous promoters. Our results highlight the importance of a careful evaluation of the promoter sequence used in gene delivery vectors, suggesting a neuron-targeted therapy leading to high levels of enzymatic activity in the brain but lower GCase expression in the viscera, might be the optimal therapeutic strategy for nGD.

Translin facilitates RNA polymerase II dissociation and suppresses genome instability during RNase H2- and Dicer-deficiency.

The conserved nucleic acid binding protein Translin contributes to numerous facets of mammalian biology and genetic diseases. It was first identified as a binder of cancer-associated chromosomal translocation breakpoint junctions leading to the suggestion that it was involved in genetic recombination. With a paralogous partner protein, Trax, Translin has subsequently been found to form a hetero-octomeric RNase complex that drives some of its functions, including passenger strand removal in RNA interference (RNAi). The Translin-Trax complex also degrades the precursors to tumour suppressing microRNAs in cancers deficient for the RNase III Dicer. This oncogenic activity has resulted in the Translin-Trax complex being explored as a therapeutic target. Additionally, Translin and Trax have been implicated in a wider range of biological functions ranging from sleep regulation to telomere transcript control. Here we reveal a Trax- and RNAi-independent function for Translin in dissociating RNA polymerase II from its genomic template, with loss of Translin function resulting in increased transcription-associated recombination and elevated genome instability. This provides genetic insight into the longstanding question of how Translin might influence chromosomal rearrangements in human genetic diseases and provides important functional understanding of an oncological therapeutic target.

Fetal gene therapy.

Fetal gene therapy was first proposed toward the end of the 1990s when the field of gene therapy was, to quote the Gartner hype cycle, at its "peak of inflated expectations." Gene therapy was still an immature field but over the ensuing decade, it matured and is now a clinical and market reality. The trajectory of treatment for several genetic diseases is toward earlier intervention. The ability, capacity, and the will to diagnose genetic disease early-in utero-improves day by day. A confluence of clinical trials now signposts a trajectory toward fetal gene therapy. In this review, we recount the history of fetal gene therapy in the context of the broader field, discuss advances in fetal surgery and diagnosis, and explore the full ambit of preclinical gene therapy for inherited metabolic disease.

Advanced Formulation Approaches for Emerging Therapeutic Technologies.

In addition to proteins, discussed in the Chapter "Advances in Vaccine Adjuvants: Nanomaterials and Small Molecules", there are a wide range of alternatives to small molecule active ingredients. Cells, extracellular vesicles, and nucleic acids in particular have attracted increasing research attention in recent years. There are now a number of products on the market based on these emerging technologies, the most famous of which are the mRNA-based vaccines against SARS-COV-2. These advanced therapeutic moieties are challenging to formulate however, and there remain significant challenges for their more widespread use. In this chapter, we consider the potential and bottlenecks for developing further medical products based on these systems. Cells, extracellular vesicles, and nucleic acids will be discussed in terms of their mechanism of action, the key requirements for translation, and how advanced formulation approaches can aid their future development. These points will be presented with selected examples from the literature, and with a focus on the formulations which have made the transition to clinical trials and clinical products.

Comparison of Different Antiviral Regimens in the Treatment of Patients with Severe COVID-19: A Retrospective Cohort.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes respiratory disorders, with disease severity ranging from asymptomatic to critical manifestations. The current retrospective study compared the efficacies of different antiviral regimens used in patients suffering from severe COVID-19 disease from 19 January 2020 to December 2021 in a single center in Saudi Arabia. In total, 188 patients were enrolled in the current study, including 158 patients treated with different antiviral regimens, and 30 who did not receive any antiviral treatment. Different antiviral regimens, including favipiravir, remdesivir, oseltamivir, favipiravir/remdesivir, and favipiravir/oseltamivir were adopted. The effects of using different antivirals and antibiotics on the survival rate were evaluated, as well as the presence of comorbidities. Among all severely affected patients, 39/188 (20.7%) survived. Both age and comorbidities, including diabetes and hypertension, were significantly correlated with high case fatality following SARS-CoV-2 infection. Remdesivir alone and the combination of favipiravir and remdesivir increased the survival rate. Surprisingly, both imipenem and linezolid helped in the deterioration of disease outcome in the patients. A negative correlation was detected between increased mortality and the use of favipiravir and the use of either imipenem or linezolid. Among the compared antiviral regimens used in the treatment of severe COVID-19, remdesivir was found to be an effective antiviral that reduces COVID-19 case fatality. Antibiotic treatment using imipenem and/or linezolid should be carefully re-evaluated.

Systemic AAV9 gene therapy using the synapsin I promoter rescues a mouse model of neuronopathic Gaucher disease but with limited cross-correction potential to astrocytes.

Gaucher disease is caused by mutations in the GBA gene, which encodes for the lysosomal enzyme ß-glucocerebrosidase (GCase), resulting in the accumulation of storage material in visceral organs and in some cases the brain of affected patients. While there is a commercially available treatment for the systemic manifestations, neuropathology still remains untreatable. We previously demonstrated that gene therapy represents a feasible therapeutic tool for the treatment of the neuronopathic forms of Gaucher disease (nGD). In order to further enhance the therapeutic affects to the central nervous system, we systemically delivered an adeno-associated virus (AAV) serotype 9 carrying the human GBA gene under control of a neuron-specific promoter to an nGD mouse model. Gene therapy increased the life span of treated animals, rescued the lethal neurodegeneration, normalized the locomotor behavioural defects and ameliorated the visceral pathology. Together, these results provided further indication of gene therapy as a possible effective treatment option for the neuropathic forms of Gaucher disease.

Neonatal brain-directed gene therapy rescues a mouse model of neurodegenerative CLN6 Batten disease.

The neuronal ceroid lipofuscinoses (NCLs), more commonly referred to as Batten disease, are a group of inherited lysosomal storage disorders that present with neurodegeneration, loss of vision and premature death. There are at least 13 genetically distinct forms of NCL. Enzyme replacement therapies and pre-clinical studies on gene supplementation have shown promising results for NCLs caused by lysosomal enzyme deficiencies. The development of gene therapies targeting the brain for NCLs caused by defects in transmembrane proteins has been more challenging and only limited therapeutic effects in animal models have been achieved so far. Here, we describe the development of an adeno-associated virus (AAV)-mediated gene therapy to treat the neurodegeneration in a mouse model of CLN6 disease, a form of NCL with a deficiency in the membrane-bound protein CLN6. We show that neonatal bilateral intracerebroventricular injections with AAV9 carrying CLN6 increase lifespan by more than 90%, maintain motor skills and motor coordination and reduce neuropathological hallmarks of Cln6-deficient mice up to 23 months post vector administration. These data demonstrate that brain-directed gene therapy is a valid strategy to treat the neurodegeneration of CLN6 disease and may be applied to other forms of NCL caused by transmembrane protein deficiencies in the future.

The inhibition of CYP1A2, CYP2C9, and CYP2D6 by pterostilbene in human liver microsomes.

This study used human liver microsomes to assess pterostilbene's effect on the metabolic activity of cytochrome P450 (CYP) 1A2, CYP2C9, and CYP2D6. The metabolism of their substrates (phenacetin, tolbutamide, and dextromethorphan) was assayed by quantifying their relevant metabolites by HPLC. The IC50 value was used to express the strength of inhibition, and the value of a volume per dose index (VDI) was used to indicate the metabolic ability of the enzyme. In this study, pterostilbene inhibited CYP1A2, CYP2C9, and CYP2D6's metabolic activities in vitro. CYP2C9's activity was most significantly inhibited by pterostilbene; its IC50 value was 0.12±0.04 µM. The IC50 value of CYP1A2 and CYP2D6 was 56.3±10.4 µM and 62.33±11.4 µM, respectively. The finding that suggests that pterostilbene has the potential to interact with CYP2C9 substrates in vivo. These results warrant clinical studies to assess the in vivo significance of these interactions.

Epilepsy Gene Therapy Using an Engineered Potassium Channel.

Refractory focal epilepsy is a devastating disease for which there is frequently no effective treatment. Gene therapy represents a promising alternative, but treating epilepsy in this way involves irreversible changes to brain tissue, so vector design must be carefully optimized to guarantee safety without compromising efficacy. We set out to develop an epilepsy gene therapy vector optimized for clinical translation. The gene encoding the voltage-gated potassium channel Kv1.1, KCNA1, was codon optimized for human expression and mutated to accelerate the recovery of the channels from inactivation. For improved safety, this engineered potassium channel (EKC) gene was packaged into a nonintegrating lentiviral vector under the control of a cell type-specific CAMK2A promoter. In a blinded, randomized, placebo-controlled preclinical trial, the EKC lentivector robustly reduced seizure frequency in a male rat model of focal neocortical epilepsy characterized by discrete spontaneous seizures. When packaged into an adeno-associated viral vector (AAV2/9), the EKC gene was also effective at suppressing seizures in a male rat model of temporal lobe epilepsy. This demonstration of efficacy in a clinically relevant setting, combined with the improved safety conferred by cell type-specific expression and integration-deficient delivery, identify EKC gene therapy as being ready for clinical translation in the treatment of refractory focal epilepsy.SIGNIFICANCE STATEMENT Pharmacoresistant epilepsy affects up to 0.3% of the population. Although epilepsy surgery can be effective, it is limited by risks to normal brain function. We have developed a gene therapy that builds on a mechanistic understanding of altered neuronal and circuit excitability in cortical epilepsy. The potassium channel gene KCNA1 was mutated to bypass post-transcriptional editing and was packaged in a nonintegrating lentivector to reduce the risk of insertional mutagenesis. A randomized, blinded preclinical study demonstrated therapeutic effectiveness in a rodent model of focal neocortical epilepsy. Adeno-associated viral delivery of the channel to both hippocampi was also effective in a model of temporal lobe epilepsy. These results support clinical translation to address a major unmet need.

AAV9 intracerebroventricular gene therapy improves lifespan, locomotor function and pathology in a mouse model of Niemann-Pick type C1 disease.

Niemann-Pick type C disease (NP-C) is a fatal neurodegenerative lysosomal storage disorder. It is caused in 95% of cases by a mutation in the NPC1 gene that encodes NPC1, an integral transmembrane protein localized to the limiting membrane of the lysosome. There is no cure for NP-C but there is a disease-modifying drug (miglustat) that slows disease progression but with associated side effects. Here, we demonstrate in a well-characterized mouse model of NP-C that a single administration of AAV-mediated gene therapy to the brain can significantly extend lifespan, improve quality of life, prevent or ameliorate neurodegeneration, reduce biochemical pathology and normalize or improve various indices of motor function. Over-expression of human NPC1 does not cause adverse effects in the brain and correctly localizes to late endosomal/lysosomal compartments. Furthermore, we directly compare gene therapy to licensed miglustat. Even at a low dose, gene therapy has all the benefits of miglustat but without adverse effects. On the basis of these findings and on-going ascendency of the field, we propose intracerebroventricular gene therapy as a potential therapeutic option for clinical use in NP-C.

A phase 2 trial of consolidation pembrolizumab following concurrent chemoradiation for patients with unresectable stage III non-small cell lung cancer: Hoosier Cancer Research Network LUN 14-179.

BACKGROUND: Five-year overall survival (OS) for patients with unresectable stage III non-small cell lung cancer (NSCLC) is poor. Until recently, a standard of care was concurrent chemoradiation alone. Patients with metastatic NSCLC treated with anti-programmed death 1 antibodies have demonstrated improved OS. This trial evaluated pembrolizumab as consolidation therapy after concurrent chemoradiation in patients with unresectable stage III disease. METHODS: Patients with unresectable stage III NSCLC received concurrent chemoradiation with cisplatin and etoposide, cisplatin and pemetrexed, or carboplatin and paclitaxel and 59.4 to 66.6 Gy of radiation. Patients with nonprogression of disease were enrolled and received pembrolizumab (200 mg intravenously every 3 weeks for up to 12 months). The primary endpoint was the time to metastatic disease or death (TMDD). Secondary endpoints included progression-free survival (PFS) and OS. RESULTS: The median follow-up for 93 patients (92 for efficacy) was 32.2 months (range, 1.2-46.6 months). The median TMDD was 30.7 months (95% confidence interval [CI], 18.7 months to not reached), which was significantly longer than the historical control of 12 months (P < .0001). The median PFS was 18.7 months (95% CI, 12.4-33.8 months), and the median OS was 35.8 months (95% CI, 24.2 months to not reached). The 1-, 2-, and 3-year OS estimates were 81.2%, 62.0%, and 48.5%, respectively. Forty patients (43.5%) completed 12 months of treatment (median number of cycles, 13.5). Symptomatic pneumonitis (grade 2 or higher) was noted in 16 patients (17.2%); these cases included 4 grade 3 events (4.3%), 1 grade 4 event (1.1%), and 1 grade 5 event (1.1%). CONCLUSIONS: Consolidation pembrolizumab after concurrent chemoradiation improves TMDD, PFS, and OS in comparison with historical controls of chemoradiation alone. Rates of grade 3 to 5 pneumonitis were similar to those reported with chemoradiation alone.

Biological and clinical implications of BIRC3 mutations in chronic lymphocytic leukemia.

BIRC3 is a recurrently mutated gene in chronic lymphocytic leukemia (CLL) but the functional implications of BIRC3 mutations are largely unexplored. Furthermore, little is known about the prognostic impact of BIRC3 mutations in CLL cohorts homogeneously treated with first-line fludarabine, cyclophosphamide, and rituximab (FCR). By immunoblotting analysis, we showed that the non-canonical nuclear factor-κB pathway is active in BIRC3-mutated cell lines and in primary CLL samples, as documented by the stabilization of MAP3K14 and by the nuclear localization of p52. In addition, BIRC3-mutated primary CLL cells are less sensitive to flu-darabine. In order to confirm in patients that BIRC3 mutations confer resistance to fludarabine-based chemoimmunotherapy, a retrospective multicenter cohort of 287 untreated patients receiving first-line FCR was analyzed by targeted next-generation sequencing of 24 recurrently mutated genes in CLL. By univariate analysis adjusted for multiple comparisons BIRC3 mutations identify a poor prognostic subgroup of patients in whom FCR treatment fails (median progression-free survival: 2.2 years, P<0.001) similar to cases harboring TP53 mutations (median progression-free survival: 2.6 years, P<0.0001). BIRC3 mutations maintained an independent association with an increased risk of progression with a hazard ratio of 2.8 (95% confidence interval 1.4-5.6, P=0.004) in multivariate analysis adjusted for TP53 mutation, 17p deletion and IGHV mutation status. If validated, BIRC3 mutations may be used as a new molecular predictor to select high-risk patients for novel frontline therapeutic approaches.

Nitrous oxide inhalant abuse and massive pulmonary embolism in COVID-19.

A patient presented to the emergency department with altered mental status and lower extremity weakness in the setting of nitrous oxide inhalant abuse and Coronavirus Disease-2019 (COVID-19) infection. He subsequently developed hypotension and severe hypoxia, found to have a saddle pulmonary embolus (PE) with right heart strain requiring alteplase (tPA).

Clinical and Molecular Features of Early Infantile Niemann Pick Type C Disease.

Niemann Pick disease type C (NPC) is a neurovisceral disorder due to mutations in NPC1 or NPC2. This review focuses on poorly characterized clinical and molecular features of early infantile form of NPC (EIF) and identified 89 cases caused by NPC1 (NPC1) and 16 by NPC2 (NPC2) mutations. Extra-neuronal features were common; visceromegaly reported in 80/89 NPC1 and in 15/16 NPC2, prolonged jaundice in 30/89 NPC1 and 7/16 NPC2. Early lung involvement was present in 12/16 NPC2 cases. Median age of neurological onset was 12 (0-24) and 7.5 (0-24) months in NPC1 and NPC2 groups, respectively. Developmental delay and hypotonia were the commonest first detected neurological symptoms reported in 39/89 and 18/89 NPC1, and in 8/16 and 10/16 NPC2, respectively. Additional neurological symptoms included vertical supranuclear gaze palsy, dysarthria, cataplexy, dysphagia, seizures, dystonia, and spasticity. The following mutations in homozygous state conferred EIF: deletion of exon 1+promoter, c.3578_3591 + 9del, c.385delT, p.C63fsX75, IVS21-2delATGC, c. 2740T>A (p.C914S), c.3584G>T (p.G1195V), c.3478-6T>A, c.960_961dup (p.A321Gfs*16) in NPC1 and c.434T>A (p.V145E), c.199T>C (p.S67P), c.133C>T (p.Q45X), c.141C>A (p.C47X) in NPC2. This comprehensive analysis of the EIF type of NPC will benefit clinical patient management, genetic counselling, and assist design of novel therapy trials.

E. coli expression and immunological assessment of expressed recombinant Newcastle disease virus hemagglutinin-neuraminidase protein in chickens.

Every year, the poultry industry experiences significant economic losses due to epidemics of Newcastle disease virus (NDV). Developing new vaccines by identifying and using the immunogenic hemagglutinin-neuraminidase (HN) protein can protect the poultry industry. In the present study, the full-length HN protein was expressed in Escherichia coli (E. coli) BL21 (DE3) cells, purified via affinity chromatography and detected via western blot analysis using His-specific antibodies. The purified HN protein was further evaluated in chickens to study the immune response against NDV. The successful production of HN-specific IgY proved the activity of the purified HN protein. IgY was present in the serum of immunized chickens. However, the immune response was higher in chickens immunized with purified HN protein along with complete and incomplete adjuvants than in chickens immunized with only the HN protein. Keywords: protein; Newcastle disease virus; poultry; infectious diseases; vaccines.

Neuroprotective exendin-4 enhances hypothermia therapy in a model of hypoxic-ischaemic encephalopathy.

Hypoxic-ischaemic encephalopathy remains a global health burden. Despite medical advances and treatment with therapeutic hypothermia, over 50% of cooled infants are not protected and still develop lifelong neurodisabilities, including cerebral palsy. Furthermore, hypothermia is not used in preterm cases or low resource settings. Alternatives or adjunct therapies are urgently needed. Exendin-4 is a drug used to treat type 2 diabetes mellitus that has also demonstrated neuroprotective properties, and is currently being tested in clinical trials for Alzheimer's and Parkinson's diseases. Therefore, we hypothesized a neuroprotective effect for exendin-4 in neonatal neurodisorders, particularly in the treatment of neonatal hypoxic-ischaemic encephalopathy. Initially, we confirmed that the glucagon like peptide 1 receptor (GLP1R) was expressed in the human neonatal brain and in murine neurons at postnatal Day 7 (human equivalent late preterm) and postnatal Day 10 (term). Using a well characterized mouse model of neonatal hypoxic-ischaemic brain injury, we investigated the potential neuroprotective effect of exendin-4 in both postnatal Day 7 and 10 mice. An optimal exendin-4 treatment dosing regimen was identified, where four high doses (0.5 µg/g) starting at 0 h, then at 12 h, 24 h and 36 h after postnatal Day 7 hypoxic-ischaemic insult resulted in significant brain neuroprotection. Furthermore, neuroprotection was sustained even when treatment using exendin-4 was delayed by 2 h post hypoxic-ischaemic brain injury. This protective effect was observed in various histopathological markers: tissue infarction, cell death, astrogliosis, microglial and endothelial activation. Blood glucose levels were not altered by high dose exendin-4 administration when compared to controls. Exendin-4 administration did not result in adverse organ histopathology (haematoxylin and eosin) or inflammation (CD68). Despite initial reduced weight gain, animals restored weight gain following end of treatment. Overall high dose exendin-4 administration was well tolerated. To mimic the clinical scenario, postnatal Day 10 mice underwent exendin-4 and therapeutic hypothermia treatment, either alone or in combination, and brain tissue loss was assessed after 1 week. Exendin-4 treatment resulted in significant neuroprotection alone, and enhanced the cerebroprotective effect of therapeutic hypothermia. In summary, the safety and tolerance of high dose exendin-4 administrations, combined with its neuroprotective effect alone or in conjunction with clinically relevant hypothermia make the repurposing of exendin-4 for the treatment of neonatal hypoxic-ischaemic encephalopathy particularly promising.

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.

Inhibition of Mitochondrial Complex I Impairs Release of α-Galactosidase by Jurkat Cells.

Fabry disease (FD) is caused by mutations in the GLA gene that encodes lysosomal α-galactosidase-A (α-gal-A). A number of pathogenic mechanisms have been proposed and these include loss of mitochondrial respiratory chain activity. For FD, gene therapy is beginning to be applied as a treatment. In view of the loss of mitochondrial function reported in FD, we have considered here the impact of loss of mitochondrial respiratory chain activity on the ability of a GLA lentiviral vector to increase cellular α-gal-A activity and participate in cross correction. Jurkat cells were used in this study and were exposed to increasing viral copies. Intracellular and extracellular enzyme activities were then determined; this in the presence or absence of the mitochondrial complex I inhibitor, rotenone. The ability of cells to take up released enzyme was also evaluated. Increasing transgene copies was associated with increasing intracellular α-gal-A activity but this was associated with an increase in Km. Release of enzyme and cellular uptake was also demonstrated. However, in the presence of rotenone, enzyme release was inhibited by 37%. Excessive enzyme generation may result in a protein with inferior kinetic properties and a background of compromised mitochondrial function may impair the cross correction process.

4EGI-1 represses cap-dependent translation and regulates genome-wide translation in malignant pleural mesothelioma.

Deregulation of cap-dependent translation has been implicated in the malignant transformation of numerous human tissues. 4EGI-1, a novel small-molecule inhibitor of cap-dependent translation, disrupts formation of the eukaryotic initiation factor 4F (eIF4F) complex. The effects of 4EGI-1-mediated inhibition of translation initiation in malignant pleural mesothelioma (MPM) were examined. 4EGI-1 preferentially inhibited cell viability and induced apoptosis in MPM cells compared to normal mesothelial (LP9) cells. This effect was associated with hypophosphorylation of 4E-binding protein 1 (4E-BP1) and decreased protein levels of the cancer-related genes, c-myc and osteopontin. 4EGI-1 showed enhanced cytotoxicity in combination with pemetrexed or gemcitabine. Translatome-wide polysome microarray analysis revealed a large cohort of genes that were translationally regulated upon treatment with 4EGI-1. The 4EGI-1-regulated translatome was negatively correlated to a previously published translatome regulated by eIF4E overexpression in human mammary epithelial cells, which is in agreement with the notion that 4EGI-1 inhibits the eIF4F complex. These data indicate that inhibition of the eIF4F complex by 4EGI-1 or similar translation inhibitors could be a strategy for treating mesothelioma. Genome wide translational profiling identified a large cohort of promising target genes that should be further evaluated for their potential significance in the treatment of MPM.

Delineating pathological pathways in a chemically induced mouse model of Gaucher disease.

Great interest has been shown in understanding the pathology of Gaucher disease (GD) due to the recently discovered genetic relationship with Parkinson's disease. For such studies, suitable animal models of GD are required. Chemical induction of GD by inhibition of acid ß-glucosidase (GCase) using the irreversible inhibitor conduritol B-epoxide (CBE) is particularly attractive, although few systematic studies examining the effect of CBE on the development of symptoms associated with neurological forms of GD have been performed. We now demonstrate a correlation between the amount of CBE injected into mice and levels of accumulation of the GD substrates, glucosylceramide and glucosylsphingosine, and show that disease pathology, indicated by altered levels of pathological markers, depends on both the levels of accumulated lipids and the time at which their accumulation begins. Gene array analysis shows a remarkable similarity in the gene expression profiles of CBE-treated mice and a genetic GD mouse model, the Gba(flox/flox) ;nestin-Cre mouse, with 120 of the 144 genes up-regulated in CBE-treated mice also up-regulated in Gba(flox/flox) ;nestin-Cre mice. We also demonstrate that various aspects of neuropathology and some behavioural abnormalities can be arrested upon cessation of CBE treatment during a specific time window. Together, our data demonstrate that injection of mice with CBE provides a rapid and relatively easy way to induce symptoms typical of neuronal forms of GD. This is particularly useful when examining the role of specific biochemical pathways in GD pathology, since CBE can be injected into mice defective in components of putative pathological pathways, alleviating the need for time-consuming crossing of mice. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.