Targeting Neuroinflammation to Combat Neurodegenerative and Rare Diseases

Tech Snapshot® captures today’s cutting-edge tools and technologies that will help drive drug discovery tomorrow. This installment was written by QPS, a global leader in clinical and preclinical research.

Drug development for the treatment of neurodegenerative diseases focusses mostly on disease-specific targets like amyloid plaques in Alzheimer’s disease (AD) or α-synuclein in Parkinson’s disease (PD). Although newly developed drugs are successfully binding such targets, the effect on disease symptoms in humans is often disappointing. New approaches thus aim at more general pathological disease features. Neuroinflammation is known to be a common pathological hallmark of many neurodegenerative and rare diseases, characterized by increased cytokine release, activated microglia and astrocytosis. In Alzheimer’s disease patients, for example, microglial activation parallels Aβ plaque burden and is, as such, a characteristic feature of AD progression. Oxidative stress can activate inflammatory processes and is thought to be one of the earliest events in the development of AD and other neurodegenerative diseases.

Modelling neuroinflammation

To support the drug development process, QPS Neuropharmacology offers preclinical research services with several in vitro and in vivo models of neuroinflammation:

General models

Neuroinflammation is in vitro and in vivo modelled by using lipopolysaccharide (LPS). In vitro modelling is performed in the microglia cell line BV-2, organotypic hippocampal or whole brain slices, as well as primary mouse microglia by stimulating cells with LPS for several hours. In vivo, LPS induces neuroinflammation in the mouse hippocampal formation after intraperitoneal injections on four consecutive days. Additionally, LPS can be directly administrated into the brain via stereotactic injections. Set up for both in vitro and in vivo models is quick, resulting in a fast turn-around time, and therefore rapidly showing first effects of test compounds.

Disease-specific models

Transgenic, knock-in, knockout and induced in vivo models are all valuable to model neuroinflammation in neurodegenerative and rare diseases. Mouse models of Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, Amyotrophic Lateral Sclerosis (ALS), Niemann-Pick disease, Gaucher disease, Pompe disease, Mucopolysaccaridosis IIIA and Multiple Sclerosis are available at QPS Neuropharmacology, and they have all been validated for their neuroinflammatory pathology. These models can be applied for in vivo efficacy studies using many different treatment routes, ranging from oral treatment via gavage, intravenous injection, stereotactic intracerebral injection and many more.

Additionally, microglia of adult mice can be isolated by Magnetic Cell Sorting (MACS) and used to test compounds ex vivo. For the majority of these models, it has already been proven, that neuroinflammatory hallmarks present a susceptible target for efficacy studies.

Evaluation of neuroinflammation

To analyze neuroinflammation, several different methods are available at QPS Neuropharmacology, depending on model and research question:

Oxidative stress

Oxidative stress and neuroinflammation are two tightly linked pathways in neuropathological processes. Oxidative stress is commonly determined by measuring reactive oxygen species (ROS) and nitric oxygen (NO) release. There are many different ROS assays available measuring oxidative stress in vitro (e.g., DCFDA assays, CellROXTM reagent). Plasma levels of thiobarbituric acid reactive substances (TBARS) represent a currently well-recognized marker for systemic oxidative stress in human subjects. The TBARS assay can be performed using brain tissue and/or plasma. Additionally, 4-hydroxynonenal (4-HNE), a common byproduct of lipid peroxidation, can be used as marker for oxidative stress.

Cytokines

Cytokines are important markers to monitor inflammatory processes. Pro- and anti-inflammatory cytokines can be measured using customized multiplex assays from Meso Scale Diagnostics, LLC (MSD®). Upon LPS stimulation in different cell systems, cell culture supernatants can be collected at different timepoints and up to 10 cytokines of choice can be analyzed in one single reaction. In in vivo studies, cytokines can be analyzed in plasma and/or brain tissue. The effect of LPS stimulation on mRNA level can be also assessed using qRT-PCR.

Neuroinflammatory markers

Typical protein markers of neuroinflammation can be analyzed using standard Western blotting or, as a more sensitive approach, by WesTM (ProteinSimple®). After LPS stimulation of organotypic slices for example, inflammasome activation can be measured by quantifying NLRP3 by WesTM technology.

Additionally, any of the following inflammation markers can be qualitatively and quantitatively evaluated by immunofluorescent labeling (e. g. Iba1, CD11b, TREM2, Tmem119, CD45, CD68, F4/80, B220/CD45R, CD3). Region-specific analyses, like plaque-associated neuroinflammation in Alzheimer’s disease mouse models, allow very precise evaluations of the target protein. In addition, up to 5-channel epifluorescence imaging, allowing co-labeling and expression analysis of target proteins can be performed. Systematic uniform random sampling, automated & comprehensive multidimensional image acquisition, as well as macro-based software solutions, guarantee a rater-independent and automated analysis of histological labeling.

Targeting neuroinflammation is a promising treatment approach for many untreatable diseases. QPS Neuropharmacology therefore offers a broad portfolio to test neuroinflammation-targeting compounds.

About QPS Neuropharmacology

QPS Neuropharmacology is a division of QPS, a GLP/GCP-compliant contract research organization (CRO) delivering the highest grade of discovery, preclinical, and clinical drug development services since 1995. QPS Neuropharmacology focuses on preclinical studies related to central nervous system (CNS) diseases, rare diseases and mental disorders. With highly predictive disease models available on site and unparalleled preclinical experience, QPS Neuropharmacology can handle most CNS drug development needs for biopharmaceutical companies of all sizes. For more information about QPS visit us on Scientist.com or www.qps.com, and for more information about QPS Neuropharmacology, visit www.qpsneuro.com.