A Multiwell Plate Format Microfluidic Immobilization Chip for High-Content Imaging of Whole Animals for in vivoNeurotoxicology Testing - Project Summary:
Neurotoxicological evaluation of new compounds intended for human use or of potential human exposure is
mandated by international regulatory bodies and largely relies on lethality testing in higher-order vertebrate
animals. High screening costs, long experimental times, and legislative requirements to reduce dependence on
animal testing have led many industries to search for alternative technologies. In vitro toxicology testing uses
isolated cells or monotypic cell culture and can only provide limited insight since these models lack biologically
relevant intact multi-typic cellular network structures. While both technologies have been augmented by in silico
technologies, there is still a non-trivial gap between what can be learned and translated from simple, fast,
inexpensive in vitro methods versus longer, complex, and costly in vivo studies in higher order animals.
Newormics’ approach to filling this gap is to enable in vivo neurotoxicological assessment in Caenorhabditis
elegans, an accepted alternative invertebrate model organism, by developing neuron-specific toxicity assays,
delivered via a proprietary high-density, large-scale microfluidic immobilization device for high-content, high
throughput analysis. Building on advances made during Phase I and important market learnings from
participation in the NIH I-Corps program, Phase II proposes several new elements of innovation to achieve our
goals in 3 specific aims. In Aim 1, we will convert our first-generation microfluidic device to a high-density (384-
well) vivoChip with improved microfabrication technologies, incorporate on-chip culture for transfer-less
exposure and testing, and integrate automation for chip loading, imaging, and analysis. These measures will
significantly increase test scale (from 80 compounds per chip to 280) and lower the consumable and labor costs
per test. In Aim 2, building on our dopaminergic neurotox assay from Phase I, we will develop four neurotox
assays with brightly fluorescently labeled dopaminergic, serotonergic, GABAergic, and cholinergic neurons
providing the unprecedented ability to assess subtle phenotypic effects of chemicals on individual intact,
functional neurons. To achieve real-time image processing, multi-parameter phenotyping, and managing the
terabytes of image data generated per test, we will build a computational platform empowered by a graphic user
interface. This platform will be used for image compilation, user-annotated phenotype definition and scoring, and
automated report generation with appropriate statistical analysis. In Aim 3, with our industry partners, we will
validate our platform and assays using reference chemicals. As more chemicals are tested, we will build a
database which can be further mined. The outcome of this work will enable many industries to reduce lethal
animal testing and get safer industrial and personal consumer products to market faster for economic benefit,
reaching regulatory compliance for reduced animal use, and improved healthcare for neurological diseases.
