Droplet-based microfluidic platform for intracellular ion channel drug discovery

New technology is widening the chance of developing new pharmacological compounds and has the potential to create new jobs and have economic and societal impact on healthcare. Economic impact: The average expenditure to develop and bring to market a new drug is estimated to be approximately $2 billion, with target identification, discovery and the development of a new chemical compound before clinical trials accounting for 30% of the total cost. Commercialization of the developed platform has potential to add considerable information over the gold standard obtained by current electrophysiological tests with automated patch-clamp technology and impact the pre-screening stages of compounds, most of which is currently provided as a service by CROs and/or CMOs. Therefore, outcomes from this project have potential to carve a unique sector in the market of drug screening instrumentation. To assess this impact and available opportunities, we have secured the participation of members from academia, healthcare technology and industry to act as an advisory board for consultancy regarding knowledge transfer and commercialisation. Industrial impact: The developed system will be tested in collaboration with project partner Apconix for the development of pharmacological assays based on combinatorial chemistry approaches. Outside the immediate benefit in identifying new active compounds for the CLIC4 channel, the potential for translating the developed procedures for other pharmacologically relevant eukaryotic ion channels present cost-saving arguments against current live-cell based, large-screening assay in the Pharmaceutical industry. In addition, the collaboration with project partner Smartox will explore a further application of the technology for use with limited resources of venom samples only available in extremely small quantities that currently elude industrial automated screening. The proposed technology has the potential to greatly impact drug discovery by identifying new efficient venom-derived drugs. Together, this double partnership will enable us to address different screening needs for both SME and the Pharmaceutical Industry. Societal impact: Identification of new drug candidates from the developed technology will expand the panel of drugs with implications in cancer, mitochondrial dysfunction and neurodegenerative diseases. Improving the efficacy of identified drugs at preclinical level is expected to improve outcomes during clinical trials. Therefore, wide societal benefits would be in the form of new medicine and their improved efficacy. Furthermore, the applicability of the proposed technology to the other identified applications (see academic beneficiaries) and healthcare related fields could impact the screening of vaccines and environmental toxins, all of which will produce massive societal benefits

Droplet-based microfluidic platform for intracellular ion channel drug discovery

New technology is widening the chance of developing new pharmacological compounds and has the potential to create new jobs and have economic and societal impact on healthcare. Economic impact: The average expenditure to develop and bring to market a new drug is estimated to be approximately $2 billion, with target identification, discovery and the development of a new chemical compound before clinical trials accounting for 30% of the total cost. Commercialization of the developed platform has potential to add considerable information over the gold standard obtained by current electrophysiological tests with automated patch-clamp technology and impact the pre-screening stages of compounds, most of which is currently provided as a service by CROs and/or CMOs. Therefore, outcomes from this project have potential to carve a unique sector in the market of drug screening instrumentation. To assess this impact and available opportunities, we have secured the participation of members from academia, healthcare technology and industry to act as an advisory board for consultancy regarding knowledge transfer and commercialisation. Industrial impact: The developed system will be tested in collaboration with project partner Apconix for the development of pharmacological assays based on combinatorial chemistry approaches. Outside the immediate benefit in identifying new active compounds for the CLIC4 channel, the potential for translating the developed procedures for other pharmacologically relevant eukaryotic ion channels present cost-saving arguments against current live-cell based, large-screening assay in the Pharmaceutical industry. In addition, the collaboration with project partner Smartox will explore a further application of the technology for use with limited resources of venom samples only available in extremely small quantities that currently elude industrial automated screening. The proposed technology has the potential to greatly impact drug discovery by identifying new efficient venom-derived drugs. Together, this double partnership will enable us to address different screening needs for both SME and the Pharmaceutical Industry. Societal impact: Identification of new drug candidates from the developed technology will expand the panel of drugs with implications in cancer, mitochondrial dysfunction and neurodegenerative diseases. Improving the efficacy of identified drugs at preclinical level is expected to improve outcomes during clinical trials. Therefore, wide societal benefits would be in the form of new medicine and their improved efficacy. Furthermore, the applicability of the proposed technology to the other identified applications (see academic beneficiaries) and healthcare related fields could impact the screening of vaccines and environmental toxins, all of which will produce massive societal benefits

Transitioning from multi-phase to single-phase microfluidics for long-term culture and treatment of multicellular spheroids

When compared to methodologies based on low adhesion or hanging drop plates, droplet microfluidics offers several advantages for the formation and culture of multicellular spheroids, such as the potential for higher throughput screening and the use of reduced cell numbers, whilst providing increased stability for plate handling. However, a drawback of the technology is its characteristic compartmentalisation which limits the nutrients available to cells within an emulsion and poses challenges to the exchange of the encapsulated solution, often resulting in short-term cell culture and/or viability issues. The aim of this study was to develop a multi-purpose microfluidic platform that combines the high-throughput characteristics of multi-phase flows with that of ease of perfusion typical of single-phase microfluidics. We developed a versatile system to upscale the formation and long-term culture of multicellular spheroids for testing anticancer treatments, creating and array of fluidically addressable, compact spheroids that could be cultured in either medium or within a gel scaffold. The work provides proof-of-concept results for using this system to test both chemo- and radio-therapeutic protocols using in vitro 3D cancer models

Perfusion based microfluidic system for pharmacological profiling of neuronal networks

This work presents the integration of a semi-automated microfluidic platform that utilizes calcium imaging to enable the pharmacological characterization of functionally connected, but environmentally isolated neuronal networks. This approach allows, for the first time, to assess the cause-effect relationship of neuronal communication following drug application, thus allowing the pharmacological characterisation of novel drugs proposed to influence communication between neuronal networks

Controlled delivery of membrane proteins to artificial lipid bilayers by nystatin-ergosterol modulated vesicle fusion

The study of ion channels and other membrane proteins and their potential use as biosensors and drug screening targets require their reconstitution in an artificial membrane. These applications would greatly benefit from microfabricated devices in which stable artificial lipid bilayers can be rapidly and reliably formed. However, the amount of protein delivered to the bilayer must be carefully controlled. A vesicle fusion technique is investigated where composite ion channels of the polyene antibiotic nystatin and the sterol ergosterol are employed to render protein-carrying vesicles fusogenic After fusion with an ergosterol-free artificial bilayer the nystatin-ergosterol channels do not dissociate immediately and thus cause a transient current signal that marks the vesicle fusion event. Experimental pitfalls of this method were identified, the influence of the nystatin and ergosterol concentration on the fusion rate and the shape of the fusion event marker was explored, and the number of different lipid was reduced. Under these conditions, the B-amyloid peptide could be delivered in a controlled manner to a standard planar bilayer. Additionally, the electrical recordings were obtained of vesicles fusing with a planar lipid bilayer in a microfabricated device, demonstrating the suitability of nystatin-ergosterol modulated vesicle fusion for protein delivery within microsystems

In-line single-mode fiber variable optical attenuator based on electrically addressable microdroplets

We report an in-line, fiber optic, broadband variable optical attenuator employing a side-polished, single-mode optical fiber integrated on a digital microfluidics platform. The system is designed to electrically translate a liquid droplet along the polished surface of an optical fiber using electrowetting forces. This fiber optic device has the advantage of no moving mechanical parts and lends itself to miniaturization. A maximum attenuation of 25 dB has been obtained in the wavelength range between 1520 nm and 1560 nm

Time-lapse measurement of single-cell response to nanomaterial : a microfluidic approach

This work presents the successful application of a single-cell microfluidic platform for high-throughput, real-time screening of nanoparticle-cell interactions. Taking vaccine delivery as a proof-of-concept application, ovalbumin-conjugated gold nanorods were produced and controllably delivered to primary dendritic cells within the device. Time-lapse imaging enabled monitoring of hundreds of single-cells during exposure to a range of concentrations of nanoparticle conjugates and simultaneous quantification of specific cellular functions. This integrated system provides throughput and statistical data comparable to that obtained with flow cytometry but also offers a novel approach to determine the dynamics of nanoparticle-cell interactions and nanoparticle-mediated antigen delivery with single-cell resolution

A microfluidics tool for high-throughput, real-time multimodal imaging of nanoparticle-cell interactions

The increasing use of nanomaterials for biomedical applications has raised the need for efficient, robust and low-cost high-throughput assessment of nanotoxicity and cell-nanoparticle interactions. Microfluidics provides the tools for high-throughput single-cell functional monitoring, while gold nanorods have unique potential for intracellular tracking and can simultaneously be used as drug carriers. Presented here is a miniaturised platform that integrates these features with a multimodal approach to cell imaging. A microfluidic device allows for trapping of an array of singlecells, followed by the controlled delivery of nanoparticles into the cell array and subsequent real-time multimodal imaging of cellular interactions with functionalised nanoparticles. This system has been successfully used to assess cellnanoparticle interactions at the single-cell level