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Non-invasive measuring of enzymatic activity and metabolite absorption in living animals using sensitive microsystem

English title Non-invasive measuring of enzymatic activity and metabolite absorption in living animals using sensitive microsystem
Applicant Dubikovskaya Elena
Number 157023
Funding scheme Interdisciplinary projects
Research institution Institut des sciences et ingénierie chimiques EPFL - SB - ISIC
Institution of higher education EPF Lausanne - EPFL
Main discipline Biochemistry
Start/End 01.11.2014 - 31.10.2018
Approved amount 707'411.00
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All Disciplines (2)

Discipline
Biochemistry
Microelectronics. Optoelectronics

Keywords (5)

Enzymatic activity; Microlectronique; metabolite absoprtion; in vivo; Non-invasive sensing

Lay Summary (French)

Lead
Ce projet de recherche vise à développer une nouvelle méthodologie pour l’évaluation en temps réel de l’activité de certains enzymes ainsi que de mesurer l’absorption de métabolites dans les animaux vivants de façon non-invasive.
Lay summary

Le projet que nous souhaitons réaliser est basé sur le développement d’une nouvelle technologie. Celle-ci permet la mesure en temps réel de l’activité de certaines protéines associées par exemple, au développement du cancer ou encore pour mesurer l’absorption de certains nutriments comme les acides gras ou les sucres. Cette technologie nous permettra d’évaluer l’efficacité des médicaments en temps réel ou encore de comprendre de manière plus efficace notre métabolisme au cours du temps.

De façon très similaire aux implants contraceptifs, nous avons développés un implant contenant des cellules mammifères qui produisent une enzyme spéciale appelée luciférase. Cette enzyme permet de libérer de la lumière après interaction avec une petite substance appelée Luciferin. Cette substance  (luciferin) est libérée dans notre corps seulement si la protéine étudiée est produite. La quantité de lumière émise est proportionnelle à la quantité de protéines présente ou la quantité de nutriments qui a été consommée. Cette lumière est collectée et mesurée à l’aide d’un micro-détecteur similaire aux détecteurs des appareils photographiques.

 

Cette technologie est en théorie applicable à tous types d’animaux comme les souris, chiens, cochons, singes et potentiellement à l’homme. Nous espérons que cela nous permettra de mieux comprendre les mécanismes des métabolismes du corps humain mais aussi de pouvoir suivre le traitement des patients atteints de maladie, de façon plus efficace.

Direct link to Lay Summary Last update: 15.10.2014

Responsible applicant and co-applicants

Employees

Publications

Publication
Toward low-power PPG-based heart rate monitoring using a MOS-PN hybrid photosensor
Gosselin P., Bucella P., Koukab A., Kayal M. (2017), Toward low-power PPG-based heart rate monitoring using a MOS-PN hybrid photosensor, in 15th IEEE International New Circuits and Systems Conference (NEWCAS), 141-144.
Computing the Impact of White and Flicker Noise in Continuous-Time Integrator-Based ADCs
Gosselin P., Koukab A., Kayal M. (2016), Computing the Impact of White and Flicker Noise in Continuous-Time Integrator-Based ADCs, in Proceedings of the 23rd International Conference Mixed Design of Integrated Circuits and Systems, MIXDES 2016, 316/752975-320.
Estimating the Noise-Related Error in Continuous-Time Integrator-Based ADCs
Gosselin P., Koukab A., Kayal M. (2016), Estimating the Noise-Related Error in Continuous-Time Integrator-Based ADCs, in International Journal of Microelectronics and Computer Science, 7(2, 2016), 54-59.

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
Keystone meeting on Tumor Metabolism Poster Bioluminescent-based imaging and quantification of glucose uptake in vivo 24.02.2019 Colorado, United States of America Dubikovskaya Elena;


Communication with the public

Communication Title Media Place Year
Media relations: print media, online media Reagents track glucose uptake International 2019
Media relations: print media, online media Shedding light on cancer metabolism in real-time with bioluminescence Western Switzerland 2019

Associated projects

Number Title Start Funding scheme
150134 Molecular in vivo imaging of lipid metabolism 01.01.2014 Project funding (Div. I-III)

Abstract

Currently no efficient methodologies exist that allow real-time non-invasive sensing of enzymatic and metabolic events in living organisms. To address the unmet need we propose to develop a non-invasive bioluminescent (BL) method for evaluation of enzymatic activity and metabolite absorption in animal models of human disease. The methodology is based on bioluminescent imaging (BLI) that is widely used in the field of pre-clinical research to monitor tumor progression, metastasis and response to candidate therapeutics in animal models. The prototypical BLI experiment employs a luciferase enzyme as a reporter and enzymatic oxidation of its substrate luciferin is accompanied by photon emission. Importantly, BLI obviates several problems associated with fluorescence imaging, such as tissue-derived background signal, photobleaching, and inefficient probe excitation. Recent development of BLI probes to sense molecular signatures of target tissues relies on simple principle that luciferin "caged" on the phenolic oxygen is not a substrate for luciferase unless it gets "uncaged" by specific biological process of interest. For example, several luciferin probes have been designed to sense enzymatic activities such as those of beta-galactosidase, caspases, beta-lactamases, and furin. Another set of probes allows quantification of hydrogen peroxide and fatty acid fluxes as well as protein-protein interactions. However, despite its great sensitivity and ease of use, current BLI has serious limitations. First, it is only limited to the use in transgenic mice that genetically express luciferase enzyme either ubiquitously or in certain organs as well as animals transplanted with luciferase-expressing cells. Unfortunately, these represent only a small percentage of existing animal models of human disease. Secondly, all current technologies that allow sensing of bioluminescent signal are limited to the use in rodents (mice and rats). In the standard set-up of typical experiment the animals are placed into the "black box" of IVIS camera (PerkinElmer, USA) and optical signal is detected using "charged coupled device" (CCD). This represents another serious limitation for the use of this technology for drug development because a big fraction of in vivo tests are done on non-rodent animal models. Third, the animals must be anesthetized during the signal acquisition that drastically changes their metabolism and affects multiple biological processes in the body. This is the major reason why none of the current technologies have been adopted by pharmaceutical companies for the use in drug development research.Therefore, there is a pressing demand for more efficient in vivo models to evaluate enzymatic activities and metabolite absorption in live animals that currently do not exist. Since most of the enzymes are intracellular, they cannot be directly measured in blood, urine, or other bodily fluids. Thus, whole body monitoring of particular disease-related enzyme would require a substrate of this enzyme to be injected in order for it to be taken up by all cells of interest inside the organism, and enzymatically cleaved (metabolized). The resulting metabolite is further detected by blood/tissue sampling followed by liquid chromatography/mass spectrometry (LS/MS) analysis. This procedure is very labour-intensive because multiple blood/tissue manipulations steps are required to obtain the data. In addition, it is highly invasive for animals leading to big number being sacrificed in the course of one assay. This makes the current technology very expensive and not suitable for screening of large number of drug candidates and experimental conditions. The same thing is true for studies of metabolic fluxes where no real-time non-invasive methods currently exist for evaluation of these important biological processes.To address this unmet need for efficient in vivo models, we propose to develop a non-invasive bioluminescent (BL) method that should allow real-time longitudinal study using a highly sensitive sensor microsystem (SM) and Cell Encapsulation Device (CED). The new sensing technology has several advantages over existing methods. First, it will allow real-time non-invasive evaluation of enzymatic activity and metabolic fluxes and has the potential to become an important tool for drug discovery. Second, since the method is based on highly sensitive imaging modality, it is expected to achieve better sensitivity in comparison to the one based on tissue/blood sampling. Another significant advantage of the new methodology is the ability to obtain kinetic longitudinal readout of enzymatic activity, without any need of tissue manipulation steps. Third, the new method aims to significantly reduce the number of animals because the technique is highly non-invasive and animals can be re-used used for other assays. In addition, the new sensing technology would reduce the work load associated with tissue manipulation steps therefore making this procedure a lot less expensive. Forth, it can be extended to studies of metabolite absorption in real-time non-invasive manner, for which not good methods exist. The new methodology will be immediately adapted for non-invasive evaluation of activities of enzymes for which suitable reagents described above already exist (ex. caspase 3/7, b-lactamase, b-galactosidase, fatty acid absorption, etc). In addition, we propose to expend the list of existing imaging reagents by making new probes for evaluation of activity of glucose transporters that play important role in many human diseases. Our preliminary data on some of the novel probes for studies of glucose uptake provide good foundation for their further investigation. This project aims to investigate the following main points:•Development of novel "Cell Encapsulation Device" (CED) that would consist of polymeric "hollow fiber" implanted with cells expressing luciferase reporter that will survive at high density for several weeks. Once the imaging reagent gets "uncaged" by corresponding enzyme, the CED will produce light, proportional to the activity of the enzyme or flux of metabolite. •Development of a new sensor microsystem for detection of bioluminescent signal produced as the result of activity of certain enzyme and determination of optimal measurement parameters with this novel device.•Analysis of the microelectronics approach able to efficiently support performance of high accuracy detector.•Design of the microelectronic circuits and corresponding algorithm for cost-efficient ultra-high sensitivity device.•Bench-marking and validation of this new approach using existing enzymatic activity imaging reagents such as probes for measurements of caspase 3/7 or beta-galactosidase activity.•Extension of this novel sensing technology for imaging of metabolite absorption. Validation with existing fatty acid uptake probes and development of novel probes to study activity of glucose transporters.
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