Breaking the penetration limit of microscopy – Photoswitching Optoacoustics

Project Information

SWOPT

Grant agreement ID: 101046667

DOI

10.3030/101046667

EC signature date 15 March 2022

Start date 1 September 2022

End date 31 August 2027

Funded under

• The European Innovation Council (EIC)

Total cost

€ 3 536 935,75

EU contribution

€ 3 536 935,00

Coordinated by

HELMHOLTZ ZENTRUM MUENCHEN DEUTSCHES FORSCHUNGSZENTRUM FUER GESUNDHEIT UND UMWELT GMBH
Germany

Periodic Reporting for period 2 - SWOPT (Breaking the penetration limit of microscopy – Photoswitching Optoacoustics)

Reporting period: 2023-09-01 to 2025-02-28

Summary of the context and overall objectives of the project

SWOPT aims to solve one of the great challenges in visualising biology: break through the penetration limits of optical microscopy and image cells and their function in vivo at depths of several millimetres to centimetres, while retaining high resolution and sensitivity of single cells. The ability to visualize few cells in a live organism is important because many biological phenomena, especially in the immune system, the onset of tumours, or fundamental developmental biology, rely on a small number of cells.
SWOPT will achieve this by combining the principles of photoswitching with optoacoustic imaging. Optoacoustic imaging is a method that relies on reading out ultrasound signal generated by light. It already has the power to deliver a combination of higher penetration depth, higher resolution, and larger fields of view than other imaging technologies. However, for many research questions, optoacoustic needs tools like genetically encoded reporters and sensors. Photoswitchable label proteins can help here. These proteins can change their state upon illumination, like a tiny switch that changes the state of the protein from ON to OFF and with that, the signal it generates. In optoacoustic, we use the light switchable signal to make the label blink, which enables the visualization of small numbers of cells against a strong background of other signals, which shows a constant signal. One can imagine the effect, like the blinking of a lighthouse in a stormy dark night at sea.
SWOPT will enable examination of whole tissues in vivo with the same ease, flexibility and, eventually, abundance of tools, paralleling fluorescence microscopy, thus bringing research and understanding of living organisms to the next level. As an affordable imaging technology, SWOPT aspires to become routine in life science and bio-medical research.
The overall objective of SWOPT is to create a technology based on optoacoustic imaging and photoswitching. To this aim, a new imaging instrumentation will be developed, and SWOPT-tailored contrast agents (protein-based and synthetic) will be created. SWOPT’s abilities will be benchmarked by visualizing cells and their anatomical and chemical environment in vivo in whole tumours and in 3D models based on spheroids and organoids.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

Since the start of the project, SWOPT has made large progress on the development of the new instrumentation, algorithms and the photoswitchable agents.
We have set up a dedicated imaging system based on raster scanning optoacoustic mesoscopy coupled to an optical microscope that helps localising the regions of interest. The system can be adapted to incorporate the innovations developed in the frame of the project, including new concepts of ultrasound high frequency transducers that have never been explored so far in this context.
A crucial step to acquire a three-dimensional image of the different photoswitchable protein labeled cells in tissue is to develop suitable mathematical algorithms. Towards this end, we have already obtained a model that includes the successive steps from light excitation to ultrasound measurements and allows successful unmixing.
Regarding the development of photoswitching agents, several architectures of synthetic dyes have been characterised, and selected candidates are currently being tested for their optoacoustic signal generation. The development of genetically engineered sensors and reporters is ongoing. Some of the developed sensors have already been expressed in cultured cells, which has enabled assessing their performance in biological contexts. The results of these preliminary experiments are being analysed to fine tune the development of further improved agents.
The setup of the validation system has already started with the exploration of the best suitable in vitro models, including cultured cells and 2D/3D models based on organoids. This is a crucial step that will help designing the final validation studies in mice.
Exploitation is an important dimension of the project, and IP monitoring is done regularly. In this last project stage, an iteration of the exploitation plan has been done to concrete the potential routes to commercialization based on the most recent results.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The development of the SWOPT technology will set a new level of imaging whole opaque organism in vivo with direct impact on biomedical research. We aim to primarily target SWOPT towards the pre-clinical research market, for which our proof-of-concept experiments in cells and in mice become crucial. Upon validation and exploitation, SWOPT may be used in future studies to visualize cells, their functionality, and their chemical surrounding in context with the complex tissue anatomy. This will contribute to understanding cell specific metabolic pathways that underlie pathological mechanisms of a wide range of diseases.
Additionally, the use of SWOPT is not restricted to live animal tissues, but also expands, with further research, to the emerging large tissue organoids market or explanted organs mimicking disease states. In this sense, SWOPT aspires to become a game-changer for biomedical research, contributing to the development of new disease models and therapeutics.

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