Home SynCell2022 Abstracts for SynCell2022 Abstracts for parallel session (Wed): Applications in medicine

Abstracts for parallel session on Wednesday May 18th

Applications in medicine: drug delivery and hybrid cultures

1.1: Temperature controlled release of in-situ synthesised proteins from synthetic cells

Razia Chowdhry, Michael Booth

University of Oxford, United Kingdom

Thermosensitive liposomes have been produced for therapeutic controlled delivery of small molecules in tumours with local hyperthermia. This project focuses on producing lysolipid thermosensitive synthetic cells (LTS-SC) using the hydration method of lyophilized vesicles, to encapsulate proteins of a cell-free protein synthesis (CFPS) system and UV light-activated DNA. Synthetic cells (SC) are lipid-based compartments that contain CFPS system that offer promise as drug delivery systems, as they will be able to synthesise therapeutic molecules in-situ at the target site. Spatiotemporal controlled expression of protein by UV-light and then using mild hyperthermia as a control for the release of the proteins can be applied. The performance of both mechanisms is characterised via released protein (e.g. β – galactosidase) action on reporter substrates.

1.2: Bottom-up assembly of biomimetic cell-cell
interaction modules using lipid-based synthetic cells

Martin Schröter, Jacqueline De Lora, Ilja Platzman, Joachim Spatz

Max Planck Institute for Medical Research, Germany

Research on bottom-up assembly of hybrid tissues comprising mixtures of synthetic and natural cells is indeed a high priority field in synthetic biology. Nevertheless, considerable challenges still loom at many steps in the process of stable integration of functional fully synthetic cells into the natural cell culture. To fulfil this gap, we describe an approach that integrates giant unilamellar vesicle (GUV)-based synthetic cell adhesion modules with natural cells by exploiting the natural cell-cell adhesion protein, E-cadherin. The proposed model sets the base to construct interconnected, cell-like communication networks by means of chemical and mechanical signals as mediated by E-cadherin. For bottom-up generation of E-cadherin-functionalized GUVs, we use droplet-based microfluidics. Then we construct a hybrid system by coculturing functionalized GUVs with epithelial cells and measure their interactions using a high-throughput approach combining fluorescence microscopy and image analysis. In the future, this system will be used to study the role of cell-cell adhesions during complex biological processes like collective cell migration during wound healing.

1.3: Temperature controlled release of in-situ
synthesised proteins from synthetic cells

Gal Chen1, Rotem Levin1, Shira Landau2, Maya Kaduri1, Omer Adir1, Iris Ianovici2, Jeny Shklover1, Janna Shanisky-Roitman1, Shulamit Levenberg2, Avi Schroeder1

1: Department of Chemical Engineering, Technion – Israel Institute of Technology, Israel;  2: Department of Biomedical Engineering, Technion, Israel;

Technological advances in synthetic biology push the development of synthetic cells (SCs), autonomous protein-manufacturing particles, as promising mimetic replacements for dysfunctional cells inside the body. Here we report for the first time, that SCs genetically encoded to produce proangiogenic factors stimulate the physiological process of neovascularization in mice. The SCs were constructed from giant lipid vesicles with optimal membrane composition sustaining an enhanced protein production capacity. When introduced with the appropriate genetic and chemical design, the SCs were able to synthesize a recombinant, basic fibroblast growth factor (bFGF) under physiological conditions, reaching the expression of up to 9·10^6 protein copies per SC. Next, we confirmed the proangiogenic activity of the SCs, showing their ability to induce endothelial cells’ proliferation, migration, tube formation, and angiogenesis-related intracellular signalings. Integrating the SCs with bioengineered constructs bearing endothelial and mural cells promoted the remodeling of a mature vascular network supported by a collagen-IV, basement-membrane-like matrix. Finally, prolonged local administration of the SCs in mice triggered the infiltration of blood vessels towards implanted gel plugs, without recording systemic immunogenicity. These findings emphasize the prospect of SCs as a therapeutic platform to activate vital physiological processes, by autonomously producing biological drugs inside the body.

1.4: Membrane machines to interact with living matter

Cesar Rodriguez-Emmenegger1,2,3

1: DWI Leibniz, Germany;  2: Institute for Bioengineering of Catalonia (IBEC), Spain;  3: Institució Catalana de Reserca I Estudis Avançats (ICREA), Spain

Nature achieves unmatched functionality by the self-assembly of (macro)molecular building blocks in a hierarchical manner. All information necessary for the function is encoded at the molecular level. This talk will focus on developing Membrane Machines, tailor-made synthetic vesicles capable of recapitulating some fundamental biological properties and performing tasks beyond nature. To tackle this, we have designed and synthesized new families of amphiphiles —comb-polymers and Janus dendrimers— that self-assemble into cell-mimetic vesicles termed combisomes and dendrimersomes. Although these molecules do not exist in nature, the formed vesicles closely mimic cell membranes’ thickness, flexibility, and lateral 2D organization. The unparalleled matching of biophysical properties enabled the harboring of functional components of natural membranes and even fusion with living cells to “hijack” their periphery. This provides an almost inexhaustible palette to design the chemical and biological makeup of the synthetic cells. Finally, I will demonstrate the use of this synthetic platform to generate macrophage-mimetic phagocytic synthetic cells capable of specifically recognizing, engulfing and destroying pathogens. The phagocytic synthetic cells may serve as a new paradigm to fight antibiotic resistant bacteria and viral infections.

1.5: Synthetic Tumour Immune Microenvironments

Oskar Staufer

University of Oxford, United Kingdom

The microenvironment of tumors comprises multiple types of immune cells, rendering the tumour immune-microenvironment (TIME) exceedingly complex in structure and function. Although some deceptively simple signaling axis (e.g. PD 1, LAG3 etc) have been pinpointed, empiric investigations of this multipartite system have proven to be ineffective, currently impending improvement of immune-targeted cancer therapies. Systematically combining the mosaic of functional immune parts for bottom-up engineering of an artificial TIME (ART-TIME), that exhibits key characteristics of tumor-immune interactions, opens up new perspectives towards rational analysis of TIME and its influence on tumor initiation, progression and treatment.

I will present how immune cells, the defining elements of a TIME, can be recreated as synthetic cells by bottom-up assembly. The programmable synthetic cells are introduced into tumor organoids to function as lifelike leukocyte mimics inside in vitro tumors. By this, a molecularly defined immune environment is created. Multi-parametric screening is applied to assess organoid development as well as immunotherapy response as a function of the ART-TIME configuration. This links TIME architectures to cancer adaptation and therapy resistance. ART-TIME strives to de-convolute the dynamic complexity of immune microenvironments towards a rational dissection. Moreover, ART-TIME contributes concepts for the assembly of hybrid biomaterials and insights on tumour immunology using programmable man-made materials. This interdisciplinary approach opens up perspectives for synthetic cells capable of manipulating tissue patterns by creating hybrid materials at the vanishing boarders between the living and non-living world.

1.6: Synthetic magnetic cells (syMcells): exploring forces during hybrid collective cell migration

Jacqueline De Lora1,2, Lucie Motyckova1, Sadaf Pashapour1,2, Ilia Platzman1,2, Peer Fischer1, Joachim Spatz1,2,3

1: Max Planck Institute for Medical Research, Germany;  2: Institute of Molecular Systems Engineering (IMSE), Germany;  3: Heidelberg University, Max Planck School Matter to Life, Germany

The functioning of synthetic cells that are designed to execute predetermined biophysical tasks could be directly evaluated by interfacing with natural cells. This biophysical approach would validate built-in operations such as synthetic cell-natural cell adhesion interactions. It follows that bioinspired synthetic cells in hybrid cultures with natural cells can be used as mechanical sensors or they may even trigger unpredicted emergent behaviors. In this work, a hybrid cell migration system that emulates E-cadherin-mediated cellular interactions is built where the remote control of SyMcells is achieved with magnetic fields. SyMcells are formed using an emulsion to create a lipid-derived chassis that encapsulates magnetic cytoplasm comprising an oil-based ferrofluid. SyMcells are then functionalized with recombinant E-cadherin proteins by polyhistidine-Ni-NTA complexation. Finally, syMcells are cultured with natural cells that express E-cadherin-EGFP and within side-by-side confinement chambers. Upon releasing the hybrid cultures from their confinement, hybrid cell migration dynamics are observed by time-lapse fluorescence microscopy. Parameters such as cell migration velocity, E-cadherin clustering, and actin cytoskeleton organization are measured as a function of the presence/manipulation of syMcells. The morphologies of syMcells are actuated during the migration period by applying a switching homogeneous magnetic field with a custom-built coil integrated in our microscope. We use this system to investigate the manipulation of hybrid cell migration characteristics including to gauge forces within migrating monolayer hybrid cultures. Future work will examine control and manipulation of leader vs follower phenotypes.

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