Research Bert Van den Bogerd

Abstract

This study aims to advance the cornea-on-chip (CoC), a microfluidic device designed as a microscopy slide, featuring eight corneal constructs to mimic human corneal barriers and physiology. The objective is to validate this prototype via a comprehensive approach as an in vitro model with pre-clinical value of the human cornea, focusing on drug permeability and corneal toxicity testing. By collaborating with Ghent University's Centre for Microsystems Technology, we combine our respective expertise's in corneal tissue engineering and microfluidics with my background in GMP manufacturing to assess the CoC's performance. The CoC platform's efficacy as a drug absorption model is confirmed by assessing corneal permeation and small molecule permeability. Anticipating potential limitations in mimicking the physiology of the cornea in vivo, we will explore biocompatible materials to improve cell behavior and drug permeation, laying the foundation for the model's second and improved version. Furthermore, through multi-omics analysis, I will investigate the interactions between cells and biomaterials to gain valuable insights. Finally, as a proof of concept regarding the application of the model, the cornea-on-chip is used to identify strategies to bypass the corneal barrier via the use of penetration enhancers, to boost drug delivery effectiveness.

Researcher(s)

• Promoter: Koppen Carina
• Co-promoter: Van den Bogerd Bert
• Fellow: De Vos Evelyne

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

Refractive errors are a collective definition for problems with focusing light accurately on the retina due to an aberrant shape of the eye and/or cornea. This leads to visual impairment if left untreated. They are currently treated with either wearables such as contact lenses or spectacles that may pose several disadvantages for patients such as discomfort, inadequate correction or (for lenses) an increased risk for eye infection. That is why there is a increasing trend towards refractive surgery for permeant vision correction, which range from laser ablative therapies to intraocular lens implantation. However, refractive surgeries are currently invasive, irreversible and limited to a certain degree of correction. We have developed a new treatment modality that can permanently correct refractive errors in a way that is personalized, non-invasive and reversible. In a previous IOF POC DEVELOP project, we confirmed our hypothesis on an in vitro level and initiated in vivo experiments, while this IOF POC LAUNCH project serves to deliver an in vivo safety and functional proof-of-concept rat animal model with improved chemical formulation. Furthermore, a financial and regulatory strategic plan will be established to increase the commercial readiness level.

Researcher(s)

• Promoter: Van den Bogerd Bert
• Co-promoter: Koppen Carina

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

This innovation mandate aims to validate essential material properties of an innovative medical device that are crucial during surgery, both in the short and long term after the procedure. In this project, the focus is on basic research to further support the preliminary proof-of-concept. We have planned various qualification and validation experiments that the product must undergo, related to three phases: the procedure itself, short-term post-operation (the first four weeks), and long-term (months to years). Furthermore, market research is also conducted on two fronts. A roadmap is outlined for the preclinical development of the technology from the establishment of the spin-off, and a go-to-market strategy is being formulated. Using a SMART analysis, five parameters crucial to the functioning of the permanent contact lens were identified: shelf life, sterilization, laser impact, reversibility, and stability. This analysis forms the basis of the work packages of the basic research discussed in the project description.

Researcher(s)

• Promoter: Koppen Carina
• Fellow: Van den Bogerd Bert

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

The corneal endothelium is the innermost layer of the human cornea, the eye's transparent window. A dysfunctional endothelium leads to corneal opacification, which is a common cause of corneal blindness worldwide and results in an unavoidable need for a transplantation. Unfortunately, the global donor shortage causes very long waiting lists. A pharmacological compound to stimulate in vivo corneal endothelial regeneration therefore is a very interesting alternative treatment to reduce the reliance on donor corneas. However, there are many limitations regarding the traditional drug discovery pipelines such as high cost and long development times. Moreover, the corneal anatomy hampers drug permeation. The aim of my PhD therefore is to tackle both these limitations so to deliver an innovative pharmacological treatment. Hence, this project proposes a high throughput biological screening of repurposed drugs, i.e. the screening of formerly approved compounds. This biological data will serve to develop and train a virtual compound screening model to predict additional potential lead compounds. The high throughput screenings together with a thorough characterization and optimalization of the physico-chemical properties of the main hits, will lead to the identification of one repurposed lead compound. Eventually, I will assemble this compound together with a corneal permeability enhancer into an inventive eye drop that facilitates corneal penetration to reach the endothelium.

Researcher(s)

• Promoter: Koppen Carina
• Co-promoter: Deben Christophe
• Co-promoter: Van den Bogerd Bert
• Fellow: Witters Charissa

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

The corneal endothelium lies on the interior of the cornea, which is the window of our body. Proper function of this corneal endothelium is essential to obtain a crystal-clear cornea. Current consensus holds that endothelial cells do not display any significant regenerative capacity. Damaged or non-functional cells may consequently lead to corneal blindness. In this regard, stimulation of the regenerative capacity of the corneal endothelium is of exceptional importance in the search for new therapeutic possibilities. Furthermore, for in vitro biomedical research the culture of primary corneal endothelium is an extremely time-intensive process without full guarantee of cell expansion. In this project we intend to provide a solution for these issues by pharmacological stimulation of the regenerative capacity of corneal endothelial cells through re-orientation of available drugs. During this project various molecules will be subject to a tri-fold screening method (cellular, subcellular and molecular), which leads to the selection of a regenerative compound for corneal endothelial purposes. On the one hand, specific ROCK-inhibitors will be tested, given their growth potential within tissue regeneration. On the other hand, commercially available drug libraries will be screened for compounds with known activity for regenerative capacities.

Researcher(s)

• Promoter: Koppen Carina
• Co-promoter: Deben Christophe
• Co-promoter: Van den Bogerd Bert

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

The cornea is the transparent window to the eye and as such, eye drops are an interesting route of drug administration. In this project, we propose the development of a 3D corneal cell model that can be used to investigate corneal drug interaction. Studies that aim to simulate the pharmacokinetics and toxicological properties of drugs are mainly based on oversimplified 2D monocultures or animal studies that suffer from interspecies differences. These limitations skew proper predictive power during preclinical drug investigation. We propose the development of a 3D corneal model that includes all three cell layers and integrated microfluidics to simulate relevant physiological flows such as the tear film. In this way, we reduce the need for animal studies, while simultaneously introducing the in vivo complexity of 3D cell environments. The materials used will be fabricated (photocrosslinkable biopolymers) and printed in-chip using 2PP bioprinting that can simultaneously integrate corneal cells. The interfaces between materials and corneal cells are characterized in depth using molecular spectroscopy techniques, while detailed protein expression of cells in the 3D ECM are benchmarked to ex vivo cadaveric donor corneas with proteomics. TEER, flow rate and cell viability will be measured real-time by integrated sensors. The project finally aims for a proof-of-concept to determine permeability coefficients of common ocular drugs in the fully assembled 3D cell culture chip.

Researcher(s)

• Promoter: Koppen Carina
• Co-promoter: Van den Bogerd Bert

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

The cornea is a barrier that protects the eye from the outside world and likewise hampers drug absorption. Most ophthalmic medicine is washed away during instillation and is absorbed into systemic circulation. This is especially relevant when considering the main target population, the elderly, which also have an increased susceptibility to adverse drug reactions due to polypharmacy and decreased renal function. Hence, new topical formulations require rigorous testing to ensure therapeutic efficacy, while keeping local and systemic toxicity to a minimum. However, promising preclinical results are often not corroborated during human testing because available corneal models have poor predictive power. Traditional 2D culture fails to recapitulate complex tissues such as the cornea while animal models exhibit interspecies differences that limit translatability of results to humans. Organs-on-chips, which are rationally-designed microfluidic chips that contain artificial tissue, hold the promise of improving the status quo. While organ-on-chip technology has already proven its merits in certain fields, in the context of the cornea it remains relatively unexplored. This project proposal outlines the development of the first cornea-on-chip that comprises every cellular layer – epithelium, stroma and endothelium – of the cornea, exposed to the dynamics of an artificial tear film on one side and connected to an artificial anterior chamber on the other.

Researcher(s)

• Promoter: Koppen Carina
• Co-promoter: Van den Bogerd Bert
• Fellow: Van Meenen Joris

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

Keratoconus is a progressive disease of the cornea that is characterized by corneal thinning and outward bulging in the shape of a cone. This leads to visual impairment if left untreated. A range ofsoft to hard contact lenses are commercially available for visual corrections, but they become inadequate or uncomfortable after prolonged wear or in case of disease progression. The second-line treatment consist of corneal crosslinking or transplantation, but these often still require corrective lenses afterwards. We have developed a new treatment modality for keratoconus patients that can permanently correct their vision and is non-invasive and reversible. In a previous IOF POC CREATE project, we confirmed our hypothesis on an in vitro level, while this IOF POC DEVELOP project serves to deliver an in vivo safety and functional proof-of-concept in a mouse and rabbit animal model respectively. Furthermore, a thorough business plan will be further drafted with the emphasis on a regulatory roadmap and a quantitative market analysis.

Researcher(s)

• Promoter: Van den Bogerd Bert
• Co-promoter: Ni Dhubhghaill Sorcha

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

The corneal endothelium is the most inner layer of the human cornea, the window of the human eye. This cell layer is of utmost importance, since it functions to maintain corneal transparency. Upon damage to the endothelium, corneal oedema ensues and subsequently, visual impairment. The only solution for these patients is to transplant a new, functional corneal endothelium sourced from a donor eye. However, on a worldwide scale there are not enough donor corneas to meet for the patients in need. Not only does this shortage result in long waiting lists, but this donor scarcity also hinders corneal endothelial research. The limited tissue supply make research time-consuming, but also the corneal endothelial cells are very difficult to bring in culture. That is why until this very day the pathophysiology of corneal endothelial diseases remains to be elucidated. In this project I propose the reversible immortalization of corneal endothelial cells to overcome both the worldwide donor shortage and the intricate ex vivo growth. The process of reversible immortalization consists of four different steps: 1) insertion of removable genes to stimulate proliferation, (2) positive selection of transfected cells, (3) gene excision and (4) an additional negative selection to eliminate any residual engineered cells. In this small research project, I will investigate to establish the first three steps using either a lentiviral vector or a transposon system. The final aim is to effectively use the transposon system, while lentiviruses are part of this application as a positive control, since protocols and results are found throughout the literature, but are scarce for PiggyBac. However, the PiggyBac displays the intrinsic advantage such as a finger-print free excision of a transgene. Upon success, we can easily create vast amounts of corneal endothelial cells in our laboratory. These cells can will feed into our established projects of developing a corneal endothelial cell therapy or the development of a so-called cornea-on-a-chip.

Researcher(s)

• Promoter: Van den Bogerd Bert

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project

Abstract

Human corneal endothelial cells (HCEnC) regulate fluid and solute transport across the posterior surface of the human cornea and actively maintain the cornea in a dehydrated state, which is crucial for optical transparency.The dual function of the corneal endothelium is described as the "pump-leak hypothesis" which is essential to allow nutrition to the cornea whilst maintaining its avascularity and transparency. There is no evidence that human endothelial cells divide under normal circumstances as they are arrested in G1 phase of the cell cycle, although they can be induced to divide in vitro. When the amount of corneal endothelial cells decreases below a certain threshold, this cell layer can no longer pump sufficient fluid back to the anterior chamber, resulting in an irreversibly swollen, cloudy cornea. Despite its success, corneal transplantation (either full-thickness or partial) is limited worldwide by the shortage of suitable donor corneas incurring long waiting times. Initial progress to overcome this global shortage is the use of one donor cornea for multiple partial keratoplasties ("split-cornea transplantations"), by using one donor cornea for a partial endothelial and a stromal transplantation. This project aims to investigate ex vivo expansion of corneal endothelial cells to develop a cell sheet based therapy. This would overcome donor deficit that limits the treatment of corneal endotheliopathies. The principle is to expand primary human corneal endothelial cells isolated from human cadavers and to seed them on an ideal scaffolding material to introduce these cells in the patient. Specifically in this project we propose the expansion of human corneal endothelial cells (HCEnC) on human lens capsules to obtain a composite graft. The final goal of this project is a proof-of-principle of this functional cell sheet in a rabbit corneal endotheliopathy model.

Researcher(s)

• Promoter: Koppen Carina
• Promoter: Tassignon Marie-Jose
• Co-promoter: Zakaria Nadia
• Fellow: Van den Bogerd Bert

Research team(s)

• Translational Neurosciences (TNW)

Project type(s)

• Research Project