MICA: Proof of Concept for Real Architecture for 3D Tissues in the Cornea

The cornea on the front surface of the eye is our window to the world. If transparency is compromised, visual impairment and even blindness can occur. There are 10 million people worldwide who are blinded by scarring of the cornea and are denied a sight saving corneal graft due to insufficient donor tissue availability. The ability to 'grow' components of a cornea in the laboratory would represent a very significant scientific and medical advance that could improve quality of life. As a first step towards this goal, we are working to address a condition called limbal epithelial stem cell (LESC) deficiency of the cornea, which causes painful, blinding corneal surface failure. An estimated 240 new cases of LESC deficiency occur annually in the UK. We have cultured and transplanted sheets of LESCs to improve vision in patients with chemical burns. However, many patients have very difficult to manage diseases requiring treatment with a substrate to aid cell survival, in our case human amnion. This approach can significantly improve quality of life and productivity in the workplace (patient personal communications) however, we have found the long-term therapeutic benefit to be variable. In part, we suspect, because cultured LESC sheets do not restore the normal LESC microenvironment or 'niche' destroyed by disease. Using a novel tissue engineering approach we have recently made simple constructs of the human corneal surface using protein, epithelial cells (including LESCs) and the fibroblasts that support them. Novel technology (Real Architecture for 3D Tissues - RAFT) is used to make a fibroblast-seeded collagen construct with surface toplology that mimics the in vivo stem cell niche, then corneal epithelial cells are seeded on the surface. Our data are very promising, the epithelial cells grow successfully and produce an epithelium. RAFT constructs are reproducible and have significant advantages over human amnion for LESC culture. Amnion is biologially variable and often (40% of cases) does not support the growth of healthy LESCs. We now wish to a) asses the physical and functional properties of RAFT and b) perform safety and efficacy studies so that we may proceed to first in man studies at Moorfields Eye Hospital (MEH) following this project. We already know that RAFT is strong enough to be handled and that text can be read through it. Here we will test actual mechanical strength as this must be sufficient to withstand surgery. Transparency will also be measured as RAFT should ideally be at least as transparent as the amnion we aim to replace if not better. The key driver behind this proposal is the need to establish if RAFT is safe to use and if it can improve the surface of a cornea with LESC deficiency. Without this information we cannot test RAFT in humans. Here we will work with colleagues to assess RAFT safety and efficacy in an established model of LESC deficiency. Our techniques and methods for producing RAFT constructs, presenting them to the surgeon in an accessible device and measuring clinical outcome in our model will be rigorously validated. If during this pre-clinical testing RAFT constructs can restore corneal transparency and maintain a healthy ocular surface, our data will be used to develop regulatory compliant standard operating procedures for the production of RAFT for future testing in man. This project is therefore essential to progress our research findings into clinical practice. If the project goes as planned, IoO have the experience and capacity to manufacture RAFT constructs in the 'Cells for Sight Cell Therapy Research Unit'. This new state of the art cleanroom facility, led by the applicant, is used to produce cell therapies for patients. If the project is successful, IoO and MEH will design a phase I/II safety and efficacy clinical trial to compare RAFT with LESC cultured on amnion.

10 million people worldwide are blinded by scarring of the cornea. The ability to engineer biomimetic corneal components could improve quality of life. As a first step, we are working to treat limbal epithelial stem cell (LESC) deficiency of the cornea, which causes painful, blinding surface failure. We have transplanted cultured LESCs to improve vision in patients with chemical burns. However, many patients have challenging diseases requiring a substrate to aid cell survival, in our case human amnion. This approach can improve vision, however, we have found the long-term therapeutic benefit to be variable. In part, we suspect, because cultured LESC sheets do not restore the normal LESC 'niche' destroyed by disease. We have made simple constructs of the human corneal surface. Novel technology (Real Architecture for 3D Tissues - RAFT) is used to make a fibroblast-seeded collagen I construct with surface topology that mimics the in vivo stem cell niche, with an epithelium on the surface. Our data are very promising, the epithelial cells grow successfully and produce an epithelium. RAFT constructs are reproducible and have significant advantages over human amnion for LESC culture. Amnion is biologially variable and often (40% of cases) does not support the growth of healthy LESCs. Here we will quantify RAFT mechanical strength, using a culture force monitor, as this must be sufficient to withstand surgery. Spectral transmission (transparency) will be measured as RAFT should ideally be at least as transparent amnion. We will test if RAFT is safe to use and if it can improve the surface of a cornea with LESC deficiency with our Japanese collaborators using an established rabbit model of LESC deficiency. If successful this data will inform GMP compliant standard operating procedures for therapeutic RAFT production. IoO have the experience and capacity to manufacture GMP compliant RAFT constructs in the 'Cells for Sight Cell Therapy Research Unit'.

Academics, clinicians, commercial private sector, policy makers, patients, charities will benefit from this research.
Academics interested in the research areas of regenerative medicine, corneal biology, stem cell regulation, cell-cell interactions, tissue engineering and reducing the use of animals in experiments will benefit from having a simple, easy to make and reliable model of human tissue to adapt for their own research.
Clinical academics interested in enhancing healthspan, alleviating the effects of ocular surface failure and corneal disease will have a model in which to test new concepts and therapies.
The concept of Advanced Therapy Medicinal Products (ATMP), of which RAFT is one, is still relatively new. This project will give the MHRA an opportunity to examine a novel ATMP from an academic / SME partnership that may first be approved along the hospital 'Specials' route before becoming an Investigational Medicinal Product (IMP) in a clinical trial. The path that we cut together may influence the advise the MHRA gives to other researchers proposing to take this regulatory route.
Any IP and knowhow generated could be licensed by the commercial private sector to generate future economic return for the University and the companies involved. We already have an excellent track record of collaboration with TAP Biosystems (formerly The Automation Partnership) that we aim to continue here.
Availability of reliable models of human tissue and a demonstration of their therapeutic potential could inform policy makers with regards to the direction of future research investment.
Patients could benefit from this research if it leads to the demonstration that RAFT is a suitable technology for the transfer and survival of stem cells on the ocular surface. Such data would support the need for a clinical trial with the prospect of future product development. If this project is successful, it will provide early safety data for the use of RAFT in the human eye. This will increase our drive to develop additional ocular therapies, using RAFT, to treat diseases with much larger indications. These could include replacement of scarred corneal tissue, delivery of corneal endothelial cells, delivery of cells for retinal degeneration. Success of such approaches would improve quality of life, and make a positive contribution to the economy by alleviating the financial burden on the NHS and by generating new jobs. Such advances would strengthen the UK's position in terms of international research leadership that would in turn attract new economic investment.
Charities may also benefit by having a tangible cause against which to raise funds.
The researchers working on the project will benefit from engaging with scientists from different backgrounds. This will seed new ideas, forge further new collaborations and secure future research funding.