UCL NKH Research Update – Oct 2024

Gene therapy exploring vector production and safety studies, and RNA treatment study launched!

Highlights:

• Significant progress has been made on understanding NKH in GLDC mice models, particularly with a detailed analysis of gene expression changes in the brain and in brain structure.
• There are now 5 AMT mouse models, and UCL are checking biomarkers, so they can be used in treatment studies.
• GLDC gene therapy in both the brain and liver continues, and are in the process of testing vector production and obtaining regulatory approvals.
• RNA treatment study launched

Overall strategy

The UCL research strategy continues to focus on two main themes: They aim (i) to better understand the
NKH disease process, and (ii) to use this information to develop new treatments and evaluate how
well they work.

Understanding NKH – experimental models of NKH

Analysis of experimental models of NKH continues to be a major activity, aiming to both understand the disease process (including identification of new targets for treatment) and to provide outcome measures to evaluate treatments, across both AMT and GLDC.

Analysis of NKH mouse models

GLDC

• UCL have continued with several projects analysing our two main mouse models which have impaired function of GLDC (the main gene affected in NKH).
• They have adapted their models to allow rescue or knockout of GLDC function at selected times and in selected tissues. They previously used this to rescue GLDC expression just in the liver (published in 2020) and have more recently again used this approach to rescue expression in selected cell types in the brain. These experiments allow ‘proof of principle’ for gene therapy approaches by telling us which tissues they should prioritise for gene replacement.
• UCL have made significant progress with their project which has involved detailed analysis of gene expression changes in the brain, in combination with detailed analysis of brain structure. Current work is focused on validating the gene expression changes that they have found and evaluating the significance of these findings in the disease.
• In their previous report they highlighted a study which has examined changes in biochemistry that happen in brain and liver in NKH, and which identified metabolic alterations that were previously unrecognised in children with NKH. The paper to report this data is in progress.

AMT

• AMT is the causative gene in about 1/5 of children with NKH. They have now generated 3 mouse models with differing changes in the AMT gene and imported a further 2 models that were generated elsewhere. They are establishing these mice at UCL and have begun to characterise them as models of AMT-related NKH. The mouse strains will be an important resource for testing new treatments and for understanding how and why changes in AMT lead to NKH.
• They now have access to mouse models which carry AMT gene changes that are found in children with NKH. UCL are now checking the effect on biomarkers such as glycine. They also have two models in which the AMT is completely disrupted. In one of these they have replaced the protein-coding part of the gene with the sequence for a fluorescent protein – enabling them to track AMT-knockout cells.

NKH cell and organoid models

• Work with the tissues and organoids derived from induced pluripotent stem (iPS) cells has continued in projects being carried out by two PhD students. The aims are to analyse effects of AMT and GLDC loss of function in various human cell types with the current focus on nervous system and liver cells.

Towards new treatments for NKH

UCL continue to work on several different projects with two main strategies which comprise, (i) gene therapy (one-time treatment) and (ii) other treatments which aim to lower glycine and/or improve folate metabolism.

Gene therapy

These projects aim to place a working copy of the GLDC or AMT gene into the body. The two main GLDC projects aim to target either the brain and liver (using an AAV vector), or primarily the liver (using a lentiviral vector). UCL showed that both these approaches effectively lower glycine.

The activity over recent months has involved planning the next steps in moving the work from mouse models towards clinical trial. This will need scale-up and production of a ‘clinical-grade’ vector (in a specialist facility), confirmation that this vector is active, and safety testing as required by regulatory bodies before a clinical trial can be carried out.

1. AAV gene therapy for GLDC

UCL have identified a production facility and developed a plan for pilot work to test feasibility of vector production – they have completed initial steps in our lab and the out-sourced work is starting this month. In parallel, they are preparing a major grant application for funding to support the next phase of the project (manufacture, safety etc). UCL have now completed a set of experiments to test whether AAV gene therapy could be used together with other glycine lowering approaches (e.g. benzoate).

2. Lentiviral gene therapy for GLDC (targeting the liver)

UCL have planned the next steps of the project to obtain regulatory approvals including production of clinical grade vectors, testing their effectiveness and safety, and planning for a clinical trial. Since the last report they have completed the deep molecular biology analysis of treated human liver organoids which has provided important information to support safety.  

3. AAV gene therapy for AMT

UCL have made good progress on the new project aiming to develop gene therapy for NKH caused by mutation of the AMT gene. They tested vectors in cells and production of vector for treatment of the AMT mouse model(s) is in progress.

Additional approaches for development of new therapies

UCL are testing several different approaches using either small molecules, nutrients or drugs which may prevent or correct specific features of NKH. This includes studies on using cinnamate and folate-related nutrients.

Cinnamate treatment to control glycine levels

UCL are working to finish a paper describing effects of benzoate and cinnamate – both of which can lower glycine in the NKH (GLDC) mouse model. Additional current work is in progress to repeat the key biochemical tests using alternative methodology to validate the findings

Small molecules for normalization of folate one-carbon metabolism

Testing of folate-related molecules is ongoing in the embryo model, building on their previous findings of protective effects of formate or methionine. This work initiated was in the GLDC mouse models and we hope to extend this to the AMT models soon.

RNA-based treatment for NKH?

UCL would particularly like to highlight a new project for which we obtained funding from the MRC with additional support from the Mikaere Foundation. This project aims to develop an RNA-based treatment, which may be beneficial – as it can be used alongside Gene Therapy and has the potential for repeated dosing.