Development of rAAV-mediated gene therapy for a severe paediatric metabolic liver disease: Ornithine Transcarbamylase deficiency.

Lead Research Organisation: University College London
Department Name: Institute of Child Health


The urea cycle is an essential pathway in the liver which is necessary for the detoxification of ammonia, produced by the breakdown of proteins. Ammonia is highly neurotoxic, and defects in any of the urea cycle enzymes leads to high rates of mortality and morbidity. Ornithine transcarbamylase (OTC) deficiency is the most common of the urea cycle defects. In the most severe forms of the disease, symptoms present early in the newborn period progressing to coma and death if untreated. Patients are managed by pharmacological and dietary intervention but ultimately require a liver transplant for long-term survival. However, liver transplantation is associated with its own risks - mortality, morbidity and life-long need for immunosuppression.
Gene therapy offers an attractive alternative to liver transplantation, as the patient's own cells are repaired by transfer of a functional gene. Adeno-associated viruses (AAV) are emerging as a highly effective gene delivery system, and the ability to alter the outer coat (capsid) of the virus allows a variety of cells to be targeted. AAV8 is currently yielding promising results in the liver in a clinical trial for Haemophilia B (Factor IX deficiency), led by UCL. While exciting, this success is made possible by the low number of liver cells that need to be genetically repaired in this condition, as the Factor IX protein is released (secreted) into the bloodstream with as little as 1-2% of normal levels providing therapeutic benefit. In many metabolic conditions involving the liver, such as OTC deficiency, where the gene product is not secreted, a much higher proportion of cells must be modified for clinical success. Prof. Alexander has successfully cured OTC-deficient mice using an AAV8-based gene therapy approach, however, AAV8 is far less effective for gene delivery to human liver cells. An exciting collaboration with colleagues at Stanford University has led to the development of a novel AAV, referred to as LK03, which targets human liver cells with a much higher efficiency that AAV8 (12 times). We therefore propose a first-in-man use of an AAV gene delivery system based on AAV/LK03 for the treatment of OTC deficiency in paediatric patients.
This application describes pivotal preclinical studies required to support authorisation of a phase I/II UK-based AAV/LK03-mediated clinical trial for OTC deficiency, and builds on already funded studies seeking to refine AAV/LK03 for clinical trial use. The proposed preclinical studies include immunological investigation of the target paediatric population, quality control tests (potency, safety and purity) on clinical-grade reagent, and toxicology and biodistribution studies in non-human primates. Non-human primates are the chosen animal model for these studies as the gene therapy delivery system is highly specific for human liver cells, with very little ability to function in mouse liver cells.
Initially, a survey of the immunological status (seroprevalence) will be carried out on the target paediatric population. The presence of antibodies (humoral immunity) to AAV/LK03 can impede the effectiveness of the gene delivery. Given that previous surveys have shown that humoral immunity to AAV does not generally develop before the age of two, and the majority of trial participants will be younger than this, this is unlikely to be a significant problem.
The gene therapy reagent will be produced in the UCL Vector Core Facility and subjected to standard safety, purity and sterility testing. The reagent will then be tested in Cynologmus macaques for toxicology and biodistribution. At the conclusion of the pre-clinical testing, the results will be presented to the Medicines and Healthcare Products Regulatory Agency (MHRA) in the form of an Investigational Medicinal Product Dossier (IMPD) to request Clinical Trial Authorisation. These results will also support an Orphan Designation application of the gene therapy reagent at the European Medicine Agency.

Technical Summary

The urea cycle is an essential hepatic pathway involved in detoxification of ammonia. Urea Cycle Defects (UCDs) share high rates of mortality and neurodisability. Ornithine Transcarbamylase Deficiency (OTCD) is the commonest UCD (60% of all UCDs). As X-linked inherited disorders, males are severely affected with presentation in the newborn period resulting in coma and death if untreated. Milder cases can present with acute hyperammonaemia at any age particularly in females. Despite best-accepted treatment with ammonia scavengers and protein-restricted diet, acute hyperammonaemic decompensations can occur. Liver transplantation cures UCDs with a risk of mortality and morbidity, including lifelong immunosuppression. The first successful clinical trial of liver-directed GT was led by UCL for Haemophilia B, using AAV2/8. Prof Alexander has shown correction of OTC-deficient mice with an AAV-based GT approach and collaborated in the development of a novel capsid, AAV-LK03, which targets human hepatocytes 12-fold better than AAV2/8.
We propose to set up the first paediatric liver-directed GT clinical trial using AAV-LK03 for OTCD. An NHMRC grant (2015-2016) has been awarded to Professor's Alexander & Gissen to optimise vector configuration, and determine efficacy, minimal effective dosing and investigate insertional mutagenesis in a xenograft mouse model engrafted with OTC-deficient human hepatocytes.
In this application, we request funding for preclinical studies to determine AAV-LK03 vector pre-existing immunity, biodistribution and formal toxicology. The data will allow preparation of an Investigational Medicinal Product Dossier to support Clinical Trial Authorisation from the Medicines and Healthcare Products Regulatory Agency. This data, along with the proof-of-concept studies by Prof Alexander, will allow an Orphan Designation application and apply for a Protocol Assistance to ensure a complete package for Marketing Authorisation to the European Medicine Agency.

Planned Impact

Who will benefit from this research?
In addition to the specific academic beneficiaries that have been listed in the academic beneficiaries section above, the pharmaceutical industry, patients with rare and common liver diseases, the general public and the wider academic and clinical community will benefit from development of novel vector which is highly hepatotropic for human cells. Benefits will occur as follows :
1. Benefits for Industry: The research outlined in this proposal is likely to be important to the pharmaceutical industry in two specific ways:
(i) There is currently renewed interest by the pharmaceutical industry, in development of gene therapy vectors, because of the current vectors still not able to provide high enough transduction of human hepatocytes. The examples of such interest is the substantial interest from companies such as Dimension Therapeutics and Uniqure, which is reflected in their letters of support. Further PI and Co-Is had numerous meetings with scientists from other companies who are also keen to know the results from our investigation. As new findings emerge about the properties of the novel vector they will be shared with scientists at different companies and more widely as other interest develops.

(ii) Identifying and characterizing the novel hepatotropic vector in this project will not only provide therapy for OTC deficiency therapy but also for any other liver based single gene disorder. Furthermore AAV-LK03 may be able to transduce non-liver cells and may be used to treat other human diseases. Moreover application of gene transfer to liver cells has been proposed in many common liver diseases and could be useful in diseases such as hepatitis, liver cirrhosis and liver cancer.
We have already engaged UCL Business in protecting potential intellectual property arising from this research.

2. Benefits for the General Public: The general public will benefit from this research, because it will help our understanding of the use of novel potentially curative approach to treatment of liver based metabolic disorders. The findings of biodistribution and toxicology experiments will have an educational impact. We will engage the UCL Public Engagement Unit as well as our Institute website to provide educational information about liver directed gene therapy as a way to get our new findings across to the general public including patients with liver diseases.

3. Benefits for the wider academic and clinical community: In the long term health care professionals, both academic and clinical as well as patients will benefit from the work undertaken in this study especially if new knowledge about the biodistribution and toxicity of AAV-LK03 vector can lead to better drugs that can treat patients with liver diseases. In the long term there may be impact on the well-being and quality of life of patients as well as economic benefits by saving health care costs also initiating activities in the UK-based pharmaceutical industry.

4. Benefits for the regulators as gaining experience with the first paediatric AAV-mediated liver-directed human trial.


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Cozmescu AC (2021) Gene therapies targeting the liver. in Journal of hepatology

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Baruteau J (2017) Expanding the phenotype in argininosuccinic aciduria: need for new therapies. in Journal of inherited metabolic disease

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Baruteau J (2019) Argininosuccinic aciduria: Recent pathophysiological insights and therapeutic prospects. in Journal of inherited metabolic disease

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Baruteau J (2019) Fetal gene therapy for neurodegenerative lysosomal storage diseases. in Journal of inherited metabolic disease

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Forny P (2019) Liver neoplasms in methylmalonic aciduria: An emerging complication. in Journal of inherited metabolic disease

Title Innovations in gene therapy for the treatment of OTC deficiency 
Description In collaboration with SciAni we created an animation that describes this project. It is posted on YouTube and targets general population. 
Type Of Art Film/Video/Animation 
Year Produced 2019 
Impact The movie has been seen by >1000 viewers. 
Description BRC Catalyst Fellowship for Dr Julien Baruteau
Amount £80,000 (GBP)
Organisation Great Ormond Street Hospital (GOSH) 
Department NIHR Great Ormond Street Biomedical Research Centre
Sector Academic/University
Country United Kingdom
Start 04/2019 
End 04/2020
Description Children's Hospital of Philadelphia 
Organisation Children's Hospital of Philadelphia
Country United States 
Sector Hospitals 
PI Contribution This is a collaboration on gene therapy in OTC Deficiency. We provide the clinical trial expertise to this collaboration.
Collaborator Contribution The partners provide expertise in gene therapy vector generation.
Impact Partner contributes to the project on gene therapy in urea cycle disorders
Start Year 2017
Description Sydney University, Australia 
Organisation University of Sydney
Department Central Clinical School
Country Australia 
Sector Hospitals 
PI Contribution We are collaborating with Professor Alexander on the liver directed gene therapy project. We have a joint grant from the MRC DPFS where I am the PI and Prof Alexander is the co-PI. My group's contribution is the expertise in running clinical trial in rare paediatric diseases.
Collaborator Contribution Prof Alexander's group have the expertise in adeno associated vectors and specifically vector design for gene therapy in urea cycle disorders.
Impact 2 grants: NHMRC grant to Prof Alexander where I am a co-PI and the MRC DPFS grant currently.
Start Year 2014
Title Preclinical development of a gene therapy product for OTC deficiency 
Description Gene therapy product to be tested in non human primates 
Type Therapeutic Intervention - Cellular and gene therapies
Current Stage Of Development Initial development
Year Development Stage Completed 2017
Development Status Under active development/distribution
Impact No impact yet 
Title Received funding for a Phase I/II clinical trial evaluating AAV-mediated gene therapy for a severe paediatric metabolic liver disease: Ornithine Transcarbamylase deficiency 
Description We received funding from the MRC to convene a Phase I/II clinical trial evaluating AAV-mediated gene therapy for a severe paediatric metabolic liver disease: Ornithine Transcarbamylase deficiency. This next stage will start in 2019. 
Type Therapeutic Intervention - Cellular and gene therapies
Current Stage Of Development Refinement. Clinical
Year Development Stage Completed 2019
Development Status Under active development/distribution
Impact The development of this product will lead to the first in child trial of a novel AAV capsid specific for liver. 
Description CLIMB conference 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Patients, carers and/or patient groups
Results and Impact Dr Baruteau made a presentation at the 2017 CLIMB conference
Year(s) Of Engagement Activity 2017
Description Presentation on liver directed gene therapy at the British Society of Gene and Cell Therapy 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Presentation at the British Society of Gene and Cell Therapy which took place in London but with international participants.
Year(s) Of Engagement Activity 2018