Gene therapy explained.
Introduction.
The worldwide Rett community is excited as two biotechnology companies have recently either started- or announced plans to start- gene therapy clinical trials. However, gene therapy has different meanings including gene editing (by editing DNA or RNA by CRISPR technology), gene addition (by delivery of genes that encode, for example, antibodies to specific pathogens e.g. influenza), gene silencing (used to reverse the effect of mutant genes which contain toxic elements) and finally, gene replacement (as applied to Rett Syndrome).
As a reminder, a gene is a stretch of DNA with a specific nucleotide sequence that contains (encodes) the necessary components to result in the expression of a functional protein.
Gene replacement to treat Rett Syndrome.
It has been known for some time that Rett Syndrome results from mutations in a single gene namely, MeCP2, and that replacement of the defective gene in Rett model mice can alleviate many of the symptoms and characteristics of Rett Syndrome in these mice. This led to the realisation that similar gene replacement in Rett patients might also alleviate symptoms or potentially result in a cure. Thus, in the context of Rett Syndrome, gene replacement aims to replace a mutant (defective) MeCP2 gene with a functional MeCP2 gene, resulting in expression of a functional MeCP2 protein in neurons with the potential to correct any underlying deficit.
Consequently, the strategy is clear, to deliver a functional MeCP2 gene to neurons, the most common cell type in the brain, but several obstacles need to be overcome. Although the gene can easily be generated in the laboratory, naked DNA is not readily taken up by cells in the body so that injection of the MeCP2 gene in the form of purified naked DNA is unsuccessful. To overcome this, the MeCP2 gene (DNA) is contained within (encapsidated by) a small virus called Adeno-associated virus (AAV) generating a recombinant AAV (rAAV-MeCP2). AAV naturally infects humans without causing any disease and is considered the delivery vehicle (vector) of choice. However, because AAV is a very small virus and the MeCP2 gene is large, it was necessary to reduce the length of the gene to ensure that it was packaged into the rAAV. Taysha and Neurogene used different strategies to achieve this; Taysha used a MeCP2 minigene while Neurogene used a shortened version of a regulatory element of the gene.
Delivery of the rAAV-MeCP2.
Preliminary studies with rAAV-MeCP2 in animal models used systemic delivery via intravenous injection to deliver the gene to the brain, but this resulted in uptake of the product by the liver and in toxicity. In addition, the blood brain barrier (BBB), a physical barrier between circulating blood and the brain composed of specialised cells, prevents the entry of rAAV from the blood to the brain. To overcome this, the rAAV-MeCP2 is delivered in patients by direct injection into the central nervous system (CNS) either by intrathecal delivery (into the spinal column) or by intracerebroventricular (ICV) delivery directly into the brain, performed after sedation or under general anaesthetic. These delivery methods appear to be distressing but are considered routine by the clinical practitioners and are necessary to ensure reliable delivery of the rAAV-MeCP2 to neurons distributed throughout the brain. Because many humans have previously been infected with AAV, direct injection of the rAAV-MeCP2 will also overcome any existing anti-AAV immunity and is usually accompanied by some immunosuppression to ensure that the patient has a minimal immune response to the rAAV-MeCP2 that might reduce the efficiency of the procedure.
After the MeCP2 gene is delivered to neurons by rAAV-MeCP2 it appears to be quite stable for many years (meaning, at least in theory, that a single dose is sufficient), with the capacity to direct the expression of a functional MeCP2 protein. To ensure that the dose of MeCP2 protein is correct to avoid MeCP2 duplication syndrome, both Taysha and Neurogene included unique molecular control elements in the rAAV-MeCP2, a strategy that I have previously discussed (https://rettaustralia.org.au/blog/a-game-changer-for-gene-therapy-for-rett-syndrome/).
The outcome of rAAV-MeCP2 gene replacement.
Although the preclinical experiments in mice provided hope that expression of a functional MeCP2 protein after gene replacement might reduce or alleviate the symptoms of Rett Syndrome in patients, we cannot be certain of this. This is why clinical trials are necessary, and details of the Rett clinical trials, including entry and exclusion criteria, are available on the US NIH website, https://clinicaltrials.gov.
As yet, two patients have received rAAV-MeCP2 therapy; preliminary data from the first patient 6 weeks after therapy were reported in a press release available on the internet (https://ir.tayshagtx.com/news-releases/news-release-details/taysha-gene-therapies-reports-initial-clinical-data-first-adult). A recent press release from Taysha reported on the results from these two patients, 12 weeks and 4 weeks after therapy. As noted by the principal investigator; “The two adult patients dosed with TSHA-102 have different mutations in their MECP2 gene that manifest in different phenotypes and clinical severity. Following treatment, both patients experienced improvement in key clinical domains impacting activities of daily living, including breathing dysrhythmia, autonomic function, socialization, and gross and fine motor skills. Both patients display significantly reduced breathing dysrhythmia, with less breath holding spells and infrequent hyperventilation, improved limb perfusion and vastly improved interest in social communication and activities. In addition, the first patient experienced sustained and new improvements, with restored movement in her legs and the gained ability to sit unassisted for up to 15 minutes for the first time in over a decade. Further, her hand function improved with the gained ability to grasp objects with her non-dominant hand and transfer them to her dominant hand for the first time since infancy. Following treatment, the second patient’s posture, gait and stability improved, resulting in straighter posture and smoother movements when walking. Her hand stereotypies also improved for the first time since regression at age three: she now displays less forceful hand wringing and her hands are often open and relaxed, providing new opportunities for fine motor skill learning. In addition, her seizures are much less frequent. I’m encouraged by the early positive signals and consistent improvement seen in both patients following treatment.” The full press release of November 14th is available on the internet;
https://ir.tayshagtx.com/news-releases/news-release-details/taysha-gene-therapies-reports-third-quarter-2023-financial
These data are incredibly exciting, not only showing that the patients showed no adverse events, but also showed clinical improvement over a broad range of measures. This leads us to consider that gene replacement may well have a future in the treatment of Rett Syndrome.
Is gene therapy safe?
Nevertheless, it is too soon to determine if gene replacement therapy can alter the quality of life for Rett patients and many questions will be addressed by the planned clinical trials. However, many parents will seek reassurance of the safety of the procedure. A review published in 2019 noted that there were 145 international clinical trials using AAV vectors listed on the NIH website, including late-stage trials for other monogenic diseases. These include lipoprotein lipase (LPL) deficiency and retinal dystrophy, treated by direct injection into the muscle and the eye respectively with Glybera and Luxturna, the trade names for licensed gene replacement products. However, gene replacement therapy for spinal muscular atrophy (SMA) is most similar to the proposed therapy for Rett Syndrome as the SMA gene product (Zolgensma) is delivered directly into the CNS of young children to target neurons. Most importantly, the therapy was shown to be effective. However, despite all precautions and the collective AAV expertise, two children in Russia and Kazakhstan died from acute liver failure after Zolgensma therapy showing that risks cannot be totally eliminated. However, more than 2300 patients have been successfully treated and it was suggested that these two patients may not have met the strict criteria for treatment.
In summary, clinical trials are designed primarily to examine the safety of a new therapy but are generally also designed to assess efficacy. Despite the encouraging results noted above, it is too soon to know if gene replacement therapy for Rett Syndrome will result in any side effects or alleviate symptoms in individual treated patients, but we remain optimistic that progress in the next five years will be similarly staggering to that in the past five years.
Caveat.
This article is intended to present a brief general description of gene replacement therapy and is not intended to provide details that might apply to specific individuals. It is also intended to provide broad background information only as it is impossible to present details of complex clinical trials and strategies in this brief article.
Eric Gowans
November, 2023
