A Boston-based company reports exciting results on a new approach to treat genetic diseases.
Phenylketonuria (PKU) is a genetic metabolic disorder, where the patients cannot properly metabolize the amino acid phenylalanine (Phe). It is an inherited condition, caused by low levels of the enzyme phenylalanine hydroxylase (PAH) (or non-functional recycling pathway of the co-factor THB, required by phenylalanine hydroxylase).
PKU is a serious condition. If untreated, PKU symptoms include mental and behaviour disorders, seizures, light skin, and musty smell. Newborn screening can reveal very quickly the imbalance in the amino acid metabolism, and patients can lead a normal life if they have extremely low-phenylalanine diet. Maintain a lifelong prescription diet is challenging, associated with high costs and social limitations.
There is a very limited amount of non-dietary treatment options and no cure is available. One approved therapy is the continuous injection of pegylated phenylalanine ammonia lyase (PAL), an enzyme that degrades phenylalanine; however this treatment comes at a high cost and with severe anaphylaxis risk. KUVAN is a PAH activator—effective in patients with poor availability of THB—is not applicable to many patients and cannot replace low-phenylalanine diet. Gene therapy could solve the problem once and for all, replacing the patient’s non-functional enzyme at the DNA level. However, there is no such treatment approved yet.
Synlogic, a therapeutics company from Cambridge, MT, decided to take a different approach. Instead of attempting to modify the genome of the patient, the company decided to focus on the microbiome. Their idea is to modify Escherichia coli Nissle, a bacterium naturally occurring in the human microbiome, to break down phenylalanine before it damages the host.
The results of this effort were recently reported in Nature Biotechnology, while a clinical trial is underway. The researchers engineered E. coli with two Phe-catabolising pathways: in the first instance, they expressed PAL (which is a cytosolic enzyme and requires the co-expression of phenylalanine importer); in the second instance, they expressed L-amino acid deaminase (LAAD) that converts Phe to phenylpyruvate (it is a membrane localised enzyme, but requires oxygen). Both pathways can reduce Phe in the assays, but displayed reduced growth.
Creating the final strain required some clever engineering. The researchers wanted a strain that would activate the Phe-degrading pathways just before administration (with induction) and during its life in the gut, but not during production. They also wanted the LAAD enzyme to be expressed in microaerobic environment (where it has activity). Moreover, the strains would need to harbour no antibiotic resistance cassettes (to be used in humans) and have the transgenes incorporated in the genome (for stability). All these design specifications were achieved in the final E. coli strain SYNB1618. SYNB1618 has three and two copies of PAL and LAAD respectively, to account for potential loss-of-function caused by random mutations. Also, SYNB1618 requires exogenous diaminopimelate to form its cell wall, a feature added as a biocontainment device.
The final strain’s efficiency as a phenylalanine regulator was then experimentally tested in mice with PKU and in healthy monkeys. In both cases, the researchers detected degradation products caused by the heterologous enzymes secreted in the urine, without any side effect. Also, they report that unhealthy mice injected with the probiotic strain reduced their phenylalanile blood levels by 38%.
Things looks promising so far, so I am looking forward to the clinical results. SYNB1618 could act as a regulator of Phe, especially in adult PKU patients that have trouble maintaining a prescription diet. Synlogic has a rich portfolio of probiotic strains as treatment to metabolic disorders (trademarked as Synthetic Biotics™), in various stages of trial as therapeutics. Exploiting the microbiome for therapeutic applications is a fascinating and promising direction, and I expect to see more in the future.