Mucopolysaccharidoses (MPS) are a group of very rare disorders, also known as orphan diseases. They belong to the group of lysosomal storage diseases which are caused by a deficiency of one of the enzymes involved in the degradation of mucopolysaccharides (the acid glycosaminoglycans or GAGs). The enzymes are coded by genes which produce deficient gene products due to gene variants in each of the two gene-alleles.

Children of two carriers as parents have a 25% risk to suffer from MPS. For many families, the birth of the first affected child is a shock and a disaster. The disease is continuously progressing, and life spans are dramatically decreased without therapy. As a result, extensive efforts are put into the cure of these fatal disorders.

Enzymes are relatively small proteins, produced in the endoplasmatic reticulum of each cell. Before reaching the locus of their function, the lysosomes, additional modifications with special sugars are performed in the Golgi apparatus (glycosylation). Via mannose-6-phosphate marker, they connect to the mannose-6-phosphat receptor on the lysosomal membrane and can reach the final locus of their function. In the lysosomes, enzymes degrade the GAG chains into the smallest molecules for recycling or excretion. Any disturbance in this process leads to the accumulation of non-degraded material, which affects many other cell functions such as homeostasis, calcium metabolism, accelerates apoptosis and induces inflammation processes.

As lysosomes are ubiquitous, any disturbance leads to storage in many different tissues and organs. MPSs are a good example for chronic progressive multi-systemic disorders. The best theoretical option for treatment of any patient is to supplement the missing enzyme which could reach any organ via blood flow and get inside the lysosomes continuing the interrupted degradation processes.

The enzymes are ubiquitous and have some tissue specific compositions. Enzymes produced in the different cells and tissues have their own characteristics and are available on site. The production of recombinant enzymes means that the artificial glycosylation is created in a uniform composition for intravenous substitution with the aim to reach the organs with the blood-flow. There is no doubt that the therapeutic efficacy is ideal for many organs, such as liver, spleen, lung, and skin. All these organs have a good blood circulation and some ability to regenerate.

However, after years of treatment with the already available enzymes, it is shown that some organs are poorly supplied with blood and renewal cycles are slow, the ability to regenerate is decreased. Organs such as bones, cartilage, muscles, cornea, heart valves, meninges or the brain do not show the hope-for effect. All MPS types with brain involvement (neuronopathic forms of MPS types I, II and VII) or predominant skeletal dysplasia (MPS types IVA and B) cannot benefit from enzyme-replacement therapy and do not show the desired improvement.

In animal studies, modifications of glycosylation can change the ability to pass into organs not yet sufficiently reached such as cartilage or bones, but tissue-specific features cannot be sufficiently considered in any artificial production of the enzymes.

Avascular cartilage, heart valves and corneas cannot be reached by blood flow. Also, between blood vessels and brain tissue, several specialised cells form the blood-brain-barrier (BBB) to protect the brain from any unwanted substances in the blood. Therefore, new strategies are necessary to improve the therapeutic efficiency and to provide better outcomes for the affected patients. If patients with MPS I are diagnosed at a very young age, the best option is to treat them with haematopoietic stem cell transplantation (HSCT). Migrating stem cells can reach the brain and other organs, and then differentiate into organ-specific cells producing the missing lysosomal enzymes.

A straightforward method to overcome BBB is the direct injection of a recombinant enzyme into the cerebral fluid. This can be by lumbar puncture (intra-thecal) or intra-ventricular injections in the brain ventricles. Effects can be observed, however unfortunately not all challenges can currently be solved. The liquor flow can be reduced by thickened meninges with storage and vertebral deformities, which are typical for the disease. However, the barrier between cerebral fluid and brain tissue has still not been fully studied. The half-life of enzymes is limited, and the procedure has to be repeated regularly. The clinical trials for patients with MPS I, II, IIIA and IIIB could show some reduced or reversed progression of CNS pathology but long-term effects remain unclear.

Another possibility to overcome BBB is to fuse the enzyme proteins with macromolecules which enter the brain through receptor mediated active transport systems. This physiological transport is known for hormones, neurotransmitters and many other proteins (such as transferrin and insulin). They are transported through the BBB directly into the brain via specific receptores, so, the strategy is to fuse the natural proteins with the artificial enzymes needed in the MPS patient. It is important to note that clinical trials could potentially still show some improvement in affected MPS patients.

Another method is to conjugate the therapeutic enzymes with nano-capsules and to then ferry them across BBB via transcytosis or other transport mechanisms directly into brain cells. Pharmacological chaperones have been proven to be effective in other lysosomal storage diseases such as Gaucher or Fabry disease. Chaperones are able to stabilise three-dimensional conformation of misfolded proteins, such as enzymes. This would be the case of genetic variants causing missense mutation and exchange of only one amino acid in the protein chain. The misfolding pathology reduces stability, half-life and effect of the genetically conditioned enzyme, whereas the chaperone can reverse this disadvantage and increase the activity and efficacy of the enzyme. As a result, pharmacological chaperones are a good option for some diseases and could therefore be an option for some MPS patients in the future.

Some genetic variants cause stop-codons and the production of truncated dysfunctional peptides without any enzymatic activity and degradation within the cell. Stop-codon read through therapy aims for the genetic correction on an RNA level, resulting in the production of a sufficiently functioning gene product. It is already used for some specific mutation for patients with Duchenne muscular dystrophy, but it is too early to predict positive results for patients with MPS I.

Another possibility in the future might be the use of GAG-reducing small molecules such as Genistein, Pentosam polysulfate or Rhodamine B. They are able to influence and/or reduce the synthesis of GAGs which cannot be degraded sufficiently by the genetically changed enzymes with reduced function.

To reduce the GAGs as substrate, could be a chance to create a better relation between substrate and the impaired substrate reducing enzyme. As a result, lysosomal storage could therefore be reduced. Substrate reduction therapy is an established therapeutic concept in some of the other lysosomal storage diseases, but the usefulness in MPS disorders still needs to be proven.

The genetic corrections of DNA sequences in patient cells are no longer only future options as they have now become a reality. Gene variants causing missing or impaired functioning gene products could be replaced by correct genetic sequences and genes. This can be made as an ex vivo approach, where stem cells or fibroblast are removed from the patient and are then cultured in vitro, genetically corrected and consecutively re-injected into the patient.

The genetically corrected DNA in the re-transplanted autologous cells is able to produce correct gene products (in terms of MPS, this is the specific enzyme). The amounts of newly produced enzymes might be sufficient to positively influence the disease course of the treated patients.

An in vivo approach utilises viral vectors which invade cells, and even cell nuclei. Such viruses used are adeno-associated-viruses or lenti-viruses. Such manipulated viruses with the corrective genetic material are directly injected into the patient where they are internalised into deficient cells and are then able to produce the missing gene product. In the case of MPS, the aim is to produce enzyme proteins with sufficient concentrations and activity to prevent the storage of GAGs. Furthermore, clinical trials are underway for several MPS types and therefore, might offer a therapeutic opportunity in early life for affected patients. However, larger studies and a longer follow-up is still needed.

To conclude, MPS are rare genetic disorders and for a long time, they were linked with the myth of being untreatable diseases. Although some of the new therapeutic options are still in clinical trials and not routinely used, the present shows that many of the patients can benefit from the yet available options of HSCT and enzyme replacement therapies. These therapies have an undoubted effect for some of the MPS patients, especially if any form of therapy is started early or if the course of the disease does not affect the nervous system.

However, in the future, new therapeutic options will hopefully bring benefits to those that are not sufficiently improved; the decision of the best therapy will be made on the basis of factors such as the genetic defect, the type of MPS, and the age during treatment. This individualised and personalised therapy will improve the success of MPSs therapies.

Susanne Gerit KircherMedical University of Vienna, AustriaCenter of Pathobiochemistry and Geneticssusanne.kircher@meduniwien.ac.atwww.mps-austria.at

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Mucopolysaccharidoses: future therapies and perspectives - SciTech Europa