- Published on 18 September 2013
Knowledge of underlying mechanisms, advances in biotechnology, and progress in exploratory methods that make it possible to evaluate the effects of new treatments, are all coming together to provide novel treatments to restore cochlear structure and function, and some patients are already benefiting from these innovations.
For many years, scientific progress and innovation in the areas of hearing and hearing disorders have been major topics on the agenda of the annual national conference of the French Union for Hearing Aid Specialists (UNSAF). And the 35th edition of the event, held in Paris in mid-April, proved to be no exception. Given recent progress and the fact that novel approaches will soon be arriving in clinical practice, it seems that the time has come for the various professions to work side by side in new ways, because, as Professor Paul Avan explains, ”Treating deafness by gene therapy will not mean that there will no longer be a need for hearing aids! The medical scientific approach and the field of audioprosthetics are entirely compatible. To make sure that these complementary areas bring as much benefit to patients as possible, it is important that all the players move forward together, sharing knowledge as we go.” A strategy to ensure that “knowledge makes its way out of universities", which was an integral part of the scientific sessions at the conference, during which the speakers were able to provide an audience of ENT practitioners and hearing aid specialists with all the latest news on research in gene therapy applied to the area of deafness, and on new avenues currently being explored for drug therapy.
Gene therapy: has the cochlea’s time arrived?
Professor Lawrence (Larry) Lustig, ENT surgeon specializing in cochlear implants and director of a research laboratory at the University of California focusing on gene therapy, is a good example of someone whose work in both worlds helps to turn advances in research into practical applications. Prof. Lustig was invited to the congress by Paul Avan and Éric Bizaguet as a guest speaker to talk about the progress made so far in fundamental and clinical research, and the advances that are showing promise as new solutions for patients with deafness in the near future.
A better understanding of the intricate mechanisms that enable us to hear
Will gene therapy make good on its promises concerning restoration of cochlear function? This question is worth posing, particularly since there is such a diverse range of disorders that could theoretically benefit from progress in genetics for the development of treatments. Of course, the greatest advances have been made in the field of monogenic deafness, since their underlying pathophysiology is better understood. A classic example of a disorder in this group is deafness occurring in Usher syndrome. Many multidisciplinary research teams are working on this syndrome which includes early hearing loss and late-onset blindness. In this disorder, like in other forms of deafness, the groundwork has been laid for the launch of clinical trials in the near future. However, as Aziz El Amraoui, research director at the Institut Pasteur’s Unit for Genetics and Physiology of Hearing, explains, "More than two thirds of cases of childhood deafness are of genetic origin, but of the 200 to 300 genes we think may be involved, only 70 have been identified and the product is known for only 40 of them. Each of these genes is an entry point when trying to understand the mechanisms that underlie the chain of events that makes it possible to understand messages through sound.”
Other forms of deafness that are more multifactorial are far less well understood. Their pathogenesis and course involve multiple pathophysiological cascades in which genetic determinants intervene as modulators of responses to a wide range of environmental factors. This is the area that includes the most frequent forms of deafness, for instance those related to age or so-called "acquired” deafness. But many of these disorders are also dependent on genetic susceptibility factors. Much more work needs to be done to breakdown precisely the mechanisms that lead to dysfunction of physiological systems as diverse as those involved in the areas of inflammation, but also immunity or even antioxidant or antiapoptotic defenses. Each of these systems is complex and partially understood, and all are interlinked as a result of common mediators, many of which remain to be discovered. There are still many obstacles to overcome, even for this very first, but essential, step in elucidating the fundamental mechanisms that result in deafness. Beyond that, the perimeters of the work still need to be defined, especially if we follow the line of thought of Aziz El Amraoui, who says that "Research is not so much about finding an answer to a question, it’s more about finding the next question that needs to be asked.”
He and his research team are currently focusing on a fundamental part of the system, the cilia of ciliated cells. More specifically, the scientists at Institut Pasteur are trying to understand how stereocilia grow and are organized during embryonic development. They have already demonstrated that the central primary cilium, attached to an actin base, migrates to the periphery to organize coherent and orderly growth of the three rows of stereocilia, with the final structure being anchored through adhesins and connexin 26.
Vectors, promoters and routes of administration
Alongside efforts to elucidate the fundamental mechanisms of hearing and their genetic determinants, there are also highly technical aspects that need to be examined to insert “medicinal” genetic material into defective cells. This mainly involves identifying the best vectors and promoters able to carry a foreign gene and to ensure its expression in the target organ.
In this area, adenoviruses and adeno-associated viruses (AAVs) are the most effective and most commonly used when developing treatments intended for clinical use in humans. As of the late 1990s, it was demonstrated that these viruses are able to induce stable protein expression from a foreign gene in cells of the inner ear in mammals. These vectors are still the most widely used in current research. The promoters that need to be included with the "genetic drug" are also of primary importance to ensure that new protein production effectively takes place in the target organ. There are ubiquitous promoters such as CMV that can be used, but there are also more specific ones. For example, platelet-derived growth factor (PDGF) will direct expression towards cochlear vessels, neuron-specific enolase (NSE) to nerve fibers, and elongation factor-1 alpha (EF-1 alpha) towards certain cells in the spiral limbus.
From fundamental research to first in man trials
Research carried out in the early 2000s using knock-out mice models for ATOH1 (a major gene in the differentiation of ciliated cells) proved that it was experimentally possible to restore functional ciliated cells leading to correction of hearing, and that the level of correction was related to the number of restored cells. Similar successes have been achieved in the vestibule. Complete cochlear restoration, with functional internal ciliated cells, has even been obtained in genetic transfer models in mice pups in utero. Investigations continue and the most recent relate to non-syndromic deafness, in which the GJB2 gene appears to play a key role. The glutamate transporter 3 protein is the topic of research at the unit headed by Larry Lustig, who explains "MicroRNA transfers have been found to be capable of inducing expression of this molecule throughout the organ of Corti, enabling restoration of hearing in the treated animals.”
New drug therapy approaches for presbycusis and tinnitusProfessor Jean-Luc Puel, Director of the Montpellier Neurosciences Institute, is also Director of the Montpellier School for Hearing Aid Specialists. Links between training and research are the center of his day-to-day activities, making him an ideal speaker for these scientific sessions. In a presentation with a title that he himself admits is somewhat provocative "A pill to hear and no longer to hear "them"", he presented an overview of pharmacological treatment options for hearing loss and tinnitus, a phenomenon frequently associated with hearing impairment. “Presbycusis does not exist, but rather presbycuses. In the same way as there isn’t one tinnitus, but various forms of tinnitus," says Prof. Puel, highlighting the complexity of developing these new treatments.
Current obstacles limiting gene therapy regenerationIn utero, during embryonic development, cochlear cells all stem from early cells that are poorly differentiated, known as progenitors. Subsequent differentiation and specialization into some type of cell take place under the influence of various genetic factors. For example, a progenitor cell expressing the Math1 gene will differentiate into a ciliated cell, while another expressing p27 will evolve into a supporting cell. If ciliated cells are lost, is it possible to "reprogram" supporting cells to transform them into ciliated cells?
Jean-Luc Puel has two answers to this question: "Gene therapy approaches that aim to promote expression of the genes needed for reprogramming by the remaining cells appear to have reached proof of concept, but until now they have only been able to yield cells that are quite primitive and not particularly functional. This type of strategy also brings up the issue of a possible deficiency of supporting cells following this reprogramming process." Other avenues are also being explored, for instance first reprogramming remaining cells to become progenitors, and then redirecting them toward a differentiation route, either ciliated cells or supporting cells. Prof. Puel’s team has tried to do just that, in vitro then in vivo. An adenovirus was used as a vector for the genes of interest. Initial results show that the infected cells divide and then one week later, 10 to 12% of them express ciliated cell markers “but remain very archaic…,” he explains.
Drug therapy: a step closer to clinical applications
What are the signaling pathways that lead to apoptosis of ciliated cells when exposed to noise? Understanding these mechanisms would help to identify therapeutic targets, be they enzymes, transcription factors, etc., and to develop compounds able to counteract the expression of a specific signaling pathway, with the aim of protecting the cochlea from noise.
But an effective methodology to investigate these questions still needs to be found. Animal models of noise-related trauma are difficult to work with because sensitivity to noise shows high interindividual variability. Prof. Puel adds "I prefer experimentation in which each animal is their own control, studying one ear versus the contralateral ear as a control.”
An antiapoptotic compound for ciliated cells has been tested, with application to the round window. It provides a protective effect, but the therapeutic interval is short, at about 24 hours. A result that is "Hard to apply in clinical practice," says Prof. Puel. Mouse models with accelerated aging are well known in studies concerning Alzheimer’s disease and can also be used to study age-related changes in hearing. These animals have a combination of changes that reflect the range of pathways that lead to hearing loss: loss of external ciliated cells (sensory presbycusis), neuronal loss with preservation of internal ciliated cells (neural presbycusis), and changes in the stria vascularis (striatal presbycusis). This model has been used in particular to assess the effects of oxidative stress and free radicals on the cochlea, and then antioxidant effects. The findings showed that this type of compound could be able to slow the course of presbycusis.
What about a "tinnitus drug"?
A model for tinnitus in rats with induction by aspirin or noise has been used to test the effects of a glutamate inhibitor. The test proved to be conclusive in animals, with antiglutamates blocking persistent tinnitus. Animal models however have their limits, making long-term efficacy impossible to evaluate. The first clinical trials with antiglutamates were carried out in Germany. Treatment was applied using an external pump to the round window. During application, tinnitus decreased, but unfortunately recurred as soon as treatment was stopped and were even exacerbated is some cases.
The current approach is therefore to develop an implantable minipump, able to administer the drug continuously via a catheter applied to the oval window. A prototype of this kind should soon be available through a collaborative project between INSERM (French National Institute for Health and Medical Research), the CEA (French Atomic Energy Commission), the European Membranes Institute, and the Australian company Cochlear. The results of these studies are highly awaited because the prevalence of tinnitus in the general population is known to be high, particularly in young subjects exposed to the risk factor of amplified music.
Arielle Le Masne, editor-in-chief, Audiology infos France