Bernardi is a pioneer in the field of mitochondrial channels and their role in cellular physiopathology. In particular, he has focused on the permeability transition pore (PTP), a high-conductance channel increasingly recognised as a key element in cell death. In the early 1990s, he defined the key points of PTP regulation in isolated mitochondria. Subsequently, he developed tools to reliably monitor mitochondrial function in situ, addressing mechanistic questions regarding PTP as a target in degenerative diseases and cancer (see article).
His studies became extended to in vivo models and showed that early adaptation of mitochondria plays a key role in hepatocarcinogenesis (a tumour caused by an uncontrolled development of liver tissue cells) and in the onset of the Warburg effect (read more); and that PTP-mediated mitochondrial dysfunction unexpectedly causes muscular dystrophy in collagen VI deficiency. These studies have paved the way towards a potential therapy for Ullrich Congenital Muscular Dystrophy and Bethlem Myopathy with NIM811 (read more), a non-immunosuppressive analogue of cyclosporine A. The recent identification of PTP, which appears to originate from ATP synthase FOF1 dimers, offers significant promises for further molecular definition of the pore and its function in health and disease. The development of new chemical inhibitors of PTP with potential use in degenerative diseases, funded by the NIH – National Institute of Health, is currently one of the most successful research programmes in Bernardi’s laboratory.
Paolo Bernardi, in 1992, was one of the few to understand the importance of mitochondria in cell death, long before cytochrome c release was shown to be a key event in apoptosis. He pioneered the field, quickly achieving international recognition.
Dear Professor Bernardi, we are aware that from 1st October you will be retiring and will work for a period of two months at New York University. For this reason, unfortunately, you will not be present at the meeting on 19th October 2024. How are you coping with this new stage of life?
We researchers will never stop working! We do such an exciting and stimulating job that we are unaware of the passing of time and, when your project has a serious impact on other people’s health, you become even more committed. It is true, however, that turnover is also indispensable in the academic field. However, my group is still very active and comprises a very talented doctor, Silvia Castagnaro, who trained in Professor Bonaldo’s group, with whom she has published some excellent articles. Silvia is the main leader of my Telethon-funded project, which aims to test new active molecules in animal models in both type 6 collagenopathy and Duchenne dystrophy. We have also considered extending the range of applicability to other neuromuscular diseases, with a view to widening their therapeutic potential and appeal for the pharmaceutical industry.
We have also initiated a collaboration with a group in Lausanne, Switzerland, which has considerable experience in pharmacokinetics and pharmacodynamics, in order to study the biological availability and distribution of a new experimental drug. As a prerequisite for the project to go ahead, it is in fact essential to have reliable data on mammalian models, i.e. mice. We achieved great results with zebrafish, but a mammalian model is required by regulators and sponsors. This team is the same one that will be working on the Alisporivir – Debio 025 trial.
Tell us, first of all, about your role and that of your team in the Alisporivir – Debio 025 trial at Telethon.
The situation should be heading towards a conclusion, in the sense that all the necessary documents required have been completed. We, the laboratory researchers in Padua, are ready to work on this trial, which will follow the Cyclosporin A trial organised by Dr Luciano Merlini in 2008. From a biopsy taken before treatment and one after, we will study the mitochondrial function ex vivo to verify that we have the phenotype we have always found and that it responds to treatment in vitro. A sort of baseline. But importantly, the sample will also be used to make a definition of apoptosis, i.e. the number of damaged fibres. On the other hand, we will examine the regenerating fibres, having discovered from the Cyclosporin A study that the treatment not only improves performance and decreases the number of cells that die, but improves regeneration in the samples taken especially in children and young people. So, we will perform this test to obtain a baseline of each patient prior to treatment.
After treatment, we will then repeat the test to check whether the drug ‘hits the target’ as in the case of Cyclosporin A, observing the results in the regeneration and reduction of apoptosis, i.e. the loss of cells. This is objective data, because we can accurately measure the number of cells and thus the effect of the drug in the same patient. Dr Silvia Castagnaro has the necessary expertise to perform and interpret the results. I will remain in the Department as a consultant, I promise you, and this trial will remain one of my priorities! I consider it a moral responsibility.
What is your opinion as regards the shelf life of Alisporivir – Debio 025, in the sense that it was produced years ago and is still sitting there in storage?
In our experience the drug, which has a cyclic structure, is very stable and is not prone to degradation over time. My laboratory has been using samples of this drug for years and I honestly don’t think there is any problem. As soon as the one that will be used in the Trial is available, we will still conduct a one-day trial to confirm the efficacy in vitro. In vitro we have seen that it can be used even years later and I still have in the lab the original samples from Debiopharm that we started the study with!
How was the Alisporivir – Debio 025 molecule discovered?
It all started with HIV in 1992, when a very talented researcher discovered that administration of Cyclosporin A in vitro made it very difficult for HIV to replicate. The issue was that Cyclosporine could not be given to HIV patients because they were already immunosuppressed. Someone at Debiopharm came up with the idea of producing Cyclosporine without an immunosuppressive effect to see if it could be used in HIV therapy. It was already known that it was possible to separate immunosuppression from the desired effect in this case, i.e. the inhibitory effect on cyclophilins, which is also at the basis of protection in collagen VI diseases through mitochondrial cyclophilin. In the case of HIV, this is another cyclophilin found in the cytoplasm, but the importance lies in the fact that all cyclophilins react with these non-immunosuppressive derivatives.
Debiopharm had therefore produced this drug to test it in a trial with HIV patients, only to discover that the same patients all also had hepatitis B or C. The trial showed some efficacy of the drug against HIV, but it was not as effective as existing retrovirus inhibitor therapies. Instead, surprisingly, it specifically cured hepatitis C! In 2007, a trial was then carried out by Novartis in patients with HCV, i.e. hepatitis C, with excellent results, which were, however, superseded by a new discovery, i.e. the viral protease inhibitor that became the standard for hepatitis C therapy. This led to Alisporivir’s therapeutic indication for hepatitis C being dropped and, in 2015, its potential use in the therapy of collagen 6 diseases was re-launched.
All in all, this has been positive for us, in the sense that the steps and all the safety controls have been completed, allowing us to now test it for collagen 6. We are now in an optimum situation to be able to conduct the trial.
Can you make a comment on gene therapy for neuromuscular diseases? What is your opinion?
Following the latest discoveries, I have read extraordinary things. However, we have to think about where the ‘sick’ proteins are and in what kind of cells gene therapy is possible. In the case of Collagen 6, it would be necessary to reconstruct the entire global network that is formed at birth. This is a very complex matter. Replacing a stable extracellular protein, especially in the presence of variants, is a very difficult challenge. In other diseases, however, considerable progress has been made, especially for cells with a short life span and a high regenerative potential, such as red blood cell precursors but also epithelia affected by cystic fibrosis, for example.
In the case of collagen 6 and the extracellular matrix, the picture is far more complex. The muscle is in an intermediate situation because we know that it can regenerate; in fact one of our hopes is to replace the satellite cells so that they give rise to normal cells. The problem is that the collagen 6 defect is outside the muscle. But how do we replace collagen 6? Genetically, it is difficult because you would have to rebuild the tissue around the muscle in its entirety. This is why I find it somewhat short-sighted that attempts at pharmacological therapies are not promoted when they exist. I am thinking for example of molecules that can be administered that act on the muscle as if they were collagen 6 or, as in the case of Alisporivir, that help prevent the negative effects of the lack of collagen 6 on the muscle.
Would you please say a few words about the new molecules called TR001 and TR002, which are being studied in an official Telethon-funded project ‘Mitochondrial Therapy for Muscular Dystrophies’?
I will leave as a legacy new molecules, which are even more effective in vitro than Alisporivir and which I hope in the future can be used for combination therapy. These molecules (TR001 and TR002; TR is derived from triazole because this group was used to increase the stability of the molecule, which was originally obtained by Professor Forte and myself in a project funded by the NIH-National Institute of Health, are molecules that inhibit the mitochondrial permeability transition pore. They have shown excellent results in cells and in zebrafish models (see article) of both collagen 6 disorders and Duchenne dystrophy (see article). The project aims to evaluate their efficacy in mouse models of these diseases. The target is always the pore, as in the case of Alisporivir-Debio 025.
The next step will be to find someone to fund us with a view to creating a biotechnology initiative, and this is a project I may dedicate my time to even after retirement! I will have a free hand as my own agent for drug development.
Professor Paolo Bernardi, bernardi@bio.unipd.it
As an expert professor of collagen 6 on our Medical-Scientific Committee (Maybe you didn’t know this, but by accepting this interview you have given your consent for your appointment to the MSC of the Col6 Association), we definitely want you to keep us updated on upcoming projects especially with regard to the TR001-002 molecules.
We sincerely thank you, Professor Paolo Bernardi, for taking the time to conduct this interview and for your valuable contribution to scientific research to date. Your work has had and will continue to have a significant impact on the understanding and treatment of neuromuscular diseases.
We wish you all the best in your new phase of life and in your future research projects.