Mitochondrial diseases are a clinically, biochemically and genetically heterogeneous group of disorders. Signs, symptoms and the organs involved can differ from one person to another, making the diagnosis of mitochondrial diseases and the search for potential therapies complex. Here we outline the heterogeneity surrounding mitochondrial diseases and explore some of the challenges biopharma companies active in this field, including Khondrion, face in developing much needed treatments for patients.

Mitochondrial disease patients can present with a broad spectrum of clinical expression, with symptoms starting at any age and involving virtually any, and in many cases, multiple, organs or tissues. Affected organs can also cause symptoms across other organs and tissues – for example, altered brain function can impair motor function.

Many of the chemical processes taking place in our bodies can be impacted by mitochondrial diseases. All kinds of combinations of enzyme-complex deficiencies have been reported, caused by single or multiple oxidative phosphorylation enzyme-complex deficiencies, resulting in heterogeneity at the biochemical level.

Another aspect, when it comes to the many different characteristics of mitochondrial disease, is the diversity of genetic mutations that can affect the proteins involved in the assembly and maintenance of the body’s oxidative phosphorylation system. This system drives the key biochemical process (called oxidative phosphorylation) that our cells use to generate energy, and which is associated with many forms of mitochondrial disease. Currently around 400 different mutations have been described and more are frequently being discovered.

To add to this genetic complexity, some known mitochondrial diseases are not caused by the same gene mutation in every patient but can be caused by different mutations in different genes, which go on to affect single or multiple proteins in our oxidative phosphorylation system. Leigh disease is the most prominent example of a mitochondrial disease that has one common pathology but can be caused by different genetic defects, with approximately 75 genes involved. Moreover, a single mutation in a single gene does not always lead to the same mitochondrial disease characteristics in patients. For example, the much studied 3243A>G mutation in the MT-TL1 gene may give rise to variable clinical phenotypes (characteristics) like classic MELAS (mitochondrial encephalopathy with lactic acidosis and stroke-like episodes) and maternally inherited diabetes and deafness (MIDD) syndromes, and a variety of other single or multi-organ involvement (mixed) phenotypes.

The reasons for these differences in the age of onset at which symptoms present and why there is so much variability in symptoms among patients with the same gene mutation are unfortunately poorly understood, which complicates the research into new drugs. One factor causing this heterogeneity is mitochondrial DNA heteroplasmy, the phenomenon that not all cells carry the same quantities of the genetic defect. A cell can have some mitochondria that have a mutation in its mitochondrial DNA and some that do not. The influence of other, still unknown, factors are under investigation in the mitochondrial research field.

Unravelling mitochondrial disease heterogeneity in drug development
As scientists search for the urgently needed treatments for mitochondrial disease, several avenues are currently being explored to correct underlying gene defects (e.g. Leber’s hereditary optic neuropathy, LHON) or the cellular consequences of hampered oxidative phosphorylation. In light of mitochondrial disease heterogeneity, when undertaking such research, it is important to carefully define which patients to study and how to evaluate the effects of investigational drugs in clinical trials, e.g. which symptom measures to evaluate and how long to treat and follow patients.

Deciding what patients to study
At Khondrion, we decided that the best place to start our clinical studies was in adults with the m.3243A>G mutation. This followed an in-depth analysis of the cellular consequences of mitochondrial diseases and positive effects from our lead drug candidate sonlicromanol in various in vitro, ex vivo and in vivo cell and animal models. This particular mutation underlies one of the most frequently encountered mitochondrial diseases, MELAS spectrum disorders. With their specific disease-causing mutation, these patients represented a first group for us to start our clinical research, supported by a wealth of natural history data for this population from leading mitochondrial disease centers worldwide.

Deciding what outcomes to measure
Given the variable nature of mitochondrial diseases, including MELAS spectrum disorders, predicting the clinical effects of new drug interventions based on preclinical studies is difficult. As a result, early clinical efficacy and safety studies involving patients are largely exploratory even when, due to statistical analysis rules, primary and secondary endpoints need to be predefined. It is therefore important that when studying mitochondrial disease in such heterogeneous patient populations, the design of such studies takes into account a well-considered primary endpoint and a broader set of secondary, disease-related parameters to provide much needed clinical information to optimize subsequent decision-making.

Following the science
Extensive preclinical research with sonlicromanol showed, amongst others, improvements in brain parameters related to cognition. In our Phase 2a trial (the KHENERGY study), the first indications of a beneficial effect for patients on cognition were confirmed. Following more research by the broader mitochondrial community in this area in recent years, it has become increasingly clear that cognitive impairment is an important, clinically relevant symptom in MELAS spectrum disorders, having a significant negative impact on daily activities and quality of life for patients (read more here about the growing understanding between mitochondrial disease and cognition).

Following up on the data from the KHENERGY study, that is why in our ongoing Phase 2b trial (the KHENERGYZE study, a placebo-controlled, double-blind dose-finding trial) we are now studying the dose-effect of sonlicromanol on patients’ cognitive functioning. We have also further refined the patient inclusion criteria for this study, to reduce the heterogeneity of the patient population and better ‘match’ the patient group with the cognition-oriented functional outcome measures. It is our hope that this study will reconfirm a possible effect of sonlicromanol so that we can explore this further by following up with the included patients in a long-term extension study as well as advancing our innovative medicine into a pivotal Phase 3 study.

We believe this stepwise trial approach is critical for getting new treatments for mitochondrial diseases approved, particularly given these diseases’ heterogeneous nature and the complexity of how patients are affected. Together with an active dialogue with regulators, the continued persistence of patient groups in raising awareness and, most importantly, all of the patients participating in and supporting our studies, we are of the opinion that this multi-faceted approach to mitochondrial disease research and drug development is best suited to bring much needed effective treatments to patients in a timely manner.