STAC3 disorder: gene therapy and malignant hyperthermia – STAC3

The project is divided into two main work packages. The investigations on normal and pathologic STAC3 function are essentially performed on a cellular model by the Austrian partner in Innsbruck. All mouse work (generation, characterization, therapeutic approaches) is largely conducted by the French partner in Strasbourg.

For the cellular work, we generated myotubes expressing STAC3 with or without the most common mutation. We also identified different STAC3 isoforms (= different proteins encoded by the same gene) and generated STAC3 constructs composed of different domains. We will investigate the capacity of the different STAC3 isoforms and constructs to ensure normal ECC in myotubes by recording calcium transients. In the next step, we will expose the myotubes to halothane (an anesthetics triggering MH) and caffeine (triggering intracellular calcium movements) and perform the same set of experiments. We will also investigate the expression of different genes implicated in the regulation of calcium balance. In parallel, we will examine the pathogenic effect of other mutations leading to MH to identify common and diverging pathomechanisms underlying the different disease forms.

The animal work involves two different mouse models - mice carrying the most common STAC3 mutation (needs to be generated and characterized), and mice completely lacking STAC3 in muscle (already available). Following the systematic characterization of the murine model with the frequent STAC3 mutation (measurement of muscle mass, behavioral tests, quantification of muscle force and fatigability, microscopic examination of myofiber morphology), both models will receive a single dose of a viral vector encapsulating STAC3. We will assess vector distribution in different tissues, perform echocardiography to conclude on heart function, determine muscle function and structure 4 and 8 weeks post-injection, and isolate single myofibers for calcium measurements.

For the work package managed by the French partner: STAC3 mice carrying the most common mutation have been successfully generated. The animals are alive and fertile, and we currently produce cohorts of sufficient size for systematic characterization.

The cellular work conducted by the Austrian partner is ongoing and a description of the first results will be provided soon.

Currently, STAC3 disorder is barely understood, and no therapy exists for this debilitating disease. This is a significant burden for affected families.

Investigations on the role of STAC3 in the ECC pathway and on the pathogenic effect of the STAC3 mutations will help to decipher the molecular factors mediating the development of MH. Moreover, the analysis of the cellular role of different STAC3 domains will help to design prospective therapeutic approaches with specific modes of action. The complementary work in Austria and France bridges the gap between cellular mechanisms and whole organism disease development, providing a more comprehensive view on STAC3 disorder and MH pathophysiology, and helps to enhance patient safety during surgeries requiring anesthesia.

Finally, the establishment of a reliable mouse model reproducing the main signs of the disorder is an asset and a prerequisite for the testing and validation of therapeutic strategies, which may be translated to STAC3 patients in the frame of clinical trials.

Wider research context: The adaptor protein STAC3 is essential for skeletal muscle excitation-contraction coupling and establishes two distinct interactions with the voltage sensor of EC coupling CaV1.1. A missense mutation in STAC3, W284S, disrupts one of the interactions with CaV1.1 and causes STAC3 disorder (OMIM # 255995). This rare recessively inherited congenital myopathy, for which no cure or treatment is available, is characterized by severe muscle weakness and susceptibility to malignant hyperthermia (MHS).
Hypotheses and objectives: Because affected patients carry two copies of mutated STAC3, whereas in heterozygous individuals the presence of one functional copy prevents the manifestation of symptoms, we hypothesize that introducing a functional copy of STAC3 to complement the genetic mutation associated with this disorder will alleviate the associated symptoms. Furthermore, we hypothesize that investigating the molecular mechanisms leading to MHS in STAC3 disorder will improve our understanding of not only the pathophysiology of MHS, but also of normal STAC3 function.
Approach: First, we will attempt gene therapy in two STAC3 disorder mouse models, an already available Stac3 cKO and a novel Stac3W280S cKI, with different STAC3 constructs. Secondly, we will investigate the molecular basis of MHS in Stac3W280S cKI fibers and reconstituted double Stac3/CaV1.1 KO myotubes.
Originality and innovation: STAC3 disorder is a debilitating disease for which no cure or treatment is available. Establishing a STAC3 disorder mouse model will allow validation of gene therapy in vivo, a first important step towards clinical trial. Additionally, investigating the molecular determinants of how STAC3 disorder causes MHS will further elucidate both the genetic and molecular factors contributing to malignant hyperthermia and STAC3 function in skeletal muscle excitation-contraction coupling. Finally, combining our expertise in both cell culture (Austria) and in vivo models (France) will allow us to bridge the gap between cellular mechanisms and whole organism responses, providing a more comprehensive perspective on the STAC3 disorder and MHS pathophysiology.
Primary researchers involved: Austria. The proposed work will be carried out by the applicant (M. Campiglio), a postdoc and a technical assistant. France: The proposed work will be carried out by the applicants (J. Laporte and J. Böhm), a postdoc and a part-time (50%) technical assistant.