Stroke Drug Development Targeting Mitochondria

Mitochondria take an important part in the pathological process of stroke via their different functions. For example, the energy provided by mitochondria is the basis for neuronal cell survival, while the reactive oxygen species produced by mitochondria promote cell death. In addition to energy supply and oxidative stress, mitochondria also influence the course of stroke through various mechanisms such as autophagy, apoptosis, and mitochondrial permeability transition pores. However, the specific mechanisms of action are still shrouded in mystery and remain to be uncovered. In view of this, Ace Therapeutics offers a comprehensive service for the development of drugs for stroke targeting mitochondrial function.

Virtual Screening of Mitochondrial Modulators

Discovery of drug candidates is the first step in the development of new drugs. Quickly and reliably finding a large number of modulators of mitochondrial function is the first step in targeting mitochondria for stroke drug development studies, and then, further evaluating the pharmacological effects of candidates on stroke to obtain potential drugs for the treatment of stroke. Virtual screening is undoubtedly the fastest way to screen a large number of mitochondrial modulators. We have an advanced and validated virtual screening platform for screening drug candidates that modulate mitochondrial dysfunction. Our platform allows virtual screening by targeting proteins associated with different mitochondrial functions, such as ATP-dependent channels, mitochondrial autophagy, mitochondrial apoptosis, oxidative stress, etc. We have an ultra-large compound library, natural product library, and fragment library to maximize diversity and successful hits.

• For the function of mitochondrial autophagy, we have selected specific target proteins including but not limited to PINK, Parkin, NIX, BNIP3, FUNDC1, etc.
• For mitochondria-associated apoptotic pathways, we select some signature proteins such as Bcl-2, BH3-2, Noxa, and PARP1.
• For mitochondria-associated oxidative stress, we select ROS, SOD2, NADPH oxidase, and other signature proteins.
• For ATP-dependent channels, we select signature proteins, such as Na+/K+ ATPase.

Validation of Mitochondrial Modulatory Activity of Candidates

For the mitochondrial modulators obtained from virtual screening, Ace Therapeutics has established abundant and stable in vitro and in vivo models of mitochondrial dysfunction to validate the modulatory effects of drug candidates on mitochondrial function, and those confirmed to be effective can then be used for subsequent pharmacological evaluation against stroke. These models include but are not limited to PINK1/Parkin activation-induced autophagy model, autophagy model induced by autophagy inducer, AIF activation-induced mitochondria-mediated apoptosis model, caspase3 activation-induced mitochondria-mediated apoptosis model, Bax activation-induced mitochondria-mediated apoptosis model, and p38/MK2 complex activation-induced mitochondria oxidative stress model.

We also provide various assays to verify the modulatory effects of drug candidates on mitochondrial function, such as examination of autophagosomes by transmission electron microscopy, detection of autophagic flow by fluorescent indicator systems, observation of chromatin condensation by DNA dye staining, and detection of mitochondrial membrane potential. Here, as an example of the detection of characteristic biomarkers, for the role of ATP function, we can detect Na+/K+ ATPase, serving the ATP content in mitochondria, and for mitochondrial autophagy, we offer the detection of PINK, Parkin, NIX, BNIP3, FUNDC1 and other proteins. For mitochondria-related apoptosis, we offer assays for caspase-3, Bcl-2, Bcl-xL, Cyt C, and others. For oxidative stress, we can detect ROS (reactive oxygen species), RNS (reactive nitrogen species), and HSP60.

Preclinical Evaluation of Drug Candidates for the Treatment of Stroke

After identifying suitable mitochondrial modulators through two screens, it is necessary to further confirm whether the drug candidates can alleviate stroke by modulating the function of mitochondrial dysregulation. To explore the pharmacological effects of these drug candidates for stroke, Ace Therapeutics has established mitochondrial-deficient stroke models, such as the post-ischemia-reperfusion model, mitochondrial apoptosis protein knockout stroke model, ATP synthesis-deficient stroke model, and other disease models. We also offer testing services for relevant biomarkers such as antioxidant enzymes, ATP levels, ROS, pro-apoptotic and anti-apoptotic factors to help you find the right drug for mitochondrial dysfunction to treat your stroke faster. These services include but are not limited to

• Development of cell-based assays
• Biomarker analysis
• Stroke models of mitochondrial dysfunction
• Mitochondrial ROS assays

Fig. 1 Workflow for mitochondria-targeted stroke drug development.

Depending on the design of the experiment, we offer personalized and customized services for the development of stroke drugs targeting mitochondria. If you would like to learn more about our services, please feel free to contact us.

1. Guan, R., et al. Mitophagy, a potential therapeutic target for stroke. J Biomed Sci, 2018. 25(1): p. 87.
2. Nguyen, H., et al. Understanding the role of dysfunctional and healthy mitochondria in stroke pathology and its treatment. Int J Mol Sci, 2018. 19(7).