Nat Commun and Qi Xin's group reveal the molecular mechanism of compound CHIR99021 in the treatment of Huntington's disease by regulating mitochondrial function

Written by Di Hu, editor in charge, Wang Sizhen Mitochondria (mitochondria) are important organelles responsible for the synthesis of ATP
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In neurons, mitochondria are responsible for lipid metabolism, Ca2+ buffering, and redox regulation [1]
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Therefore, mitochondria directly affect the function of neurons, and mitochondrial dysfunction can lead to neuron death [1]
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In a variety of neurodegenerative diseases (neurodegenerative diseases), mitochondrial dysfunction (mitochondrial dysfunction) has been widely regarded as an important cause of neuronal death [2]
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Therefore, enhancing and regulating mitochondrial function may become an effective treatment for neurodegenerative diseases
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 Huntington's disease (Huntington's disease, HD) is a hereditary malignant neurodegenerative disease.
Its typical neuropathological features include the death of neutral spiny neurons in the striatum of the brain (medium spiny neuronal death) And the aggregation of mutant huntingtin (mtHtt) [3]
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 Patients suffering from this disease will have emotional and intellectual problems in the early stage, followed by uncoordinated movement, as the movement ability gradually deteriorates, causing movement difficulties, and severe cases eventually die [3]
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There is currently no cure for HD, and existing treatments can only delay the progression of the disease
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Because mtHtt can damage a variety of molecules and cell functions in neurons, the discovery of new targets for HD and drug development have become difficult [4]
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Recent studies have clarified that mitochondrial dysfunction plays a key role in the pathogenesis of HD
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 On September 6, 2021, Xin Qi's research group from Case Western Reserve University School of Medicine and Drew Adams' research group jointly published an online publication entitled "Small -molecule suppression of calpastatin degradation reduces neuropathology in models of Huntington's disease" research paper reveals that enhancing mitochondrial function is an important mechanism for a potential treatment of HD
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Di Hu is the first author of the paper
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In this study, the authors used high-throughput screening to find the small molecule compound CHIR99021 with mitochondrial function as the target, and verified its therapeutic effect in a variety of cell and animal models
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The author clarified the signal pathways by which the compound regulates mitochondrial function
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This research has promoted the understanding of the pathogenesis of HD and the corresponding drug development
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In the striatal nerve cells (HdhQ111 cells) derived from HD mice, CHIR99021 treatment can significantly enhance mitochondrial membrane potential, improve cell viability, and effectively enhance mitochondrial respiration
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In the neutral spiny neurons differentiated from induced pluripotent stem cells (iPSC) in HD patients, CHIR99021 treatment also effectively enhanced mitochondrial function and significantly improved neuronal morphology
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These results indicate that CHIR99021 has a protective effect on neurons affected by HD
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Next, the author used two HD mouse models, R6/2 (acute transgenic model) and YAC128 (chronic transgenic model) to verify the protective effect of CHIR99021
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Through long-term intraperitoneal injection of CHIR99021, the authors observed that the survival of R6/2 HD mice was significantly improved, the accumulation of mtHtt protein in the striatum was significantly reduced, and the number of neutral spiny neurons was significantly increased
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Long-term injection of CHIR99021 also significantly reduced the neuropathological characteristics of YAC128 HD mice
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In addition, CHIR99021 treatment also significantly improved the spontaneous activity and motor coordination ability of mice
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These results further prove that CHIR99021 can effectively protect neurons and alleviate the pathological development of HD
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At the same time, CHIR99021 injection has no obvious side effects on normal mice
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 Furthermore, after excluding glycogen synthase kinase 3 (GSK3) as the molecular target of CHIR99021 through a series of experiments, the authors used proteomic analysis to find that CHIR99021 treatment in HdhQ111 cells down-regulated Calpain I (Calpain I) And up-regulate the expression of calpastatin (Calpastatin, CAST)
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It is known that CAST is an endogenous specific inhibitor of Calpain I [5]
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In many neurodegenerative diseases, the overactivation of Calpain I is importantly related to nerve damage [6]
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Therefore, the authors hypothesized that CHIR99021 protects mitochondrial function by regulating the CAST-Calpain I molecular pathway
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In a variety of cells, mouse models, and brain tissues of HD patients, the authors observed that the protein level of CAST was consistently reduced, and the protein cleavage level of Calpain I substrate α-Spectrin was significantly increased
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However, the author did not detect the direct effect of CHIR99021 on Calpain I enzyme activity, which indicates that CHIR99021 indirectly inhibits Calpain I by up-regulating the level of CAST protein
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In HdhQ111 cells and HD iPSC differentiated neurons, CAST protein knockdown prevented CHIR99021 treatment no longer enhancing the mitochondrial membrane potential and cell survival, proving that the mitochondrial protection provided by CHIR99021 depends on the CAST protein
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The authors further found that in HD cells, the degradation rate of CAST protein and the level of ubiquitination increased significantly
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CHIR99021 treatment greatly reduced the ubiquitination level of CAST protein, so that the protein level returned to the normal level
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It can be seen that CHIR99021 stabilizes CAST protein levels by inhibiting CAST degradation.
In other words, CHIR99021 treatment can inhibit the ubiquitin-lysosomal protein degradation pathway of CAST in HD
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 Finally, the authors found that CHIR99021 can inhibit mitochondrial division and restore mitochondrial morphology by regulating the activity of dynamin-related protein Drp1
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The phosphorylation of Drp1 promotes excessive mitochondrial fragmentation, leading to mitochondrial fragmentation, respiratory depression, and a decrease in membrane potential
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The authors tested the expression of a series of proteins related to mitochondrial quality regulation and found that the protein level of phosphorylated Drp1 was significantly increased in HD cells, while CHIR99021 treatment reduced it to a normal level
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In addition, in the HD mouse model, the authors also observed that CHIR99021 treatment significantly reduced the phosphorylation level of Drp1
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This shows that CHIR99021 regulates mitochondrial morphology and function by regulating the activity of Drp1 protein
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Similarly, CAST protein knockdown makes CHIR99021 treatment no longer regulate mitochondrial morphology, which further proves that the mitochondrial protection of CHIR99021 depends on CAST protein
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Figure 1 Summary of work: The mechanism of CHIR99021 regulating mitochondrial function (picture quoted from: Hu et al.
, Nat Comm 2021; 12: 5305) Conclusion and discussion of the article, inspiration and outlook The author discovered CAST by studying the mechanism of CHIR99021 to enhance mitochondrial function -Drp1 acts as a signaling pathway for regulating mitochondrial function (Figure 1)
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Decreased CAST protein levels cause Drp1 phosphorylation and lead to mitochondrial fragmentation and mitochondrial dysfunction
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CHIR99021 treatment can effectively inhibit the ubiquitination and degradation of CAST protein, thereby inhibiting mitochondrial division and restoring mitochondrial morphology
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Long-term injection of CHIR99021 significantly alleviated the neuropathological characteristics of HD mice and improved their exercise performance
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This shows that enhancing mitochondrial function can effectively alleviate the pathological development of HD
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Therefore, CHIR99021 may be a potential HD treatment drug
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In addition, this study also suggests that in addition to HD, in other neurodegenerative diseases, such as Parkinson's disease and Alzheimer's syndrome, nerve damage caused by the over-activation of Calpain I has also been repeatedly demonstrated
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Therefore, CAST can be used as a universal target protein, and CHIR99021 may be widely used
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 Of course, there are still some unresolved problems in this research
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For example, although CHIR99021 can inhibit the ubiquitination and degradation of CAST, the corresponding regulatory molecules have not yet been discovered, and the possibilities include specific ubiquitin or deubiquitinating enzymes
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Secondly, although CHIR99021 does not regulate mitochondrial function by inhibiting GSK3, GSK3 is still an important target protein of CHIR99021.
Therefore, it is necessary to develop CHIR99021 derivatives or chemically modified small molecules that can selectively enhance CAST activity
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Link to the original text: https:// Qi Xin (Xin Qi) Professor Xin Qi's research group is committed to studying the role of mitochondrial quality control and metabolism in neurodegenerative diseases
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The research group uses cell biology, molecular biology, stem cell culture, animal models and other related experimental techniques to comprehensively analyze the pathological mechanism of neurological diseases, and through high-throughput screening of small molecule compounds and self-design and synthesis of specific peptides to develop mitochondria Targeted treatment methods
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The research team is recruiting post-doctoral fellows and researchers, please contact us if you are interested
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 Selected articles from previous issues [1] JAMA Neurol︱ Attention! Young people are more likely to suffer from "Alzheimer's disease"? [2] Cereb Cortex︱ Ku Yixuan team reveals the ipsilateral sensory cortex representation model of working memory [3] Brain | For the first time! PAX6 may be a key factor in the pathogenesis of Alzheimer's disease and a new therapeutic target [4] Sci Adv︱ blockbuster! DNA methylation protein DNMT1 mutation can induce neurodegenerative diseases [5] Cell︱ new discovery! New enlightenment of midbrain-regulated movement phenomenon for the treatment of Parkinson’s disease [6] Cereb Cortex︱MET tyrosine kinase signal transduction timing abnormality is a key mechanism affecting the development and behavior of normal cortical neural circuits in mice [7] Nat Biomed Eng︱ The team of academician Ye Yuru develops a new strategy for whole-brain gene editing-mediated treatment of Alzheimer's disease [8] Luo Liqun Science's heavy review System Interpretation ︱ Neural circuit structure-a system that makes the brain "computer" run at high speed [9] Sci Adv ︱Important discovery! The calcium homeostasis regulatory protein Calhm2 regulates microglia activation and participates in the process of Alzheimer's disease [10] EMBO J︱ new discovery! AGHGAP11B promotes the expansion of the neocortex into adulthood and improves cognitive ability [11] Cell Death Differ︱ Qi Yitao/Wu Hongmei and others cooperate to reveal the molecular mechanism of SUMO modification regulating neurogenesis in adult mice [12] Cereb Cortex︱A2A receptor antagonist can Reversal of sequence learning impairment induced by abnormal aggregation of α-Syn (slide up and down to view) [1] Misgeld, T.
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