- Nrf2 activators bind to Keap1, a protein that coordinates the cellular response to reactive oxygen species (ROS). Binding to Keap1 activates Nrf2, a transcription factor that promotes normal mitochondrial function by making reducing equivalents available for ATP production and increases cellular antioxidant content. This availability reduces mitochondrial ROS production and ROS-mediated activation of inflammatory signaling complexes. Binding to Keap1, and activation of Nrf2, also inhibits NF-κB, the primary transcription factor producing proteins that promote inflammation and the production of ROS.
- Nrf2 activators restore mitochondrial production of ATP, increase production of antioxidants, reduce oxidative stress, and reduce pro-inflammatory signaling. Since mitochondrial dysfunction, oxidative stress, and inflammation are features of many diseases, Nrf2 activators have many potential clinical applications and have been the subject of hundreds of peer-reviewed scientific papers. Mitochondrial dysfunction is directly implicated in a number of diseases and, in many diseases, is seen in conjunction with chronic inflammation. In a number of diseases, chronic inflammation can lead, in the longer term, to organ fibrosis and remodeling. Reducing this chronic inflammation may lead to reduction of fibrosis and remodeling. The effects of Nrf2 activators are described below in more detail.
Promoting Energy Metabolism and Mitochondrial Function: Mitochondria are often described as the power plants of the cell because they generate energy through the production of ATP, the primary unit of cellular energy. Mitochondrial dysfunction, which is manifested through decreased cellular energy production and increased production of ROS, is a feature of many chronic inflammatory diseases1. Nrf2 activation reduces mitochondrial ROS, promotes the availability of fatty acids and glucose for mitochondrial ATP production, and increases mitochondrial biogenesis2,3,4.
Reducing Oxidative Stress, Inflammation, and Inflammatory Signaling:
Inflammation is a protective response of the body to harmful stimuli such as invading pathogens, damaged cells, or irritants. It evolved to neutralize the initial cause of injury, eliminate dead cells, and initiate repair of damaged tissues. A central feature of inflammation is mitochondrial dysfunction and the production of ROS, which promotes activation of inflammatory pathways. In many diseases, inflammation does not resolve normally and is associated with chronic excessive ROS, organ fibrosis, remodeling or other tissue damage, and impaired ATP production5,6,7.
The foundational biology of Nrf2 underlies our two lead product candidates, bardoxolone methyl and omaveloxolone, and certain of our preclinical programs.
Many diseases have three features in common: inflammation, oxidative stress, and mitochondrial dysfunction. Healthy cells normally sense the presence of pathogenic organisms and other toxic stimuli. When these danger/damage signals are sensed, cells rapidly respond by increasing the production of reactive oxygen species (ROS) and pro-inflammatory cytokines. To facilitate production of these inflammatory mediators, mitochondria adopt an “inflamed” state, wherein normal mitochondrial function is temporarily suppressed. This shift-in-state allows mitochondrial resources to be diverted away from ATP production and toward other pathways that orchestrate normal cellular defense and repair processes. Reorientation of mitochondria from energy production to ROS production allows the affected cells to kill invading pathogens with molecules such as hydrogen peroxide and superoxide. Some of the ROS produced by inflamed mitochondria are released from the mitochondria into the cytoplasm and promote the activity of multiple key pro-inflammatory signaling complexes, including the IκBα/NF-κB complex and the NLRP3 inflammasome. In a normal disease process, after the pathogens have been eliminated the resolution phase of inflammation can begin, and naturally occurring molecules promote the resolution of inflammation, in part, by activating the Keap1/Nrf2 pathway. As a result, pro-inflammatory mediators are reduced, ROS are neutralized, and normal mitochondrial function is restored. However, in many chronic inflammatory and genetic diseases, this resolution process does not occur or is inadequate, leading to mitochondrial dysfunction, oxidative stress, and chronic inflammation, all of which can ultimately lead to tissue damage.
Nrf2 activators mimic the activity of the endogenous molecules that promote the resolution of inflammation and restore homeostasis by binding to Keap1, a protein that coordinates the cellular response to ROS and other stimuli, each of which can cause cellular damage (generally referred to as oxidative stress). Binding to Keap1 activates Nrf2, a transcription factor that increases the levels of antioxidants and transporters, thereby reducing the levels of oxidative stress caused by excess ROS. Nrf2 also restores normal mitochondrial function by increasing the availability of substrates and reducing equivalents that are required to support ATP production. Nrf2 activation inhibits inflammation by reducing ROS levels, restoring normal mitochondrial function, and directly inhibiting pro-inflammatory signaling.
Because mitochondrial dysfunction, oxidative stress, and inflammation are features of many diseases, including diseases of genetic mutation, fibrotic cardiovascular diseases, and chronic inflammatory diseases, Nrf2 activators have many potential clinical applications and have been the subject of hundreds of peer-reviewed scientific papers.
Promoting Energy Metabolism and Mitochondrial Function
- ATP production: ROS are produced in the mitochondria as a byproduct of ATP production. Nrf2 activation improves mitochondrial efficiency by making antioxidant enzymes available to reduce or neutralize ROS2. The management of these reducing equivalents is a constant and critical balancing act within the mitochondria. Disease processes that increase ROS deplete reducing equivalents available for ATP production1. Accordingly, the induction of antioxidant proteins through Nrf2 activation augments mitochondrial ATP production2,8.
- Efficient consumption of fats and sugars: Nrf2 activation promotes the transport of fatty acids to the mitochondria where they are converted into reducing equivalents used to produce ATP1. Nrf2 also promotes the transport of glucose from the bloodstream into the cells where it is converted into reducing equivalents used to produce ATP. Nrf2 activators have been shown to promote glucose uptake and oxygen consumption in animal models of diet-induced obesity and diabetes9,10.
- Mitochondrial biogenesis: PGC1α is a protein that increases the number of mitochondria in a cell. Activation of Nrf2 has been shown to increase PGC1alpha expression in skeletal muscle, which may increase ATP production4,11.
Reducing Oxidative Stress, Inflammation, and Inflammatory Signaling
- Reactive oxygen species: ROS are chemically reactive molecules that contain oxygen and have important roles in cell signaling and balancing cellular systems. ROS are formed during mitochondrial ATP production and by a variety of other cellular processes. ROS increase inflammatory signaling, and excessive ROS can cause cellular damage to tissues in critical organs including the muscles, lung, heart, liver, brain, and eyes. Excessive ROS and chronic inflammation have been shown to be the cause of cellular damage in many diseases7. Nrf2 activation increases the cellular content of antioxidant, which makes reducing equivalents available to neutralize ROS12. This suppresses the pro-inflammatory signaling effects of ROS and protects tissues from the damaging effects of excessive ROS.
- Inhibition of inflammatory signaling: Mitochondrial ROS, NF-κB, and the NLRP3 inflammasome are important activators and regulators of the inflammatory response. Nrf2 activators, through reduction of ROS and inhibition of NF-κB and the inflammasome, suppress production of TNFα, IL-6, IL-1, and other inflammatory cytokines, or cellular messengers. The suppression of these inflammatory cytokines inhibits their downstream pro-inflammatory signaling pathways12,13,14.
- Reduction of enzymes associated with fibrosis and tissue remodeling: Tissue remodeling and fibrosis can be caused by chronic inflammation due to deposition of collagen and other factors. In a variety of models and settings, suppression of ROS and inhibition of NF-κB has been observed to reduce the expression of enzymes associated with tissue remodeling that are implicated in the progression of PH, certain types of cancer, arthritis, and many other diseases5,12.
- Inhibition of cellular proliferative pathways: Inhibition of NF-kB and the NLRP3 inflammasome by Nrf2 activators inhibits a number of signaling pathways that promote hyperproliferation (harmful excessive growth) of cells. The antiproliferative effects of Nrf2 activators have been demonstrated in a variety of models of cancer and inflammatory or fibrotic diseases15,16,17.
Effects in Immuno-Oncology Settings
New agents that improve anti-tumor immune responses have recently shown significant clinical activity, and this approach shows great promise as a source of improved cancer treatment. Nrf2 activators are uniquely positioned to be useful in this emerging field, because they inhibit a fundamental mechanism (induction of antigen-specific T cell tolerance) used by tumors to evade immune surveillance. A pivotal series of studies has shown that certain immature white blood cells, known as myeloid-derived suppressor cells (MDSCs), play a critical role in the induction of T cell tolerance, and production of ROS/RNS by MDSCs is required for this process. High levels of ROS/RNS induce nitrosylation of tumor antigens, effectively “cloaking” these antigens from T cell recognition18. Nrf2 activators inhibit this process and restore anti-tumor immune responses in vivo19. Nrf2 activators also promote the maturation of dendritic cells, which are required for presentation of antigens to the immune system20. Reduced expression of inducible nitric oxide synthase (iNOS), an enzyme that plays a major role in the generation of ROS/RNS, has been documented in tumor biopsies from cancer patients treated with Nrf2 activators21. Elevated iNOS levels are implicated in MDSC-mediated immune evasion and are correlated with poor clinical outcomes in cancer patients22.
The immunomodulatory effects of Nrf2 activators have the potential to be complementary to other modes of cancer immunotherapy, positioning Nrf2 activators to be applicable in a wide array of cancer therapy settings. In animal studies, treatment with Nrf2 activators improved immune cell infiltration of tumors and enhanced the anti-tumor effects of a PD-1 inhibitor and a cancer vaccine19,21.
- Nrf2 activators inhibit cancer cell proliferation in vitro and in vivo, and promote apoptosis of tumor cells by reducing the expression of Bcl-XL and other anti-apoptotic proteins.
- Nrf2 activators inhibit inflammatory signaling pathways, reduce the expression of proteins that promote tissue invasion and metastasis, and potently inhibit tumor-related angiogenesis.
- An extensive body of literature has documented the ability of Nrf2 activators to prevent carcinogenesis and inhibit early tumor formation and growth. These effects have potential clinical relevance for the prevention of post-therapy recurrence and suppression of micrometastases.
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- Uruno et al. The Keap1-Nrf2 system prevents onset of diabetes mellitus. Mol Cell Biol. 2013 Aug;33(15):2996-3010.
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