Ferrostatin-1 - Ferroptosis inhibitor

CAS #347174-05-4 - InvitroFit™ - Cell culture-tested

Ferrostatin-1 chemical structure

Ferrostatin-1 (Fer-1) is a strong, synthetic inhibitor of ferroptosis, a form of non-apoptotic cell death triggered by lipid peroxidation. It is an active radical-scavenger antioxidant that traps lipid reactive oxygen species (ROS) and thus, serves as a potential inhibitor of ferroptosis [1].

 More details

Mode of action

Ferrostatin-1 functions by scavenging lipid peroxyl radicals and preventing oxidative membrane damage, thereby protecting cells from iron-dependent oxidative stress and subsequent ferroptotic cell death [1]. 

Upon stimulation of our Ferroptosis reporter HT-1080 cells using the ferroptosis activator RSL3, the cell membrane ruptures and the reporter protein HMGB1::Lucia is released in the extracellular milieu. Levels of HMGB1::Lucia in the supernatant can be readily monitored by measuring the light signal produced after the addition of QUANTI-Luc™ 4 Lucia/Gaussia, a Lucia® luciferase detection reagent. Ferrostatin-1 successfully inhibits RSL3-induced ferroptosis and HMGB1::Lucia release (see figure). Moreover, ferroptosis inhibition in this cell line can be assessed using the classic cytotoxic lactate dehydrogenase (LDH) assay (see figure). 

Key features

• Each lot is functionally tested and validated.
• The absence of endotoxin is determined using the EndotoxDetect™ assay.
• InvitroFit™ grade: each lot is highly pure (≥95%) and functionally tested

InvivoGen's products are for research use only, and not for human or veterinary use.

Figures

Ferrostatin-1 inhibits RSL3-induced HMGB1::Lucia release in a dose-dependent manner. HT1080-HMGB1-Lucia™ cells were incubated with increasing concentrations of Ferrostatin-1 (0.03 nM - 3 µM) for 30 min followed by the addition of the ferroptosis inducer RSL3 (1 µM final concentration). After 48 hours, the inhibition of cell death was quantified by measuring the levels of HMGB1::Lucia in the supernatant using the QUANTI-Luc™ detection reagent. Data are shown as (A) fold induction over non-induced cells or (B) percentage (%) activity (mean ± SEM).

Ferrostatin-1 inhibits RSL3-induced LDH release in a dose-dependent manner. HT1080-HMGB1-Lucia™ cells were incubated with increasing concentrations of Ferrostatin-1 (0.03 nM - 3 µM) for 30 min followed by the addition of the ferroptosis inducer RSL3 (1 µM final concentration). After 48 hours, the inhibition of cell death was quantified using the lactate dehydrogenase (LDH) assay. Data is shown as a percentage of cell death (mean ± SEM).

Specifications

Applications: Ferroptosis inhibition

Target: Ferroptosis

Synonym: Fer-1, Ethyl 3-amino-4-(cyclohexylamino) benzoate

CAS number: 347174-05-4

Working concentration: 10 nM - 10 µM

Solubility: DMSO (20 mg/ml)

Purity: ≥95%

Physical form: Dried powder

Chemical formula: C₁₅H₂₂N₂O₂ 

Molecular weight: 262.35 g/mol

Quality control: Each lot is functionally tested and validated.

Contents

Ferrostatin-1 is provided as a dried powder. 

• inh-fers1: 10 mg
   

Ferrostatin-1 is shipped at room temperature.

Upon receipt, store at -20°C for up to 3 years.

 Resuspended product is stable for up to 1 month at -20 °C or up to 1 year at -80 °C when properly stored.

 Avoid repeated freeze-thaw cycles. Protect from light.

Details

Ferroptosis inhibition by Ferrostatin-1

Ferroptosis overview

In 2012, Dixon et al. [1] discovered a novel form of non-apoptotic cell death characterized by iron overload and accumulation of lethal lipid peroxidation [2]. Because iron chelators were shown to block this type of regulated cell death, it was originally defined as iron-dependent and was called "Ferroptosis" [1]. In contrast to apoptosis, necrosis, and autophagy, ferroptosis is characterized by the build-up of lipid reactive oxygen species (ROS) triggering membrane damage. Cells undergoing ferroptosis do not exhibit classic apoptosis-like features, such as chromatin condensation or membrane blebbing. Instead, mitochondrial shrinkage and increased membrane density are often observed [1-2]. 

Ferroptosis is regulated by a complex network involving iron homeostasis, lipid metabolism, and glutathione-dependent oxidative-reductive balance [3-4]. Iron metabolism plays a central role, as excessive intracellular iron promotes the Fenton reaction, generating lethal amounts of ROS that drive lipid peroxidation [4]. Moreover, the enzyme glutathione peroxidase 4 (GPX4) is a key regulator that protects cells from ferroptosis by reducing lipid peroxides. GPX4 activity is dependent on Glutathione (GSH) which is synthesized using cystine imported via the cystine-glutamate antiporter, System Xc−. When System Xc− is inhibited, cystine uptake decreases, leading to GSH depletion and GPX4 inactivation. As a result, lipid peroxides accumulate in the cells ultimately leading to ferroptotic cell death [2-4]. 
 

Ferroptosis in disease

Ferroptosis is involved in various diseases, including neurodegenerative disorders, organ injury-related conditions, and cancer [2-3]. In neurodegenerative diseases, such as Parkinson's and Alzheimer's, ferroptosis contributes to neuronal loss through iron deposition in the brain and oxidative stress [3]. It has also been identified as a major pathogenic driver in other organ injury-related disorders including acute kidney injury and COVID-19-induced myocarditis [3]. Studies have shown that using ferroptosis-specific inhibitors can protect against severe tissue damage in these cases [3]. On the other hand, triggering ferroptosis is being explored as a potential therapeutic approach for treating therapy-resistant cancers [1-4]. Certain types of cancer, especially those with high iron levels or deficiencies in GPX4, are particularly sensitive to ferroptosis-inducing substances. Compounds like RSL3 or Erastin hold promise to overcome drug resistance in chemotherapy and immunotherapy [2]. Advancing our understanding of ferroptosis will continue to drive progress in cell death research and therapeutic development.

References

1. Dixon SJ, et al., 2012. Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell. 2012 May 25;149(5):1060-72.
2. Du Y, Guo Z. 2022. Recent progress in ferroptosis: inducers and inhibitors. Cell Death Discov. 8(1):501.
3. Sun S, et al., 2023. Targeting ferroptosis opens new avenues for the development of novel therapeutics. Signal Transduct Target Ther. 8(1):372.
4. Li J, et al., 2020. Ferroptosis: past, present and future. Cell Death Dis. 11(2):88.