Chemotherapeutic xCT inhibitors sorafenib and erastin unraveled with the synaptic optogenetic function analysis tool

Abstract

In the search for new potential chemotherapeutics, the compounds’ toxicity to healthy cells is an important factor. The brain with its functional units, the neurons, is especially endangered during the radio- and chemotherapeutic treatment of brain tumors. The effect of the potential compounds not only on neuronal survival but also neuronal function needs to be taken into account. Therefore, in this study we aimed to comprehend the biological effects of chemotherapeutic xCT inhibition on healthy neuronal cells with our synaptic optogenetic function analysis tool (SOFA). We combined common approaches, such as investigation of morphological markers, neuronal function and cell metabolism. The glutamate-cystine exchanger xCT (SLC7A11, system Xc−) is the main glutamate exporter in malignant brain tumors and as such a relevant drug target for treating deadly glioblastomas (WHO grades III and IV). Recently, two small molecules termed sorafenib (Nexavar) and erastin have been found to efficiently block xCT function. We investigated neuronal morphology, metabolic secretome profiles, synaptic function and cell metabolism of primary hippocampal cultures (containing neurons and glial cells) treated with sorafenib and erastin in clinically relevant concentrations. We found that sorafenib severely damaged neurons already after 24 h of treatment. Noteworthy, also at a lower concentration, where no morphological damage or metabolic disturbance was monitored, sorafenib still interfered with synaptic and metabolic homeostasis. In contrast, erastin-treated neurons displayed mostly inconspicuous morphology and metabolic rates. Key parameters of proper neuronal function, such as synaptic vesicle pool sizes, were however disrupted following erastin application. In conclusion, our data revealed that while sorafenib and erastin effectively inhibited xCT function they also interfered with essential neuronal (synaptic) function. These findings highlight the particular importance of investigating the effects of potential neurooncological and general cancer chemotherapeutics also on healthy neuronal cells and their function as revealed by the SOFA tool.

Cellular internalization of bystander nanomaterial induced by TAT-nanoparticles and regulated by extracellular cysteine

“…We utilized inhibitors to investigate whether Cys regulation depends on the transport of Cys or cystine into cells. After pre-incubation with glutamate or erastin (system Xc- inhibitors, which blocks the cellular uptake of Cys and cystine) 36,37 , CHO cells were incubated with T-Ag and AgNPs for internalization. The bystander uptake of AgNPs was significantly inhibited by erastin and glutamate, while these inhibitors had little effect on T-Ag uptake (Fig.…”

Primary allogeneic mitochondrial mix (PAMM) transfer/transplant by MitoCeption to address damage in PBMCs caused by ultraviolet radiation

“…The metabolic activity of PBMCs was measured with spectrophotometry using the MTT assay. Mitochondrial mass data was analyzed using flow cytometry and MTT assay for metabolic activity and subsequently normalized using sample/control-average transformation [17]. Finally, we stained PBMCs with Trypan Blue to determine the percentage of dead cells.…”

Genetic Ablation of the Cystine Transporter xCT in PDAC Cells Inhibits mTORC1, Growth, Survival, and Tumor Formation via Nutrient and Oxidative Stresses

“…Taking advantage of the cells with genetically deleted xCT, we tested the sensitivity and specificity of the widely accepted xCT inhibitor-erastin (4,9,10,29). WT and xCT-KO cells from both PDAC cell lines were treated with increasing inhibitor concentrations (from 0 to 5 mmol/L).…”