Project description:Cisplatin is a widely-used cancer metallodrug which induces cytotoxicity by targeting DNA and chelating cysteines in proteins. Here we applied a competitive activity-based protein profiling strategy to identify cisplatin-binding cysteines in cancer proteomes. A novel cisplatin target, MetAP1, was found and functionally validated to contribute to cisplatin’s cytotoxicity.
Project description:Here we report that piroxicam/cisplatin combined treatment exerts an apoptotic effect on mesothelioma cells. Genome-wide transcriptome analyses lead us to identify p21 as the possible apoptosis mediator acting as downstream target of the piroxicam/cisplatin treatment. To analyze the potential therapeutic effect of the piroxicam/cisplatin treatment at molecular level, and to identify gene expression pattern modifications following the combined treatment, we performed a transcriptional profiling on HGU133A arrays, using cells treated with piroxicam, cisplatin or with piroxicam and cisplatin, choosing the time exposures in which apoptosis induction was absent or evident (8 and 24 hours). Biological duplicates or triplicates were generated for each prototypic situation and data were analyzed using the oneChannelGUI Bioconductor package (Sanges et al., 2007), comparing untreated cells with cells treated in single or combined treatment. After quality controls, the complexity of the data set was reduced removing the non-significant probe sets, resulting in a total of 4,247 out of the 22,283 probe sets present in the microarray. To assess differential expression, linked to piroxicam, cisplatin- and the combined treatment we used an empirical Bayes method together with a false discovery rate (FDR) correction of the P-value. Specifically genes were selected using a corrected p-value M-bM-^IM-$0.05 and |log2(fc)| M-bM-^IM-%1. We detect a total of 536 differentially expressed genes.
Project description:Cisplatin is a common anticancer drug, but its frequent nephrotoxicity limits its clinical use. Small GTP-binding protein GDP dissociation stimulator (smgGDS), a small GTPase chaperone protein, was considerably downregulated during cisplatin-induced acute kidney injury (CDDP-AKI), especially in renal tubular epithelial cells. SmgGDS-knockdown mice was established and found that smgGDS knockdown promoted CDDP-AKI, as demonstrated by an increase in serum creatine, blood urea nitrogen levels and the appearance of tubular patterns. RNA sequencing suggested that protein kinase RNA-like ER kinase (PERK), which bridges mitochondria-associated ER membranes, was involved in smgGDS knockdown following CDDP-AKI, and then identified that smgGDS knockdown increased phosphorylated-PERK in vivo and in vitro. Furthermore, we confirmed that smgGDS deficiency aggravated apoptosis and ER stress in vivo and in vitro. And the ER stress inhibitor 4-Phenylbutyric acid and the inhibition of PERK phosphorylation mitigated smgGDS deficiency-induced ER stress related apoptosis following cisplatin treatment, while the eIF2α phosphorylation inhibitor could not reverse the smgGDS deficiency accelerated cell death. Furthermore, the over-expression of smgGDS could reverse the ER stress and apoptosis caused by CDDP. Overall, smgGDS regulated PERK-dependent ER stress and apoptosis, thereby influencing renal damage. This study identified a target for diagnosing and treating cisplatin-induced acute kidney injury.
Project description:The aim of this study is to select a cisplatin resistant Saccharomyces cerevisiae strain as a model organism to look for new molecular markers of cisplatin resistance and the identification of mechanisms/interactions involved. A cisplatin resistant S. cerevisiae strain was obtained after continuous exposure to the drug during 80 days. Then, total protein extraction, purification and identification were carried out, in wild type (wt) and resistant strains, by tandem mass spectrometry using a "nano HPLC-ESI-MS/MS" ion trap system. The increase in emPAI (resistant vs wt strains) was calculated to study the increase in protein expression. "Genemania" software (http://www.Genemania.org/) was used to compare the effects, functions and interactions of proteins. The selected cisplatin resistant strain showed 2.5 times more resistance than the wt strain (for the ID50 value) and 2.78 times more resistant for the ID90 value. The long-term exposure to cisplatin induced resistance in S. cerevisiae, obtaining an increased expression of QCR2, QCR1, ALDH4, ATPB, ATPA, SCW10, HSP26, ATPG, and PCKA proteins. The overexpression of the above-mentioned proteins suggests that they could be involved in cisplatin resistance. The resistance acquisition process is complex and involves the activation of multiple mechanisms that interact together.
Project description:Cisplatin is a widely used anti-tumor agent for the treatment of testicular and ovarian cancers. Carboplatin is used extensively for small cell, non small cell lung cancer and ovarian cancer. Oxaliplatin has recently been approved in the United States (US) for treatment of colorectal cancer. A large portion (in the range of 65% to 98%) of cisplatin in the blood plasma was bound to protein within a day after intravenous administration. The binding of cisplatin and other analogues to proteins and enzymes is generally believed to be the cause of several severe side effects such as ototoxicity and nephrotoxicity. The interactions between platinum based chemotherapy drugs and proteins is proposed to play important roles in both drug activity and toxicity. Therefore, a better understanding of the molecular mechanism of platinum-protein interactions may have an impact on optimization of strategies for treatment. The objective is to develop novel approaches and techniques to provide detailed mechanistic, kinetic and high-resolution structural information on the binding of platinum analogues to blood proteins, and to improve treatment efficacy and reduce side effects.
Project description:Cisplatin is used in chemotherapy of prostate, ovary and other cancer types but unfortunately the efficiency of cisplatin treatment is frequently hampered by acquired resistance. The cytotoxic effect of cisplatin has been attributed to its binding to DNA. HMGB1 is able to bind to cisplatin-DNA adducts with high affinity and its expression levels are associated with cisplatin resistance. This study reports the interactome of HMGB1 in prostate and ovarian cancer cells treated with cisplatin.
Project description:Cisplatin, one of the most widely used anticancer drugs, crosslinks DNA and ultimately induces cell death. However, the genomic pattern of cisplatin-DNA adducts remains unknown, due to the lack of a reliable and sensitive genome-wide method. Here we present “cisplatin-seq” to identify genome-wide cisplatin crosslinking sites at base-resolution. Cisplatin-seq reveals that mitochondrial DNA is a preferred target of cisplatin. For nuclear genome, cisplatin-DNA adducts are enriched within promoters and regions harboring transcription termination sites. While the density of GG dinucleotide determines the initial crosslinking of cisplatin, binding of proteins to the genome largely contributes to the accumulative pattern of cisplatin-DNA adducts.
Project description:Non-canonical DNA structures such as G-quadruplex (G4) and i-motif (iM) are formed at the guanine- and cytosine-rich sequences, respectively, and prohibit DNA replication and transcription. The formation and resolution of these non-canonical structures are therefore required to be dynamically regulated by either physiological conditions or factors able to bind the G4 and iM structures. Although many G4 binding proteins responsible for tuning of the G4 structure have been discovered, understanding of structural regulation of the iM structure by iM binding proteins is far less behind. Here, we developed a protein-labeling DNA probe bearing an alkyne moiety through a reactive tosylate linker for proximity labeling of nucleic acid-binding proteins and searched for iM binding proteins. The proteome analyses of the captured proteins suggested new candidates that potentially bind the iM structure, in addition to the known iM binders.