Celastrol Potentiates Radiotherapy by Impairment of DNA Damage Processing in Human Prostate Cancer

Abstract

Purpose Celastrol is an active ingredient of traditional herbal medicine and has recently been identified as a potent natural proteasome inhibitor. In this study, we evaluated the radiosensitizing potential of Celastrol in the human prostate cancer PC-3 model. Materials and Methods Clonogenic assays were performed to determine the radiosensitizing effect of Celastrol. Apoptosis was examined by flow cytometry using Annexin V and propidium iodide staining and by a caspase-3 activation assay. DNA damage processing was examined by immunofluorescent staining and Western blot for phosphorylated H2AX (γH2AX). PC-3 xenograft model in athymic nude mouse was exployed for in vivo efficacy of Celastrol in combination with radiation. The tumor samples were also analyzed for apoptosis and angiogenesis. Results Celastrol sensitized PC-3 cells to ionizing radiation (IR) in a dose- and schedule-dependent manner, where pretreatment of Celastrol for 1 h followed by IR achieved maximum radiosensitization. Celastrol significantly prolonged the presence of IR-induced γH2AX and increased IR-induced apoptosis. Celastrol combined with fractionated radiation significantly inhibited PC-3 tumor growth in vivo without obvious systemic toxicity. The combination treatment increased γH2AX levels and apoptosis, induced cleavage of poly(ADP-ribose)polymerase (PARP) and Mcl-1, as well as reduced angiogenesis in vivo, as compared with either treatment alone. Conclusions Celastrol sensitizes PC-3 cells to radiation both in vitro and in vivo, by impairing DNA damage processing as well as by augmenting apoptosis. Celastrol may represent a promising new adjuvant regime for the treatment of hormone-refractory prostate cancer.

ATF2 contributes to cisplatin resistance in non‐small cell lung cancer and celastrol induces cisplatin resensitization through inhibition of JNK/ATF2 pathway

“…1 middle panel) respectively, being Table 1). IC50 24 ranged from 264 to 87 mM, being H522 and H1395 the most resistant, while H596 and Calu1 showed IC50 24 values 2-3 times lower than H522. A correlation between ATF2 protein expression (both total -ATF2-and phosphorylated -pATF2-forms) and CDDP resistance was identified (Fig.…”

“…These data indicate that Celastrol-enhanced sensibilization of cells to Gemcitabine. In particular, in the H1395 cell line the Gemcitabine IC50 24 decreased from 190 mM, when used alone, to 0.84 mM when used in combination with Figure 4. Celastrol affects ATF2-responsive modulation and increases the effect mediated by platinum in resistant cell lines.…”

“…Technical quadruplicates were performed for each drug conditions and the experiment was replicated at least 2 times. The half maximal inhibitory concentration at 24h (IC50 24 ) is a measure of the effectiveness of a molecule/drug inhibiting a specific biological or biochemical function. In our analysis, this is the concentration of a single drug, or combinations of drugs, endowing viability reduction of 50% after 24h of drug exposure.…”

“…In our analysis, this is the concentration of a single drug, or combinations of drugs, endowing viability reduction of 50% after 24h of drug exposure. IC50 24 values were calculated by evaluating the viability data with Sigmoidal Dose Response analysis using GraphPad software and Combination Index (CI) calculated using mutually non-exclusive formula: RNA extraction, cDNA synthesis and qPCR:…”

“…15,20,21 In addition, it synergistically enhances the cytotoxicity of ionizing radiation, gambogic acid, TRAIL/APO-2L and temozolomide. [22][23][24] Although several hypothetical cellular targets for celastrol have been reported, its mechanism of action has not been fully elucidated. Celastrol affects several biological processes through the inhibition of ATF2, nuclear factor-KB, inducible NOS, VEGF receptors, HSP90, potassium/calcium channel and eventually increases the reactive oxygen species (ROS) in cancer cells, thus in turn activating apoptosis pathways.…”