Background: Ailanthone (Aila) is a natural active compound isolated from the Ailanthus altissima,-inhibitory effect against several cancer cell lines. Advanced bladder cancer is a common disease characterized by a frequent onset of resistance to cisplatin-based therapy. The cisplatin (CDDP) resistance is accompanied by an increase in Nrf2 protein expression which contributes to conferring resistance. Recently, we demonstrated a crosstalk between Nrf2 and YAP. YAP has also been demonstrated to play an important role in chemoresistance of bladder cancer. Purpose: We analyzed the antitumor effect of Aila in sensitive and CDDP-resistant bladder cancer cells and the molecular mechanisms involved in Aila activity. Study design: Sensitive and CDDP-resistant 253J B-V and 253J bladder cancer cells, and intrinsically CDDP-resistant T24 bladder cancer cells were used. Cells were treated with diverse concentrations of Aila and proliferation, cell cycle, apoptosis and gene expressions were determined. Methods: Aila toxicity and proliferation were determined by MTT and colony forming methods, respectively. Cell cycle was determined at cytofluorimeter by PI staining method Apoptosis was detected using Annexin V and PI double staining followed by quantitative flow cytometry. Expressions of Nrf2, Yap, c-Myc, and housekeeping genes were determined by western blot with specific antibodies. Cell migration was detected by wound healing and Boyden chamber analysis. Results: Aila inhibited growth of sensitive and CDDP-resistant bladder cancer cells with the same effectiveness, and reduced cell migration with higher effectiveness in CDDP resistant cells. Interestingly, Aila strongly reduced Nrf2 expression in all cell lines. Cell cycle analysis revealed an accumulation of Aila-treated cells in G0/G1 phase. Moreover, Aila significantly reduced YAP and c-Myc protein expression. The random and the oriented migration were strongly inhibited by Aila treatment, in particular in CDDP-resistant cells. Conclusions: Aila, inhibited proliferation and invasiveness of bladder cancer cells. Its high effectiveness in CDDP resistant cells could be related to the inhibition of Nrf2 , YAP and c-Myc expressions. Aila could represent a new tool to overcome CDDP resistance in bladder cancer.

Chemoresistance represents the main obstacle to cancer treatment with both conventional and targeted therapy. Beyond specific molecular alterations, which can lead to targeted therapy, metabolic remodeling, including the control of redox status, plays an important role in cancer cell survival following therapy. Although cancer cells generally have a high basal reactive oxygen species (ROS) level, which makes them more susceptible than normal cells to a further increase of ROS, chemoresistant cancer cells become highly adapted to intrinsic or drug-induced oxidative stress by upregulating their antioxidant systems. The antioxidant response is principally mediated by the transcription factor Nrf2, which has been considered the master regulator of antioxidant and cytoprotective genes. Nrf2 expression is often increased in several types of chemoresistant cancer cells, and its expression is mediated by diverse mechanisms. In addition to Nrf2, other transcription factors and transcriptional coactivators can participate to maintain the high antioxidant levels in chemo and radio-resistant cancer cells. The control of expression and function of these molecules has been recently deepened to identify which of these could be used as a new therapeutic target in the treatment of tumors resistant to conventional therapy. In this review, we report the more recent advances in the study of Nrf2 regulation in chemoresistant cancers and the role played by other transcription factors and transcriptional coactivators in the control of antioxidant responses in chemoresistant cancer cells.

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Melanoma is a highly aggressive cancer with the poorest prognosis, representing the deadliest form of skin cancer. Activating mutations in BRAF are the most frequent genetic alterations, present in approximately 50% of all melanoma cases. The use of specific inhibitors towards mutant BRAF variants and MEK, a downstream signaling target of BRAF in the MAPK pathway, has significantly improved progression-free and overall survival in advanced melanoma patients carrying BRAF mutations. Nevertheless, despite these improvements, resistance still develops within the first year of therapy in around 50% of patients, which is a significant problem in managing BRAF-mutated advanced melanoma. Understanding these mechanisms is one of the mainstreams of the research on BRAFi/MEKi acquired resistance. Both genetic and epigenetic mechanisms have been described. Moreover, in recent years, oxidative stress has emerged as another major force involved in all the phases of melanoma development, from initiation to progression until the onsets of the metastatic phenotype and chemoresistance, and has thus become a target for therapy. In the present review, we discuss the current knowledge on oxidative stress and its signaling in melanoma, as well as the oxidative stress-related mechanisms in the acquired resistance to targeted therapies.

The transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) is considered as the master regulator of antioxidant and cytoprotective gene expressions. Moreover, it plays a pivotal role in cancer progression. Nrf2 mediates the adaptive response which contributes to the resistance to chemotherapeutic pro-oxidant drugs, such as cisplatin (CDDP), in various tumors, including bladder cancers. For this reason, Nrf2 could be a promising target to overcome chemoresistance. There are several known Nrf2 pharmacological inhibitors; however, most of them are not specific. The use of a specific small interfering RNA (siRNA) targeting the Nrf2 gene (siNrf2) loaded into nanovehicles is an attractive alternative, since it can increase specificity. This study aimed to evaluate the biological activity of siNrf2 loaded on guanidine-terminated carbosilane dendrimers (GCDs) in overcoming CDDP resistance in bladder cancer cells with a high level of Nrf2. Parameters such as viability, proliferation, apoptosis, migration, and oxidative stress level were taken into account. Results demonstrated that siNrf2-GCD treatment sensitized CDDP-resistant cells to CDDP treatment. Moreover, data obtained by treating the non-cancerous human kidney HK-2 cell line strongly suggest a good safety profile of the carbosilane dendrimers loaded with siNrf2. In conclusion, we suggest that siNrf2-GCD is a promising drug delivery system for gene therapy to be used in vivo; and it may represent an important tool in the therapy of CDDP-resistant cancer.