Publications by authors named "Patrizia Limonta"

49 Publications

The multifaceted roles of mitochondria at the crossroads of cell life and death in cancer.

Free Radic Biol Med 2021 11 28;176:203-221. Epub 2021 Sep 28.

Department of Pharmacological and Biomolecular Sciences, Università Degli Studi di Milano, Milano, Italy. Electronic address:

Mitochondria are the cytoplasmic organelles mostly known as the "electric engine" of the cells; however, they also play pivotal roles in different biological processes, such as cell growth/apoptosis, Ca and redox homeostasis, and cell stemness. In cancer cells, mitochondria undergo peculiar functional and structural dynamics involved in the survival/death fate of the cell. Cancer cells use glycolysis to support macromolecular biosynthesis and energy production ("Warburg effect"); however, mitochondrial OXPHOS has been shown to be still active during carcinogenesis and even exacerbated in drug-resistant and stem cancer cells. This metabolic rewiring is associated with mutations in genes encoding mitochondrial metabolic enzymes ("oncometabolites"), alterations of ROS production and redox biology, and a fine-tuned balance between anti-/proapoptotic proteins. In cancer cells, mitochondria also experience dynamic alterations from the structural point of view undergoing coordinated cycles of biogenesis, fusion/fission and mitophagy, and physically communicating with the endoplasmic reticulum (ER), through the Ca flux, at the MAM (mitochondria-associated membranes) levels. This review addresses the peculiar mitochondrial metabolic and structural dynamics occurring in cancer cells and their role in coordinating the balance between cell survival and death. The role of mitochondrial dynamics as effective biomarkers of tumor progression and promising targets for anticancer strategies is also discussed.
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http://dx.doi.org/10.1016/j.freeradbiomed.2021.09.024DOI Listing
November 2021

δ-Tocotrienol sensitizes and re-sensitizes ovarian cancer cells to cisplatin via induction of G1 phase cell cycle arrest and ROS/MAPK-mediated apoptosis.

Cell Prolif 2021 Nov 14;54(11):e13111. Epub 2021 Sep 14.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milan, Italy.

Objectives: Among gynaecologic malignancies, ovarian cancer (OC) represents the leading cause of death for women worldwide. Current OC treatment involves cytoreductive surgery followed by platinum-based chemotherapy, which is associated with severe side effects and development of drug resistance. Therefore, new therapeutic strategies are urgently needed. Herein, we evaluated the anti-tumour effects of Vitamin E-derived δ-tocotrienol (δ-TT) in two human OC cell lines, IGROV-1 and SKOV-3 cells.

Materials And Methods: MTT and Trypan blue exclusion assays were used to assess δ-TT cytotoxicity, alone or in combination with other molecules. δ-TT effects on cell cycle, apoptosis, ROS generation and MAPK phosphorylation were investigated by flow cytometry, Western blot and immunofluorescence analyses. The synergism between δ-TT and chemotherapy was evaluated by isobologram analysis.

Results: We demonstrated that δ-TT could induce cell cycle block at G1-S phase and mitochondrial apoptosis in OC cell lines. In particular, we found that the proapoptotic activity of δ-TT correlated with mitochondrial ROS production and subsequent JNK and p38 activation. Finally, we observed that the compound was able to synergize with cisplatin, not only enhancing its cytotoxicity in IGROV-1 and SKOV-3 cells but also re-sensitizing IGROV-1/Pt1 cell line to its anti-tumour effects.

Conclusions: δ-TT triggers G1 phase cell cycle arrest and ROS/MAPK-mediated apoptosis in OC cells and sensitizes them to platinum treatment, thus representing an interesting option for novel chemopreventive/therapeutic strategies for OC.
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http://dx.doi.org/10.1111/cpr.13111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8560608PMC
November 2021

In Vitro 3D Cultures to Model the Tumor Microenvironment.

Cancers (Basel) 2021 Jun 13;13(12). Epub 2021 Jun 13.

Department of Pharmacological and Biomolecular Sciences, University of Milan, Via Balzaretti 9, 20133 Milan, Italy.

It is now well established that the tumor microenvironment plays a key role in determining cancer growth, metastasis and drug resistance. Thus, it is fundamental to understand how cancer cells interact and communicate with their stroma and how this crosstalk regulates disease initiation and progression. In this setting, 3D cell cultures have gained a lot of interest in the last two decades, due to their ability to better recapitulate the complexity of tumor microenvironment and therefore to bridge the gap between 2D monolayers and animal models. Herein, we present an overview of the 3D systems commonly used for studying tumor-stroma interactions, with a focus on recent advances in cancer modeling and drug discovery and testing.
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http://dx.doi.org/10.3390/cancers13122970DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8231786PMC
June 2021

Dissecting the Hormonal Signaling Landscape in Castration-Resistant Prostate Cancer.

Cells 2021 05 7;10(5). Epub 2021 May 7.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, 20133 Milano, Italy.

Understanding the molecular mechanisms underlying prostate cancer (PCa) progression towards its most aggressive, castration-resistant (CRPC) stage is urgently needed to improve the therapeutic options for this almost incurable pathology. Interestingly, CRPC is known to be characterized by a peculiar hormonal landscape. It is now well established that the androgen/androgen receptor (AR) axis is still active in CRPC cells. The persistent activity of this axis in PCa progression has been shown to be related to different mechanisms, such as intratumoral androgen synthesis, AR amplification and mutations, AR mRNA alternative splicing, increased expression/activity of AR-related transcription factors and coregulators. The hypothalamic gonadotropin-releasing hormone (GnRH), by binding to its specific receptors (GnRH-Rs) at the pituitary level, plays a pivotal role in the regulation of the reproductive functions. GnRH and GnRH-R are also expressed in different types of tumors, including PCa. Specifically, it has been demonstrated that, in CRPC cells, the activation of GnRH-Rs is associated with a significant antiproliferative/proapoptotic, antimetastatic and antiangiogenic activity. This antitumor activity is mainly mediated by the GnRH-R-associated Gαi/cAMP signaling pathway. In this review, we dissect the molecular mechanisms underlying the role of the androgen/AR and GnRH/GnRH-R axes in CRPC progression and the possible therapeutic implications.
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http://dx.doi.org/10.3390/cells10051133DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8151003PMC
May 2021

Ca overload- and ROS-associated mitochondrial dysfunction contributes to δ-tocotrienol-mediated paraptosis in melanoma cells.

Apoptosis 2021 06 3;26(5-6):277-292. Epub 2021 Apr 3.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milan, Italy.

Melanoma is an aggressive tumor with still poor therapy outcomes. δ-tocotrienol (δ-TT) is a vitamin E derivative displaying potent anti-cancer properties. Previously, we demonstrated that δ-TT triggers apoptosis in human melanoma cells. Here, we investigated whether it might also activate paraptosis, a non-canonical programmed cell death. In accordance with the main paraptotic features, δ-TT was shown to promote cytoplasmic vacuolization, associated with endoplasmic reticulum/mitochondrial dilation and protein synthesis, as well as MAPK activation in A375 and BLM cell lines. Moreover, treated cells exhibited a significant reduced expression of OXPHOS complex I and a marked decrease in oxygen consumption and mitochondrial membrane potential, culminating in decreased ATP synthesis and AMPK phosphorylation. This mitochondrial dysfunction resulted in ROS overproduction, found to be responsible for paraptosis induction. Additionally, δ-TT caused Ca homeostasis disruption, with endoplasmic reticulum-derived ions accumulating in mitochondria and activating the paraptotic signaling. Interestingly, by using both IP3R and VDAC inhibitors, a close cause-effect relationship between mitochondrial Ca overload and ROS generation was evidenced. Collectively, these results provide novel insights into δ-TT anti-melanoma activity, highlighting its ability to induce mitochondrial dysfunction-mediated paraptosis. δ-tocotrienol induces paraptotic cell death in human melanoma cells, causing endoplasmic reticulum dilation and mitochondrial swelling. These alterations induce an impairment of mitochondrial function, ROS production and calcium overload.
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http://dx.doi.org/10.1007/s10495-021-01668-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8197726PMC
June 2021

Cancer Stem Cells-Key Players in Tumor Relapse.

Cancers (Basel) 2021 Jan 20;13(3). Epub 2021 Jan 20.

Department of Pharmacological and Biomolecular Sciences, University of Milan, via Balzaretti 9, 20133 Milano, Italy.

Tumor relapse and treatment failure are unfortunately common events for cancer patients, thus often rendering cancer an uncurable disease. Cancer stem cells (CSCs) are a subset of cancer cells endowed with tumor-initiating and self-renewal capacity, as well as with high adaptive abilities. Altogether, these features contribute to CSC survival after one or multiple therapeutic approaches, thus leading to treatment failure and tumor progression/relapse. Thus, elucidating the molecular mechanisms associated with stemness-driven resistance is crucial for the development of more effective drugs and durable responses. This review will highlight the mechanisms exploited by CSCs to overcome different therapeutic strategies, from chemo- and radiotherapies to targeted therapies and immunotherapies, shedding light on their plasticity as an insidious trait responsible for their adaptation/escape. Finally, novel CSC-specific approaches will be described, providing evidence of their preclinical and clinical applications.
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http://dx.doi.org/10.3390/cancers13030376DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7864187PMC
January 2021

Gonadotropin-Releasing Hormone Receptors in Prostate Cancer: Molecular Aspects and Biological Functions.

Int J Mol Sci 2020 Dec 14;21(24). Epub 2020 Dec 14.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, via Balzaretti 9, 20133 Milan, Italy.

Pituitary Gonadotropin-Releasing Hormone receptors (GnRH-R) mediate the activity of the hypothalamic decapeptide GnRH, thus playing a key role in the regulation of the reproductive axis. Early-stage prostate cancer (PCa) is dependent on serum androgen levels, and androgen-deprivation therapy (ADT), based on GnRH agonists and antagonists, represents the standard therapeutic approach for PCa patients. Unfortunately, the tumor often progresses towards the more aggressive castration-resistant prostate cancer (CRPC) stage. GnRH receptors are also expressed in CRPC tissues, where their binding to both GnRH agonists and antagonists is associated with significant antiproliferative/proapoptotic, antimetastatic and antiangiogenic effects, mediated by the Gαi/cAMP signaling cascade. GnRH agonists and antagonists are now considered as an effective therapeutic strategy for CRPC patients with many clinical trials demonstrating that the combined use of these drugs with standard therapies (i.e., docetaxel, enzalutamide, abiraterone) significantly improves disease-free survival. In this context, GnRH-based bioconjugates (cytotoxic drugs covalently linked to a GnRH-based decapeptide) have been recently developed. The rationale of this treatment is that the GnRH peptide selectively binds to its receptors, delivering the cytotoxic drug to CRPC cells while sparing nontumor cells. Some of these compounds have already entered clinical trials.
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http://dx.doi.org/10.3390/ijms21249511DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7765031PMC
December 2020

Three-Dimensional Cell Cultures as an In Vitro Tool for Prostate Cancer Modeling and Drug Discovery.

Int J Mol Sci 2020 Sep 16;21(18). Epub 2020 Sep 16.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, via Balzaretti 9, 20133 Milan, Italy.

In the last decade, three-dimensional (3D) cell culture technology has gained a lot of interest due to its ability to better recapitulate the in vivo organization and microenvironment of in vitro cultured cancer cells. In particular, 3D tumor models have demonstrated several different characteristics compared with traditional two-dimensional (2D) cultures and have provided an interesting link between the latter and animal experiments. Indeed, 3D cell cultures represent a useful platform for the identification of the biological features of cancer cells as well as for the screening of novel antitumor agents. The present review is aimed at summarizing the most common 3D cell culture methods and applications, with a focus on prostate cancer modeling and drug discovery.
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http://dx.doi.org/10.3390/ijms21186806DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7554845PMC
September 2020

Mitochondrial functional and structural impairment is involved in the antitumor activity of δ-tocotrienol in prostate cancer cells.

Free Radic Biol Med 2020 11 29;160:376-390. Epub 2020 Jul 29.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milan, Italy. Electronic address:

The therapeutic options for castration-resistant prostate cancer (CRPC) are still limited. Natural bioactive compounds were shown to possess pro-death properties in different tumors. We previously reported that δ-tocotrienol (δ-TT) induces apoptosis, paraptosis and autophagy in CRPC cells. Here, we investigated whether δ-TT might exert its activity by impairing mitochondrial functions. We demonstrated that, in PC3 and DU145 cells, δ-TT impairs mitochondrial respiration and structural dynamics. In both cell lines, δ-TT triggers mitochondrial Ca and ROS overload. In PC3 cells, both Ca and ROS mediate the δ-TT-related anticancer activities (decrease of cell viability, apoptosis, paraptosis, autophagy and mitophagy). As expected, in autophagy-defective DU145 cells, Ca overload was involved in δ-TT-induced pro-death effects but not in autophagy and mitophagy. In this cell line, we also demonstrated that ROS overload is not involved in the anticancer activities of δ-TT, supporting a low susceptibility of these cells to ROS-related oxidative stress. Taken together, these data demonstrate that, in CRPC cells, δ-TT triggers cell death by inducing mitochondrial functional and structural impairments, providing novel mechanistic insights in its antitumor activity.
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http://dx.doi.org/10.1016/j.freeradbiomed.2020.07.009DOI Listing
November 2020

Natural Compounds in Prostate Cancer Prevention and Treatment: Mechanisms of Action and Molecular Targets.

Cells 2020 02 18;9(2). Epub 2020 Feb 18.

Department of Pharmacological and Biomolecular Sciences, University of Milano, 20122 Milano, Italy.

Prostate cancer (PCa) represents a major cause of cancer mortality among men in developed countries. Patients with recurrent disease initially respond to androgen-deprivation therapy, but the tumor eventually progresses into castration-resistant PCa; in this condition, tumor cells acquire the ability to escape cell death and develop resistance to current therapies. Thus, new therapeutic approaches for PCa management are urgently needed. In this setting, natural products have been extensively studied for their anti-PCa activities, such as tumor growth suppression, cell death induction, and inhibition of metastasis and angiogenesis. Additionally, numerous studies have shown that phytochemicals can specifically target the androgen receptor (AR) signaling, as well as the PCa stem cells (PCSCs). Interestingly, many clinical trials have been conducted to test the efficacy of nutraceuticals in human subjects, and they have partially confirmed the promising results obtained in vitro and in preclinical models. This article summarizes the anti-cancer mechanisms and therapeutic potentials of different natural compounds in the context of PCa prevention and treatment.
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http://dx.doi.org/10.3390/cells9020460DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7072821PMC
February 2020

The emerging role of paraptosis in tumor cell biology: Perspectives for cancer prevention and therapy with natural compounds.

Biochim Biophys Acta Rev Cancer 2020 04 3;1873(2):188338. Epub 2020 Jan 3.

Department of Pharmacological and Biomolecular Sciences, University of Milano, Milano, Italy. Electronic address:

Standard anti-cancer therapies promote tumor growth suppression mainly via induction of apoptosis. However, in most cases cancer cells acquire the ability to escape apoptotic cell death, thus becoming resistant to current treatments. In this setting, the interest in alternative cell death modes has recently increased. Paraptosis is a new form of programmed cell death displaying endoplasmic reticulum (ER) and/or mitochondria dilation, generally due to proteostasis disruption or redox and ion homeostasis alteration. Recent studies have highlighted that several natural compounds can trigger paraptosis in different tumor cell lines. Here, we review the molecular mechanisms underlying paraptotic cell death, as well as the natural products inducing this kind of cell death program. A better understanding of paraptosis should facilitate the development of new therapeutic strategies for cancer prevention and treatment.
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http://dx.doi.org/10.1016/j.bbcan.2020.188338DOI Listing
April 2020

New insights in melanoma biology: Running fast towards precision medicine.

Semin Cancer Biol 2019 12 25;59:161-164. Epub 2019 Sep 25.

Medical Oncology of Melanoma, Sarcoma and Rare Tumors, IEO, European Institute of Oncology IRCCS, Via Ripamonti 435, 20141 Milano, Italy. Electronic address:

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http://dx.doi.org/10.1016/j.semcancer.2019.09.019DOI Listing
December 2019

Heterologous and cross-species tropism of cancer-derived extracellular vesicles.

Theranostics 2019 9;9(19):5681-5693. Epub 2019 Aug 9.

Department of Oncology and Hemato-Oncology, Center of Excellence on Neurodegenerative Diseases, University of Milan, Milan, Italy.

Extracellular vesicles (EVs) are naturally occurring cargo delivery vesicles that have recently received considerable attention for their roles in intercellular communication in many physiological and pathological processes, including tumourigenesis. EVs generated by different tissues demonstrated specific homing: in particular, cancer-derived EVs showed a selective tropism for the tumor tissue from which the vesicles originated. For this property, EVs have been proposed as drug delivery tools for anti-cancer therapies, although the limited knowledge about their tropism hinders their therapeutic applications. The current study aimed to characterize the targeting properties of cancer-derived EVs and their biodistribution , by using an imaging approach. : EVs were generated from: i) murine lung (LL/2) and colon (MC-38) cancer lines, ii) human lung cancer cell line (A549) and iii) human liver biopsy samples from healthy individuals. EVs were loaded with fluorescent dyes alone or in combination with a biopharmaceutical agent, the oncolytic adenovirus (OV), characterized for charge and size and tested for their activity in cancer cell lines. Finally, optical imaging was extensively applied to study and the biodistribution of EVs originated from different sources in different mouse models of cancer, including xenograft, syngeneic graft and the MMTV-NeuT genetically modified animal. : We initially demonstrated that even loading EVs even with a large biopharmaceutical oncolytic viruses (OVs) did not significantly change their charge and dimension properties, while increasing their anti-neoplastic activity compared to the virus or EVs alone. Interestingly, this activity was observed even if the EVs derived from lung cancer were applied to colon carcinoma cell lines and , suggesting that the EV uptake occurred without any specificity for the cancer cells from which the vesicles originated. When administered i.v (intravenously) to the mouse models of cancer, the tumour-derived EVs, but not the EVs derived from a healthy tissue, demonstrated a selective accumulation of the fluorescence at the tumour site 24 h after injection; adding OVs to the formulation did not change the tumour-specific tropism of the EVs also . Most interestingly, the experiments confirmed the observation of the generalized tropism of tumour-derived EVs for any neoplastic tissue, independent of the tumour type or even the species originating the vesicles. : Taken together, our and data demonstrate for the first time a heterologous, cross-species tumour-tropism for cancer-derived EVs. This finding challenges our current view on the homing properties of EVs and opens new avenues for the selective delivery of diagnostic/therapeutic agents to solid tumours.
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http://dx.doi.org/10.7150/thno.34824DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6735396PMC
August 2020

Beneficial effects of δ-tocotrienol against oxidative stress in osteoblastic cells: studies on the mechanisms of action.

Eur J Nutr 2020 Aug 6;59(5):1975-1987. Epub 2019 Jul 6.

Department of Medical Biotechnology and Translational Medicine, Università degli Studi di Milano, Via Vanvitelli, 32, 20129, Milan, Italy.

Purpose: Natural antioxidants are considered as promising compounds in the prevention/treatment of osteoporosis. We studied the ability of purified δ-tocotrienol (δ-TT) isolated from a commercial palm oil (Elaeis guineensis) fraction to protect osteoblast MC3T3-E1 and osteocyte MLO-Y4 cells against tert-butyl hydroperoxide (t-BHP)-induced oxidative damage and the mechanisms involved in its protective action in MC3T3-E1.

Methods: MC3T3-E1 and MLO-Y4 cells were treated with δ-TT (1.25-20 µg/ml for 2 h) followed by t-BHP at 250 µM or 125 µM for 3 h, respectively. MTT test was used to measure cell viability. Apoptotic cells were stained with Hoechst-33258 dye. Intracellular ROS levels were measured by dichlorofluorescein CM-DCFA. The OPT fluorimetric assay was used to detect the reduced glutathione to oxidized glutathione ratio (GSH/GSSG) contents.

Results: δ-TT significantly prevented the effects of t-BHP on cell viability and apoptosis reaching a maximum protective activity at 10 and 5 µg/ml in MC3T3-E1 and MLO-Y4 cells, respectively. This protective effect was due to a reduction of intracellular ROS levels and an increase in the defense systems shown by the increase in the GSH/GSSG. GSH loss induced by an inhibitor of GSH synthesis significantly reduced the δ-TT-positive effect on ROS levels. δ-TT prevention of oxidative damage was completely removed by combined treatment with the specific inhibitors of PI3K/AKT (LY294002) and Nrf2 (ML385).

Conclusions: The δ-TT protective effect against oxidative damage in MC3T3-E1 cells is due to a reduction of intracellular ROS levels and an increase of the GSH/GSSG ratio, and involves an interaction between the PI3K/Akt-Nrf2 signaling pathways.
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http://dx.doi.org/10.1007/s00394-019-02047-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7351870PMC
August 2020

Cellular and molecular biology of cancer stem cells in melanoma: Possible therapeutic implications.

Semin Cancer Biol 2019 12 29;59:221-235. Epub 2019 Jun 29.

Department of Pharmacological and Biomolecular Sciences, University of Milano, Milano, Italy. Electronic address:

Malignant melanoma is a tumor characterized by a very high level of heterogeneity, responsible for its malignant behavior and ability to escape from standard therapies. In this review we highlight the molecular and biological features of the subpopulation of cancer stem cells (CSCs), well known to be characterized by self-renewal properties, deeply involved in triggering the processes of tumor generation, metastasis, progression and drug resistance. From the molecular point of view, melanoma CSCs are identified and characterized by the expression of stemness markers, such as surface markers, ATP-binding cassette (ABC) transporters, embryonic stem cells and intracellular markers. These cells are endowed with different functional features. In particular, they play pivotal roles in the processes of tumor dissemination, epithelial-to-mesenchymal transition (EMT) and angiogenesis, mediated by specific intracellular signaling pathways; moreover, they are characterized by a unique metabolic reprogramming. As reported for other types of tumors, the CSCs subpopulation in melanoma is also characterized by a low immunogenic profile as well as by the ability to escape the immune system, through the expression of a negative modulation of T cell functions and the secretion of immunosuppressive factors. These biological features allow melanoma CSCs to escape standard treatments, thus being deeply involved in tumor relapse. Targeting the CSCs subpopulation is now considered an attractive treatment strategy; in particular, combination treatments, based on both CSCs-targeting and standard drugs, will likely increase the therapeutic options for melanoma patients. The characterization of CSCs in liquid biopsies from single patients will pave the way towards precision medicine.
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http://dx.doi.org/10.1016/j.semcancer.2019.06.019DOI Listing
December 2019

Unraveling the molecular mechanisms and the potential chemopreventive/therapeutic properties of natural compounds in melanoma.

Semin Cancer Biol 2019 12 22;59:266-282. Epub 2019 Jun 22.

Department of Pharmacological and Biomolecular Sciences, University of Milano, Milano, Italy. Electronic address:

Melanoma is the most fatal form of skin cancer. Current therapeutic approaches include surgical resection, chemotherapy, targeted therapy and immunotherapy. However, these treatment strategies are associated with development of drug resistance and severe side effects. In recent years, natural compounds have also been extensively studied for their anti-melanoma effects, including tumor growth inhibition, apoptosis induction, angiogenesis and metastasis suppression and cancer stem cell elimination. Moreover, a considerable number of studies reported the synergistic activity of phytochemicals and standard anti-melanoma agents, as well as the enhanced effectiveness of their synthetic derivatives and novel formulations. However, clinical data confirming these promising effects in patients are still scanty. This review emphasizes the anti-tumor mechanisms and potential application of the most studied natural products for melanoma prevention and treatment.
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http://dx.doi.org/10.1016/j.semcancer.2019.06.011DOI Listing
December 2019

Role of Endoplasmic Reticulum Stress in the Anticancer Activity of Natural Compounds.

Int J Mol Sci 2019 Feb 22;20(4). Epub 2019 Feb 22.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, 20133 Milano, Italy.

Cancer represents a serious global health problem, and its incidence and mortality are rapidly growing worldwide. One of the main causes of the failure of an anticancer treatment is the development of drug resistance by cancer cells. Therefore, it is necessary to develop new drugs characterized by better pharmacological and toxicological profiles. Natural compounds can represent an optimal collection of bioactive molecules. Many natural compounds have been proven to possess anticancer effects in different types of tumors, but often the molecular mechanisms associated with their cytotoxicity are not completely understood. The endoplasmic reticulum (ER) is an organelle involved in multiple cellular processes. Alteration of ER homeostasis and its appropriate functioning originates a cascade of signaling events known as ER stress response or unfolded protein response (UPR). The UPR pathways involve three different sensors (protein kinase RNA(PKR)-like ER kinase (PERK), inositol requiring enzyme1α (IRE1) and activating transcription factor 6 (ATF6)) residing on the ER membranes. Although the main purpose of UPR is to restore this organelle's homeostasis, a persistent UPR can trigger cell death pathways such as apoptosis. There is a growing body of evidence showing that ER stress may play a role in the cytotoxicity of many natural compounds. In this review we present an overview of different plant-derived natural compounds, such as curcumin, resveratrol, green tea polyphenols, tocotrienols, and garcinia derivates, that exert their anticancer activity via ER stress modulation in different human cancers.
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http://dx.doi.org/10.3390/ijms20040961DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6412802PMC
February 2019

Epithelial-To-Mesenchymal Transition Markers and CD44 Isoforms Are Differently Expressed in 2D and 3D Cell Cultures of Prostate Cancer Cells.

Cells 2019 02 11;8(2). Epub 2019 Feb 11.

Department of Biomedical Sciences for Health, Università degli Studi di Milano, via Mangiagalli 31, 20133 Milan, Italy.

Three-dimensional (3D) cell cultures allow the mimic of functions of living tissues andprovide key information encoded in tissue architecture. Considered the pivotal role of epithelial-tomesenchymaltransition (EMT) in carcinoma progression, including prostate cancer (PCa), weaimed at investigating the effect of the 3D arrangement on the expression of some key markers ofEMT in cultured human prostate cancer (PCa) cells, to better understand PCa cell behavior. PC3 andDU145 PCa cells were cultured in RPMI cell culture medium either in 2D-monolayers or in 3Dspheroids.The main EMT markers E-cadherin, N-cadherin, α-smooth muscle actin (αSMA),vimentin, Snail, Slug, Twist and Zeb1 were evaluated by confocal microscopy, real-time PCR andWestern blot. Confocal microscopy revealed that E-cadherin was similarly expressed at the cellboundaries on the plasma membrane of PCa cells grown in 2D-monolayers, as well as in 3Dspheroids,but resulted up-regulated in 3D-spheroids, compared to 2D-monolayers, at the mRNAand protein level. Moreover, markers of the mesenchymal phenotype were expressed at very lowlevels in 3D-spheroids, suggesting important differences in the phenotype of PCa cells grown in 3Dspheroidsor in 2D-monolayers. Considered as a whole, our findings contribute to a clarification ofthe role of EMT in PCa and confirm that a 3D cell culture model could provide deeper insight intothe understanding of the biology of PCa.
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http://dx.doi.org/10.3390/cells8020143DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6406374PMC
February 2019

δ-Tocotrienol induces apoptosis, involving endoplasmic reticulum stress and autophagy, and paraptosis in prostate cancer cells.

Cell Prolif 2019 May 4;52(3):e12576. Epub 2019 Feb 4.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, Italy.

Objectives: Prostate cancer, after the phase of androgen dependence, may progress to the castration-resistant prostate cancer (CRPC) stage, with resistance to standard therapies. Vitamin E-derived tocotrienols (TTs) possess a significant antitumour activity. Here, we evaluated the anti-cancer properties of δ-TT in CRPC cells (PC3 and DU145) and the related mechanisms of action.

Materials And Methods: MTT, Trypan blue and colony formation assays were used to assess cell viability/cell death/cytotoxicity. Western blot, immunofluorescence and MTT analyses were utilized to investigate apoptosis, ER stress and autophagy. Morphological changes were investigated by light and transmission electron microscopy.

Results: We demonstrated that δ-TT exerts a cytotoxic/proapoptotic activity in CRPC cells. We found that in PC3 cells: (a) δ-TT triggers both the endoplasmic reticulum (ER) stress and autophagy pathways; (b) autophagy induction is related to the ER stress, and this ER stress/autophagy axis is involved in the antitumour activity of δ-TT; in autophagy-defective DU145 cells, only the ER stress pathway is involved in the proapoptotic effects of δ-TT; (c) in both CRPC cell lines, δ-TT also induces an intense vacuolation prevented by the ER stress inhibitor salubrinal and the protein synthesis inhibitor cycloheximide, together with increased levels of phosphorylated JNK and p38, supporting the induction of paraptosis by δ-TT.

Conclusions: These data demonstrate that apoptosis, involving ER stress and autophagy (in autophagy positive PC3 cells), and paraptosis are involved in the anti-cancer activity of δ-TT in CRPC cells.
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http://dx.doi.org/10.1111/cpr.12576DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6536411PMC
May 2019

Tocotrienols and Cancer: From the State of the Art to Promising Novel Patents.

Recent Pat Anticancer Drug Discov 2019 ;14(1):5-18

Department of Excellence, Department of Pharmacological and Biomolecular Sciences, University of Milan, Milan, Italy.

Background: Tocotrienols (TTs) are vitamin E derivatives naturally occurring in several plants and vegetable oils. Like Tocopherols (TPs), they comprise four isoforms, α, β, γ and δ, but unlike TPs, they present an unsaturated isoprenoid chain. Recent studies indicate that TTs provide important health benefits, including neuroprotective, anti-inflammatory, anti-oxidant, cholesterol lowering and immunomodulatory effects. Moreover, they have been found to possess unique anti-cancer properties.

Objective: The purpose of this review is to present an overview of the state of the art of TTs role in cancer prevention and treatment, as well as to describe recent patents proposing new methods for TTs isolation, chemical modification and use in cancer prevention and/or therapy.

Methods: Recent literature and patents focusing on TTs anti-cancer applications have been identified and reviewed, with special regard to their scientific impact and novelty.

Results: TTs have demonstrated significant anti-cancer activity in multiple tumor types, both in vitro and in vivo. Furthermore, they have shown synergistic effects when given in combination with standard anti-cancer agents or other anti-tumor natural compounds. Finally, new purification processes and transgenic sources have been designed in order to improve TTs production, and novel TTs formulations and synthetic derivatives have been developed to enhance their solubility and bioavailability.

Conclusion: The promising anti-cancer effects shown by TTs in several preclinical studies may open new opportunities for therapeutic interventions in different tumors. Thus, clinical trials aimed at confirming TTs chemopreventive and tumor-suppressing activity, particularly in combination with standard therapies, are urgently needed.
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http://dx.doi.org/10.2174/1574892814666190116111827DOI Listing
December 2019

Dual role of autophagy on docetaxel-sensitivity in prostate cancer cells.

Cell Death Dis 2018 08 30;9(9):889. Epub 2018 Aug 30.

Department of Excellence: Dipartimento di Scienze Farmacologiche e Biomolecolari (DiSFeB), Università degli Studi di Milano, Milano, Italy.

Prostate cancer (PC) is one of the leading causes of death in males. Available treatments often lead to the appearance of chemoresistant foci and metastases, with mechanisms still partially unknown. Within tumour mass, autophagy may promote cell survival by enhancing cancer cells tolerability to different cell stresses, like hypoxia, starvation or those triggered by chemotherapic agents. Because of its connection with the apoptotic pathways, autophagy has been differentially implicated, either as prodeath or prosurvival factor, in the appearance of more aggressive tumours. Here, in three PC cells (LNCaP, PC3, and DU145), we tested how different autophagy inducers modulate docetaxel-induced apoptosis. We selected the mTOR-independent disaccharide trehalose and the mTOR-dependent macrolide lactone rapamycin autophagy inducers. In castration-resistant PC (CRPC) PC3 cells, trehalose specifically prevented intrinsic apoptosis in docetaxel-treated cells. Trehalose reduced the release of cytochrome c triggered by docetaxel and the formation of aberrant mitochondria, possibly by enhancing the turnover of damaged mitochondria via autophagy (mitophagy). In fact, trehalose increased LC3 and p62 expression, LC3-II and p62 (p62 bodies) accumulation and the induction of LC3 puncta. In docetaxel-treated cells, trehalose, but not rapamycin, determined a perinuclear mitochondrial aggregation (mito-aggresomes), and mitochondria specifically colocalized with LC3 and p62-positive autophagosomes. In PC3 cells, rapamycin retained its ability to activate autophagy without evidences of mitophagy even in presence of docetaxel. Interestingly, these results were replicated in LNCaP cells, whereas trehalose and rapamycin did not modify the response to docetaxel in the ATG5-deficient (autophagy resistant) DU145 cells. Therefore, autophagy is involved to alter the response to chemotherapy in combination therapies and the response may be influenced by the different autophagic pathways utilized and by the type of cancer cells.
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http://dx.doi.org/10.1038/s41419-018-0866-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6117300PMC
August 2018

Anticancer properties of tocotrienols: A review of cellular mechanisms and molecular targets.

J Cell Physiol 2019 02 1;234(2):1147-1164. Epub 2018 Aug 1.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, Italy.

Vitamin E is composed of two groups of compounds: α-, β-, γ-, and δ-tocopherols (TPs), and the corresponding unsaturated tocotrienols (TTs). TTs are found in natural sources such as red palm oil, annatto seeds, and rice bran. In the last decades, TTs (specifically, γ-TT and δ-TT) have gained interest due to their health benefits in chronic diseases, based on their antioxidant, neuroprotective, cholesterol-lowering, anti-inflammatory activities. Several in vitro and in vivo studies pointed out that TTs also exert a significant antitumor activity in a wide range of cancer cells. Specifically, TTs were shown to exert antiproliferative/proapoptotic effects and to reduce the metastatic or angiogenic properties of different cancer cells; moreover, these compounds were reported to specifically target the subpopulation of cancer stem cells, known to be deeply involved in the development of resistance to standard therapies. Interestingly, recent studies pointed out that TTs exert a synergistic antitumor effect on cancer cells when given in combination with either standard antitumor agents (i.e., chemotherapeutics, statins, "targeted" therapies) or natural compounds with anticancer activity (i.e., sesamin, epigallocatechin gallate (EGCG), resveratrol, ferulic acid). Based on these observations, different TT synthetic derivatives and formulations were recently developed and demonstrated to improve TT water solubility and to reduce TT metabolism in cancer cells, thus increasing their biological activity. These promising results, together with the safety of TT administration in healthy subjects, suggest that these compounds might represent a new chemopreventive or anticancer treatment (i.e., in combination with standard therapies) strategy. Clinical trials aimed at confirming this antitumor activity of TTs are needed.
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http://dx.doi.org/10.1002/jcp.27075DOI Listing
February 2019

GnRH in the Human Female Reproductive Axis.

Vitam Horm 2018 12;107:27-66. Epub 2018 Feb 12.

Università degli Studi di Milano, Milano, Italy. Electronic address:

Gonadotropin-releasing hormone (GnRH) is recognized as the central regulator of the functions of the pituitary-gonadal axis. The increasing knowledge on the mechanisms controlling the development and the function of GnRH-producing neurons is leading to a better diagnostic and therapeutic approach for hypogonadotropic hypogonadisms and for alterations of the puberty onset. During female life span, the function of the GnRH pulse generator may be affected by a number of inputs from other neuronal systems, offering alternative strategies for diagnostic and therapeutic interventions. Moreover, the identification of a GnRH/GnRH receptor system in both human ovary and endometrium has widened the spectrum of action of the peptide outside its hypothalamic functions. The pharmacological use of GnRH itself or its synthetic analogs (agonists and antagonists) provides a valid tool to either stimulate or block gonadotropin secretion and to modulate the female fertility in several reproductive disorders and in assisted reproduction technology. The use of GnRH agonists in young female patients undergoing chemotherapy is also considered a promising therapeutic approach to counteract iatrogenic ovarian failure.
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http://dx.doi.org/10.1016/bs.vh.2018.01.003DOI Listing
November 2018

Targeting melanoma stem cells with the Vitamin E derivative δ-tocotrienol.

Sci Rep 2018 01 12;8(1):587. Epub 2018 Jan 12.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, 20133, Italy.

The prognosis of metastatic melanoma is very poor, due to the development of drug resistance. Cancer stem cells (CSCs) may play a crucial role in this mechanism, contributing to disease relapse. We first characterized CSCs in melanoma cell lines. We observed that A375 (but not BLM) cells are able to form melanospheres and show CSCs traits: expression of the pluripotency markers SOX2 and KLF4, higher invasiveness and tumor formation capability in vivo with respect to parental adherent cells. We also showed that a subpopulation of autofluorescent cells expressing the ABCG2 stem cell marker is present in the A375 spheroid culture. Based on these data, we investigated whether δ-TT might target melanoma CSCs. We demonstrated that melanoma cells escaping the antitumor activity of δ-TT are completely devoid of the ability to form melanospheres. In contrast, cells that escaped vemurafenib treatment show a higher ability to form melanospheres than control cells. δ-TT also induced disaggregation of A375 melanospheres and reduced the spheroidogenic ability of sphere-derived cells, reducing the expression of the ABCG2 marker. These data demonstrate that δ-TT exerts its antitumor activity by targeting the CSC subpopulation of A375 melanoma cells and might represent a novel chemopreventive/therapeutic strategy against melanoma.
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http://dx.doi.org/10.1038/s41598-017-19057-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5766483PMC
January 2018

Semi-preparative HPLC purification of δ-tocotrienol (δ-T3) from Elaeis guineensis Jacq. and Bixa orellana L. and evaluation of its in vitro anticancer activity in human A375 melanoma cells.

Nat Prod Res 2018 May 24;32(10):1130-1135. Epub 2017 Apr 24.

b Department of Pharmacological and Biomolecular Sciences , Università degli Studi di Milano , Milano , Italy.

In this work, we report a rapid and convenient HPLC-UV-DAD method for the isolation of δ-T3 on semi-preparative scale from two different vitamin E rich processed, commercially available products obtained from the fruits of Elaeis guineensis Jacq. (oil palm) and from the seeds of Bixa orellana L. (achiote tree). Chromatography was run using reverse phase (RP) C-18 columns and HPLC-grade acetonitrile as mobile phase. The purity of the isolated δ-T3, assessed by GC-MS and H NMR was above 98%. The δ-T3 cytotoxic activity found in vitro against the proliferation of human A375 melanoma cells compared to that of the other δ-T3 free tocols mixture suggest its primary role in the experimental anticancer activity observed for palm oil derived products. Taken altogether, the results of this study highlight the importance of the application of suitable purification systems for the preparations of tocotrienols prior to their experimental or clinical testing.
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http://dx.doi.org/10.1080/14786419.2017.1320793DOI Listing
May 2018

Estrogen Receptor β in Melanoma: From Molecular Insights to Potential Clinical Utility.

Front Endocrinol (Lausanne) 2016 26;7:140. Epub 2016 Oct 26.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano , Milano , Italy.

Cutaneous melanoma is an aggressive tumor; its incidence has been reported to increase fast in the past decades. Melanoma is a heterogeneous tumor, with most patients harboring mutations in the BRAF or NRAS oncogenes, leading to the overactivation of the MAPK/ERK and PI3K/Akt pathways. The current therapeutic approaches are based on therapies targeting mutated BRAF and the downstream pathway, and on monoclonal antibodies against the immune checkpoint blockade. However, treatment resistance and side effects are common events of these therapeutic strategies. Increasing evidence supports that melanoma is a hormone-related cancer. Melanoma incidence is higher in males than in females, and females have a significant survival advantage over men. Estrogens exert their effects through estrogen receptors (ERα and ERβ) that affect cancer growth in an opposite way: ERα is associated with a proliferative action and ERβ with an anticancer effect. ERβ is the predominant ER in melanoma, and its expression decreases in melanoma progression, supporting its role as a tumor suppressor. Thus, ERβ is now considered as an effective molecular target for melanoma treatment. 17β-estradiol was reported to inhibit melanoma cells proliferation; however, clinical trials did not provide the expected survival benefits. studies demonstrate that ERβ ligands inhibit the proliferation of melanoma cells harboring the NRAS (but not the BRAF) mutation, suggesting that ERβ activation might impair melanoma development through the inhibition of the PI3K/Akt pathway. These data suggest that ERβ agonists might be considered as an effective treatment strategy, in combination with MAPK inhibitors, for NRAS mutant melanomas. In an era of personalized medicine, pretreatment evaluation of the expression of ER isoforms together with the concurrent oncogenic mutations should be considered before selecting the most appropriate therapeutic intervention. Natural compounds that specifically bind to ERβ have been identified. These phytoestrogens decrease the proliferation of melanoma cells. Importantly, these effects are unrelated to the oncogenic mutations of melanomas, suggesting that, in addition to their ERβ activating function, these compounds might impair melanoma development through additional mechanisms. A better identification of the role of ERβ in melanoma development will help increase the therapeutic options for this aggressive pathology.
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http://dx.doi.org/10.3389/fendo.2016.00140DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5080294PMC
October 2016

Vitamin E δ-tocotrienol triggers endoplasmic reticulum stress-mediated apoptosis in human melanoma cells.

Sci Rep 2016 07 27;6:30502. Epub 2016 Jul 27.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, 20133, Italy.

Malignant melanoma is the leading cause of death from skin cancer. Drug toxicity and resistance represent a serious challange for melanoma treatments. Evidence demonstrates that natural compounds may play a crucial role in cancer prevention, growth and progression. Vitamin E tocotrienols (TT) were shown to possess antitumor activity. Here, we analyzed the effects of δ-TT on melanoma cell growth and the involvement of the endoplasmic reticulum (ER) stress in this activity. The experiments were performed on human melanoma cell lines, BLM and A375. δ-TT exerted a significant proapoptotic effect on both cell lines, involving the intrinsic apoptosis pathway; importantly, this compound did not affect the viability of normal human melanocytes. In melanoma cells, δ-TT exerted its antitumor effect through activation of the PERK/p-eIF2α/ATF4/CHOP, IRE1α and caspase-4 ER stress-related branches. Salubrinal, an inhibitor of the ER stress, counteracted the cytotoxic activity of δ-TT. In vivo experiments performed in nude mice bearing A375 xenografts evidenced that δ-TT reduces tumor volume and tumor mass; importantly, tumor progression was significantly delayed by δ-TT treatment. In conclusion, δ-TT exerts a proapoptotic activity on melanoma cells, through activation of the ER stress-related pathways. δ-TT might represent an effective option for novel chemopreventive/therapeutic strategies for melanoma.
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http://dx.doi.org/10.1038/srep30502DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4996065PMC
July 2016

GnRH and GnRH receptors in the pathophysiology of the human female reproductive system.

Hum Reprod Update 2016 04 29;22(3):358-81. Epub 2015 Dec 29.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Via G. Balzaretti, 9, 20133 Milano, Italy.

Background: Human reproduction depends on an intact hypothalamic-pituitary-gonadal (HPG) axis. Hypothalamic gonadotrophin-releasing hormone (GnRH) has been recognized, since its identification in 1971, as the central regulator of the production and release of the pituitary gonadotrophins that, in turn, regulate the gonadal functions and the production of sex steroids. The characteristic peculiar development, distribution and episodic activity of GnRH-producing neurons have solicited an interdisciplinary interest on the etiopathogenesis of several reproductive diseases. The more recent identification of a GnRH/GnRH receptor (GnRHR) system in both the human endometrium and ovary has widened the spectrum of action of the peptide and of its analogues beyond its hypothalamic function.

Methods: An analysis of research and review articles published in international journals until June 2015 has been carried out to comprehensively summarize both the well established and the most recent knowledge on the physiopathology of the GnRH system in the central and peripheral control of female reproductive functions and diseases.

Results: This review focuses on the role of GnRH neurons in the control of the reproductive axis. New knowledge is accumulating on the genetic programme that drives GnRH neuron development to ameliorate the diagnosis and treatment of GnRH deficiency and consequent delayed or absent puberty. Moreover, a better understanding of the mechanisms controlling the episodic release of GnRH during the onset of puberty and the ovulatory cycle has enabled the pharmacological use of GnRH itself or its synthetic analogues (agonists and antagonists) to either stimulate or to block the gonadotrophin secretion and modulate the functions of the reproductive axis in several reproductive diseases and in assisted reproduction technology. Several inputs from other neuronal populations, as well as metabolic, somatic and age-related signals, may greatly affect the functions of the GnRH pulse generator during the female lifespan; their modulation may offer new possible strategies for diagnostic and therapeutic interventions. A GnRH/GnRHR system is also expressed in female reproductive tissues (e.g. endometrium and ovary), both in normal and pathological conditions. The expression of this system in the human endometrium and ovary supports its physiological regulatory role in the processes of trophoblast invasion of the maternal endometrium and embryo implantation as well as of follicular development and corpus luteum functions. The GnRH/GnRHR system that is expressed in diseased tissues of the female reproductive tract (both benign and malignant) is at present considered an effective molecular target for the development of novel therapeutic approaches for these pathologies. GnRH agonists are also considered as a promising therapeutic approach to counteract ovarian failure in young female patients undergoing chemotherapy.

Conclusions: Increasing knowledge about the regulation of GnRH pulsatile release, as well as the therapeutic use of its analogues, offers interesting new perspectives in the diagnosis, treatment and outcome of female reproductive disorders, including tumoral and iatrogenic diseases.
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http://dx.doi.org/10.1093/humupd/dmv059DOI Listing
April 2016

Estrogen Receptor β Agonists Differentially Affect the Growth of Human Melanoma Cell Lines.

PLoS One 2015 30;10(7):e0134396. Epub 2015 Jul 30.

Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, Italy.

Background: Cutaneous melanoma is an aggressive malignancy; its incidence is increasing worldwide and its prognosis remains poor. Clinical observations indicate that estrogen receptor β (ERβ) is expressed in melanoma tissues and its expression decreases with tumor progression, suggesting its tumor suppressive function. These experiments were performed to investigate the effects of ERβ activation on melanoma cell growth.

Methods And Results: Protein expression was analyzed by Western blot and immunofluorescence assays. Cell proliferation was assessed by counting the cells by hemocytometer. ERβ transcriptional activity was evaluated by gene reporter assay. Global DNA methylation was analyzed by restriction enzyme assay and ERβ isoforms were identified by qRT-PCR. We demonstrated that ERβ is expressed in a panel of human melanoma cell lines (BLM, WM115, A375, WM1552). In BLM (NRAS-mutant) cells, ERβ agonists significantly and specifically inhibited cell proliferation. ERβ activation triggered its cytoplasmic-to-nuclear translocation and transcriptional activity. Moreover, the antiproliferative activity of ERβ agonists was associated with an altered expression of G1-S transition-related proteins. In these cells, global DNA was found to be hypomethylated when compared to normal melanocytes; this DNA hypomethylation status was reverted by ERβ activation. ERβ agonists also decreased the proliferation of WM115 (BRAF V600D-mutant) cells, while they failed to reduce the growth of A375 and WM1552 (BRAF V600E-mutant) cells. Finally, we could observe that ERβ isoforms are expressed at different levels in the various cell lines. Specific oncogenic mutations or differential expression of receptor isoforms might be responsible for the different responses of cell lines to ERβ agonists.

Conclusions: Our results demonstrate that ERβ is expressed in melanoma cell lines and that ERβ agonists differentially regulate the proliferation of these cells. These data confirm the notion that melanoma is a heterogeneous tumor and that genetic profiling is mandatory for the development of effective personalized therapeutic approaches for melanoma patients.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0134396PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4520550PMC
May 2016

Oxime bond-linked daunorubicin-GnRH-III bioconjugates exert antitumor activity in castration-resistant prostate cancer cells via the type I GnRH receptor.

Int J Oncol 2015 Jan 24;46(1):243-53. Epub 2014 Oct 24.

Department of Pharmacological and Biomolecular Sciences, University of Milan, I-20133 Milan, Italy.

It is well established that gonadotropin-releasing hormone receptors (GnRH-R) are expressed in different types of cancers, including castration-resistant prostate cancer (CRPC) and mediate the antiproliferative effect of GnRH analogs. Thus, these compounds are employed as targeting moieties to selectively deliver chemotherapeutic agents to cancer cells. GnRH-III, the decapeptide isolated from the sea lamprey brain, has lower potency than GnRH in stimulating gonadotropin secretion, but it exerts antiproliferative effects on many tumors expressing the GnRH-R. GnRH-III-based peptides are considered promising targeting moieties for the preparation of anticancer drug delivery systems. These studies were aimed at i) evaluating the antitumor activity of two cytotoxic oxime bond-linked daunorubicin (Dau)-GnRH-III derivative bioconjugates (Dau-GnRH-III, in which daunorubicin was coupled to the 8Lys in the native form of GnRH-III, and Dau-[4Lys(Ac)]-GnRH-III, in which daunorubicin was attached to the 8Lys of a GnRH-III derivative where 4Ser was replaced by an acetylated lysine) on CRPC cells; and ii) to elucidate the involvement of the classical GnRH-R (type I GnRH-R) in this antitumor activity. Our results demonstrated that both Dau-GnRH-III and Dau-[4Lys(Ac)]-GnRH-III were rapidly internalized into DU145 prostate cancer cells and exerted a significant cytostatic effect. Both bioconjugates increased the levels of the active form of caspase-3, indicating the involvement of apoptosis in their antitumor activity. The antiproliferative effect of both Dau-GnRH-III and Dau-[4Lys(Ac)]-GnRH-III was counteracted by the simultaneous treatment of the cells with Antide, an antagonist of the GnRH-R. Moreover, after silencing the type I GnRH-R the antitumor activity of both bioconjugates was completely abolished. These data demonstrate that in CRPC cells, daunorubicin-GnRH-III derivative bioconjugates: i) inhibit tumor cell proliferation, by triggering the apoptosis process; ii) exert their antitumor effect through the activation of the type I GnRH-R expressed on these cells. Cytotoxic-GnRH-III derivative may represent promising targeted chemotherapeutics for the treatment of CRPC patients.
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http://dx.doi.org/10.3892/ijo.2014.2730DOI Listing
January 2015
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