Filamin A in triple negative breast cancer.

Triple-negative breast cancer is a rare but highly heterogeneous breast cancer subtype with a limited choice of specific treatments. Chemotherapy remains the only efficient treatment, but its side effects and the development of resistance consolidate the urgent need to discover new targets. In TNBC, filamin A expression correlates to grade and TNM stage. Accordingly, this protein could constitute a new target for this BC subtype. Even if most of the data indicates its direct involvement in cancer progression, some contrasting results underline the need to deepen the studies. To elucidate a possible function of this protein as a TNBC marker, we summarized the main characteristic of filamin A and its involvement in physiological and pathological processes such as cancer. Lastly, we scrutinized its actions in triple-negative breast cancer and highlighted the need to increase the number of studies useful to better clarify the role of this versatile protein as a marker and target in TNBC, alone or in "collaboration" with other proteins with a relevant role in this BC subgroup.

RIZ2 at the crossroad of the EGF/EGFR signaling in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is the third most deadly and fourth most diagnosed cancer worldwide. Despite the progress in early diagnosis and advanced therapeutic options, CRC shows a poor prognosis with a 5 year survival rate of ~ 45%. PRDM2/RIZ, a member of PR/SET domain family (PRDM), expresses two main molecular variants, the PR-plus isoform (RIZ1) and the PR-minus (RIZ2). The imbalance in their expression levels in favor of RIZ2 is observed in many cancer types. The full length RIZ1 has been extensively investigated in several cancers where it acts as a tumor suppressor, whereas few studies have explored the RIZ2 oncogenic properties. PRDM2 is often target of frameshift mutations and aberrant DNA methylation in CRC. However, little is known about its role in CRC. METHODS: We combined in-silico investigation of The Cancer Genome Atlas (TCGA) CRC datasets, cellular and molecular assays, transcriptome sequencing and functional annotation analysis to assess the role of RIZ2 in human CRC. RESULTS: Our in-silico analysis on TCGA datasets confirmed that PRDM2 gene is frequently mutated and transcriptionally deregulated in CRC and revealed that a RIZ2 increase is highly correlated with a significant RIZ1 downregulation. Then, we assayed several CRC cell lines by qRT-PCR analysis for the main PRDM2 transcripts and selected DLD1 cell line, which showed the lowest RIZ2 levels. Therefore, we overexpressed RIZ2 in these cells to mimic TCGA datasets analysis results and consequently to assess the PRDM2/RIZ2 role in CRC. Data from RNA-seq disclosed that RIZ2 overexpression induced profound changes in CRC cell transcriptome via EGF pathway deregulation, suggesting that RIZ2 is involved in the EGF autocrine regulation of DLD1 cell behavior. Noteworthy, the forced RIZ2 expression increased cell viability, growth, colony formation, migration and organoid formation. These effects could be mediated by the release of high EGF levels by RIZ2 overexpressing DLD1 cells. CONCLUSIONS: Our findings add novel insights on the putative RIZ2 tumor-promoting functions in CRC, although additional efforts are warranted to define the underlying molecular mechanism.

Exosomes: Emerging Modulators of Pancreatic Cancer Drug Resistance.

Pancreatic cancer (PaC) is one of the most lethal tumors worldwide, difficult to diagnose, and with inadequate therapeutical chances. The most used therapy is gemcitabine, alone or in combination with nanoparticle albumin-bound paclitaxel (nab-paclitaxel), and the multidrug FOLFIRINOX. Unfortunately, PaC develops resistance early, thus reducing the already poor life expectancy of patients. The mechanisms responsible for drug resistance are not fully elucidated, and exosomes seem to be actively involved in this phenomenon, thanks to their ability to transfer molecules regulating this process from drug-resistant to drug-sensitive PaC cells. These extracellular vesicles are released by both normal and cancer cells and seem to be essential mediators of intercellular communications, especially in cancer, where they are secreted at very high numbers. This review illustrates the role of exosomes in PaC drug resistance. This manuscript first provides an overview of the pharmacological approaches used in PaC and, in the last part, focuses on the mechanisms exploited by the exosomes released by cancer cells to induce drug resistance.

The nerve growth factor-delivered signals in prostate cancer and its associated microenvironment: when the dialogue replaces the monologue.

Prostate cancer (PC) represents the most diagnosed and the second most lethal cancer in men worldwide. Its development and progression occur in concert with alterations in the surrounding tumor microenvironment (TME), made up of stromal cells and extracellular matrix (ECM) that dynamically interact with epithelial PC cells affecting their growth and invasiveness. PC cells, in turn, can functionally sculpt the TME through the secretion of various factors, including neurotrophins. Among them, the nerve growth factor (NGF) that is released by both epithelial PC cells and carcinoma-associated fibroblasts (CAFs) triggers the activation of various intracellular signaling cascades, thereby promoting the acquisition of a metastatic phenotype. After many years of investigation, it is indeed well established that aberrations and/or derangement of NGF signaling are involved not only in neurological disorders, but also in the pathogenesis of human proliferative diseases, including PC. Another key feature of cancer progression is the nerve outgrowth in TME and the concept of nerve dependence related to perineural invasion is currently emerging. NGF released by cancer cells can be a driver of tumor neurogenesis and nerves infiltrated in TME release neurotransmitters, which might stimulate the growth and sustainment of tumor cells.In this review, we aim to provide a snapshot of NGF action in the interactions between TME, nerves and PC cells. Understanding the molecular basis of this dialogue might expand the arsenal of therapeutic strategies against this widespread disease.

Androgens and NGF Mediate the Neurite-Outgrowth through Inactivation of RhoA.

Steroid hormones and growth factors control neuritogenesis through their cognate receptors under physiological and pathological conditions. We have already shown that nerve growth factor and androgens induce neurite outgrowth of PC12 cells through a reciprocal crosstalk between the NGF receptor, TrkA and the androgen receptor. Here, we report that androgens or NGF induce neuritogenesis in PC12 cells through inactivation of RhoA. Ectopic expression of the dominant negative RhoA N19 promotes, indeed, the neurite-elongation of unchallenged and androgen- or NGF-challenged PC12 cells and the increase in the expression levels of ßIII tubulin, a specific neuronal marker. Pharmacological inhibition of the Ser/Thr kinase ROCK, an RhoA effector, induces neuritogenesis in unchallenged PC12 cells, and potentiates the effect of androgens and NGF, confirming the role of RhoA/ROCK axis in the neuritogenesis induced by androgen and NGF, through the phosphorylation of Akt. These findings suggest that therapies based on new selective androgen receptor modulators and/or RhoA/ROCK inhibitors might exert beneficial effects in the treatment of neuro-disorders, neurological diseases and ageing-related processes.

Agri-Food By-Products in Cancer: New Targets and Strategies.

The globalization and the changes in consumer lifestyles are forcing us to face a deep transformation in food demand and in the organization of the entire food production system. In this new era, the food-loss and food-waste security nexus is relevant in the global debate and avoiding unsustainable waste in agri-food systems as well as the supply chain is a big challenge. "Food waste" is useful for the recovery of its valuable components, thus it can assume the connotation of a "food by-product". Sustainable utilization of agri-food waste by-products provides a great opportunity. Increasing evidence shows that agri-food by-products are a source of different bioactive molecules that lower the inflammatory state and, hence, the aggressiveness of several proliferative diseases. This review aims to summarize the effects of agri-food by-products derivatives, already recognized as promising therapeutics in human diseases, including different cancer types, such as breast, prostate, and colorectal cancer. Here, we examine products modulating or interfering in the signaling mediated by the epidermal growth factor receptor.

Inhibition of Vps34 and p110δ PI3K Impairs Migration, Invasion and Three-Dimensional Spheroid Growth in Breast Cancer Cells.

Breast cancer is a heterogeneous disease that represents the most common cancer around the world; it comprises 12% of new cases according to the World Health Organization. Despite new approaches in early diagnosis and current treatment, breast cancer is still the leading cause of death for cancer mortality. New targeted therapies against key signalling transduction molecules are required. Phosphoinositide 3-kinase (PI3K) regulates multiple biological functions such as proliferation, survival, migration, and growth. It is well established that PI3K isoform-selective inhibitors show fewer toxic side effects compared to broad spectrum inhibition of PI3K (pan-PI3K inhibitors). Therefore, we tested the PI3K p110δ-selective inhibitor, IC87114, and Vps34-selective inhibitor, Vps34-IN1, on the breast cancer cell lines MCF-7 and MDA-MB-231, representing hormone-responsive and triple-negative breast cancer cells, respectively. Our data show that both inhibitors decreased migration of MCF-7 and MDA-MB-231 cells, and Vps34 also significantly impacted MCF-7 cell proliferation. Three-dimensional (3D) in vitro culture models show that IC87114 and Vps34-IN1 treatment reduced the growth of MCF-7 and MDA-MB-231 cells in 3D tumour spheroid cultures. This study identifies IC87114 and Vps34-IN1 as potential therapeutic approaches in breast cancer.

The Role of Curcumin in Prostate Cancer Cells and Derived Spheroids.

A major challenge in the clinical management of prostate cancer (PC) is to inhibit tumor growth and prevent metastatic spreading. In recent years, considerable efforts have been made to discover new compounds useful for PC therapy, and promising advances in this field were reached. Drugs currently used in PC therapy frequently induce resistance and PC progresses toward metastatic castration-resistant forms (mCRPC), making it virtually incurable. Curcumin, a commercially available nutritional supplement, represents an attractive therapeutic agent for mCRPC patients. In the present study, we compared the effects of chemotherapeutic drugs such as docetaxel, paclitaxel, and cisplatin, to curcumin, on two PC cell lines displaying a different metastatic potential: DU145 (moderate metastatic potential) and PC-3 (high metastatic potential). Our results revealed a dose-dependent reduction of DU145 and PC-3 cell viability upon treatment with curcumin similar to chemotherapeutic agents (paclitaxel, cisplatin, and docetaxel). Furthermore, we explored the EGFR-mediated signaling effects on ERK activation in DU145 and PC-3 cells. Our results showed that DU145 and PC-3 cells overexpress EGFR, and the treatment with chemotherapeutic agents or curcumin reduced EGFR expression levels and ERK activation. Finally, chemotherapeutic agents and curcumin reduced the size of DU145 and PC-3 spheroids and have the potential to induce apoptosis and also in Matrigel. In conclusion, despite different studies being carried out to identify the potential synergistic curcumin combinations with chemopreventive/therapeutic efficacy for inhibiting PC growth, the results show the ability of curcumin used alone, or in combinatorial approaches, to impair the size and the viability of PC-derived spheroids.

New TRPM8 blockers exert anticancer activity over castration-resistant prostate cancer models.

TRPM8 has recently emerged as a druggable target in prostate cancer (PC) and TRPM8 modulators have been proposed as potential anticancer agents in this pathology. We have recently demonstrated their effectiveness in a castration-resistant prostate cancer (CRPC) model that is usually resistant to androgen deprivation therapy (ADT) and is considered the most aggressive form of PC. This is why the discovery of selective, effective, and potent TRPM8 modulators would improve the molecular arsenal in support of PC standard-of-care treatments. In the present paper we describe the design and the synthesis of a new series of TRPM8 antagonists, preliminarily characterized in vitro for their potency and selectivity by fluorimetric calcium assays. The preliminary screening allowed the identification of several potent (0.11 µM < IC50 < 0.49 µM) and selective compounds. The most potent derivatives were further characterized by patch-clamp electrophysiology assays, confirming their noteworthy activity. Moreover, the behavior of these compounds was investigated in 2D and 3D models of PC. These TRPM8 antagonists showed remarkable efficacy in inhibiting the growth induced by androgen in various PC cells as well as in CRPC models, confirming their potential as anticancer agents.

New Insights and Emerging Therapeutic Approaches in Prostate Cancer.

Prostate cancer is the second most frequently diagnosed cancer in men and several therapeutic approaches are currently available for patient's care. Although the androgen receptor status represents a good predictor of response to androgen deprivation therapy, prostate cancer frequently becomes resistant to this approach and spreads. The molecular mechanisms that contribute to progression and drug-resistance of this cancer remain still debated. However, few therapeutic options are available for patient's management, at this stage. Recent years have seen a great expansion of the studies concerning the role of stromal-epithelial interactions and tumor microenvironment in prostate cancer progression. The findings so far collected have provided new insights into diagnostic and clinical management of prostate cancer patients. Further, new fascinating aspects concerning the intersection of the androgen receptor with survival factors as well as calcium channels have been reported in cultured prostate cancer cells and mouse models. The results of these researches have opened the way for a better understanding of the basic mechanisms involved in prostate cancer invasion and drug-resistance. They have also significantly expanded the list of new biomarkers and druggable targets in prostate cancer. The primary aim of this manuscript is to provide an update of these issues, together with their translational aspects. Exploiting the power of novel promising therapeutics would increase the success rate in the diagnostic path and clinical management of patients with advanced disease.

Therapeutic potential of TRPM8 antagonists in prostate cancer.

Transient receptor potential melastatin-8 (TRPM8) represents an emerging target in prostate cancer, although its mechanism of action remains unclear. Here, we have characterized and investigated the effects of TRPM8 modulators in prostate cancer aggressiveness disclosing the molecular mechanism underlying their biological activity. Patch-clamp and calcium fluorometric assays were used to characterize the synthesized compounds. Androgen-stimulated prostate cancer-derived cells were challenged with the compounds and the DNA synthesis was investigated in a preliminary screening. The most effective compounds were then employed to inhibit the pro-metastatic behavior of in various PC-derived cells, at different degree of malignancy. The effect of the compounds was then assayed in prostate cancer cell-derived 3D model and the molecular targets of selected compounds were lastly identified using transcriptional and non-transcriptional reporter assays. TRPM8 antagonists inhibit the androgen-dependent prostate cancer cell proliferation, migration and invasiveness. They are highly effective in reverting the androgen-induced increase in prostate cancer cell spheroid size. The compounds also revert the proliferation of castrate-resistant prostate cancer cells, provided they express the androgen receptor. In contrast, no effects were recorded in prostate cancer cells devoid of the receptor. Selected antagonists interfere in non-genomic androgen action and abolish the androgen-induced androgen receptor/TRPM8 complex assembly as well as the increase in intracellular calcium levels in prostate cancer cells. Our results shed light in the processes controlling prostate cancer progression and make the transient receptor potential melastatin-8 as a 'druggable' target in the androgen receptor-expressing prostate cancers.

Communication between cells: exosomes as a delivery system in prostate cancer.

Despite the considerable efforts in screening and diagnostic protocols, prostate cancer still represents the second leading cause of cancer-related death in men. Many patients with localized disease and low risk of recurrence have a favourable outcome. In a substantial proportion of patients, however, the disease progresses and becomes aggressive. The mechanisms that promote prostate cancer progression remain still debated. Many findings point to the role of cross-communication between prostate tumor cells and their surrounding microenvironment during the disease progression. Such a connection fosters survival, proliferation, angiogenesis, metastatic spreading and drug-resistance of prostate cancer. Recent years have seen a profound interest in understanding the way by which prostate cancer cells communicate with the surrounding cells in the microenvironment. In this regard, direct cell-to-cell contacts and soluble factors have been identified. Increasing evidence indicates that PC cells communicate with the surrounding cells through the release of extracellular vesicles, mainly the exosomes. By directly acting in stromal or prostate cancer epithelial cells, exosomes represent a critical intercellular communication system. By querying the public database ( https://pubmed.ncbi.nlm.nih.gov ) for the past 10 years, we have found more than four hundred papers. Among them, we have extrapolated the most relevant about the role of exosomes in prostate cancer malignancy and progression. Emerging data concerning the use of these vesicles in diagnostic management and therapeutic guidance of PC patients are also presented. Video Abstract.

Targeting the Nerve Growth Factor Signaling Impairs the Proliferative and Migratory Phenotype of Triple-Negative Breast Cancer Cells.

Triple-negative breast cancer is a heterogeneous disease that still lacks specific therapeutic approaches. The identification of new biomarkers, predictive of the disease's aggressiveness and pharmacological response, is a challenge for a more tailored approach in the clinical management of patients. Nerve growth factor, initially identified as a key factor for neuronal survival and differentiation, turned out to be a multifaceted molecule with pleiotropic effects in quite divergent cell types, including cancer cells. Many solid tumors exhibit derangements of the nerve growth factor and its receptors, including the tropomyosin receptor kinase A. This receptor is expressed in triple-negative breast cancer, although its role in the pathogenesis and aggressiveness of this disease is still under investigation. We now report that triple-negative breast cancer-derived MDA-MB-231 and MDA-MB-453 cells express appreciable levels of tropomyosin receptor kinase A and release a biologically active nerve growth factor. Activation of tropomyosin receptor kinase by nerve growth factor treatment positively affects the migration, invasion, and proliferation of triple-negative breast cancer cells. An increase in the size of triple-negative breast cancer cell spheroids is also detected. This latter effect might occur through the nerve growth factor-induced release of matrix metalloproteinase 9, which contributes to the reorganization of the extracellular matrix and cell invasiveness. The tropomyosin receptor kinase A inhibitor GW441756 reverses all these responses. Co-immunoprecipitation experiments in both cell lines show that nerve growth factor triggers the assembly of the TrkA/ß1-integrin/FAK/Src complex, thereby activating several downstream effectors. GW441756 prevents the complex assembly induced by nerve growth factor as well as the activation of its dependent signaling. Pharmacological inhibition of the tyrosine kinases Src and FAK (focal adhesion kinase), together with the silencing of ß1-integrin, shows that the tyrosine kinases impinge on both proliferation and motility, while ß1-integrin is needed for motility induced by nerve growth factor in triple-negative breast cancer cells. The present data support the key role of the nerve growth factor/tropomyosin receptor kinase A pathway in triple-negative breast cancer and offer new hints in the diagnostic and therapeutic management of patients.

The androgen receptor/filamin A complex as a target in prostate cancer microenvironment.

Prostate cancer represents the major cause of cancer-related death in men and patients frequently develop drug-resistance and metastatic disease. Most studies focus on hormone-resistance mechanisms related to androgen receptor mutations or to the acquired property of prostate cancer cells to over-activate signaling pathways. Tumor microenvironment plays a critical role in prostate cancer progression. However, the mechanism involving androgen/androgen receptor signaling in cancer associated fibroblasts and consequences for prostate cancer progression still remains elusive. We now report that prostate cancer associated fibroblasts express a transcriptional-incompetent androgen receptor. Upon androgen challenging, the receptor co-localizes with the scaffold protein filamin A in the extra-nuclear compartment of fibroblasts, thus mediating their migration and invasiveness. Cancer-associated fibroblasts move towards epithelial prostate cancer cells in 2D and 3D cultures, thereby inducing an increase of the prostate cancer organoid size. Androgen enhances both these effects through androgen receptor/filamin A complex assembly in cancer-associated fibroblasts. An androgen receptor-derived stapled peptide, which disrupts the androgen receptor/filamin A complex assembly, abolishes the androgen-dependent migration and invasiveness of cancer associated fibroblasts. Notably, the peptide impairs the androgen-induced invasiveness of CAFs in 2D models and reduces the overall tumor area in androgen-treated 3D co-culture. The androgen receptor in association with ß1 integrin and membrane type-matrix metalloproteinase 1 activates a protease cascade triggering extracellular matrix remodeling. The peptide also impairs the androgen activation of this cascade. This study offers a potential new marker, the androgen receptor/filamin A complex, and a new therapeutic approach targeting intracellular pathways activated by the androgen/androgen receptor axis in prostate cancer-associated fibroblasts. Such a strategy, alone or in combination with conventional therapies, may allow a more efficient treatment of prostate cancer.