Rita Mancini

Ovarian carcinoma (OVCA) is the most lethal gynaecological malignancy, and platinum resistance sustains its mortality, creating a need for cytotoxic strategies that act independently of apoptosis. OVCA is characterized by an "iron-addicted" phenotype that can render it susceptible to ferroptosis; however, this vulnerability has been tested almost exclusively with non-clinical GPX4/system Xc- inhibitors, and the determinants of intrinsic sensitivity remain undefined. Here, we tested parenteral iron formulations already in clinical use, namely ferric gluconate, ferric ammonium citrate and iron dextran, as ferroptosis inducers across the PEA1, COV362, KURAMOCHI, and PEO1 OVCA lines. Iron loading killed PEA1 and COV362 through ferroptosis, rescued by ferrostatin-1 but not by inhibitors of apoptosis, necroptosis or autophagy, and accompanied by lipid peroxidation, cytoplasmic reactive oxygen species (ROS) and GPX4 loss, while sparing non-malignant ovarian epithelial cells; sensitivity was independent of both canonical GPX4/system Xc- inhibition and platinum-response status. Mechanistically, the resistant PEO1 and KURAMOCHI cells mounted the canonical protective response to iron overload, by restricting import through transferrin-receptor (TfR1) downregulation, enhancing iron export via ferroportin upregulation, and promoting mitochondria clearance through PARKIN-mediated mitophagy. In contrast, the sensitive PEA1 and COV362 cells failed to activate those mechanisms and instead preferentially directed iron to the mitochondria via mitoferrin-1 (MFRN1), resulting in mitochondrial ROS accumulation and bioenergetic failure. Silencing MFRN1 protected sensitive cells and abolished mitochondrial iron loading without lowering the cytosolic iron pool, identifying the mitochondrial iron pool as the lethal determinant. In vivo, iron dextran suppressed the growth of sensitive but not resistant cells-derived xenografts, with induction of ferroptosis markers (PTGS2, MFRN1) and GPX4 downregulation, and no caspase activation. Overall, we identified mitochondrial iron routing through MFRN1 as determinant of ferroptotic vulnerability in OVCA and propose repurposed iron formulations as a therapeutic strategy for a molecularly defined subset of this disease.

The advent of immune checkpoint inhibitors (ICIs) has transformed the treatment landscape of several malignancies, but durable clinical benefits are only observed in a subset of patients. Primary and acquired resistance to immunotherapy, largely driven by an immunosuppressive tumor immune microenvironment (TIME), remains among the most critical hurdles to overcome. Amid the molecular regulators orchestrating immune evasion, tumor-derived microRNAs (miRNAs) have emerged as pivotal players. These small non-coding RNAs reshape the TIME through diverse mechanisms, including impairment of antigen presentation, modulation of immune checkpoint pathways, suppression of cytotoxic T and NK cell activity, induction of regulatory immune populations, and reprogramming of stromal and metabolic networks. This review explores the multifaceted contribution of miRNAs to ICI resistance and discusses innovative therapeutic strategies aimed at targeting dysregulated miRNAs, either through inhibition of oncomiRs or restoration of tumor-suppressors, with the goal of reprogramming the TIME toward an immunocompetent state. The clinical implementation of combination regimens of miRNA therapeutics with ICIs remains limited, highlighting the need to test these combinations in advanced preclinical models that faithfully recapitulate the human TIME. Such models could provide an optimal platform to integrate miRNA therapeutics into immuno-oncology strategies, ultimately expanding the benefits of ICIs to a broader patient population.

Ferroptosis, a regulated form of cell death driven by iron-dependent lipid peroxidation, has recently emerged as a promising therapeutic vulnerability in cancer. Among its key modulators, stearoyl-CoA desaturase 1 (SCD1) plays a pivotal role in ferroptosis resistance by converting saturated fatty acids into monounsaturated fatty acids (MUFAs), thereby limiting the accumulation of highly peroxidizable polyunsaturated fatty acids (PUFAs) and stabilizing membrane integrity under oxidative stress. Multiple oncogenic, transcriptional, epigenetic, and microenvironmental cues enhance tumor survival by promoting SCD1-dependent ferroptosis resistance. Despite this diversity of regulatory inputs, a unifying principle emerges across tumor types: SCD1 activity preserves lipid desaturation as a dominant metabolic strategy to suppress ferroptotic cell death. In this review, we critically analyze how SCD1-dependent ferroptosis resistance is shaped by tumor-specific metabolic states, microenvironmental pressures, and regulatory hierarchies across multiple malignancies. We identify recurring mechanistic themes through which SCD1 integrates redox control, lipid metabolism, stemness, and therapy resistance, while highlighting how these processes are differentially regulated across tissues. Preclinical evidence indicates that targeting SCD1, particularly in rational combination with ferroptosis inducers, chemotherapy, radiotherapy, or immunotherapy, can lower the ferroptotic threshold and overcomes treatment resistance. Finally, we discuss translational challenges and emerging strategies, including tumor-selective delivery, adaptive dosing, and context-specific combinations, that may enable safe and effective exploitation of the SCD1-ferroptosis axis in cancer therapy.

Circulating miRNAs (cmiRNAs) are emerging as valuable non-invasive biomarkers for monitoring disease progression and therapeutic response in cancer. Their stability in biological fluids, tissue-specific expression, and functional roles in tumor biology make them particularly suitable for liquid biopsy approaches. However, challenges related to quantification accuracy and assay standardization have limited their clinical translation. Digital PCR (dPCR) offers a highly sensitive and reproducible solution for absolute quantification of low-abundance transcripts, addressing many of these limitations. We developed and analytically validated the first duplex dPCR assay for the simultaneous detection of miR-4488 and miR-579-3p in serum samples from patients with BRAF-mutant metastatic melanoma receiving MAPK inhibitor therapy. These two cmiRNAs were previously identified by our group as biomarkers predictive of treatment response. Using fluorescently labelled probes, both targets were co-amplified in a single reaction. The assay was tested for analytical performance, including comparison with singleplex formats and quantitative Real-Time PCR (qRT-PCR). We then applied the duplex assay to assess the prognostic potential of the expression ratio between the two miRNAs, termed miRatio, at baseline and over treatment timepoints. The duplex assay maintained analytical performance comparable to singleplex reactions while reducing sample and reagent use. Compared to qRT-PCR, dPCR showed superior sensitivity, particularly for detecting low-abundance miRNAs like miR-4488. miRatio effectively predicts disease outcome when measured at baseline prior to MAPKi therapy and exhibits dynamic changes during treatment, supporting its potential as a longitudinal biomarker. ROC analysis demonstrated strong prognostic value, with improved accuracy over previous qRT-PCR-based evaluations. This study highlights duplex dPCR as a robust, sensitive, and scalable technology for circulating miRNA quantification in liquid biopsy applications. By enabling absolute and simultaneous detection of miR-4488 and miR-579-3p, the assay provides a technically advanced platform for real-time monitoring in metastatic melanoma. While miRatio remains a promising biomarker, the key innovation of this work is the development of a duplex assay suitable for clinical implementation in precision oncology.

Metabolic reprogramming and epigenetic alterations are fundamental hallmarks of cancer cells, contributing to adaptation, progression, and resistance. In melanoma, high metabolic-epigenetic plasticity enables the rapid modulation of cell states in response to environmental and therapeutic pressures. Recent studies have highlighted a bidirectional crosstalk between cellular metabolism and epigenetic regulation. Epigenetic modifications influence the transcriptional control of metabolic genes, thereby shaping metabolic phenotypes. Conversely, specific metabolites are essential cofactors or substrates for epigenetic enzymes, directly modulating the epigenome. Understanding the intricate mechanisms of this interaction offers opportunities for the development of innovative tumor management that combines epigenetic, metabolic, and therapy interventions. In this review, we summarize the latest evidence on the role of the metabolism–epigenetics axis in melanoma and discuss its potential clinical implications, aiming to provide a comprehensive overview of metabolic/epigenetic interconnections.

Myxedema is a potentially life-threatening condition typically observed in severe hypothyroidism. However, localized or diffuse myxedema is also observed in hyperthyroidism. The exact cause and mechanism of this paradoxical situation is not clear. We report here the analysis of body fluid distribution by bioelectrical impedance analysis (BIA) in 103 thyroid patients, subdivided according to their functional status. All BIA parameters measured in subclinical thyroid dysfunctions did not significantly differ from those observed in euthyroid controls. On the contrary, they were clearly altered in the two extreme, opposite conditions of thyroid dysfunctions, namely overt hyperthyroidism and severe hypothyroidism, indicating the occurrence of a typical hormetic condition. Surprisingly, differences in BIA parameters related to fluid body composition were even more evident in hyperthyroidism than in hypothyroidism. A hormetic response to thyroid hormone (TH)s was previously reported to explain the paradoxical, biphasic, time- and dose-dependent effects on other conditions. Our results indicate that myxedema, observed in both hypothyroid and hyperthyroid conditions, represents another example of a hormetic-type response to THs. BIA offers no additional valuable information in evaluating fluid body composition in subclinical thyroid dysfunctions, but it represents a valuable method to analyze and monitor body fluid composition and distribution in overt and severe thyroid dysfunctions.

Malignant pleural effusion (MPE) from patients with advanced non-small-cell lung cancer (NSCLC) has been proven valuable for molecular analysis; however, simultaneous detection of driver fusions in MPE is still challenging. In this study, we investigated the Idylla™ GeneFusion Panel, a stand-alone test in tissue samples, in the evaluation of ALK, ROS1, RET and MET ex14 skipping mutations in MPE and compared its performance with routine reference methods (Real-time-based and Next-generation Sequencing—NGS). The inclusion criteria for sample selection were as follows: advanced NSCLC harboring ALK, ROS1, RET fusions or MET exon-skipping alterations and the availability of MPE collected at diagnosis or disease progression. Molecular alterations have been investigated on tissue by fluorescence in situ hybridization (FISH) or Real-time PCR or NGS. For molecular profiling with the Idylla™ GeneFusion, 200 µL of MPE supernatants combined with 50 µL of RNA Later solution were loaded into the Idylla™ cartridge without cfRNA extraction. The Idylla™ GeneFusion Assay performed on MPEs was able to confirm molecular profile, previously diagnosed with conventional methods, in all cases. Our data confirm that MPE are suitable material for investigating fusion alterations. The Idylla™ GeneFusion, although indicated for investigation of tissue samples, offers the possibility of performing a molecular characterization of supernatants without undertaking the entire cfRNA extraction procedure providing a rapid and reliable strategy for the detection of actionable genetic alterations.

Scientific literature supports the evidence that cancer stem cells (CSCs) retain inside low reactive oxygen species (ROS) levels and are, therefore, less susceptible to cell death, including ferroptosis, a type of cell death dependent on iron-driven lipid peroxidation. A collection of lung adenocarcinoma (LUAD) primary cell lines derived from malignant pleural effusions (MPEs) of patients was used to obtain 3D spheroids enriched for stem‐like properties. We observed that the ferroptosis inducer RSL3 triggered lipid peroxidation and cell death in LUAD cells when grown in 2D conditions; however, when grown in 3D conditions, all cell lines underwent a phenotypic switch, exhibiting substantial resistance to RSL3 and, therefore, protection against ferroptotic cell death. Interestingly, this phenomenon was reversed by disrupting 3D cells and growing them back in adherence, supporting the idea of CSCs plasticity, which holds that cancer cells have the dynamic ability to transition between a CSC state and a non-CSC state. Molecular analyses showed that ferroptosis resistance in 3D spheroids correlated with an increased expression of antioxidant genes and high levels of proteins involved in iron storage and export, indicating protection against oxidative stress and low availability of iron for the initiation of ferroptosis. Moreover, transcriptomic analyses highlighted a novel subset of genes commonly modulated in 3D spheroids and potentially capable of driving ferroptosis protection in LUAD-CSCs, thus allowing to better understand the mechanisms of CSC-mediated drug resistance in tumors.

Therapy of melanoma has improved dramatically over the last years thanks to the development of targeted therapies (MAPKi) and immunotherapies. However, drug resistance continues to limit the efficacy of these therapies. Our research group has provided robust evidence as to the involvement of a set of microRNAs in the development of resistance to target therapy in BRAF-mutated melanomas. Among them, a pivotal role is played by the oncosuppressor miR-579-3p. Here we show that miR-579-3p and the microphthalmia-associated transcription factor (MITF) influence reciprocally their expression through positive feedback regulatory loops. In particular we show that miR-579-3p is specifically deregulated in BRAF-mutant melanomas and that its expression levels mirror those of MITF. Luciferase and ChIP studies show that MITF is a positive regulator of miR-579-3p, which is located in the intron 11 of the human gene ZFR (Zink-finger recombinase) and is co-transcribed with its host gene. Moreover, miR-579-3p, by targeting BRAF, is able to stabilize MITF protein thus inducing its own transcription. From biological points of view, early exposure to MAPKi or, alternatively miR-579-3p transfection, induce block of proliferation and trigger senescence programs in BRAF-mutant melanoma cells. Finally, the long-term development of resistance to MAPKi is able to select cells characterized by the loss of both miR-579-3p and MITF and the same down-regulation is also present in patients relapsing after treatments. Altogether these findings suggest that miR-579-3p/MITF interplay potentially governs the balance between proliferation, senescence and resistance to therapies in BRAF-mutant melanomas.

Many factors have contributed to rendering frailty an emerging, relevant, and very popular concept. First, many pandemics that have affected humanity in history, including COVID-19, most recently, have had more severe effects on frail people compared to non-frail ones. Second, the increase in human life expectancy observed in many developed countries, including Italy has led to a rise in the percentage of the older population that is more likely to be frail, which is why frailty is much a more common concern among geriatricians compared to other the various health-care professionals. Third, the stratification of people according to the occurrence and the degree of frailty allows healthcare decision makers to adequately plan for the allocation of available human professional and economic resources. Since frailty is considered to be fully preventable, there are relevant consequences in terms of potential benefits both in terms of the clinical outcome and healthcare costs. Frailty is becoming a popular, pervasive, and almost omnipresent concept in many different contexts, including clinical medicine, physical health, lifestyle behavior, mental health, health policy, and socio-economic planning sciences. The emergence of the new “science of frailty” has been recently acknowledged. However, there is still debate on the exact definition of frailty, the pathogenic mechanisms involved, the most appropriate method to assess frailty, and consequently, who should be considered frail. This narrative review aims to analyze frailty from many different aspects and points of view, with a special focus on the proposed pathogenic mechanisms, the various factors that have been considered in the assessment of frailty, and the emerging role of biomarkers in the early recognition of frailty, particularly on the role of mitochondria. According to the extensive literature on this topic, it is clear that frailty is a very complex syndrome, involving many different domains and affecting multiple physiological systems. Therefore, its management should be directed towards a comprehensive and multifaceted holistic approach and a personalized intervention strategy to slow down its progression or even to completely reverse the course of this condition.