Plasma-activated Ringer's Lactate Solution Displays a Selective Cytotoxic Effect on Ovarian Cancer Cells

Epithelial Ovarian Cancer (EOC) is one of the leading causes of cancer-related deaths among women and is characterized by the diffusion of nodules or plaques from the ovary to the peritoneal surfaces. Conventional therapeutic options cannot eradicate the disease and show low efficacy against resistant tumor subclones. The treatment of liquids via cold atmospheric pressure plasma enables the production of plasma-activated liquids (PALs) containing reactive oxygen and nitrogen species (RONS) with selective anticancer activity. Thus, the delivery of RONS to cancer tissues by intraperitoneal washing with PALs might be an innovative strategy for the treatment of EOC. In this work, plasma-activated Ringer's Lactate solution (PA-RL) was produced by exposing a liquid substrate to a multiwire plasma source. Subsequently, PA-RL dilutions are used for the treatment of EOC, non-cancer and fibroblast cell lines, revealing a selectivity of PA-RL, which induces a significantly higher cytotoxic effect in EOC with respect to non-cancer cells

Molecular mechanisms underlying the acquisition of cisplatin resistance in ovarian cancer: from stemness to lipid metabolism

Il carcinoma ovarico (OC) è la neoplasia ginecologica con il maggiore tasso di mortalità, in parte dovuto all’acquisizione della resistenza al chemioterapico, che si manifesta nel 75% dei casi. In diversi tipi di tumori la sopravvivenza delle cellule tumorali trattate con chemioterapici è stata alla riprogrammazione del metabolismo. Date queste premesse l’obiettivo di questo studio è verificare se trattamenti a base di platino possano indurre un’alterazione del profilo metabolico e come questa riprogrammazione metabolica influisca sul fenotipo cellulare. L’analisi del profilo OCR in due linee cellulari di OC (OV90 e OC314) resistenti al farmaco ha mostrato una riprogrammazione metabolica verso una riduzione del profilo OXPHOS e un aumento del metabolismo glicolitico rispetto alle cellule sensibili . Inoltre, è stato verificato che l’acquisizione della resistenza al farmaco è associata ad un’aumentata espressione dei trasportatori ABCC2 e ABCG2 e ad un rallentamento della proliferazione cellulare. La resistenza è, inoltre, associata all’'acquisizione del fenotipo staminale. È stato dimostrato un arricchimento della popolazione CSC nelle linee resistenti rispetto alle loro controparti sensibili. Infatti, la sovraespressione dei fattori di Yamanaka e di ABCG2 supportano l'acquisizione di un fenotipo stem-like nelle linee resistenti. Pertanto il trattamento cronico con cisplatino ha favorito l’arricchimento delle CSC. Quindi, al fine di investigare il legame meccanicistico fra la riprogrammazione metabolica e l’acquisizione della chemioresistenza e di caratteristiche staminali, è stata effettuata un'analisi bioinformatica: gli elevati livelli di espressione dei marcatori CSC correlano con la sovraespressione dei geni coinvolti nel mantenimento dell’omeostasi del metabolismo lipidico. Questa analisi ha mostrato il coinvolgimento della via PPAR, come via di regolazione, e la β-ossidazione come via metabolica. Le cellule resistenti, con caratteristiche stem-like, costituiscono una sotto-popolazione con un profilo metabolico glicolitico e lipidico. Tali cellule presentano un vantaggio selettivo in un microambiente ricco di lipidi, quale l’omento, e potrebbero pertanto concorrere all’insorgenza della recidiva.Ovarian cancer (OC) is the gynecological malignancy with the highest mortality rate, partly due to the acquisition of resistance to chemotherapy, which occurs in 75% of cases. In several types of tumors the survival of cancer cells treated with chemotherapy has been due to metabolic reprogramming. Given these premises, the objective of this study is to verify whether platinum-based treatments could induce an alteration of metabolic profile and how this metabolic reprogramming affects the cellular phenotype. Analysis of the OCR profile in two drug-resistant OC cell lines (OV90 and OC314) showed metabolic reprogramming towards a reduction in the OXPHOS profile and an increase in glycolytic metabolism compared to sensitive cells. Furthermore, it has been verified that the acquisition of drug resistance is associated with an increased expression of the ABCC2 and ABCG2 transporters and a slowdown in cell proliferation. Resistance is also associated with the acquisition of the stemness phenotype. An enrichment in CSC population in resistant cells was demonstrated compared to their sensitive counterparts. Indeed, the overexpression of Yamanaka factors and ABCG2 support the acquisition of a stem-like phenotype in resistant cell lines. Therefore, chronic treatment with cisplatin has favored the enrichment of CSCs. Then, in order to investigate the mechanistic link between metabolic reprogramming, the acquisition of chemoresistance and stemness characteristics, a bioinformatic analysis was performed: the high levels of expression of the CSC markers correlate with the overexpression of the genes involved in the maintenance of homeostasis of lipid metabolism. This analysis showed the involvement of the PPAR pathway, as a regulatory pathway, and β-oxidation as a metabolic pathway. Resistant cells, with stem-like characteristics, constitute a sub-population with a glycolytic and lipid metabolic profile. These cells have a selective advantage in a lipid-rich microenvironment, such as the omentum, and could therefore contribute to the onset of relapse

Potential for Mitochondrial DNA Sequencing in the Differential Diagnosis of Gynaecological Malignancies

In the event of multiple synchronous gynecological lesions, a fundamental piece of information to determine patient management, prognosis, and therapeutic regimen choice is whether the simultaneous malignancies arise independently or as a result of metastatic dissemination. An example of synchronous primary tumors of the female genital tract most frequently described are ovarian and endometrial cancers. Surgical findings and histopathological examination aimed at resolving this conundrum may be aided by molecular analyses, although they are too often inconclusive. High mitochondrial DNA (mtDNA) variability and its propensity to accumulate mutations has been proposed by our group as a tool to define clonality. We showed mtDNA sequencing to be informative in synchronous primary ovarian and endometrial cancer, detecting tumor-specific mutations in both lesions, ruling out independence of the two neoplasms, and indicating clonality. Furthermore, we tested this method in another frequent simultaneously detected gynecological lesion type, borderline ovarian cancer and their peritoneal implants, which may be monoclonal extra-ovarian metastases or polyclonal independent masses. The purpose of this review is to provide an update on the potential use of mtDNA sequencing in distinguishing independent and metastatic lesions in gynecological cancers, and to compare the efficiency of molecular analyses currently in use with this novel method

Mitochondrial DNA analysis efficiently contributes to the identification of metastatic contralateral breast cancers

Purpose In daily practice, a contralateral breast cancer (CBC) is usually considered as a new independent tumor despite the indications of several studies showing that the second neoplasia may be a metastatic spread of the primary tumor. Recognition of clonal masses in the context of multiple synchronous or metachronous tumors is crucial for correct prognosis, therapeutic choice, and patient management. Mitochondrial DNA (mtDNA) sequencing shows high informative potential in the diagnosis of synchronous neoplasms, based on the fact that somatic mtDNA mutations are non-recurrent events, whereas tumors sharing them have a common origin. We here applied this technique to reveal clonality of the CBC with respect to the first tumor. Methods We analyzed 30 sample pairs of primary breast cancers and synchronous or metachronous CBCs with detailed clinical information available and compared standard clinico-pathological criteria with mtDNA sequencing to reveal the metastatic nature of CBCs. Results MtDNA analysis was informative in 23% of the cases, for which it confirmed a clonal origin of the second tumor. In addition, it allowed to solve two ambiguous cases where histopathological criteria had failed to be conclusive and to suggest a clonal origin for two additional cases that had been classified as independent by pathologists. Conclusion Overall, the mtDNA-based classification showed a more accurate predictive power than standard histopathology in identifying cases of metastatic rather than bilateral breast cancers in our cohort, suggesting that mtDNA sequencing may be a more precise and easy-to-use method to be introduced in daily routine to support and improve histopathological diagnoses

Pathogenic Mitochondrial DNA Mutation Load Inversely Correlates with Malignant Features in Familial Oncocytic Parathyroid Tumors Associated with Hyperparathyroidism-Jaw Tumor Syndrome

While somatic disruptive mitochondrial DNA (mtDNA) mutations that severely affect the respiratory chain are counter-selected in most human neoplasms, they are the genetic hallmark of indolent oncocytomas, where they appear to contribute to reduce tumorigenic potential. A correlation between mtDNA mutation type and load, and the clinical outcome of a tumor, corroborated by functional studies, is currently lacking. Recurrent familial oncocytomas are extremely rare entities, and they offer the chance to investigate the determinants of oncocytic transformation and the role of both germline and somatic mtDNA mutations in cancer. We here report the first family with Hyperparathyroidism-Jaw Tumor (HPT-JT) syndrome showing the inherited predisposition of four individuals to develop parathyroid oncocytic tumors. MtDNA sequencing revealed a rare ribosomal RNA mutation in the germline of all HPT-JT affected individuals whose pathogenicity was functionally evaluated via cybridization technique, and which was counter-selected in the most aggressive infiltrating carcinoma, but positively selected in adenomas. In all tumors different somatic mutations accumulated on this genetic background, with an inverse clear-cut correlation between the load of pathogenic mtDNA mutations and the indolent behavior of neoplasms, highlighting the importance of the former both as modifiers of cancer fate and as prognostic markers

MicroRNA and Metabolic Profiling of a Primary Ovarian Neuroendocrine Carcinoma Pulmonary-Type Reveals a High Degree of Similarity with Small Cell Lung Cancer

Small cell neuroendocrine carcinoma is most frequently found in the lung (SCLC), but it has been also reported, albeit with a very low incidence, in the ovary. Here, we analyze a case of primary small cell carcinoma of the ovary of pulmonary type (SCCOPT), a rare and aggressive tumor with poor prognosis, whose biology and molecular features have not yet been thoroughly investigated. The patient affected by SCCOPT had a residual tumor following chemotherapy which displayed pronounced similarity with neuroendocrine tumors and lung cancer in terms of its microRNA expression profile and mTOR-downstream activation. By analyzing the metabolic markers of the neoplastic lesion, we established a likely glycolytic signature. In conclusion, this in-depth characterization of SCCOPT could be useful for future diagnoses, possibly aided by microRNA profiling, allowing clinicians to adopt the most appropriate therapeutic strategy

Inducing respiratory complex I impairment elicits an increase in PGC1α in ovarian cancer

Anticancer strategies aimed at inhibiting Complex I of the mitochondrial respiratory chain are increasingly being attempted in solid tumors, as functional oxidative phosphorylation is vital for cancer cells. Using ovarian cancer as a model, we show that a compensatory response to an energy crisis induced by Complex I genetic ablation or pharmacological inhibition is an increase in the mitochondrial biogenesis master regulator PGC1α, a pleiotropic coactivator of transcription regulating diverse biological processes within the cell. We associate this compensatory response to the increase in PGC1α target gene expression, setting the basis for the comprehension of the molecular pathways triggered by Complex I inhibition that may need attention as drawbacks before these approaches are implemented in ovarian cancer care