LVI and DI-SPME Combined with GC/MS and GC/MS for Volatile Chemical Profile Investigation and Cytotoxic Power Evaluation of Essential Oil and Hydrolate from Cannabis sativa L. cv. Carmagnola.

Cannabis sativa L. is a plant that has been cultivated since ancient times thanks to its various uses. Even its extraction products, such as essential oil and hydrolate, having a varied chemical composition and rich in bioactive components, find wide use in different sectors, gathering ever-increasing interest over time. In this work, the essential oil of Cannabis sativa L. cv. Carmagnola was characterized by using Gas Chromatography/Mass Spectrometry (GC/MS) and, for the first time, the chemical profile of the hydrolate was also described through different analytical techniques such as Large-Volume Injection Gas Chromatography/Mass Spectrometry (LVI-GC/MS) and Direct Immersion-Solid Phase Microextraction-Gas Chromatography/Mass spectrometry (DI-SPME-GC/MS), in order to provide a more complete compositional profile. The results of the analyses conducted on the hydrolate highlighted a high content of α-terpineol; on the other side, in the essential oil, a prevalence of monoterpenes, with α-pinene and limonene as the characterizing components, was detected. Both matrices were also investigated to evaluate their cytotoxic activity by using a panel of cancer cell lines derived from different histotypes such as melanoma (A375, LOX IMVI), non-small cell lung cancer (H1299, A549), colon (HT29) and pancreatic (L3.6) cancer cell lines. The obtained data demonstrated that essential oil was more effective than hydrolate in terms of reduction in cell viability.

Notch3-regulated microRNAs impair CXCR4-dependent maturation of thymocytes allowing maintenance and progression of T-ALL.

Malignant transformation of T-cell progenitors causes T-cell acute lymphoblastic leukemia (T-ALL), an aggressive childhood lymphoproliferative disorder. Activating mutations of Notch, Notch1 and Notch3, have been detected in T-ALL patients. In this study, we aimed to deeply characterize hyperactive Notch3-related pathways involved in T-cell dynamics within the thymus and bone marrow to propose these processes as an important step in facilitating the progression of T-ALL. We previously generated a transgenic T-ALL mouse model (N3-ICtg) demonstrating that aberrant Notch3 signaling affects early thymocyte maturation programs and leads to bone marrow infiltration by CD4+CD8+ (DP) T cells that are notably, Notch3highCXCR4high. Newly, our in vivo results suggest that an anomalous immature thymocyte subpopulation, such as CD4-CD8- (DN) over-expressing CD3ɛ, but with low CXCR4 expression, dominates N3-ICtg thymus-resident DN subset in T-ALL progression. MicroRNAs might be of significance in T-ALL pathobiology, however, whether required for leukemia maintenance is not fully understood. The selection of specific DN subsets demonstrates the inverse correlation between CXCR4 expression and a panel of Notch3-deregulated miRNAs. Interestingly, we found that within DN thymocyte subset hyperactive Notch3 inhibits CXCR4 expression through the cooperative effects of miR-139-5p and miR-150-5p, thus impinging on thymocyte differentiation with accumulation of DNCD3ɛ+CXCR4- cells. These data point out that deregulation of Notch3 in T-ALL, besides its role in sustaining dissemination of abnormal DP T cells, as we previously demonstrated, could play a role in selecting specific DN immature T cells within the thymus, thus impeding T cell development, to facilitate T-ALL progression inside the bone marrow.

Overview of BH3 mimetics in ovarian cancer.

Ovarian carcinoma is the leading cause of gynecological cancer-related death, still with a dismal five-year prognosis, mainly due to late diagnosis and the emergence of resistance to cytotoxic and targeted agents. Bcl-2 family proteins have a key role in apoptosis and are associated with tumor development/progression and response to therapy in different cancer types, including ovarian carcinoma. In tumors, evasion of apoptosis is a possible mechanism of resistance to therapy. BH3 mimetics are small molecules that occupy the hydrophobic pocket on pro-survival proteins, allowing the induction of apoptosis, and are currently under study as single agents and/or in combination with cytotoxic and targeted agents in solid tumors. Here, we discuss recent advances in targeting anti-apoptotic proteins of the Bcl-2 family for the treatment of ovarian cancer, focusing on BH3 mimetics, and how these approaches could potentially offer an alternative/complementary way to treat patients and overcome or delay resistance to current treatments.

Bcl-2 dependent modulation of Hippo pathway in cancer cells.

INTRODUCTION: Bcl-2 and Bcl-xL are the most studied anti-apoptotic members of Bcl-2 family proteins. We previously characterized both of them, not only for their role in regulating apoptosis and resistance to therapy in cancer cells, but also for their non-canonical functions, mainly including promotion of cancer progression, metastatization, angiogenesis, and involvement in the crosstalk among cancer cells and components of the tumor microenvironment. Our goal was to identify transcriptional signature and novel cellular pathways specifically modulated by Bcl-2. METHODS: We performed RNAseq analysis of siRNA-mediated transient knockdown of Bcl-2 or Bcl-xL in human melanoma cells and gene ontology analysis to identify a specific Bcl-2 transcriptional signature. Expression of genes modulated by Bcl-2 and associated to Hippo pathway were validated in human melanoma, breast adenocarcinoma and non-small cell lung cancer cell lines by qRT-PCR. Western blotting analysis were performed to analyse protein expression of upstream regulators of YAP and in relation to different level of Bcl-2 protein. The effects of YAP silencing in Bcl-2 overexpressing cancer cells were evaluated in migration and cell viability assays in relation to different stiffness conditions. In vitro wound healing assays and co-cultures were used to evaluate cancer-specific Bcl-2 ability to activate fibroblasts. RESULTS: We demonstrated the Bcl-2-dependent modulation of Hippo Pathway in cancer cell lines from different tumor types by acting on upstream YAP regulators. YAP inhibition abolished the ability of Bcl-2 to increase tumor cell migration and proliferation on high stiffness condition of culture, to stimulate in vitro fibroblasts migration and to induce fibroblasts activation. CONCLUSIONS: We discovered that Bcl-2 regulates the Hippo pathway in different tumor types, promoting cell migration, adaptation to higher stiffness culture condition and fibroblast activation. Our data indicate that Bcl-2 inhibitors should be further investigated to counteract cancer-promoting mechanisms.

Correction: Zwergel et al. Novel Quinoline Compounds Active in Cancer Cells through Coupled DNA Methyltransferase Inhibition and Degradation. Cancers 2020, 12, 447.

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