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Several single nucleotide polymorphisms (SNPs) of the vitamin D receptor (VDR) have been observed in association with susceptibility to various pathologies, including autism, major depression, age-related changes in cognitive functioning, and Parkinson's and Alzheimer's diseases. This study aimed to establish the association between Fok1 / Apa1 polymorphic variants and anxious/depressive symptoms in nonclinical adolescents from central Italy, with the goal of identifying the risk of developing both symptoms. We found no significant difference in genotype distribution or dominant/recessive models of Fok1 / Apa1 VDR polymorphic variants between subjects with anxious/depressive symptoms and controls. HN9.10e cell lines carrying the AA genotype for Fok1 and the CC genotype for Apa1 responded better to treatment with vitamin D3 than cell lines carrying the AG genotype for Fok1 and CA genotype for Apa1 . Cell lines carrying the GG genotype for Fok1 and the AA genotype for Apa1 did not respond at all, suggesting avenues for future studies in both the general population and individuals with mental and/or neuropsychiatric disorders. These studies suggest that the level of response to vitamin D3 administered to prevent and/or treat mental or neurological disorders could depend on the polymorphic variants of the vitamin D receptor.

The S100B protein is abundant in the nervous system, mainly in astrocytes, and is also present in other districts. Among these, the adipose tissue is a site of concentration for the protein. In the light of consistent research showing some associations between S100B and adipose tissue in the context of obesity, metabolic disorders, and diabetes, this review tunes the possible role of S100B in the pathogenic processes of these disorders, which are known to involve the adipose tissue. The reported data suggest a role for adipose S100B in obesity/diabetes processes, thus putatively re-proposing the role played by astrocytic S100B in neuroinflammatory/neurodegenerative processes.

The role of sphingomyelin metabolism and vitamin C in cancer has been widely described with conflicting results ranging from a total absence of effect to possible preventive and/or protective effects. The aim of this study was to establish the possible involvement of sphingomyelin metabolism in the changes induced by vitamin C in breast cancer cells. The MCF7 cell line reproducing luminal A breast cancer and the MDA-MB-231 cell line reproducing triple-negative breast cancer were used. Cell phenotype was tested by estrogen receptor, progesterone receptor, human epidermal growth factor receptor 2 expression, and proliferation index percentage. Sphingomyelin was localized by an EGFP-NT-Lys fluorescent probe. Sphingomyelin metabolism was analyzed by RT-PCR, Western blotting and UFLC-MS/MS. The results showed that a high dose of vitamin C produced reduced cell viability, modulated cell cycle related genes, and changed the cell phenotype with estrogen receptor downregulation in MCF7 cell. In these cells, the catabolism of sphingomyelin was promoted with a large increase in ceramide content. No changes in viability and molecular expression were observed in MB231 cells. In conclusion, a high dose of vitamin C induces changes in the luminal A cell line involving sphingomyelin metabolism.

Indoleamine 2,3-dioxygenase 2 (IDO2) is a paralog of Indoleamine 2,3-dioxygenase 1 (IDO1), a tryptophan-degrading enzyme producing immunomodulatory molecules. However, the two proteins are unlikely to carry out the same functions. IDO2 shows little or no tryptophan catabolic activity and exerts contrasting immunomodulatory roles in a context-dependent manner in cancer and autoimmune diseases. The recently described potential non-enzymatic activity of IDO2 has suggested its possible involvement in alternative pathways, resulting in either pro- or anti-inflammatory effects in different models. In a previous study on non-small cell lung cancer (NSCLC) tissues, we found that IDO2 expression revealed at the plasma membrane level of tumor cells was significantly associated with poor prognosis. In this study, the A549 human cell line, basally expressing IDO2, was used as an in vitro model of human lung adenocarcinoma to gain more insights into a possible alternative function of IDO2 different from the catalytic one. In these cells, immunocytochemistry and isopycnic sucrose gradient analyses confirmed the IDO2 protein localization in the cell membrane compartment, and the immunoprecipitation of tyrosine-phosphorylated proteins revealed that kinase activities can target IDO2. The different localization from the cytosolic one and the phosphorylation state are the first indications for the signaling function of IDO2, suggesting that the IDO2 non-enzymatic role in cancer cells is worthy of deeper understanding.

In the last decade, cholesterol level has been implicated in several types of cancer, including breast cancer. In the current study, we aimed to investigate the condition of lipid depletion, hypocholesterolemia or hypercholesterolemia reproduced in vitro to analyze the response of different human breast cancer cells. Thus, MCF7 as the luminal A model, MB453 as the HER2 model and MB231 as the triple-negative model were used. No effect on cell growth and viability was detected in MB453 and MB231 cells. In MCF7 cells, hypocholesterolemia (1) reduced cell growth, and Ki67 expression; (2) increased ER/PgR expression; (3) stimulated the 3-Hydroxy-3-Methylglutaryl-CoA reductase and neutral sphingomyelinase and; (4) stimulated the expression of CDKN1A gene coding cyclin-dependent kinase inhibitor 1A protein, GADD45A coding growth arrest and DNA-damage-inducible alpha protein and, PTEN gene coding phosphatase and tensin homolog. All these effects were exacerbated by the lipid-depleted condition and reversed by the hypercholesterolemic condition. The relationship between cholesterol level and sphingomyelin metabolism was demonstrated. In summary, our data suggest that cholesterol levels should be controlled in luminal A breast cancer.

Vitamin D3, known to regulate bone homeostasis, has recently been shown to have many pleiotropic effects in different tissues and organs due to the presence of its receptor in a wide range of cells. Our previous study demonstrated that vitamin D3 was able to increase the wound healing respect to the control sample, 24 h after cutting, without however leading to a complete repair. The aim of the study was to combine vitamin D3 with silver nanoparticles to possibly enable a faster reparative effect. The results showed that this association was capable of inducing a complete wound healing only after 18 h. Moreover, a treatment of vitamin D3 + silver nanoparticles yielded a small percentage of keratinocytes vimentin-positive, suggesting the possibility that the treatment was responsible for epithelial to mesenchymal transition of the cells, facilitating wound healing repair. Since vitamin D3 acts via sphingolipid metabolism, we studied the expression of gene encoding for the metabolic enzymes and protein level. We found an increase in neutral sphingomyelinase without involvement of neutral ceramidase or sphingosine kinase2. In support, an increase in ceramide level was identified by Ultrafast Liquid Chromatography-Tandem Mass Spectrometry, suggesting a possible involvement of ceramides in wound healing process.

Immune checkpoint inhibitors (ICIs) have revolutionized the treatment of lung cancer. However, their clinical benefit is limited to a minority of patients. To unravel immune-related factors that are predictive of sensitivity or resistance to immunotherapy, we performed a gene expression analysis by RNA-Seq using the Oncomine Immuno Response Assay (OIRRA) on a total of 33 advanced NSCLC patients treated with ICI evaluating the expression levels of 365 immune-related genes. We found four genes (CD1C, HLA-DPA1, MMP2, and TLR7) downregulated ( p < 0.05) and two genes (IFNB1 and MKI67) upregulated ( p < 0.05) in ICI-Responders compared to ICI-Non-Responders. The Bayesian enrichment computational analysis showed a more complex interaction network that involved 10 other genes (IFNA1, TLR4, CD40, TLR2, IL12A, IL12B, TLR9, CD1E, IFNG, and HLA-DPB1) correlated with different functional groups. Five main pathways were identified (FDR < 0.0001). High TLR7 expression levels were significantly associated with a lack of response to immunotherapy ( p < 0.0001) and worse outcome in terms of both PFS ( p < 0.001) and OS ( p = 0.03). The multivariate analysis confirmed TLR7 RNA expression as an independent predictor for both poor PFS (HR = 2.97, 95% CI, 1.16-7.6, p = 0.023) and OS (HR = 2.2, 95% CI, 1-5.08, p = 0.049). In conclusion, a high TLR7 gene expression level was identified as an independent predictor for poor clinical benefits from ICI. These data could have important implications for the development of novel single/combinatorial strategies TLR-mediated for an efficient selection of "individualized" treatments for NSCLC in the era of immunotherapy.

Metastatic prostate cancer (mPCa) is a disease for which to date there is not curative therapy. Even the recent and attractive immunotherapeutic approaches targeting PD-L1, an immune checkpoint protein which helps cancer cells to escape from immunosurveillance, have proved ineffective. A better understanding of the molecular mechanisms contributing to keep an immunosuppressive microenvironment associated with tumor progression and refractoriness to PD-L1 inhibitors is urgently needed. In the present study, by using gene silencing and specific activators or scavengers, we demonstrated, in mPCa cell models, that methylglyoxal (MG), a potent precursor of advanced glycation end products (AGEs), especially 5-hydro-5-methylimidazolone (MG-H1), and its metabolizing enzyme, glyoxalase 1 (Glo1), contribute to maintain an immunosuppressive microenvironment through MG-H1-mediated PD-L1 up-regulation and to promote cancer progression. Moreover, our findings suggest that this novel mechanism might be responsible, at least in part, of mPCa resistance to PD-L1 inhibitors, such as atezolizumab, and that targeting it may sensitize cells to this PD-L1 inhibitor. These findings provide novel insights into the mechanisms of mPCa immunosurveillance escape and help in providing the basis to foster in vivo research toward novel therapeutic strategies for immunotherapy of mPCa.

Besides platinum-based chemotherapy, no established treatment option exists for advanced non-small-cell lung cancer (NSCLC) patients with EGFR exon 20 (Ex20ins) insertion mutations. We sought to determine the clinical outcome of patients with this EGFR mutation subtype in the immunotherapy era. Thirty NSCLCs with EGFR Ex20ins mutations were identified, of whom 15 had received immune checkpoint blockade (ICB) treatment as monotherapy (N = 12), in combination with chemotherapy (N = 2) or with another immunotherapeutic agent (N = 1). The response rate was observed in 1 out of 15 patients (6.7%), median progression-free survival (PFS) was 2.0 months and median overall survival (OS) was 5.3 months. A trend towards an inferior outcome in terms of PFS and OS was observed for patients receiving ICB treatment in the first versus second line setting (PFS: 1.6 months versus 2.7 months, respectively, p = 0.16-OS: 2.0 months versus 8.1 months, respectively, p = 0.09). Median OS from the time of diagnosis of advanced disease was shorter for patients treated with ICB versus those who did not receive immunotherapy (12.9 months versus 25.2 months, respectively, p = 0.08), which difference remained associated with a worse survival outcome at multivariate analysis ( p = 0.04). Treatment with ICB is poorly effective in NSCLCs with EGFR Ex20ins mutations, especially when given in the first-line setting. This information is crucial in order to select the optimal treatment strategy for patients with this subtype of EGFR mutation.

The enzyme indoleamine 2,3-dioxygenase 1 (IDO1) degrade tryptophan (Trp) into kynurenine (Kyn) at the initial step of an enzymatic pathway affecting T cell proliferation. IDO1 is highly expressed in various cancer types and associated with poor prognosis. Nevertheless, the serum Kyn/Trp concentration ratio has been suggested as a marker of cancer-associated immune suppression. We measured Kyn and Trp in blood samples of a wide cohort of non-small-cell lung cancer (NSCLC) patients, before they underwent surgery, and analyzed possible correlations of the Kyn/Trp ratio with either IDO1 expression or clinical-pathological parameters. Low Kyn/Trp significantly correlated with low IDO1 expression and never-smoker patients; while high Kyn/Trp was significantly associated with older (≥68 years) patients, advanced tumor stage, and squamous cell carcinoma (Sqcc), rather than the adenocarcinoma (Adc) histotype. Moreover, high Kyn/Trp was associated, among the Adc group, with higher tumor stages (II and III), and, among the Sqcc group, with a high density of tumor-infiltrating lymphocytes. A trend correlating the high Kyn/Trp ratio with the probability of recurrences from NSCLC was also found. In conclusion, high serum Kyn/Trp ratio, associated with clinical and histopathological parameters, may serve as a serum biomarker to optimize risk stratification and therapy of NSCLC patients.

Programmed death ligand 1 (PD-L1) expression is a predictive biomarker of the success of PD-1/PD-L1 inhibitor therapy for patients with advanced non-small cell lung cancer (NSCLC) but its role as a prognostic marker for early-stage resectable NSCLC remains unclear. We studied gene expression levels of immune-related genes PD-1, PD-L1, PD-L2, IDO-1, IDO-2 and INFγ in tumor tissue of surgically resected NSCLC and correlated the finding with clinicopathological features and patient outcomes. A total of 191 consecutive early-stage NSCLC patients who underwent curative pulmonary resection were studied. The mRNA expression levels of immune-related genes were evaluated by quantitative reverse transcription polymerase chain reaction (qRT-PCR) using RT 2 Profiler PCR Arrays (Qiagen). PD-1, PD-L2 and IDO-2 gene expression levels were significantly higher in patients with squamous histology ( p = 0.001, p = 0.021 and p < 0.001; respectively). PD-1, PD-L1 and IDO-2 gene expression levels were significantly higher in patients with higher stage ( p = 0.005, p = 0.048 and p = 0.002, respectively). The univariate analysis for recurrence-free survival (RFS) and overall survival (OS) showed that patients with higher levels of three-genes (PD-L1/PD-L2/INFγ) (hazard ratio (HR)) 1.90 (95% confidence interval (CI), 1.13-3.21), p = 0.015) were associated with a worse RFS, while patients with higher levels of both genes (PD-L1/IDO-2) or (PD-L2/IDO-1) were associated with a worse OS (HR 1.63 95% CI, 1.06-2.51, p = 0.024; HR 1.54 95% CI, 1.02-2.33, p = 0.04; respectively). The multivariate interaction model adjusted for histology and stage confirmed that higher levels of three genes (PD-L1/PD-L2/INFγ) were significantly associated with worse RFS (HR 1.98, p = 0.031) and higher levels of both genes (PD-L1/IDO-2) and (PD-L2/IDO-1) with worse OS (HR 1.98, p = 0.042, HR 1.92, p = 0.022). PD-L1/IDO-2 and PD-L2/IDO-1 co-expression high levels are independent negative prognostic factors for survival in early NSCLC. These features may have important implications for future immune-checkpoint therapeutic approaches.

Aryl hydrocarbon receptor (AhR) is expected to promote initiation, progression and invasion of cancer cells regulating proliferation, differentiation, gene expression, inflammation, cell motility and migration. Furthermore, an immunosuppressant function of AhR has been recognized. This study evaluated AhR expression and its role in thyroid cancer progression. AhR expression was assessed by qPCR in 107 thyroid cancer samples (90 PTCs, 11 MTCs, 6 ATCs), and by immunohistochemistry in 41 PTCs. To estimate receptor activation, the expression of target genes CYP1A1 and CYP1B1 was measured. AhR functional effects were evaluated in kynurenine-stimulated FTC-133 and BcPap cell lines by analyzing the expression of genes involved in EMT and cell motility. AhR mRNA expression resulted significantly higher in all the analyzed thyroid cancer samples compared to normal thyroid and a statistically significant correlation with CYP1B1 was detected. Kynurenine-stimulated FTC-133 and BcPap showed the activation of a specific AhR-driven EMT program characterized by E-cadherin decrease and SLUG, N-cadherin and fibronectin increase, resulting in boost of cell motility and invasion. This study confirmed the importance of the IDO1-Kyn-AhR pathway in thyroid cancer tumorigenesis, suggesting an AhR pivotal role in mediating an immunosuppressive microenvironment and favoring the acquisition of a mesenchymal phenotype that could promote invasiveness and metastasis.