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Meperidine (pethidine) is a µ-opioid receptor (MOR) agonist widely used in the treatment of cancer pain. While MOR agonists in experimental models have demonstrated both pro- and antitumorigenic properties, meperidine has unique features which may be predominantly anticancer in nature. Meperidine both inhibits NMDA (N-methyl-D-Aspartate) receptors, which are involved in the progression of glioblastoma, and blocks NADH:Ubiquinone Oxidoreductase, which may hinder mitochondrial respiration. In the developing embryonic neural tissue, meperidine reduces cell proliferation around the neural tube and lowers the expression of the B RE (brain and reproductive organ-expressed). This is notable given that the B RE gene is implicated in cancer chemoresistance and gliomagenesis. Further, meperidine inhibits P-glycoprotein, which is involved in cancer multidrug resistance and the degradation of the sphingolipid backbone, ceramide. By enhancing the pro-autophagic and pro-apoptotic ceramide levels in cancer cells, meperidine stimulates cell death and reverses multidrug resistance. Tamoxifen, a safe medication employed in the treatment of breast cancer, directly blocks P-glycoprotein and boosts levels of ceramide both via inhibition of glycosylceramide synthase and ceramidase. Further, tamoxifen blocks NMDA-neurotoxicity and therefore it may act synergistically with meperidine in reducing glioblastoma progression associated with NMDA-activation. Finally, tamoxifen blocks glycolysis which may enhance the mitochondrial-blocking activity of meperidine to shut down energy metabolism of glioblastoma cells. Because of these properties, we believe that the combination of meperidine and tamoxifen merits study in cell culture and animal models to investigate a potential synergistic relationship in the treatment of glioblastoma.

Objectives: To define ultrastructural features accompanying to antitumor effects of gemcitabine, vinorelbine and cyclooxygenase inhibitors in C6 glioma cells in vitro. Vinorelbine is a semisynthetic vinca alkaloid and recent studies showed its antitumor activity in pediatric optic and pontine gliomas. Vinorelbine infusion induces a severe tumor site-pain in systemic cancers, but it is unknown whether algesia and inflammation contribute to its antitumor effects. Gemcitabine is a nucleoside-chemotherapeutic which was recently shown to act as a radiosensitizer in high-grade glioma. Some studies showed synergism of anti-inflammatory cyclooxygenase-inhibitors with microtubule inhibitors and gemcitabine. DMSO is a solvent and blocks both cylooxygenase and ribonucleotide reductase, another target of gemcitabine. Rofecoxib is withdrawn from the market, yet we used it for investigational purposes, since it blocks cylooxygenase-2 1000-times more potently than cylooxygenase-1 and is also a selective inhibitor of crinophagy.Methods: Plating efficacy, 3D-spheroid S-phase analysis with BrdU labelling and transmission electron micro-scopical analyses were performed.Results: Vinorelbine induced frequent mitotic slippage/apoptosis and autophagy. Despite both DMSO and rofecoxib induced autophagy alone and in synergy, they reduced mitotic catastrophe and autophagy triggered by vinorelbine, which was also reflected by reduced inhibition of spheroid S-phase. Gemcitabine induced karyolysis and margination of coarse chromatin towards the nuclear membrane, abundant autophagy, gutta adipis formation and decrease in mitochondria, which were enhanced by DMSO and rofecoxib.Conclusions: Detailed ultrastructural analysis of the effects of chemotherapeutic drugs may provide a broader insight about their actions and pave to develop better strategies in treatment of glioblastoma.

While pregnancy may accelerate glioblastoma multiforme (GBM) growth, parity and progesterone (P4) containing treatments (ie, hormone replacement therapy) reduce the risk of GBM development. In parallel, low and high doses of P4 exert stimulating and inhibitory actions on GBM growth, respectively. The mechanisms behind the high‐dose P4‐suppression of GBM growth is unknown. In the present study, we assessed the changes in growth and proteomic profiles when high‐dose P4 (100 and 300 µM) was administered in human U87 and A172 GBM cell lines. The xCELLigence system was used to examine cell growth when different concentrations of P4 (20, 50, 100, and 300 µM) was administered. The protein profiles were determined by two‐dimensional gel electrophoresis in both cell lines when 100 and 300 µM P4 were administered. Finally, the pathways enriched by the differentially expressed proteins were assessed using bioinformatic tools. Increasing doses of P4 blocked the growth of both GBM cells. We identified 26 and 51 differentially expressed proteins (fc > 2) in A172 and U87 cell lines treated with P4, respectively. Only the pro‐tumorigenic mitochondrial ornithine aminotransferase and anti‐apoptotic mitochondrial 60 kDa heat shock protein were downregulated in A172 cell line and U87 cell line when treated with P4, respectively. Detoxification of reactive oxygen species, cellular response to stress, glucose metabolism, and immunity‐related proteins were altered in P4‐treated GBM cell lines. The paradox on the effect of low and high doses of P4 on GBM growth is gaining attention. The mechanism related to the high dose of P4 on GBM growth can be explained by the alterations in detoxification mechanisms, stress, and immune response and glucose metabolism. P4 suppresses GBM growth and as it is nontoxic in comparison to classical chemotherapeutics, it can be used as a new strategy in GBM treatment in the future., GBM cell line impedance alterations when different concentrations of Progesterone were administered.

High grade glial tumors (HGGs) including anaplastic astrocytoma (WHO Grade-III) and glioblastoma multiforme (GBM, WHO Grade-IV) are among the most malignant cancers known to man. Due to their defective mitochondria, HGG cells consume glucose via glycolysis even in the presence of oxygen. Overall survival is worse in HGG patients that are hyperglycemic. Unlike normal neural cells, HGG cells cannot efficiently metabolize ketone bodies for energy. Thus, a metabolic treatment based on therapeutic ketosis (reduced glucose with elevated ketone bodies) was proposed to treat GBM and was supoported from preclinical studies. Caprylic (octanoic) acid, a monocarboxylated saturated fatty acid, is among the best producers of ketone bodies and induces necrosis of experimental tumors at high dose. Caprylic acid is enriched in coconut and in goat's milk. It is also a post-translational modifier of the ghrelin hormone and is produced in trace amounts in human tissues. Caprylic acid is a straight-chain isomer of the antiepileptic valproic acid, which is used in treatment of HGG-associated seizures and which may increase survival in GBM patients according to epidemiological observations. Among the valproic acids analogs tested, caprylic acid is the most potent molecule to block C6 astrocytoma cell growth in vitro and accumulates selectively within glial cells as shown by Positron Emission Tomography in vivo. Caprylic acid blocks glycolysis both in healthy liver and in malignant liver cells, which is more prominent in the latter and also lowers blood glucose. Noteworthy, caprylic acid exerts neuroprotective- and mitochondria-protective effects in several models of neurodegenerative diseases. Boost injections of caprylic acid at non-toxic levels during classical ketogenic metabolic therapy may fortify antitumor actions and reduce systemic toxicity by differential programming of mitochondrial and other metabolic pathways.

Glioblastomas (GBMs) are primary brain tumors with extremely bad prognosis and therefore; discovery of novel regulators of their pathology is of immense importance. LncRNAs (long noncoding RNAs) regulate nuclear structure, embryonic pluripotency, cell differentiation, development and carcinogenesis. Many lncRNAs have weak evolutionary conservation; however, a nuclear lncRNA, MALAT1 (metastasis-associated lung adenocarcinoma transcript 1), is exceptionally conserved and is among the most abundant lncRNAs in benign tissues. The majority of cell culture studies and clinico-epidemiological studies demonstrated that MALAT1 acts a tumor promoter in GBMs and inhibition of MALAT1 suppressed tumor growth in various preclinical models of GBM. MALAT1 involves in stemness of GBM cells by regulating SOX2, nestin and members of WNT pathway. MALAT1 induces protective autophagy and suppresses apoptosis in GBM cells via sponging miRNA-101 and increases temozolomide chemoresistance via enhancing epithelial-mesenchymal transition, suppressing miR-203 and promoting thymidilate synthase. Moreover, knockdown of MALAT1 expression enhances blood-brain tumor barrier permeability via miR-140, which may provide a double benefit of MALAT1 suppression by increasing the delivery of chemotherapy agents into the GBM tissues. On the other hand, there also exist some cell culture and animal studies showing that MALAT1 acts as a tumor suppressor in GBMs by suppression of ERK/MAPK and MMP2 signaling and by repression of miR-155 with subsequent increase of FBXW7. Whether protective or detrimental, MALAT1 seems to be an important component of GBM pathogenesis and hence; novels studies are needed in versatile models, including many different primary GBM cultures, orthotopic and xenogreft in vivo models and transgenic mice.

Objective Glial tumor growth may accelerate during gestation, but epidemiological studies consistently demonstrated that parousity reduces life long risk of glial tumors. Pregnancy may also accelerate growth of medulloblastoma and meningioma, but parousity does not confer protection against these tumors. We were the first to show that medroxyprogesterone acetate (MPA) reduces rat C6 glioma growth in vitro. Now we aimed to determine the effects of MPA on human brain cancers (particularly glioblastoma) in vitro and C6 glioma in vivo. Patients and Methods We evaluated the effects of MPA on: i) monolayer growth of human U87 and U251 glioblastoma, ii) 3D-spheroid growth and invasion of C6 rat glioma and human U251 glioma, iii) interactions with PI3-Kinase inhibitors and coxsackie-adenovirus receptor (CAR) in modifying 3D-spheroid invasion of glioma. Results MPA at low doses (3.25–13 μM) insignificantly stimulated and at high doses (above 52 μM) strongly suppressed the growth of human U87 and U251 cells in vitro. MPA also binds to glucocorticoid receptors similar to dexamethasone (Dex) and unexpectedly, PI3-Kinase inhibitors at low doses suppressed anti-invasive efficacies of MPA and Dex. MPA exerted higher invasion-inhibitory effects on CAR-expressing human glioma cells. Lastly, MPA suppressed growth of C6 glioma implanted into rat brain. Conclusion Progesterone analogues deserve to be studied in future experimental models of high grade glial brain tumors.

Background Epidemiological studies indicate that aspirin consumption reduces the risk of tumors, which is especially relevant for colonic adenoma and carcinoma. Similar observations were made for glial brain tumors and breast cancers, yet the results are inconsistent. Gentisic acid (GA) is a minor catabolite of aspirin; yet humans carrying CYP2C9-variants incapable to catabolize aspirin to GA do not benefit from aspirin in prevention against colonic adenoma. GA blocks binding of Fibroblastic Growth Factor to its receptor and its sulphonate metabolite dobesilic acid blocks growth of C6 glioblastoma in vivo. GA is also an endogenously produced siderophore in mammalians for the transport of iron, a trace element which stimulates tumor growth and enhances anthracycline cardiotoxicity. Materials and Methods In this study, we assessed whether GA exerts direct antitumor activity on C6 glioma cells in vitro (cytotoxicity, colony growth, 3H-thymidine labeling analysis of DNA synthesis); and whether it can modify growth of Ehrlich breast ascites carcinoma (EAC) and solid tumors (EST) in vivo. GA and antitumoral trace element selenium block 12-lipoxygenase activity and aspirin's paradoxical inflammatory effects are seen in selenium-deficient humans; thus, we also investigated antitumor interactions between GA and sodium selenite. Lastly, we evaluated whether GA could protect against doxorubicin cardiotoxicity due to its function to chelate iron. Results Clinically achievable doses of GA blocked growth, colony formation and DNA synthesis of C6 glioma in vitro with high significance. GA enhanced the survival of EAC-bearing mice at a dosage of 0.4 mg/mice/day, in which 33% of the treated animals survived more than 3-weeks, when all untreated mice succumbed to their tumors. Selenium decreased EST volumes initially, yet increased tumor volumes at later stages in surviving mice. GA alone reduced solid tumor growth and did not modify selenite antineoplasticity initially, but blocked the late tumor-stimulating effects of selenite. Lastly, doxorubicin-induced cardiac myofibrillary and endothelial damage and hyalinization necrosis were attenuated with GA treatment. Conclusions GA highly merits to be studied in further animal models as an anticancer and chemoprotective drug.

Graphical abstract, Highlights • IQGAP proteins demonstrated differential expression between CV-PTC and FV-PTC. • Proteome similarities between benign and FV-PTC hampered accurate unsupervised clustering. • Changes in actin cytoskeleton proteins play predominant roles in thyroid pathology., Introduction Fine Needle Aspiration Biopsy (FNAB) allows the cytological differentiation of benign and malignant thyroid nodules. However, the method itself is not adequate in determining some cases. For example, the diagnosis of Follicular Variant Papillary Thyroid Carcinoma (FV-PTC) can be challenging. In the current study we investigate the protein profiles of FV-PTC and classical variant PTC (CV-PTC) with no lymph node metastasis and compare it with benign thyroid tissue. Method We used CV-PTC (n = 6), FV-PTC (n = 6) and benign thyroid tissues (n = 6) to prepare tissue lysates. Proteins from each group were trypsin and lys-C digested. The samples were analyzed on a Q Exactive Orbitrap mass spectrometer. Results We identified 2560 proteins across all 18 specimens. Protein profiles revealed that there was no clear distinction between benign and FV-PTC samples. However, further examination of our data showed that proteins in energy metabolism have altered in FV-PTC. Proteomic pathway analysis showed marked alteration of the actin cytoskeleton proteins, especially several members of Arp2/3 complex were significantly increased in CV-PTC. We made the novel observation that IQGAP1 protein was significantly increased in CV-PTC, whereas IQGAP2 protein was highly expressed in FV-PTC lesions, suggesting differential roles of IQGAP proteins in thyroid pathology. Conclusion In the present study, mass spectrometry based label free quantification approach was applied to investigate the protein profiles of FV-PTC, CV-PTC and benign thyroid tissues. This study pointed out that actin cytoskeleton proteins, IQGAP proteins and changes in energy metabolism play predominant roles in thyroid pathology.

25 (OH) vitamin D3 (25(OH)D) and parathyroid hormone (PTH) are important regulators of calcium homeostasis. The aim of this study was to retrospectively determine the cut-off for sufficient 25(OH)D in a four-season region and the influence of age, seasons, and gender on serum 25(OH)D and PTH levels.Laboratory results of 9890 female and 2723 male individuals aged 38.8±22.1 years who had simultaneous measurements of 25(OH)D and PTH were retrospectively analyzed by statistical softwares. Serum 25(OH)D and PTH levels were measured by a mass spectrometry method and by an electrochemiluminescence immunoassay, respectively.Mean serum 25(OH)D levels showed a sinusoidal fluctuation throughout the year and were significantly (p0.01) higher in summer and autumn. On the other hand, PTH levels were significantly higher (p0.01) in women and showed an opposite response to seasonal effects relative to 25(OH)D. Lowest levels of 25(OH)D were detected in people aged between 20 and 40 years whereas PTH hormone levels were gradually increasing in response to aging. The significant exponential inverse relationship that was found between PTH and 25(OH)D (PTH=exp(4.12-0.064Our retrospective study based on large data set supports the suitability of the currently accepted clinical cut-off of 75 nmol/L for sufficient 25(OH)D. However, the issue of assessing Vitamin D deficiency remains difficult due to seasonal variations in serum 25(OH)D. Therefore, PTH measurements should complement 25(OH)D results for diagnosing Vitamin D deficiency. It is imperative that seasonally different criteria should be considered in future.25 (OH) vitamin D3 (25(OH)D) i paratireoidni hormon (PTH) imaju važnu ulogu u regulisanju homeostaze kalcijuma. Cilj ove studije bio je da se retrospektivno odrede cut-off vrednosti za dovoljan nivo 25(OH)D u regionu sa četiri godišnja doba, kao i uticaj starosti, godišnjeg doba i pola na nivoe 25(OH)D i PTH u serumu.Laboratorijski rezultati 9890 žena i 2723 muška rca starosti 38,8±22,1 godina kod kojih su istovremeno mereni 25(OH)D i PTH retrospektivno su analizirani statističkim softverom. Nivoi 25(OH)D i PTH u serumu mereni su metodom masene spektrometrije, odnosno elektrohemiluminiscencije.Srednji nivoi 25(OH)D pokazali su sinusoidnu fluktuaciju tokom cele godine i bili su značajno viši (p0,01) u leto i jesen. S druge strane, nivoi PTH bili su značajno viši (p0,01) kod žena i pokazali su suprotan odgovor na sezonske efekte u odnosu na 25(OH)D. Najniži nivoi 25(OH)D otkriveni su kod ljudi starosti između 20 i 40 godina, dok su hormonski nivoi PTH sa starenjem bili u postepenom porastu. Značajan eksponencijalni obrnut odnos koji je utvrđen između PTH i 25(OH)D (PTH=exp(4,12–0,064Naša retrospektivna studija zasnovana na velikim skupinama podataka potvrđuje da je trenutno prihvaćena klinička cut-off vrednost od 75 nmol/L prigodna za dovoljan nivo 25(OH)D. Međutim, utvrđivanje nedostatka vitamina D ostaje teško izvodljivo usled varijacija u serumskom nivou 25(OH)D zbog godišnjih doba. Stoga, merenje PTH trebalo bi da posluži kao komplement rezultatima 25(OH)D prilikom dijagnostikovanja nedostatka vitamina D. Najvažnije je da se u budućnosti razmotre različiti kriterijumi prilagođeni različitim godišnjim dobima.

BACKGROUND: Risk of high-grade gliomas is lower in young females and its incidence enhances after menopause suggesting a likely protective role of female hormones. Hormone replacement therapy (HRT) is being employed to treat osteoporosis and some epidemiological studies showed that HRT containing progesterone analogs decrease the risk of glial tumors. Tibolone is a unique progesterone analog used in HRT with tissue-specific estrogenic effects. According to some studies, tibolone may enhance the risk of mammary and female reproductive cancers; but peculiarly, there exist several basic scientific data that it acts antitumoral at higher concentrations than which can be achieved in HRT. This feature resembles to another progesterone analog, medroxyprogesterone acetate (MPA). MPA enhances risk of breast cancer in HRT protocols; nonetheless, at high dosages, it can cause regression of far advanced breast, endometrium and renal cancers. Moreover, molecular studies have shown that MPA and tibolone induce very similar gene expression patterns. Since tibolone’s pro-estrogenic effects occur particularly in bone and brain, we have hypothesized that high dose tibolone with both progestagenic and estrogenic actions may block glioma growth alone and/or in synergy with MPA or temozolomide. MATERIAL AND METHODS: Soft agar colony assay, 3H-thymidine test to determine the S-phase (DNA synthesis phase) and transmission electron microscopy were performed to determine the effects of tibolone on C6 rat glioma. RESULTS: Tibolone dose-dependently blocked soft agar colony growth of C6 glioma. Tibolone potentiated MPA-induced growth inhibition in soft agar colonies. Tibolone potentiated temozolomide’s effect in reducing colony growth. Tibolone inhibited DNA synthesis in C6 rat glioma spheroids (as assessed by BrdU-labeling index) to similar levels which can be achieved with temozolomide. Transmission electron microscopical analyses revealed that mutual interactions between tibolone, MPA and temozolomide involve mitochondrial condensation, mitophagy and autophagy. CONCLUSION: Tibolone merits to be studied in further models of glioblastoma in vitro and in vivo.

Background: Perchlorate, thiocyanate, and nitrate can block iodide transport at the sodium iodide symporter (NIS) and this can subsequently lead to decreased thyroid hormone production and hypothyroidism. NIS inhibitor exposure has been shown to reduce iodide uptake and thyroid hormone levels; therefore we hypothesized that maternal NIS inhibitor exposure will influence both maternal and newborn thyroid function.Methods: Spot urine samples were collected from 185 lactating mothers and evaluated for perchlorate, thiocyanate, and nitrate concentrations. Blood and colostrum samples were collected from the same participants in the first 48 h after delivery. Thyroid hormones and thyroid-related antibodies (TSH, fT3, fT4, anti-TPO, anti-Tg) were analyzed in maternal blood and perchlorate was analyzed in colostrum. Also, spot blood samples were collected from newborns (n = 185) between 48 and 72 postpartum hours for TSH measurement. Correlation analysis was performed to assess the effect of NIS inhibitors on thyroid hormone levels of lactating mothers and their newborns in their first 48 postpartum hours.Results: The medians of maternal urinary perchlorate (4.00 μg/g creatinine), maternal urinary thiocyanate (403 μg/g creatinine), and maternal urinary nitrate (49,117 μg/g creatinine) were determined. Higher concentrations of all three urinary NIS inhibitors (μg/g creatinine) at their 75th percentile levels were significantly correlated with newborn TSH (r = 0.21, p < 0.001). Median colostrum perchlorate level concentration of all 185 participants was 2.30 μg/L. Colostrum perchlorate was not significantly correlated with newborn TSH (p > 0.05); however, there was a significant correlation between colostrum perchlorate level and maternal TSH (r = 0.21, p < 0.01). Similarly, there was a significant positive association between colostrum perchlorate and maternal urinary creatinine adjusted perchlorate (r = 0.32, p < 0.001).Conclusion: NIS inhibitors are ubiquitous in lactating women in Turkey and are associated with increased TSH levels in newborns, thus signifying for the first time that co-exposure to maternal NIS inhibitors can have a negative effect on the newborn thyroid function.

The single nucleotide polymorphism rs55705857, located in a non-coding but evolutionarily conserved region at 8q24.21, is strongly associated with IDH-mutant glioma development and was suggested to be a causal variant. However, the molecular mechanism underlying this association has remained unknown. With a case control study in 285 gliomas, 316 healthy controls, 380 systemic cancers, 31 other CNS-tumors, and 120 IDH-mutant cartilaginous tumors, we identified that the association was specific to IDH-mutant gliomas. Odds-ratios were 9.25 (5.17–16.52; 95% CI) for IDH-mutated gliomas and 12.85 (5.94–27.83; 95% CI) for IDH-mutated, 1p/19q co-deleted gliomas. Decreasing strength with increasing anaplasia implied a modulatory effect. No somatic mutations were noted at this locus in 114 blood-tumor pairs, nor was there a copy number difference between risk-allele and only-ancestral allele carriers. CCDC26 RNA-expression was rare and not different between the two groups. There were only minor subtype-specific differences in common glioma driver genes. RNA sequencing and LC-MS/MS comparisons pointed to significantly altered MYC-signaling. Baseline enhancer activity of the conserved region specifically on the MYC promoter and its further positive modulation by the SNP risk-allele was shown in vitro. Our findings implicate MYC deregulation as the underlying cause of the observed association., Scientific Reports, 6, ISSN:2045-2322

In a field trial on a farm in Turkey, we tested whether specific egg-yolk antibodies had a prophylactic effect on neonatal diarrhea. Owing to its antibody spectrum, this egg powder was very suitable for this farm since mainly rotavirus was identified. Regardless of the dosage (2 g, 4 g, or 8 g egg powder), the calves ( n = 164) which received specific egg-yolk antibodies via the whole egg during the first 14 days of life showed significant improvements in risk and duration of diarrhea and in body weight gain compared with calves of the control group ( n = 80) which had not received any egg powder. These significant differences in body weight still persisted after 3 months of life. The mortality risk in the control group amounted to 8.8%, while in all egg powder groups ( n = 164) only one calf died as a result of diarrhea. Breed-dependent differences in the parameters were not observed.