Triterpenes from Pholiota populnea as Cytotoxic Agents and Chemosensitizers to Overcome Multidrug Resistance of Cancer Cells.

The detailed mycochemical analysis of the n-hexane extract of Pholiota populnea led to the isolation of four new lanostane diesters, named pholiols A-D (1-4), together with an acyclic triterpene, (3S,6E,10E,14E,18E,22S)-2,3,22,23-tetrahydroxy-2,6,10,15,19,23-hexamethyl-6,10,14,18-tetracosatetraene (5), ergosterol (6), and 3ß-hydroxyergosta-7,22-diene (7). The isolation was carried out by multistep flash chromatography, and the structures were elucidated using extensive spectroscopic analyses, including 1D and 2D NMR and MS measurements. The isolated metabolites (1-6) were investigated for cytotoxic activity against Colo205 and Colo320 colon adenocarcinoma and nontumoral MRC-5 cell lines. Among the tested compounds, ergosterol (6) showed substantial cytotoxic activity against all cell lines with IC50 values of 4.9 µM (Colo 205), 6.5 µM (Colo 320), and 0.50 µM (MRC) with no tumor cell selectivity. A P-glycoprotein efflux pump modulatory test on resistant Colo320 cells revealed that pholiols A (1) and B (2) and linear triterpene polyol 5 have the capacity to inhibit the efflux-pump overexpressed in the cells. Moreover, the drug interactions of triterpenes with doxorubicin were studied by the checkerboard method on Colo 320 cells. Pholiols B (2) and D (4) interacted in synergistic and acyclic triterpene 5 in a very strong synergistic manner; the combination index (CI) values at 50% of the growth inhibition dose (ED50) were found to be 0.348, 0.660, and 0.082, respectively. Our results indicate that P. populnea is a promising source for finding new triterpenes with significant chemosensitizing activity on cancer cells.

Fingolimod, a sphingosine-1 phosphate receptor modulator, increases BDNF levels and improves symptoms of a mouse model of Rett syndrome.

The functional relevance of brain-derived neurotrophic factor (BDNF) is beginning to be well appreciated not only in mice, but also in humans. Because reduced levels typically correlate with impaired neuronal function, increasing BDNF levels with well-tolerated drugs diffusing into the central nervous system may help in ameliorating functional deficits. With this objective in mind, we used the sphingosine-1 phosphate receptor agonist fingolimod, a drug that crosses the blood-brain barrier. In addition, fingolimod has recently been introduced as the first oral treatment for multiple sclerosis. In cultured neurons, fingolimod increases BDNF levels and counteracts NMDA-induced neuronal death in a BDNF-dependent manner. Ongoing synaptic activity and MAPK signaling is required for fingolimod-induced BDNF increase, a pathway that can also be activated in vivo by systemic fingolimod administration. Mice lacking Mecp2, a gene frequently mutated in Rett syndrome, show decreased levels of BDNF, and fingolimod administration was found to partially rescue these levels as well as the size of the striatum, a volumetric sensor of BDNF signaling in rodents. These changes correlate with increased locomotor activity of the Mecp2-deficient animals, suggesting that fingolimod may improve the functional output of the nervous system, in addition to its well-documented effects on lymphocyte egress from lymph nodes.

Disease modeling using embryonic stem cells: MeCP2 regulates nuclear size and RNA synthesis in neurons.

Mutations in the gene encoding the methyl-CpG-binding protein MECP2 are the major cause of Rett syndrome, an autism spectrum disorder mainly affecting young females. MeCP2 is an abundant chromatin-associated protein, but how and when its absence begins to alter brain function is still far from clear. Using a stem cell-based system allowing the synchronous differentiation of neuronal progenitors, we found that in the absence of MeCP2, the size of neuronal nuclei fails to increase at normal rates during differentiation. This is accompanied by a marked decrease in the rate of ribonucleotide incorporation, indicating an early role of MeCP2 in regulating total gene transcription, not restricted to selected mRNAs. We also found that the levels of brain-derived neurotrophic factor (BDNF) were decreased in mutant neurons, while those of the presynaptic protein synaptophysin increased at similar rates in wild-type and mutant neurons. By contrast, nuclear size, transcription rates, and BDNF levels remained unchanged in astrocytes lacking MeCP2. Re-expressing MeCP2 in mutant neurons rescued the nuclear size phenotype as well as BDNF levels. These results reveal a new role of MeCP2 in regulating overall RNA synthesis in neurons during the course of their maturation, in line with recent findings indicating a reduced nucleolar size in neurons of the developing brain of mice lacking Mecp2.

Isolation of the Lanostane Triterpenes Pholiols L-S from Pholiota populnea and Evaluation of Their Antiproliferative and Cytotoxic Activities.

Pholiols L-S (1−8), eight undescribed triterpenes were isolated from the sporocarps of the mushroom Pholiota populnea. Various chromatographic techniques, such as open column chromatography, flash chromatography, gel filtration, preparative thin layer chromatography, and HPLC, were applied to purify the compounds. The structure elucidation was carried out by spectroscopic analysis, including 1D (1H NMR and 13C JMOD) and 2D NMR (1H-1H COSY, HSQC, HMBC and NOESY) and HRESIMS experiments. The isolated compounds had lanostane (1−7) or trinorlanostane (8) skeletons; all of them were substituted with 3-hydroxy-3-methylglutaroyl group or its 6-methyl ester. Five compounds (1, 2, 4, 6, and 8) were investigated for their antiproliferative and cytotoxic activity in vitro by MTT assay on breast cancer (MCF-7), human colon adenocarcinoma (sensitive Colo 205, and resistant Colo 320), non-small cell lung cancer (A549), and human embryonic lung fibroblast (MRC-5) cell lines. Pholiols M (2) and O (4) showed antiproliferative activity against the MCF-7 cell line with IC50 of 2.48 and 9.95 µM, respectively. These compounds displayed tumor cell selectivity on MCF-7 cells with SI values of >40 (2) and 4.3 (4), but they did not show a cytotoxic effect, proving their action exclusively on tumor cell proliferation. Pholiols L (1) and Q (8) were found to have selective cytotoxicity on drug resistant cells in comparison to their effects on Colo320 and Colo205 cells [relative resistance values 0.84 (1) and 0.62 (8)].

Pholiols E-K, lanostane-type triterpenes from Pholiota populnea with anti-inflammatory properties.

Investigation of the chloroform and ethyl acetate extracts of the edible mushroom Pholiota populnea led to the isolation of eight triterpenes, the undescribed natural products pholiols E-K and the known (+)-clavaric acid. HRESIMS and 1D and 2D NMR spectroscopy were employed to determine the structures of the undescribed compounds. The NOESY spectra were used to assign the relative configurations of triterpenes. The isolated compounds were screened for their anti-inflammatory activity on cyclooxygenase (COX-1 and COX-2), and lipoxygenase (5-LOX and 15-LOX) inhibitory assays. Dose-response investigations revealed that lanostane derivatives exhibited moderate 5-LOX and COX-2 inhibitory activities, with pholiol F (IC50 194.5 µM against 5-LOX and 439.8 µM against COX-2) the most active among the isolated compounds. Our findings indicated that P. populnea is an abundant source of new bioactive lanostane-type triterpenes.