The neuro-steroid, 3β androstene 17α diol exhibits potent cytotoxic effects on human malignant glioma and lymphoma cells through different programmed cell death pathways

Abstract

The neuro-steroids 3b-androstene-17a-diol (17a-AED), 3b-androstene-17b-diol (17b-AED), 3b-androstene-7a,-17b-triol (7a-AET) and 3b-androstene-7b,-17b-triol (7b-AET) are metabolites of dehydroepiandrosterone and are produced in neuro-ectodermal tissue. Both epimers of androstenediols (17a-AED and 17b-AED) and androstenetriols (7a-AET and 7b-AET) have markedly different biological functions of their chemical analogue. We investigated the cytotoxic activity of these neuro-steroids on human T98G and U251MG glioblastoma and U937 lymphoma cells. Proliferation studies showed that 17a-AED is the most potent inhibitor, with an IC 50 B15 mM. For T98G glioma, 90% inhibition was achieved with 25 mM of 17a-AED. Other neuro-steroids tested only marginally suppressed cell proliferation. Reduced cell adherence and viability could be detected after 18 h of 17a-AED exposure. Treatment with 17a-AED induced a significant level of apoptosis in U937 lymphoma cells, but not in the glioma cells. Cytopathology of 17a-AED-treated T98G cells revealed the presence of multiple cytoplasmic vacuoles. Acridine orange staining demonstrated the formation of acidic vesicular organelles in 17a-AED-treated T98G and U251MG, which was inhibited by bafilomycin A1. These findings indicate that 17a-AED bears the most potent cytotoxic activity of the neuro-steroids tested, and the effectiveness may depend on the number of hydroxyls and their position on the androstene molecule. These cytotoxic effects may utilize a nonapoptotic pathway in malignant glioma cells.

Autophagy and the functional roles of Atg5 and beclin-1 in the anti-tumor effects of 3β androstene 17α diol neuro-steroid on malignant glioma cells

“…In contrast, 17␣-AED treatment did not cause T98G and U251MG malignant gliomas to undergo apoptosis although the neurosteroid did have an anti-tumor effect on these two glioma cell lines [13]. These findings indicated the functional specificity of the anti-tumor effects of the adrostenediol molecules in terms of the orientation of the hydroxyl group on C-17 (␣-or ␤-orientation) and that a different cell death pathway may be utilized in tumors of glial origins as compared to other tumors exposed to 17␣-AED.…”

“…The proliferation of tumor cells was assessed by the incorporation of 3 H-TdR (Amersham Biosciences, Piscataway, NJ) as described by Graf et al [13]. Briefly, glioma cells (1 × 10 4 /well) were cultured in a 96-well, tissue culture plate in the presence of 17␣-AED (3-200 M, provided by Dr. Loria) for 3 days.…”

“…When 3-MA was used, cells were pre-incubated for 1 h with the inhibitor before 17␣-AED treatment. After 6-72 h, non-adherent and adherent cells were collected and cell lysates were prepared and immunoblotting was conducted as previously described [13]. Briefly, total protein was resolved on a 4-12% NuPAGE MES gels (Invitrogen) and transferred to nitrocellulose membranes.…”

Temozolomide induces autophagy via ATM-AMPK-ULK1 pathways in glioma

“…Quantification of autophagy by acridine orange staining using flow cytometry was performed as described previously (16,17). Briefly, following drug treatment, acridine orange (sc-358795, Santa Cruz Biotechnology, Inc.) was added at a final concentration of 1 µg/ml for a period of 15 min.…”

Differential Effect of Wortmannolone Derivatives on MDA-MB-231 Breast Cancer Cells

“…All the furanosteroids 1–6 have derived structure from 1α,2α-epoxy-3β-hydroxyandrosta-5,8-dieno[6,5,4- bc ]furan-7,17-dione and were screened for ROS inducing activity, NF-κB inhibitory, and cytotoxic activity in hormone-independent TNBC cells (7). Furanosteroids 1 – 6 displayed several different biological activities (Table I)(8, 9). Herein, the androsta-5, 8-dienofuran structure was modified and subsequently the anti-proliferative effects were examined for the obtained derivatives.…”