Differentiation Therapy Exerts Antitumor Effects on Stem-like Glioma Cells

Abstract

Purpose: Stem-like tumor cells comprise a highly tumorigenic and therapy-resistant tumor subpopulation, which is believed to substantially influence tumor initiation and therapy resistance in glioma. Currently, therapeutic, drug-induced differentiation is considered as a promising approach to eradicate this tumor-driving cell population; retinoic acid is well known as a potent modulator of differentiation and proliferation in normal stem cells. In glioma, knowledge about the efficacy of retinoic acid-induced differentiation to target the stem-like tumor cell pool could have therapeutic implications.Experimental Design: Stem-like glioma cells (SLGC) were differentiated with all-trans retinoic acid-containing medium to study the effect of differentiation on angiogenesis, invasive growth, as well as radioresistance and chemoresistance of SLGCs. In vivo effects were studied using live microscopy in a cranial window model.Results: Our data suggest that in vitro differentiation of SLGCs induces therapy-sensitizing effects, impairs the secretion of angiogenic cytokines, and disrupts SLGCs motility. Further, ex vivo differentiation reduces tumorigenicity of SLGCs. Finally, we show that all-trans retinoic acid treatment alone can induce antitumor effects in vivo.Conclusions: Altogether, these results highlight the potential of differentiation treatment to target the stem-like cell population in glioblastoma. Clin Cancer Res; 16(10); 2715-28. ©2010 AACR.

Stem cell characteristics in glioblastoma are maintained by the ecto-nucleotidase E-NPP1

“…[36][37][38] Neural stem and progenitor cells are in particular characterised by a short G1 cell cycle phase and growing evidence suggests a crucial role for lengthening of G1 phase in differentiation. 27 In line with this hypothesis, a number of experiments in GSCs describe co-occurrence of induced differentiation and reduced proliferation, 16,34,39 in particular by the accumulation of cells in G1/G0 cell cycle phase. 15,40 In agreement with these findings, ENPP1 knockdown in GSCs was found to cause a reduction in the proliferation rate and cells were found to accumulate in G1/G0 cell cycle phase.…”

“…To account for technical variability, data were normalised to the mean of negative (non-target control cells; CD133 level set to 1) and positive (TRRAP-deficient cells; CD133 level set to 0) controls of the respective plate. Untreated GSCs (Mock; one well per plate) served as further negative control, whereas GSCs treated with 10 mM all-trans retinoic acid (ATRA; two wells per plate), previously shown to induce differentiation of GSCs and thus to reduce the CD133 level, 16 was used as a second positive control. As depicted in Supplementary Figure S1B, negative and positive controls on the screening plates (n ¼ 20) exhibit only minimal variability of CD133/1 level, underlining the high quality of the screen.…”

“…Microscopic images 4 days post-transduction showed that non-target transduced GSCs efficiently formed neurospheres with regular rim and well-defined spherical shape. In contrast, knockdown of ENPP1 led to impaired neurosphere formation, with smaller spheres appearing as cell clusters with uneven rims ( As differentiation of GSCs was shown to increase apoptotic response to chemotherapeutic agents, 15,16 we tested whether knockdown of ENPP1 sensitised the cells to the cytotoxic substance 1,3-Bis (2-chloroethyl)-1-nitrosourea (BCNU). For this purpose, non-target control cells and E-NPP1-deficient GSCs were exposed to BCNU (50 and 75 mM) for 72 h. As depicted in Figure 5e and Supplementary Figure S4C, ENPP1 knockdown alone resulted in a significant increase in caspase-3/7 activity.…”

“…Several reports implicate GSCs in therapy resistance and showed that efficacy of therapy can be improved, if stem-like cells are targeted additionally. 15,16,29 These findings emphasise the need to improve understanding of the biological behaviours of GSCs and their underlying regulatory mechanisms. In that respect, a previous study identified kinases and phosphatases relevant for the survival of GSCs and thereby potential therapeutical targets for glioblastoma.…”

“…Phenotypic and genotypic characterisations of the GSC lines were carried out in previous studies. 16,52 Patient characteristics of all samples used for RNA expression analysis are summarised in Supplementary Table S5. For passaging and plating, NCH421k and NCH441 cells were dissociated using accutase (Sigma-Aldrich, St. Louis, MO, USA) and NCH644 cells by the use of a trypsin-EDTA solution (Life Technologies, Darmstadt, Germany).…”