NOTCH Pathway Blockade Depletes CD133-Positive Glioblastoma Cells and Inhibits Growth of Tumor Neurospheres and Xenografts  

Abstract

Cancer stem cells (CSCs) are thought to be critical for the engraftment and long-term growth of many tumors, including glioblastoma (GBM). The cells are at least partially spared by traditional chemotherapies and radiation therapies, and finding new treatments that can target CSCs may be critical for improving patient survival. It has been shown that the NOTCH signaling pathway regulates normal stem cells in the brain, and that GBMs contain stemlike cells with higher NOTCH activity. We therefore used low-passage and established GBM-derived neurosphere cultures to examine the overall requirement for NOTCH activity, and also examined the effects on tumor cells expressing stem cell markers. NOTCH blockade by c-secretase inhibitors (GSIs) reduced neurosphere growth and clonogenicity in vitro, whereas expression of an active form of NOTCH2 increased tumor growth. The putative CSC markers CD133, NESTIN, BMI1, and OLIG2 were reduced following NOTCH blockade. When equal numbers of viable cells pretreated with either vehicle (dimethyl sulfoxide) or GSI were injected subcutaneously into nude mice, the former always formed tumors, whereas the latter did not. In vivo delivery of GSI by implantation of drug-impregnated polymer beads also effectively blocked tumor growth, and significantly prolonged survival, albeit in a relatively small cohort of animals. We found that NOTCH pathway inhibition appears to deplete stem-like cancer cells through reduced proliferation and increased apoptosis associated with decreased AKT and STAT3 phosphorylation. In summary, we demonstrate that NOTCH pathway blockade depletes stem-like cells in GBMs, suggesting that GSIs may be useful as chemotherapeutic reagents to target CSCs in malignant gliomas.

Cytomegalovirus infection induces a stem cell phenotype in human primary glioblastoma cells: prognostic significance and biological impact

“…In fact, inhibition of Notch signaling significantly reduces GBM growth in vitro and in vivo, 12 likely by reducing the size or functionality of the cancer stem cell population. 33 This and other 17,34 innovative strategies indicate that translational therapies targeting GCSCs could significantly improve the prognosis of patients with GBM.…”

c-Jun N-terminal kinase has a pivotal role in the maintenance of self-renewal and tumorigenicity in glioma stem-like cells

“…Inhibition of Notch pathway exhausts CD133 þ stem-like tumor cells and suppresses sphere formation in vitro and tumor formation in vivo. 29,33,48 Furthermore, Notch signaling pathway directly upregulates the stem cell marker Nestin in glioma. 49 Consistent with these previous studies, our findings show that Notch-2 expression is upregulated in spherecultured cells, compared with monolayer-cultured cells.…”

“…These results are consistent with previous studies that glioma cell lines contain a subset of glioma stem-like cells that can be enriched under serum-free, sphereforming culture conditions. 33,36 To investigate the possible role of specific kinases in stemness of glioma cells, we first examined the activation status of mitogenactivated protein kinases in sphere-cultured glioma cells. Spherecultured glioma cells exhibited enhanced phosphorylation of ERK1/2 and JNK1/2, but not p38 (Figure 1a).…”

Notch1 signaling promotes survival of glioblastoma cells via EGFR-mediated induction of anti-apoptotic Mcl-1

“…This theory is supported by the finding that treatment of glioblastomas with g-secretase inhibitors resulted in a reduction of phosphorylated STAT3. 41 Recently, Sheng et al 31 demonstrated a transcriptional regulation of Mcl-1 via Raf-1-activated ATF5. However, we did not observe changes in ATF5 expression on downregulation of Notch1 (data not shown).…”