Ganglioside GD2 identifies breast cancer stem cells and promotes tumorigenesis
Venkata Lokesh Battula, … , Sendurai A. Mani, Michael Andreeff
Venkata Lokesh Battula, … , Sendurai A. Mani, Michael Andreeff
Published May 15, 2012
Citation Information: J Clin Invest. 2012;122(6):2066-2078. https://doi.org/10.1172/JCI59735.
View: Text | PDF
Research Article Oncology

Ganglioside GD2 identifies breast cancer stem cells and promotes tumorigenesis

Abstract

Cancer stem cells (CSCs) are a small subpopulation of cancer cells that have increased resistance to conventional therapies and are capable of establishing metastasis. However, only a few biomarkers of CSCs have been identified. Here, we report that ganglioside GD2 (a glycosphingolipid) identifies a small fraction of cells in human breast cancer cell lines and patient samples that are capable of forming mammospheres and initiating tumors with as few as 10 GD2+ cells. In addition, the majority of GD2+ cells are also CD44hiCD24lo, the previously established CSC-associated cell surface phenotype. Gene expression analysis revealed that GD3 synthase (GD3S) is highly expressed in GD2+ as well as in CD44hiCD24lo cells and that interference with GD3S expression, either by shRNA or using a pharmacological inhibitor, reduced the CSC population and CSC-associated properties. GD3S knockdown completely abrogated tumor formation in vivo. Also, induction of epithelial-mesenchymal transition (EMT) in transformed human mammary epithelial cells (HMLER cells) dramatically increased GD2 as well as GD3S expression in these cells, suggesting a role of EMT in the origin of GD2+ breast CSCs. In summary, we identified GD2 as a new CSC-specific cell surface marker and GD3S as a potential therapeutic target for CSCs, with the possibility of improving survival and cure rates in patients with breast cancer.

Authors

Venkata Lokesh Battula, Yuexi Shi, Kurt W. Evans, Rui-Yu Wang, Erika L. Spaeth, Rodrigo O. Jacamo, Rudy Guerra, Aysegul A. Sahin, Frank C. Marini, Gabriel Hortobagyi, Sendurai A. Mani, Michael Andreeff

×

Figure 6

Triptolide inhibits the expression of GD3S, induces apoptosis in MDA-MB-231 cells, and blocks tumor growth in NOD/SCID mice.

Options: View larger image (or click on image) Download as PowerPoint
Triptolide inhibits the expression of GD3S, induces apoptosis in MDA-MB-...
MDA-MB-231 cells (A) or SUM-159 cells (B) (5 × 105/well) of 6-well cell culture dishes were treated with 25, 50, 75, 100, or 125 nM triptolide for 24 hours. –/–, no treatment. Total RNA was extracted, and GD3S expression was measured by qRT-PCR. (C and D) To measure GD2+ cell growth inhibition, 5 × 105 MDA-MB-231 (C) and SUM-159 (D) cells were plated in each well of 6-well cell culture dishes and treated with 25, 50, 75, 100, or 125 nM triptolide for 24 or 48 hours. After incubation, the cells were detached with trypsin and stained with anti-GD2 antibody and Sytox Red (for dead cells; Invitrogen). The stained cells were analyzed on an LSR II flow cytometer. Absolute numbers of live cells were calculated by measuring 1,000 events for Trucount beads as explained in Methods. (E) To determine the inhibition of tumor growth, 1 × 106 MDA-MB-231 cells were subcutaneously transplanted into NOD/SCID mice (n = 8; 4 mice/group). A group of the mice were treated with 0.15 mg/kg/d triptolide, and the control group was treated with PBS every day by i.p. injection. At the end of 8 weeks, mice were sacrificed, and tumors were dissected out and photographed. (F) Tumor sizes from the mice in experiment in E were measured every week after tumor engraftment, and the measurements are shown. P < 0.001, week 3. (G) The survival analysis was based on Kaplan-Meier estimation, and groups were compared by the log-rank test. Control (n = 4, black line) and triptolide (n = 4, blue line) were analyzed for cumulative survival. Survival was defined as the time (in weeks) from transplantation until death. P = 0.015.