Proapoptotic PUMA targets stem-like breast cancer cells to suppress metastasis
Qi Sun, … , David A. Cheresh, Jay S. Desgrosellier
Qi Sun, … , David A. Cheresh, Jay S. Desgrosellier
Published December 11, 2017
Citation Information: J Clin Invest. 2018;128(1):531-544. https://doi.org/10.1172/JCI93707.
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Research Article Oncology

Proapoptotic PUMA targets stem-like breast cancer cells to suppress metastasis

Abstract

Breast cancer cells with stem cell properties are key contributors to metastatic disease, and there remains a need to better understand and target these cells in human cancers. Here, we identified rare stem-like cells in patients’ tumors characterized by low levels of the proapoptotic molecule p53-upregulated modulator of apoptosis (PUMA) and showed that these cells play a critical role in tumor progression that is independent of clinical subtype. A signaling axis consisting of the integrin αvβ3, Src kinase, and the transcription factor Slug suppresses PUMA in these cells, promoting tumor stemness. We showed that genetic or pharmacological disruption of αvβ3/Src signaling drives PUMA expression, specifically depleting these stem-like tumor cells; increases their sensitivity to apoptosis; and reduces pulmonary metastasis, with no effect on primary tumor growth. Taken together, these findings point to PUMA as a key vulnerability of stem-like cells and suggest that pharmacological upregulation of PUMA via Src inhibition may represent a strategy to selectively target these cells in a wide spectrum of aggressive breast cancers.

Authors

Qi Sun, Jacqueline Lesperance, Hiromi Wettersten, Elaine Luterstein, Yoko S. DeRose, Alana Welm, David A. Cheresh, Jay S. Desgrosellier

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Figure 7

PUMA expression is a critical vulnerability of stem-like cells.

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PUMA expression is a critical vulnerability of stem-like cells. (A) Quan...
(A) Quantification of cell death by trypan blue staining after transient transfection of MCF7 and BT549 breast cancer cell lines with PUMA cDNA or empty control vector relative to mock-transfected controls. P = 0.0262 (BT549; PUMA cDNA vs. vector control), by Student’s t test. n = 3 independent experiments run in duplicate. (B and C) Dasatinib dose-response experiments with LM2-4 cells transfected with control siRNA or either of 2 different PUMA siRNAs. n = 5 independent experiments performed in duplicate. (B) Curves were fitted by nonlinear regression. (C) IC50 was calculated from the nonlinear regression curves. P = 0.0331 (siCtrl vs. siPUMA no. 1); P = 0.0141 (siCtrl vs. siPUMA no. 2), by 1-way ANOVA with Dunnett’s multiple comparisons test. (D and E) In vivo tumor studies comparing different numbers of shCtrl and shPUMA LM2-4 cells injected orthotopically into adult female mice and treated with 30 mg/kg dasatinib or vehicle (1% citric acid) once daily by oral gavage for the first 7 days after injection. Red arrows indicate the start and end of daily drug treatment. n = 4 mice per group. (D) Graphs display the tumor volume after injection of 30,000 cells. P < 0.0001 (days 31 and 34), by 2-way, repeated-measures ANOVA with Sidak’s multiple comparisons test. (A–D) Data represent the mean ± SEM. *P < 0.05 and ***P < 0.001. (E) Tumor latency after injection of 15,000 cells. P = 0.0084 (shCtrl; vehicle vs. dasatinib); P = 0.5766 (shPUMA; vehicle vs. dasatinib), by log-rank (Mantel-Cox) test. (F) Schematic depicting how PUMA expression driven by αvβ3/Src blockade can target highly vulnerable metastatic and tumor-initiating stem-like breast cancer cells compared with cells in the primary tumor. See also Supplemental Figure 7.