IL-6 mediates platinum-induced enrichment of ovarian cancer stem cells
Yinu Wang, Xingyue Zong, Sumegha Mitra, Anirban Kumar Mitra, Daniela Matei, Kenneth P. Nephew
Yinu Wang, Xingyue Zong, Sumegha Mitra, Anirban Kumar Mitra, Daniela Matei, Kenneth P. Nephew
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Research Article Inflammation Stem cells

IL-6 mediates platinum-induced enrichment of ovarian cancer stem cells

Abstract

In high-grade serous ovarian cancer (OC), chemotherapy eliminates the majority of tumor cells, leaving behind residual tumors enriched in OC stem cells (OCSC). OCSC, defined as aldehyde dehydrogenase–positive (ALDH+), persist and contribute to tumor relapse. Inflammatory cytokine IL-6 is elevated in residual tumors after platinum treatment, and we hypothesized that IL-6 plays a critical role in platinum-induced OCSC enrichment. We demonstrate that IL-6 regulates stemness features of OCSC driven by ALDH1A1 expression and activity. We show that platinum induces IL-6 secretion by cancer-associated fibroblasts in the tumor microenvironment, promoting OCSC enrichment in residual tumors after chemotherapy. By activating STAT3 and upregulating ALDH1A1 expression, IL-6 treatment converted non-OCSC to OCSC. Having previously shown altered DNA methylation in OCSC, we show here that IL-6 induces DNA methyltransferase 1 (DNMT1) expression and the hypomethylating agent (HMA) guadecitabine induced differentiation of OCSC and reduced — but did not completely eradicate — OCSC. IL-6 neutralizing antibody (IL-6-Nab) combined with HMA fully eradicated OCSC, and the combination blocked IL-6/IL6-R/pSTAT3–mediated ALDH1A1 expression and eliminated OCSC in residual tumors that persisted in vivo after chemotherapy. We conclude that IL-6 signaling blockade combined with an HMA can eliminate OCSC after platinum treatment, supporting this strategy to prevent tumor recurrence after standard chemotherapy.

Authors

Yinu Wang, Xingyue Zong, Sumegha Mitra, Anirban Kumar Mitra, Daniela Matei, Kenneth P. Nephew

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Figure 5

IL-6-Nab–guadecitabine combination eliminates ALDH+ cells by inhibiting ALDH1A1 expression.

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IL-6-Nab–guadecitabine combination eliminates ALDH+ cells by inhibiting ...
(A) FACS analysis of the percentage of Kuramochi-derived ALDH+ cells treated daily with guadecitabine (100 nM), IL-6-Nab (1 μg/ml), or IL-6-Nab– guadecitabine for 4 days. Average percentage of ALDH+ cells ± SD is shown on the side scatter graph (left), and the quantification is shown (right) (n = 3, 1-way ANOVA). (B) Average fold change of ALDH1A1 expression ± SD in Kuramochi-derived ALDH+ cells treated with guadecitabine, IL-6-Nab, or IL-6-Nab–guadecitabine compared with control cells was determined by qPCR (n = 3, 1-way ANOVA, *P < 0.05 and ***P < 0.001) (C) Kuramochi-derived ALDH+ cells were treated daily with guadecitabine (100 nM), IL-6-Nab (1 μg/ml), or IL-6-Nab–guadecitabine for 4 days. The protein expression of ALDH1A1, DNMT1, pSTAT3, STAT3, and GAPDH was determined by western blot (n = 2). (D) Spheroid formation assay of 500 conditioned Kuramochi cells, which were treated with control, IL-6-Nab (400 ng/ml), and guadecitabine (100 nM, 3 days) or in combination. Cells were directly sorted in 96 low-attached plates and cultured in stem cell condition for 14 days. Representative images and the average number of spheroids ± SD are shown in the graph. Scale bar: 100μm (n = 3, 1-way ANOVA, *P < 0.05). (E) Kuramochi_ALDH+ cells were treated daily with guadecitabine (100 nM), IL-6-Nab (1 μg/ml), or IL-6-Nab– guadecitabine for 4 days. Five hundred pretreated cells were reseeded into 6-well plates for clonogenic survival assays. Representative images and the average number and of colonies formed by pretreated Kuramochi_ALDH+ cells with the conditions described previously and pretreated Kuramochi_ALDH+ cells after exposure to cisplatin (3 μM) for 3 hours (n = 3, 2-way ANOVA, Dunnett’s and Sidak’s multiple comparisons tests used for multiple comparison, *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001).