FXYD3 functionally demarcates an ancestral breast cancer stem cell subpopulation with features of drug-tolerant persisters
Mengjiao Li, … , Theodoros Foukakis, Noriko Gotoh
Mengjiao Li, … , Theodoros Foukakis, Noriko Gotoh
Published November 15, 2023
Citation Information: J Clin Invest. 2023;133(22):e166666. https://doi.org/10.1172/JCI166666.
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FXYD3 functionally demarcates an ancestral breast cancer stem cell subpopulation with features of drug-tolerant persisters

Abstract

The heterogeneity of cancer stem cells (CSCs) within tumors presents a challenge in therapeutic targeting. To decipher the cellular plasticity that fuels phenotypic heterogeneity, we undertook single-cell transcriptomics analysis in triple-negative breast cancer (TNBC) to identify subpopulations in CSCs. We found a subpopulation of CSCs with ancestral features that is marked by FXYD domain–containing ion transport regulator 3 (FXYD3), a component of the Na+/K+ pump. Accordingly, FXYD3+ CSCs evolve and proliferate, while displaying traits of alveolar progenitors that are normally induced during pregnancy. Clinically, FXYD3+ CSCs were persistent during neoadjuvant chemotherapy, hence linking them to drug-tolerant persisters (DTPs) and identifying them as crucial therapeutic targets. Importantly, FXYD3+ CSCs were sensitive to senolytic Na+/K+ pump inhibitors, such as cardiac glycosides. Together, our data indicate that FXYD3+ CSCs with ancestral features are drivers of plasticity and chemoresistance in TNBC. Targeting the Na+/K+ pump could be an effective strategy to eliminate CSCs with ancestral and DTP features that could improve TNBC prognosis.

Authors

Mengjiao Li, Tatsunori Nishimura, Yasuto Takeuchi, Tsunaki Hongu, Yuming Wang, Daisuke Shiokawa, Kang Wang, Haruka Hirose, Asako Sasahara, Masao Yano, Satoko Ishikawa, Masafumi Inokuchi, Tetsuo Ota, Masahiko Tanabe, Kei-ichiro Tada, Tetsu Akiyama, Xi Cheng, Chia-Chi Liu, Toshinari Yamashita, Sumio Sugano, Yutaro Uchida, Tomoki Chiba, Hiroshi Asahara, Masahiro Nakagawa, Shinya Sato, Yohei Miyagi, Teppei Shimamura, Luis Augusto E. Nagai, Akinori Kanai, Manami Katoh, Seitaro Nomura, Ryuichiro Nakato, Yutaka Suzuki, Arinobu Tojo, Dominic C. Voon, Seishi Ogawa, Koji Okamoto, Theodoros Foukakis, Noriko Gotoh

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Figure 7

Knockdown of ATP1B1 sensitizes TNBC PDX tumors to paclitaxel treatment and decreases proportion of FXYD3-positive ancestor-like CSCs.

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Knockdown of ATP1B1 sensitizes TNBC PDX tumors to paclitaxel treatment a...
(A) Images of tumors generated in mice. (B) Tumor growth curves during paclitaxel treatment. n = 4 for each condition of P3 PDX. Statistical significance was determined by 2-way ANOVA with 2-stage linear step-up procedure of Benjamini, Krieger, and Yekutieli post hoc tests. Results are shown as means ± SEM. (C) Left: Immunofluorescence staining of frozen tissues of PDX tumors using antibodies against NRP1 and FXYD3; nuclei were stained using Hoechst 33342. Yellow arrows indicate cells double-positive for NRP1 and FXYD3. Scale bars: 20 μm. Right: Quantification of the ratio (percent) of NRP1 and FXYD3 double-positive cells to total NRP1-positive cells. n = 16–20 random fields were collected for each condition. Outliers were excluded with the ROUT method before statistical analysis. Statistical significance was determined by 2-way ANOVA with 2-stage linear step-up procedure of Benjamini, Krieger, and Yekutieli post hoc tests. Results are shown as means ± SEM. (D) Images of tumors generated in mice. Combo, combination of paclitaxel and ouabain. (E) Tumor growth curves during paclitaxel treatment. n = 8 for each condition of P3 PDX. Statistical significance was determined by 2-way ANOVA with 2-stage linear step-up procedure of Benjamini, Krieger, and Yekutieli post hoc tests. Results are shown as means ± SEM. (F) Left: Immunofluorescence staining of paraffin tissues of PDX tumors using antibodies against NRP1 and FXYD3; nuclei were stained using DAPI. Yellow arrows indicate cells double-positive for NRP1 and FXYD3. Scale bars: 20 μm. Right: Quantification of the ratio (percent) of NRP1 and FXYD3 double-positive cells to total NRP1-positive cells. n = 21 random fields were collected for each condition. Outliers were excluded with the ROUT method before statistical analysis. Statistical significance was determined by 2-way ANOVA with 2-stage linear step-up procedure of Benjamini, Krieger, and Yekutieli post hoc tests. Results are shown as means ± SEM. (G) Quantification of the ejection fraction (EF) by transthoracic echocardiography. n = 7 mice for each condition. Statistical significance was determined by unpaired, 2-tailed Student’s t test. Results are shown as means ± SEM.