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HSD17B4 methylation enhances glucose dependence of BT-474 breast cancer cells and increases lapatinib sensitivity

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Abstract

Purpose

HER2-positive breast cancer has a high chance of achieving pathological complete response when HSD17B4, responsible for peroxisomal β-oxidation of very long-chain fatty acids (VLCFA) and estradiol, is methylation-silenced. Here, we aimed to identify the underlying molecular mechanism.

Methods

Using a HER2-positive breast cancer cell line, BT-474, control and knock-out (KO) clones were obtained. Metabolic characteristics were analyzed using a Seahorse Flux analyzer.

Results

HSD17B4 KO suppressed cellular proliferation, and enhanced sensitivity to lapatinib approximately tenfold. The KO led to accumulation of VLCFA and a decrease of polyunsaturated fatty acids (PUFAs), such as docosahexaenoic acid (DHA) and arachidonic acid. HSD17B4 KO increased Akt phosphorylation, possibly via decreased DHA, and genes involved in oxidative phosphorylation (OxPhos) and electron transport chain (ETC) were upregulated. Increased mitochondrial ATP production in the KO cells was confirmed by extracellular flux analyzer. Increased OxPhos led to severe dependence of the KO cells on pyruvate from glycolysis. Suppression of glycolysis by lapatinib led to severe delayed suppression of OxPhos in KO cells.

Conclusion

HSD17B4 KO in BT-474 cells caused a decrease of PUFAs, increased Akt phosphorylation, enhanced glucose dependence of OxPhos, and increased sensitivity to inhibition of HER2, upstream of Akt. This mechanism may be applicable to other HER2-positive glucose-dependent breast cancer cells with HSD17B4 silencing.

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Data availability

All data generated or analyzed during this study are included in this published article and its supplementary information files.

Abbreviations

HSD17B4 :

Hydroxysteroid 17-β Dehydrogenase 4

OxPhos :

Oxidative phosphorylation

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Acknowledgements

The authors would like to thank Dr. Hideki Makinoshima for his professional advice and Ms. Kana Kimura, Ms. Asuka Dosen, and Ms. Mika Wakabayashi for their technical assistance. We also thank Mr. Edward McMahon and Mr. Robert Staunton for their kind proofreading.

Funding

This research was supported by AMED under grant numbers JP19ck0106466 and JP23ck0106832.

Author information

Authors and Affiliations

  1. Division of Epigenomics, National Cancer Center Research Institute, Tokyo, Japan

    Nobuaki Arai, Naoko Hattori, Satoshi Yamashita, Yu-Yu Liu, Takahiro Ebata, Chihiro Takeuchi, Hideyuki Takeshima & Toshikazu Ushijima

  2. Department of Surgery, Kyorin University School of Medicine, Tokyo, Japan

    Nobuaki Arai & Haruhiko Kondo

  3. Department of Epigenomics, Institute for Advanced Life Sciences, Hoshi University, Tokyo, Japan

    Naoko Hattori, Yu-Yu Liu, Takahiro Ebata, Chihiro Takeuchi, Hideyuki Takeshima & Toshikazu Ushijima

  4. Department of Molecular Pathology, Yokohama City University Graduate School of Medicine, Kanagawa, Japan

    Satoshi Fujii

  5. Department of Medical Oncology, National Cancer Center Hospital East, Chiba, Japan

    Hirofumi Mukai

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  1. Nobuaki Arai
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Contributions

Conception and design: NA, NH, SF, HK, HM, and TU. Execution of experiments: NA, NH, SY, TE, CT, and HT. Analysis and interpretation of data: NA, NH, SY, and YYL. Writing, reviewing, and/or revision the manuscript: NA, NH, and TU.

Corresponding author

Correspondence to Toshikazu Ushijima.

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Supplementary Information

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Supplementary file1 (PDF 148 kb)

Supplementary Figure 1. Establishment of HSD17B4 KO clones. Complete loss of HSD17B4 protein was confirmed in KO#7 and KO#42.

Supplementary figure 2. Variation in lactate dehydrogenase A (LDHA) mRNA expression among the clones. LDHA expression was approximately 1.5 times higher in KO#7 than in KO#42.

Supplementary figure 3. A. Rescue of HSD17B4 in HCC1954, which had HSD17B4 methylation, confirmed by Western blot analysis. B. Sensitivity of control and rescue clones to lapatinib. HSD17B4 rescue cells did not show resistance to lapatinib in either normoxic (left panel) or hypoxic condition (right panel).

Supplementary figure 4. Sensitivity of control and HSD17B4 KO cells to trastuzumab. HSD17B4 KO cells did not show resistance to trastuzumab in either normoxic (left panel) or hypoxic condition (right panel).

Supplementary file2 (XLSX 3106 kb)

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Arai, N., Hattori, N., Yamashita, S. et al. HSD17B4 methylation enhances glucose dependence of BT-474 breast cancer cells and increases lapatinib sensitivity. Breast Cancer Res Treat 201, 317–328 (2023). https://doi.org/10.1007/s10549-023-07013-y

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