Fbxo22-mediated KDM4B degradation determines selective estrogen receptor modulator activity in breast cancer
Yoshikazu Johmura, Ichiro Maeda, Narumi Suzuki, Wenwen Wu, Atsushi Goda, Mariko Morita, Kiyoshi Yamaguchi, Mizuki Yamamoto, Satoi Nagasawa, Yasuyuki Kojima, Koichiro Tsugawa, Natsuko Inoue, Yasuo Miyoshi, Tomo Osako, Futoshi Akiyama, Reo Maruyama, Jun-ichiro Inoue, Yoichi Furukawa, Tomohiko Ohta, Makoto Nakanishi
Yoshikazu Johmura, Ichiro Maeda, Narumi Suzuki, Wenwen Wu, Atsushi Goda, Mariko Morita, Kiyoshi Yamaguchi, Mizuki Yamamoto, Satoi Nagasawa, Yasuyuki Kojima, Koichiro Tsugawa, Natsuko Inoue, Yasuo Miyoshi, Tomo Osako, Futoshi Akiyama, Reo Maruyama, Jun-ichiro Inoue, Yoichi Furukawa, Tomohiko Ohta, Makoto Nakanishi
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Research Article Endocrinology

Fbxo22-mediated KDM4B degradation determines selective estrogen receptor modulator activity in breast cancer

Abstract

The agonistic/antagonistic biocharacter of selective estrogen receptor modulators (SERMs) can have therapeutic advantages, particularly in the case of premenopausal breast cancers. Although the contradictory effects of these modulators have been studied in terms of crosstalk between the estrogen receptor α (ER) and coactivator dynamics and growth factor signaling, the molecular basis of these mechanisms is still obscure. We identify a series of regulatory mechanisms controlling cofactor dynamics on ER and SERM function, whose activities require F-box protein 22 (Fbxo22). Skp1, Cullin1, F-box–containing complex (SCFFbxo22) ubiquitylated lysine demethylase 4B (KDM4B) complexed with tamoxifen-bound (TAM-bound) ER, whose degradation released steroid receptor coactivator (SRC) from ER. Depletion of Fbxo22 resulted in ER-dependent transcriptional activation via transactivation function 1 (AF1) function, even in the presence of SERMs. In living cells, TAM released SRC and KDM4B from ER in a Fbxo22-dependent manner. SRC release by TAM required Fbxo22 on almost all ER-SRC–bound enhancers and promoters. TAM failed to prevent the growth of Fbxo22-depleted, ER-positive breast cancers both in vitro and in vivo. Clinically, a low level of Fbxo22 in tumor tissues predicted a poorer outcome in ER-positive/human epidermal growth factor receptor type 2–negative (HER2-negative) breast cancers with high hazard ratios, independently of other markers such as Ki-67 and node status. We propose that the level of Fbxo22 in tumor tissues defines a new subclass of ER-positive breast cancers for which SCFFbxo22-mediated KDM4B degradation in patients can be a therapeutic target for the next generation of SERMs.

Authors

Yoshikazu Johmura, Ichiro Maeda, Narumi Suzuki, Wenwen Wu, Atsushi Goda, Mariko Morita, Kiyoshi Yamaguchi, Mizuki Yamamoto, Satoi Nagasawa, Yasuyuki Kojima, Koichiro Tsugawa, Natsuko Inoue, Yasuo Miyoshi, Tomo Osako, Futoshi Akiyama, Reo Maruyama, Jun-ichiro Inoue, Yoichi Furukawa, Tomohiko Ohta, Makoto Nakanishi

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Figure 7

Fbxo22 is required for 4-OHT–mediated inhibition of breast cancer cell growth both in vitro and in vivo.

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Fbxo22 is required for 4-OHT–mediated inhibition of breast cancer cell g...
(A) The indicated Dox-shRNA-MCF7 cells with or without doxycycline-inducible FLAG-Fbxo22 were starved of E2 in the presence of doxycycline (1 μg/ml) for 72 hours, treated with E2 (10 nM) in the presence or absence of 4-OHT (100 nM) for 6 hours, and then subjected to a quantitative colony formation assay. Representative images and results (mean ± SD) are from 3 independent experiments. (B) MCF7 cells expressing the indicated doxycycline-inducible shRNAs were treated as in A and subjected to a quantitative colony formation assay. Results are shown as the mean ± SD of 3 independent experiments. (C) Tumor growth of the control (n = 5) or Fbxo22-KO (n = 5) T47D cells transplanted into the mammary fat pads of NOD/Scid mice was measured over a 2-week period with E2 pellet supplementation and then over a 4-week period with TAM pellet supplementation. *P < 0.05, by 2-tailed Welch’s t test. (D) Mice as in C were sacrificed 6 weeks after transplantation. The tumors were then excised and weighed. *P < 0.05, by 2-tailed Welch’s t test. (E) Images of tumors excised from mice as in D are shown. The weight of each tumor is indicated below the images. (F) Paraffin-embedded tumor sections from 5 mice harboring WT or Fbxo22-KO T47D cells were subjected to immunohistochemical analyses using anti–Ki-67 antibodies and antibodies against cleaved caspase 3. Ki-67–positive and cleaved caspase 3–positive cells were counted, and their numbers were normalized to that of cell nuclei in each section. *P < 0.05 and ***P < 0.05, by2-tailed Student’s t test.