5‑Aminolevurinic acid inhibits the proliferation of bladder cancer cells by activating heme synthesis

Nakai,Yasushi; Tatsumi,Yoshihiro; Hori,Shunta; Morizawa,Yosuke; Iida,Kota; Onishi,Kenta; Miyake,Makito; Oda,Yuki; Owari,Takuya; Fujii,Tomomi; Onishi,Sayuri; Tanaka,Nobumichi; Fujimoto,Kiyohide

doi:10.3892/or.2022.8401

5‑Aminolevurinic acid inhibits the proliferation of bladder cancer cells by activating heme synthesis

Authors:
- Yasushi Nakai
- Yoshihiro Tatsumi
- Shunta Hori
- Yosuke Morizawa
- Kota Iida
- Kenta Onishi
- Makito Miyake
- Yuki Oda
- Takuya Owari
- Tomomi Fujii
- Sayuri Onishi
- Nobumichi Tanaka
- Kiyohide Fujimoto
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Affiliations: Department of Urology, Nara Medical University, Kashihara, Nara 634‑8522, Japan, Department of Pathology, Nara Medical University, Kashihara, Nara 634‑8522, Japan
Published online on: September 8, 2022 https://doi.org/10.3892/or.2022.8401
Article Number: 186
Copyright: © Nakai et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Iron is an essential nutrient that facilitates cell proliferation and growth, and it can contribute to tumor growth. Although iron chelators have shown great potential in preclinical cancer models, they can cause adverse side‑effects. The aim of the present study was to determine whether treatment with 5‑aminolevurinic acid (5‑ALA) has antitumor effects in bladder cancer, by reduction of mitochondrial iron without using an iron chelator, through activation of heme synthesis. T24 and MGH‑U3 cells were treated with 5‑ALA. Ferrochelatase uses iron to convert protoporphyrin IX into heme, thus additional groups of T24 and MGH‑U3 cells were transfected with synthesized ferrochelatase small interfering RNA (siRNA) either to silence ferrochelatase or to provide a negative siRNA control group, and then cell viability, apoptosis, mitochondrial Fe²⁺, the cell cycle, and ferritin expression were analyzed in all groups and compared. As an in vivo assessment, mice with orthotopic bladder cancer induced using N‑butyl‑N‑(4‑hydro‑oxybutyl) were treated with 5‑ALA. Bladder weight and pathological findings were evaluated, and immunohistochemical analysis was performed for ferritin and proliferating cell nuclear antigen (PCNA). In the cells treated with 5‑ALA, proliferation was decreased compared with the controls, and apoptosis was not detected. In addition, the expression of Fe²⁺ in mitochondria was decreased by 5‑ALA, expression of ferritin was also reduced by 5‑ALA, and the percentage of cells in the S phase of the cell cycle was significantly increased by 5‑ALA. In T24 and MGH‑U3 cells with silenced ferrochelatase, the inhibition of cell proliferation, decreased expression of Fe²⁺ in mitochondria, reduced expression of ferritin, and increased percentage of cells in the S phase by treatment with 5‑ALA were weakened. In vivo, no mouse treated with 5‑ALA developed muscle‑invasive bladder cancer. The expression of ferritin was weaker in mice treated with 5‑ALA and that of PCNA was higher than that in mice treated without 5‑ALA. It was concluded that 5‑ALA inhibited proliferation of bladder cancer cells by activating heme synthesis.

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