L1, a 3,3′-diindolylmethane-derivative, induced ER stress-mediated apoptosis and suppressed growth through the FLI1/AKT pathway in erythroleukemia HEL cells
Introduction3,3′-Diindolylmethane (DIM) is a major phytochemical product derived from ingestion of cruciferous vegetables. As an effective cancer chemopreventive agent, DIM has been used in preclinical and clinical trials. Recently, our group synthesized and modified a novel DIM derivative, L1, and...
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Frontiers Media S.A.
2025-08-01
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| author | Yi Kuang Yi Kuang Yong Jian Yong Jian Dinghuan Wang Dinghuan Wang Lihao Bai Lihao Bai Kunlin Yu Kunlin Yu Chunlin Wang Chunlin Wang Wuling Liu Wuling Liu Sheng Liu Sheng Liu Wan Li Yaacov Ben-David Yaacov Ben-David Xiao Xiao Xiao Xiao |
| author_facet | Yi Kuang Yi Kuang Yong Jian Yong Jian Dinghuan Wang Dinghuan Wang Lihao Bai Lihao Bai Kunlin Yu Kunlin Yu Chunlin Wang Chunlin Wang Wuling Liu Wuling Liu Sheng Liu Sheng Liu Wan Li Yaacov Ben-David Yaacov Ben-David Xiao Xiao Xiao Xiao |
| author_sort | Yi Kuang |
| collection | DOAJ |
| description | Introduction3,3′-Diindolylmethane (DIM) is a major phytochemical product derived from ingestion of cruciferous vegetables. As an effective cancer chemopreventive agent, DIM has been used in preclinical and clinical trials. Recently, our group synthesized and modified a novel DIM derivative, L1, and demonstrated its significant antileukemic activities.MethodsMTT assay was used to confirm the inhibition rates and IC50 value of L1 in erythroleukemia HEL cells. Flow cytometry analysis was used to reveal cell cycle arrest and apoptosis. RNAseq data with KEGG pathway enrichment analysis was performed to predict the anticancer mechanism of L1. RT-qPCR and Western blotting were carried out to verify the mechanism in the ER stress-mediated apoptosis and FLI1/AKT pathway. FLI1 knockdown in HEL cells was performed to confirm the mechanism of L1 in the FLI1/AKT pathway. AutoDocking analysis and PPI analysis via the STRING database were used to discover the potential target of L1. HSPA1A knockdown and treatment with HSP70 inhibitor were used to further evaluate the L1 target.ResultsL1 significantly inhibited the growth of erythroleukemia HEL cells, with an IC50 value of 1.15 ± 0.03 µM L1 induced G2/M cell cycle arrest and cell apoptosis. RNA sequencing analysis revealed that differentially expressed genes (DEGs) mainly enriched in protein processing of endoplasmic reticulum (ER). L1 increased the protein expression level of GRP78 (BIP) and the RNA transcription of XBP1 and DDIT3 to induce ER stress-mediated apoptosis. Meanwhile, PPI analysis suggested that HSP70 (HSPA1A and HSPA1B) is a pivotal gene that may be involved in the ER stress. AutoDocking analysis also revealed that L1 may bind to the HSP70 protein (HSPA1A and HSPA1B). The apoptosis rate was reduced by cotreatment of L1 and the Hsp70 inhibitor VER155008. Moreover,the inhibition rate was decreased in the HSPA1A knockdown HEL cells, suggesting that L1-induced apoptosis was related to HSP70 activity. Moreover, FLI1 is a crucial target for mediating cell differentiation, apoptosis, inflammation and displays abnormal expression in HEL cells. Here, we showed that the protein expression levels of FLI1 and AKT/p-AKT decreased with L1 treatment and that the RNA expressions of their downstream genes GATA1, TFRC, GYPA, CDKN1A and CDKN1B were also regulated by L1.ConclusionThis study revealed that the DIM-derivative molecule, L1, induced ER stress-mediated apoptosis and suppressed cell growth by inhibiting the FLI1/AKT pathway in erythroleukemia HEL cells. |
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| id | doaj-art-97df816990c64c6d8223db83fd393a8a |
| institution | DOAJ |
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| language | English |
| publishDate | 2025-08-01 |
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| series | Frontiers in Pharmacology |
| spelling | doaj-art-97df816990c64c6d8223db83fd393a8a2025-08-20T03:16:21ZengFrontiers Media S.A.Frontiers in Pharmacology1663-98122025-08-011610.3389/fphar.2025.15641991564199L1, a 3,3′-diindolylmethane-derivative, induced ER stress-mediated apoptosis and suppressed growth through the FLI1/AKT pathway in erythroleukemia HEL cellsYi Kuang0Yi Kuang1Yong Jian2Yong Jian3Dinghuan Wang4Dinghuan Wang5Lihao Bai6Lihao Bai7Kunlin Yu8Kunlin Yu9Chunlin Wang10Chunlin Wang11Wuling Liu12Wuling Liu13Sheng Liu14Sheng Liu15Wan Li16Yaacov Ben-David17Yaacov Ben-David18Xiao Xiao19Xiao Xiao20State Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaInstitute of Materia Medica, Chinese Academy of Medical Science and Peking Union Medical College, Beijing, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaState Key Laboratory of Discovery and Utilization of Functional Components in Traditional Chinese Medicine, Guizhou Medical University, Guiyang, Guizhou, ChinaNatural Products Research Center of Guizhou Province, Guiyang, Guizhou, ChinaIntroduction3,3′-Diindolylmethane (DIM) is a major phytochemical product derived from ingestion of cruciferous vegetables. As an effective cancer chemopreventive agent, DIM has been used in preclinical and clinical trials. Recently, our group synthesized and modified a novel DIM derivative, L1, and demonstrated its significant antileukemic activities.MethodsMTT assay was used to confirm the inhibition rates and IC50 value of L1 in erythroleukemia HEL cells. Flow cytometry analysis was used to reveal cell cycle arrest and apoptosis. RNAseq data with KEGG pathway enrichment analysis was performed to predict the anticancer mechanism of L1. RT-qPCR and Western blotting were carried out to verify the mechanism in the ER stress-mediated apoptosis and FLI1/AKT pathway. FLI1 knockdown in HEL cells was performed to confirm the mechanism of L1 in the FLI1/AKT pathway. AutoDocking analysis and PPI analysis via the STRING database were used to discover the potential target of L1. HSPA1A knockdown and treatment with HSP70 inhibitor were used to further evaluate the L1 target.ResultsL1 significantly inhibited the growth of erythroleukemia HEL cells, with an IC50 value of 1.15 ± 0.03 µM L1 induced G2/M cell cycle arrest and cell apoptosis. RNA sequencing analysis revealed that differentially expressed genes (DEGs) mainly enriched in protein processing of endoplasmic reticulum (ER). L1 increased the protein expression level of GRP78 (BIP) and the RNA transcription of XBP1 and DDIT3 to induce ER stress-mediated apoptosis. Meanwhile, PPI analysis suggested that HSP70 (HSPA1A and HSPA1B) is a pivotal gene that may be involved in the ER stress. AutoDocking analysis also revealed that L1 may bind to the HSP70 protein (HSPA1A and HSPA1B). The apoptosis rate was reduced by cotreatment of L1 and the Hsp70 inhibitor VER155008. Moreover,the inhibition rate was decreased in the HSPA1A knockdown HEL cells, suggesting that L1-induced apoptosis was related to HSP70 activity. Moreover, FLI1 is a crucial target for mediating cell differentiation, apoptosis, inflammation and displays abnormal expression in HEL cells. Here, we showed that the protein expression levels of FLI1 and AKT/p-AKT decreased with L1 treatment and that the RNA expressions of their downstream genes GATA1, TFRC, GYPA, CDKN1A and CDKN1B were also regulated by L1.ConclusionThis study revealed that the DIM-derivative molecule, L1, induced ER stress-mediated apoptosis and suppressed cell growth by inhibiting the FLI1/AKT pathway in erythroleukemia HEL cells.https://www.frontiersin.org/articles/10.3389/fphar.2025.1564199/fullerythroleukemia3,3′-diindoylmethane derivativesendoplasmic reticulum stress-mediated apoptosisFLI1Hsp70 |
| spellingShingle | Yi Kuang Yi Kuang Yong Jian Yong Jian Dinghuan Wang Dinghuan Wang Lihao Bai Lihao Bai Kunlin Yu Kunlin Yu Chunlin Wang Chunlin Wang Wuling Liu Wuling Liu Sheng Liu Sheng Liu Wan Li Yaacov Ben-David Yaacov Ben-David Xiao Xiao Xiao Xiao L1, a 3,3′-diindolylmethane-derivative, induced ER stress-mediated apoptosis and suppressed growth through the FLI1/AKT pathway in erythroleukemia HEL cells Frontiers in Pharmacology erythroleukemia 3,3′-diindoylmethane derivatives endoplasmic reticulum stress-mediated apoptosis FLI1 Hsp70 |
| title | L1, a 3,3′-diindolylmethane-derivative, induced ER stress-mediated apoptosis and suppressed growth through the FLI1/AKT pathway in erythroleukemia HEL cells |
| title_full | L1, a 3,3′-diindolylmethane-derivative, induced ER stress-mediated apoptosis and suppressed growth through the FLI1/AKT pathway in erythroleukemia HEL cells |
| title_fullStr | L1, a 3,3′-diindolylmethane-derivative, induced ER stress-mediated apoptosis and suppressed growth through the FLI1/AKT pathway in erythroleukemia HEL cells |
| title_full_unstemmed | L1, a 3,3′-diindolylmethane-derivative, induced ER stress-mediated apoptosis and suppressed growth through the FLI1/AKT pathway in erythroleukemia HEL cells |
| title_short | L1, a 3,3′-diindolylmethane-derivative, induced ER stress-mediated apoptosis and suppressed growth through the FLI1/AKT pathway in erythroleukemia HEL cells |
| title_sort | l1 a 3 3 diindolylmethane derivative induced er stress mediated apoptosis and suppressed growth through the fli1 akt pathway in erythroleukemia hel cells |
| topic | erythroleukemia 3,3′-diindoylmethane derivatives endoplasmic reticulum stress-mediated apoptosis FLI1 Hsp70 |
| url | https://www.frontiersin.org/articles/10.3389/fphar.2025.1564199/full |
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