Novel chemical-structure TPOR agonist, TMEA, promotes megakaryocytes differentiation and thrombopoiesis via mTOR and ERK signalings

Xueqin Jiang; Yueshan Sun; Shuo Yang; Yuesong Wu; Long Wang; Wenjun Zou; Nan Jiang; Jianping Chen; Yunwei Han; Chunlan Huang; Anguo Wu; Chunxiang Zhang; Jianming Wu

doi:10.1016/j.phymed.2022.154637

Novel chemical-structure TPOR agonist, TMEA, promotes megakaryocytes differentiation and thrombopoiesis via mTOR and ERK signalings

Phytomedicine. 2023 Feb:110:154637. doi: 10.1016/j.phymed.2022.154637. Epub 2022 Dec 28.

Authors

Xueqin Jiang¹, Yueshan Sun², Shuo Yang³, Yuesong Wu³, Long Wang³, Wenjun Zou⁴, Nan Jiang³, Jianping Chen⁵, Yunwei Han⁶, Chunlan Huang⁶, Anguo Wu⁷, Chunxiang Zhang⁸, Jianming Wu⁹

Affiliations

¹ State Key Laboratory of Biotherapy and Cancer Center, West China Medical School, Sichuan University, Chengdu, Sichuan 610041, China.
² The Third People's Hospital of Chengdu, Chengdu, Sichuan 610031, China.
³ Key Laboratory of Medical Electrophysiology of Ministry of Education of China, Medical Key Laboratory for Drug Discovery and Druggability Evaluation of Sichuan Province, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan 646000, China.
⁴ School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan 611137, China.
⁵ School of Chinese Medicine, The University of Hong Kong, Hong Kong, China.
⁶ The Affiliated Hospital of Southwest Medical University, Luzhou, Sichuan 646000, China.
⁷ Key Laboratory of Medical Electrophysiology of Ministry of Education of China, Medical Key Laboratory for Drug Discovery and Druggability Evaluation of Sichuan Province, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan 646000, China. Electronic address: wag1114@foxmail.com.
⁸ Key Laboratory of Medical Electrophysiology of Ministry of Education of China, Medical Key Laboratory for Drug Discovery and Druggability Evaluation of Sichuan Province, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan 646000, China. Electronic address: zhangchx999@163.com.
⁹ Key Laboratory of Medical Electrophysiology of Ministry of Education of China, Medical Key Laboratory for Drug Discovery and Druggability Evaluation of Sichuan Province, School of Pharmacy, Southwest Medical University, Luzhou, Sichuan 646000, China; School of Basic Medical Sciences, Southwest Medical University, Luzhou, China. Electronic address: jianmingwu@swmu.edu.cn.

PMID: 36610353
DOI: 10.1016/j.phymed.2022.154637

Abstract

Background: Non-peptide thrombopoietin receptor (TPOR) agonists are promising therapies for the mitigation and treatment of thrombocytopenia. However, only few agents are available as safe and effective for stimulating platelet production for thrombocytopenic patients in the clinic.

Purpose: This study aimed to develop a novel small molecule TPOR agonist and investigate its underlying regulation of function in megakaryocytes (MKs) differentiation and thrombopoiesis.

Methods: A potential active compound that promotes MKs differentiation and thrombopoiesis was obtained by machine learning (ML). Meanwhile, the effect was verified in zebrafish model, HEL and Meg-01 cells. Next, the key regulatory target was identified by Drug Affinity Responsive Target Stabilization Assay (DARTS), Cellular Thermal Shift Assay (CETSA), and molecular simulation experiments. After that, RNA-sequencing (RNA-seq) was used to further confirm the associated pathways and evaluate the gene expression induced during MK differentiation. In vivo, irradiation (IR) mice, C57BL/6N-TPOR^em1cyagen (Tpor^-^/^-) mice were constructed by CRISPR/Cas9 technology to examine the therapeutic effect of TMEA on thrombocytopenia.

Results: A natural chemical-structure small molecule TMEA was predicted to be a potential active compound based on ML. Obvious phenotypes of MKs differentiation were observed by TMEA induction in zebrafish model and TMEA could increase co-expression of CD41/CD42b, DNA content, and promote polyploidization and maturation of MKs in HEL and Meg-01 cells. Mechanically, TMEA could bind with TPOR protein and further regulate the PI3K/AKT/mTOR/P70S6K and MEK/ERK signal pathways. In vivo, TMEA evidently promoted platelet regeneration in mice with radiation-induced thrombocytopenia but had no effect on Tpor^-^/^- and C57BL/6 (WT) mice.

Conclusion: TMEA could serve as a novel TPOR agonist to promote MKs differentiation and thrombopoiesis via mTOR and ERK signaling and could potentially be created as a promising new drug to treat thrombocytopenia.

Keywords: 3,4,3′-tri-o-methylellagic acid; Machine learning; Megakaryocytes differentiation; Sanguisorba officinalis; Thrombocytopoiesis; Thrombopoietin receptor.

MeSH terms

Animals
Cell Differentiation
MAP Kinase Signaling System
Megakaryocytes
Mice
Mice, Inbred C57BL
Phosphatidylinositol 3-Kinases / metabolism
Receptors, Thrombopoietin / antagonists & inhibitors
Signal Transduction
TOR Serine-Threonine Kinases / metabolism
Thrombocytopenia* / drug therapy
Thrombocytopenia* / metabolism
Thrombopoiesis*
Zebrafish / metabolism

Substances

Phosphatidylinositol 3-Kinases
TOR Serine-Threonine Kinases
Receptors, Thrombopoietin
tris(2-maleimidoethyl)amine