(–)-Epigallocatechin-3-gallate induces apoptosis and diferentiation in leukaemia by targeting reactive oxygen species and PIN1

Agradecimentos: Te authors would like to thank Raquel S. Foglio for English review, and Márcio Alves da Cruz for his technical assistance with the animal’s care. Molecular structure of EGCG was produced using ChemDraw 19.0 sofware. Tis work was funded by the Conselho Nacional de Desenvolvimento Científco e Tecnológico (CNPq/MCT) [Grant Number 303405/2018-0]; the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) [Grant Numbers 11/51959-0 and 17/21801-2]; and the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—Brasil (CAPES) [Finance Code 001] Abstract: (–)-Epigallocatechin-3-gallate (EGCG), the major active polyphenol extracted from green tea, has been shown to induce apoptosis and inhibit cell proliferation, cell invasion, angiogenesis and metastasis. Herein, we evaluated the in vivo efects of EGCG in acute myeloid leukaemia (AML) using an acute promyelocytic leukaemia (APL) experimental model (PML/RARa). Haematological analysis revealed that EGCG treatment reversed leucocytosis, anaemia and thrombocytopenia, and prolonged survival of PML/RARa mice. Notably, EGCG reduced leukaemia immature cells and promyelocytes in the bone marrow while increasing mature myeloid cells, possibly due to apoptosis increase and cell diferentiation. The reduction of promyelocytes and neutrophils/monocytes increase detected in the peripheral blood, in addition to the increased percentage of bone marrow cells with aggregated promyelocytic leukaemia (PML) bodies staining and decreased expression of PML-RAR oncoprotein corroborates our results. In addition, EGCG increased expression of neutrophil diferentiation markers such as CD11b, CD14, CD15 and CD66 in NB4 cells; and the combination of all-trans retinoic acid (ATRA) plus EGCG yield higher increase the expression of CD15 marker. These fndings could be explained by a decrease of peptidyl-prolyl isomerase NIMA-interacting 1 (PIN1) expression and reactive oxygen species (ROS) increase. EGCG also decreased expression of substrate oncoproteins for PIN1 (including cyclin D1, NF-?B p65, c-MYC, and AKT) and 67 kDa laminin receptor (67LR) in the bone marrow cells. Moreover, EGCG showed inhibition of ROS production in NB4 cells in the presence of N-acetyl-L-cysteine (NAC), as well as a partial blockage of neutrophil diferentiation and apoptosis, indicating that EGCG-activities involve/or are in response of oxidative stress. Furthermore, apoptosis of spleen cells was supported by increasing expression of BAD and BAX, parallel to BCL-2 and c-MYC decrease. The reduction of spleen weights of PML/RARa mice, as well as apoptosis induced by EGCG in NB4 cells in a dose-dependent manner confrms this assumption. Our results support further evaluation of EGCG in clinical trials for AML, since EGCG could represent a promising option for AML patient ineligible for current mainstay treatments CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO (CNPQ) FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO (FAPESP) COORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPES Aberto