Erythropoietin Intensifies the Proapoptotic Activity of LFM-A13 in Cells and in a Mouse Model of Colorectal Cancer

Abstract

The Bruton’s tyrosine kinase (BTK) inhibitor LFM-A13 has been widely employed as an antileukemic agent, but applications in solid cancer have been found recently. The compound promotes apoptosis, has an antiproliferative effect, and increases cancer cell sensitivity to chemotherapy drugs. We decided to assess the impact of the simultaneous use of erythropoietin (Epo) and LFM-A13 on signal transduction in colon DLD-1 and HT-29 cells, as well as in tumor xenografts. The induction of apoptosis by Epo and LFM-A-13 in the cells was confirmed by phosphatidylserine externalization, loss of mitochondrial membrane potential, and modulation of the expression of apoptotic protein BAX and antiapoptotic protein BCL-2 in colon adenocarcinoma cells. Nude mice were inoculated with adenocarcinoma cells and treated with Epo and LFM-A13 in order to evaluate the degree of tumor regression. The simultaneous use of Epo and LFM-A13 severely inhibited cell growth, activated apoptosis, and also inhibited tumor growth in xenografts. The addition of Epo to LFM-A13 intensified the antiproliferative effect of LFM-A13, confirmed by the loss of mitochondrial membrane potential and the accumulation of apoptotic colon cancer cells with externalized phosphatidylserine (PS). These preclinical results suggest that the combination of Epo and LFM-A13 has a high proapoptotic activity and should be tested in the clinic for the treatment of solid tumors such as colon cancer.

Exploration of novel heterofused 1,2,4-triazine derivative in colorectal cancer

“…Induction of apoptosis was examined using Apoptosis Detection Kit II analysed using a flow cytometry, as previously described 28 . During programmed cell death (PCD), the phosphatidylserine (PS) is transferred from the inner cell membrane to the cell surface.…”

Clinicopathological significance of Epo, EpoR, Ki-67 and Bax expression in colorectal cancer

“…Further, we must take into consideration a fact that some investigations assessing expression of EpoR protein in tumors cells were made by using immunohistochemical methods and others using RT-PCR (at mRNA level), what could lead to different results. There is a hypothesis that in some cases detection of EpoR expression in tumor cell lines is limited by lack of specificity of available anti-EpoR antibody, which were used in previous studies [29].…”

The intensification of anticancer activity of LFM-A13 by erythropoietin as a possible option for inhibition of breast cancer

“…Flow cytometry analysis of MMP and assessment of annexin V binding have been described previously 22 . The distribution of the cell cycle phases was analysed using flow cytometry.…”