A Histone Deacetylase Inhibitor Induces Apoptosis, Differentiation and Cell Cycle Changes in STI571 Resistant CML Cell Line with No Change in BCR-ABL Expression
Authors: Shaklai Matityahu , Zimra Yael , Belyaeva Inessa , Aviram Adina , Rabizadeh Esther 
Published: April 2020

Abstract
The hallmark of Chronic-myelogenous leukemia (CML) is high tyrosine kinase (TK) activity of the chimeric protein BCR-ABL, known to contribute to cell tumorogenity, resistance to apoptosis and differentiation. The gold-standard therapy for CML is treatment with the TK inhibitor Imatinib (STI571), but in some cases STI571 resistance appears due to BCR-ABL over expression, amplification or mutation in the ABL kinase domain. However, STI571 resistance due to other mechanisms which are BCR-ABL independent may not be excluded. BCR-ABL-positive cells can evade the inhibitory effect of STI571 by different mechanisms, such as reduced intake mediated by OCT-1 organic cation transporter-1), or enhanced efflux by Pgp, and, possibly, acquisition of compensatory mutations in genes other than BCR-ABL. One possible approach to overcome STI571 resistance involves the combination of Histone Deacetylase Inhibitors (HDACI). Previously we have shown: induced apoptosis, differentiation and reduced BCR-ABL protein levels in CML cell line (K562) by Pivaloyloxymethyl butyrate (Pivanex), a HDACI. In order to study STI571 resistance and the influence of HDACI on STI571 resistant CML cells we developed an STI571 K562 resistant cell line (K562-R). Our results show that s these K562-R cells had an IC50 33 times greater than that of K562 wild type (K562-S) cells. But K562-R cells had similar BCR-ABL protein and transcript levels as the K562-S and no mutations were found in the catalytic TK domain of K562-R cells. MDR protein expression and activity in K562-R were very low and similar to those of K562-S. However, the K562-R cell line had higher spontaneous apoptosis and G2M arrest and a slight but significant lower S phase. The K562-R cell line expressed more glycophorin A although spontaneous hemoglobin synthesis in K562-R cells was lower. Study of the signaling transduction pathway have shown no differences in p70S6, STAT3, Erk 1/2, CREB, Ikβ-a and JNK proteins levels (total and phosphorylated), however phosphorylated p38 was elevated in the K562-R cells. P21, a cell cycle regulatory protein, was higher in K562-R cells. HDACI induced viability loss, apoptosis and cell cycle changes in K562-R in a similar way to its effects on K562-S cells. However, it did not influence the levels of BCR-ABL. Taken together our results suggest that the STI571 resistance mechanism in these K562-R cells does not involve BCR-ABL but other regulatory mechanisms in the cell cycle, differentiation and signal transduction pathways. The higher spontaneous G2M arrest, apoptosis as well as the variable differentiation could be related to changes in p21 and p38 signaling pathways. These K562-R cells with no change in BCR-ABL may suggest a model for STI571 resistance that does not involve BCR-ABL directly. This data suggest, the possible benefit of HDACI treatment in CML STI571 resistant patient with no mutation or changes in BCR-ABL protein.