Effect of pivaloyl-substituted pyrrole containing heterocyclic compounds on DNA repair pathways in Ewing sarcoma cells

Aim. To examine deoxyribonucleic acid (DNA) damage repair and cell cycle regulatory mechanisms of Ewing sarcoma cells exposed to pivaloyl-substituted pyrrole containing heterocyclic compounds.

Methods. The study was performed on A673 Ewing sarcoma cell line. The tumor cells were incubated for 48 h in the presence of pivaloyl-substituted pyrrole containing heterocyclic compounds (compounds №20 and №24). Western blot analysis was utilized to examine expression of the markers of DNA single-strand (phosphorylated forms of ATR and Chk1) and double-strand breaks (phosphorylated forms of H2AX, АТМ, DNA-PK, BRCA-1, Chk-2). Analysis of the cell cycle phases was performed by flow cytometry (BD FacsCanto, USA).

Results. Pivaloyl-substituted pyrrole containing heterocyclic compounds substantially increased the expression of histone 2A phosphorylated on serine 138 (γ-H2AX) that indicates DNA damage (double-strand breaks). Under exposure to pivaloyl-substituted pyrrole containing heterocyclic compounds the studied cells increased expression of phosphorylated forms of ATM-kinase and BRCA-1. Also cell cycle disorders leading to substantial G2/M arrest and enhanced apoptosis of tumor cells were observed.

Conclusion. Pivaloyl-substituted pyrrole containing heterocyclic compounds induced DNA double-strand breaks in A673 Ewing sarcoma cell line; in response to DNA damage in tumor cells, the mechanisms of DNA double-strand breaks repair were activated; despite activation of DNA repair mechanisms, A673 cells underwent cell cycle arrest in the G2/M-phase and apoptosis.