共济失调毛细血管扩张症和Rad3相关激酶抑制剂与吉西他滨协同诱导子宫平滑肌肉瘤细胞死亡

Introduction: Uterine leiomyosarcoma (uLMS) is a rare, lethal gynecological cancer. Therapeutic response to standard-of-care chemotherapy for uLMS is hindered because cancer cells can activate DNA damage response (DDR) pathways. For example, gemcitabine stalls replication forks and halts DNA synthesis. This activates Ataxia telangiectasia and Rad3 related kinase (ATR), which aids DDR by repairing stalled replication forks and delaying cell cycle progression. Therefore, we investigated the potential for ATR inhibitors to enhance the potency and efficacy of chemotherapy for uLMS in vitro . Methods: CellTitre Glo, Bliss Independence analysis, and Multidimensional Synergy of Combinations (MuSyC) assessed synergy between three ATRi (elimusertib, ceralasertib, berzosertib) and SOC chemotherapy (gemcitabine, doxorubicin, docetaxel) in three commercially available (SK-LMS-1, SK-UT-1B, SK-UT-1) and two patient-derived uLMS cell lines (ACI-44A, ACI-44B). Following treatment of gemcitabine alone, elimusertib alone, or gemcitabine + elimusertib, flow cytometry was performed to analyze cell cycle phase distribution using propidium iodine-stained cells, as well as to assess apoptosis and necrosis using annexin V and sytox blue-stained cells. Western blotting investigated protein expression of ATR pathway proteins, DNA damage markers, and apoptosis markers. Results: ATR inhibitors synergized most strongly with gemcitabine, with elimusertib + gemcitabine emerging as the most synergistic combination across all five cell lines. In contrast, combinations of ATRi with doxorubicin or docetaxel showed minimal to no synergy. ATRi enhanced gemcitabine potency in all cell lines by significantly reducing its EC50. Elimusertib also increased gemcitabine's cytoxicity. In SK-UT-1B cells, the combination increased apoptotic cells by 51.4% (14.7% vs 9.7%, p < 0.05) and necrotic cells by 120.2% (24.7% vs 11.2%, p < 0.001) compared to gemcitabine alone. Elimusertib did not prevent gemcitabine-induced S phase arrest but increased the sub-G1 population, consistent with enhanced cell death. Western blot analysis confirmed that elimusertib inhibited downstream ATR signaling, including CHK1 S345 phosphorylation blockage, and that the combination produced greater DNA damage and apoptosis - evidenced by increased p-H2AX (S139) and cleaved caspase-3 expression - than either agent alone. Conclusions: Elimusertib strongly synergizes with gemcitabine in vitro across uLMS cell lines by potentiating DNA damage and apoptosis. The lack of synergy with docetaxel or doxorubicin underscores gemcitabine as the most effective combination with ATR inhibitors. These finds support further preclinical and clinical evaluation of elimusertib + gemcitabine as a promising novel therapy for uterine leiomyosarcoma.