sincalide and Leukemia--Myelogenous--Chronic--BCR-ABL-Positive

The treatment of chronic myeloid leukemia (CML) is facing the dilemma of tyrosine kinase inhibitors (TKIs) resistance and disease recurrence. The dysfunctional DNA damage repair mechanism plays an essential role not only in the initiation and progression of hematological malignancies but also links to the development of TKI resistance. Deciphering the abnormally regulated DNA damage repair and proteins involved brings new insights into the therapy of leukemias. As a G2/M phase checkpoint kinase and a DNA damage repair checkpoint kinase engaged in the DNA damage response (DDR), along with an oncogenic driver present in various cancers, the particular involvement of Wee1 in DNA damage is far from clear. Deciphering its function and targeting it via modulating DNA repair pathways is important for improving our understanding of cancer treatment.. Wee1 expression was assessed in cell lines using RT-qPCR and western blot, and Wee1 knockdown efficacy was validated using RT-qPCR, western blot, and immunofluorescence. Wee1 function was investigated by CCK-8, colony formation, and flow cytometry assay in vitro. Wee1 role in DNA repair and its interactions with other proteins were then studied using western blot, immunofluorescence, and double plasmid-repair studies. Finally, the CCK-8 and flow cytometry assay was utilized to investigate Wee1 and imatinib's synergistic effect, and a CML mouse model was constructed to study Wee1's role in carcinogenesis in vivo.. Wee1 was reported to respond quickly to DDR in an ATM-γH2AX-MDC1-dependent way upon DNA double-strand breaks (DSBs) occurrence, and it regulated homologous recombination by stimulating the recruitment of critical proteins RAD51/BRCA1 upon DSB sites. Wee1 was also revealed to be abnormally upregulated in CML cells. Further suppression of Wee1 not only causes cell cycle arrest and inhibits the proliferation of cancer cells but also enhances CML cell sensitivity to Imatinib in vitro and in vivo, possibly through an excessive accumulation of overall DSBs.. Wee1 is extensively involved in the DRR signaling and DSB repair pathway. Inhibiting abnormally elevated Wee1 benefits CML therapy in both IM-resistant and IM-sensitive cells. Our data demonstrated that Wee1 participated in promoting cell proliferation and imatinib resistance in chronic myeloid leukemia via regulating DNA damage repair dependent on ATM-γH2AX-MDC1. In the fight against CML, Wee1's dysregulation in the DNA damage repair mechanism of CML pathogenesis makes it a viable therapeutic target in clinical applications.

Topics: Animals; Cell Proliferation; DNA Damage; DNA Repair; Drug Resistance, Neoplasm; Humans; Imatinib Mesylate; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Mice; Sincalide

We sought to determine the expression levels of hypoxia-inducible factor-1α (HIF-1α) in the bone marrow chronic myelogenous leukemia (CML) patients. We also tried to determine the roles HIF-1α in the proliferation of CML cells by small interfering RNA (siRNA) knockdown. Real-time PCR was performed to determine the expression levels of HIF-1α in the bone marrows of CML patients and healthy volunteers. HIF-1α knockdown by siRNA in K562 cells was confirmed by RT-PCR. Proliferation and colony formation of the treated cells were determined by CCK8 after HIF-1α knockdown. RT-PCR and western blotting were performed to detect mRNA and protein levels of p21 and p53 in K562 cells. HIF-1α mRNA expression in the bone marrow of CML patients was significantly higher than that in the control, which was statistically significant (P < 0.05). HIF-1α knockdown dramatically reduced the proliferation of K562 cells, which was also statistically significant (P < 0.05). HIF-1α knockdown markedly reduced the colony formation ability of K562 cells, which was also statistically significant (P < 0.05). The mRNA and protein expression of p21 were significantly reduced in K562 cell after HIF-1α knockdown with affecting the mRNA and protein levels of p53. HIF-α promotes chronic CML cell proliferation by up-regulating p21 expression.

Topics: Apoptosis; Bone Marrow; Cell Line, Tumor; Cell Proliferation; Cholecystokinin; Cyclin-Dependent Kinase Inhibitor p21; Gene Expression Profiling; Gene Expression Regulation, Leukemic; Humans; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; K562 Cells; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Peptide Fragments; RNA, Small Interfering; Tumor Suppressor Protein p53; Up-Regulation