BIRC5Gene Disruption via CRISPR/Cas9n Platform Suppress Acute Myelocytic LeukemiaProgression

Background:Acute myelocytic leukemia (AML) is a clonal malignancy resulting from the accumulation of genetic abnormalities in the cells. Human baculoviral inhibitor of apoptosis repeat-containing 5 (BIRC5), encodes survivin, is one of only a handful of genes that isdifferentially over-expressed in numerous malignant diseases including AML.Methods:The BIRC5 was silenced permanently in twoAML cell lines, HL‑60 and KG-1, via the CRISPR/Cas9n system. After transfection of CRISPR constructs, genomic DNA was extracted and amplified to assess mutation detection. To evaluate BIRC5gene expression, quantitative real-timePCR was performed. Also, MTT cell viability and Annexin‑V/propidium iodide flowcytometric staining were performed,and the data were analyzed using the Kolmogorov-Smirnov,Levene's, and ANOVA tests.Results:The results indicated that Cas9n and its sgRNAs successfully triggered site-specific cleavage and mutation in the BIRC5gene locus. Moreover, suppression of BIRC5resulted in the reduction of cell viability,and induction of apoptosis and necrosis in HL60 and KG1suggested that the permanent suppressionof BIRC5remarkablydroppedthe gene expression and cells viability. Conclusion:This study reinforcesthe idea that BIRC5disruption via Cas9n:sgRNAshas favorable effects on the AML clinical outcome. It thereby can be a promising candidate in a variety of leukemia treatments.