ABL Genomic Editing Sufficiently Abolishes Oncogenesis of Human Chronic Myeloid Leukemia Cells In Vitro and In Vivo.

Shu-Huey Chen,Chia-Hwa Lee,Ming-Jang Su,Kai-Wen Hsu,Su-Mei Lin,Yun-Shan Chiang,Yao-Yu Hsieh,Huey-En Tzeng,Wen-Shyang Hsieh,Chun-Yu Lin

doi:10.3390/cancers12061399

Abstract

Chronic myelogenous leukemia (CML) is the most common type of leukemia in adults, and more than 90% of CML patients harbor the abnormal Philadelphia chromosome (Ph) that encodes the BCR-ABL oncoprotein. Although the ABL kinase inhibitor (imatinib) has proven to be very effective in achieving high remission rates and improving prognosis, up to 33% of CML patients still cannot achieve an optimal response. Here, we used CRISPR/Cas9 to specifically target the BCR-ABL junction region in K562 cells, resulting in the inhibition of cancer cell growth and oncogenesis. Due to the variety of BCR-ABL junctions in CML patients, we utilized gene editing of the human ABL gene for clinical applications. Using the ABL gene-edited virus in K562 cells, we detected 41.2% indels in ABL sgRNA_2-infected cells. The ABL-edited cells reveled significant suppression of BCR-ABL protein expression and downstream signals, inhibiting cell growth and increasing cell apoptosis. Next, we introduced the ABL gene-edited virus into a systemic K562 leukemia xenograft mouse model, and bioluminescence imaging of the mice showed a significant reduction in the leukemia cell population in ABL-targeted mice, compared to the scramble sgRNA virus-injected mice. In CML cells from clinical samples, infection with the ABL gene-edited virus resulted in more than 30.9% indels and significant cancer cell death. Notably, no off-target effects or bone marrow cell suppression was found using the ABL gene-edited virus, ensuring both user safety and treatment efficacy. This study demonstrated the critical role of the ABL gene in maintaining CML cell survival and tumorigenicity in vitro and in vivo. ABL gene editing-based therapy might provide a potential strategy for imatinib-insensitive or resistant CML patients.

Highlights

Leukemia is classified as acute or chronic and as myelogenous or lymphocytic; the subtypes include acute myelogenous leukemia (AML), chronic myelogenous leukemia (CML), acute lymphocytic leukemia (ALL) and chronic lymphocytic leukemia (CLL), and chronic leukemia grows slowly, and progressively worsens over time
The peripheral blood of Chronic myelogenous leukemia (CML) participants and healthy controls was obtained at Shuang Ho Hospital, Taipei Medical University, New Taipei City, Taiwan, according to a protocol approved by the Institutional Review Board (N201711069)
The results showe that the viability of K562 cells was significantly inhibited by infection with the BCR-ABL sgRNA_2 virus (p < 0.001) compared to infection with the BCR-ABL sgRNA_1 virus (p = 0.011) and scramble virus (Figure 2J). These findings demonstrated that targeting the BCR-ABL gene caused a dramatic disruption in DNA sequence and protein suppression and eventually suppressed cell viability in CML cells, implying that BCR-ABL

Summary

Introduction

Leukemia is classified as acute or chronic and as myelogenous or lymphocytic; the subtypes include acute myelogenous leukemia (AML), chronic myelogenous leukemia (CML), acute lymphocytic leukemia (ALL) and chronic lymphocytic leukemia (CLL), and chronic leukemia grows slowly, and progressively worsens over time. A chromosome translocation, between the long arms of chromosomes 9 and 22, t(9;22)(q34;q11), is found in over 90% of CML patients, in a lower proportion of ALL or biphenotypic acute leukemia cases and in rare cases of de novo AML [2,3]. This well-known Philadelphia (Ph) chromosome produces the BCR-ABL oncogenic fusion protein that activates multiple signaling pathways involved in the cell cycle, adhesion and apoptosis [4,5]. Imatinib has been shown in recent years to be highly effective at increasing the life expectancy of CML patients [7]

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