Generation of chromosome 1p/19q co-deletion by CRISPR/Cas9-guided genomic editing

Chao Li; Zhong Liu; Xiaoxia Zhang; Huafeng Wang; Gregory K Friedman; Qiang Ding; Xinyang Zhao; Hu Li; Kitai Kim; Xi Yu; L Burt Nabors; Xiaosi Han; Rui Zhao

doi:10.1093/noajnl/vdac131

Generation of chromosome 1p/19q co-deletion by CRISPR/Cas9-guided genomic editing

Neurooncol Adv. 2022 Aug 18;4(1):vdac131. doi: 10.1093/noajnl/vdac131. eCollection 2022 Jan-Dec.

Authors

Chao Li¹, Zhong Liu¹, Xiaoxia Zhang^{1

2}, Huafeng Wang³, Gregory K Friedman⁴, Qiang Ding⁵, Xinyang Zhao¹, Hu Li⁶, Kitai Kim⁷, Xi Yu⁸, L Burt Nabors³, Xiaosi Han³, Rui Zhao^{1

9}

Affiliations

¹ Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, AL 35294, USA.
² Department of Genetics, University of Alabama at Birmingham, AL 35294, USA.
³ Department of Neurology, University of Alabama at Birmingham, AL 35294, USA.
⁴ Department of Pediatrics, Division of Hematology/Oncology, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
⁵ Department of Anesthesiology and Perioperative Medicine & Molecular and Translational Biomedicine, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
⁶ Department of Molecular Pharmacology & Experimental Therapeutics, Center for Individualized Medicine, Mayo Clinic College of Medicine, Rochester, MN 55904, USA.
⁷ Human Stem Cell & Genome Engineering Center and Department of Biological Chemistry, University of California, Los Angeles, CA 90095, USA.
⁸ Clinical Oncology Center, The People's Hospital of Guangxi Zhuang Autonomous Region, Nanning, Guangxi 530021, China.
⁹ Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama at Birmingham, Birmingham, AL 35294, USA.

Abstract

Background: Chromosomal translocation has been detected in many human cancers including gliomas and is considered a driving force in tumorigenesis. Co-deletion of chromosome arms 1p and 19q is a hallmark for oligodendrogliomas. On the molecular level, 1p/19q co-deletion results from t(1;19)(q10;p10), which leads to the concomitant formation of a hybrid chromosome containing the 1q and 19p arms. A method to generate 1p/19q co-deletion is lacking, which hinders the investigation of how 1p/19q co-deletion contributes to gliomagenesis.

Methods: We hypothesized that chromosomal translocation, such as t(1;19)(q10;p10) resulting in the 1p/19q co-deletion, may be induced by simultaneously introducing DNA double-strand breaks (DSBs) into chromosomes 1p and 19q using CRISPR/Cas9. We developed a CRISPR/Cas9-based strategy to induce t(1;19)(q10;p10) and droplet digital PCR (ddPCR) assays to detect the hybrid 1q/19p and 1p/19q chromosomes.

Results: After translocation induction, we detected both 1p/19q and 1q/19p hybrid chromosomes by PCR amplification of the junction regions in HEK 293T, and U-251 and LN-229 glioblastoma cells. Sequencing analyses of the PCR products confirmed DNA sequences matching both chromosomes 1 and 19. Furthermore, the 1p/19q hybrid chromosome was rapidly lost in all tested cell lines. The 1q/19p hybrid chromosome also become undetectable over time likely due to cell survival disadvantage.

Conclusion: We demonstrated that t(1;19)(q10;p10) may be induced by CRISPR/Cas9-mediated genomic editing. This method represents an important step toward engineering the 1p/19q co-deletion to model oligodendrogliomas. This method may also be generalizable to engineering other cancer-relevant translocations, which may facilitate the understanding of translocation roles in cancer progression.

Keywords: 1p/19q co-deletion; CRISPR/Cas9; IDH mutant low-grade gliomas; chromosome translocation; oligodendroglioma.

Abstract

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