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

Area of Science:

• Genomic editing
• Cancer biology
• Molecular genetics

Background:

• Chromosomal translocations are key drivers in human cancers, including gliomas.
• The 1p/19q co-deletion is a defining molecular hallmark of oligodendrogliomas.
• Current methods to engineer 1p/19q co-deletion are lacking, hindering research into its role in gliomagenesis.

Purpose of the Study:

• To develop a method for inducing the t(1;19)(q10;p10) translocation, which results in 1p/19q co-deletion.
• To establish droplet digital PCR (ddPCR) assays for detecting the resulting hybrid chromosomes.
• To investigate the feasibility of using CRISPR/Cas9 for engineering cancer-relevant chromosomal translocations.

Main Methods:

• Utilized CRISPR/Cas9 to induce simultaneous DNA double-strand breaks (DSBs) on chromosomes 1p and 19q.
• Developed and applied ddPCR assays to detect the 1p/19q and 1q/19p hybrid chromosomes.
• Confirmed translocation events through PCR amplification and sequencing in HEK 293T, U-251, and LN-229 cells.

Main Results:

• Successfully induced the t(1;19)(q10;p10) translocation in multiple cell lines.
• Detected both 1p/19q and 1q/19p hybrid chromosomes via PCR and sequencing, confirming the integration of DNA from both chromosomes.
• Observed rapid loss of the 1p/19q hybrid chromosome and eventual undetectability of the 1q/19p hybrid chromosome, suggesting a potential cell survival disadvantage.

Conclusions:

• Demonstrated the efficacy of CRISPR/Cas9-mediated genomic editing for inducing t(1;19)(q10;p10).
• This method provides a novel tool for engineering 1p/19q co-deletion to model oligodendrogliomas.
• The approach is potentially generalizable for engineering other cancer-associated translocations, advancing the understanding of their roles in cancer progression.