DNA abnormalities can disrupt normal cellular processes and give rise to cancer. Now, scientists at the La Jolla Institute for Immunology (LJI) hope their findings on the role of two unusual DNA structures in the formation of B-cell lymphoma could inform the design of new cancer drugs.
Loss of function in the TET enzymes could give rise to B-cell lymphoma, and an increase in two abnormal DNA structures called G-quadruplex and R-loop appeared to be the link between the two phenomena, LJI researchers described in a new study published in Nature Immunology.
TET-deficient B cells were vulnerable to targeting of G-quadruplex and R-loops, the scientists found. Inhibiting DNMT1, an opposing enzyme to TET in the regulation of DNA activity, led to a marked delay in lymphoma development in mice, the team showed.
TET enzymes regulate a biological process called DNA methylation, which is essential for cell development, including B cells. Previous studies have observed mutations that cause TET enzymes to lose their function in many blood cancers and solid tumors.
In the current study, the LJI scientists deleted TET2 and TET3 enzymes in B cells of mice. The rodents developed lymphoma, and the researchers observed an increase in markers that are associated with DNA damage and genomic instability.
Compared with control B cells, those TET-deficient B cells displayed increased G-quadruplexes and R-loops, the team found. R-loops happen when a piece of RNA drives a wedge between the two DNA strands that form a DNA molecule. G-quadruplexes are like knots on the DNA strands. Both structures can act as physical impediments to DNA transcription and replication.
At a genomewide level, the increases in G-quadruplexes and R-loops in the TET-deficient B cells were associated with increased DNA double-strand breaks at immunoglobulin switch regions, the researchers noted.
“These structures represent sites in the DNA that are much more fragile than other regions,” the study’s first author, Vipul Shukla, Ph.D., said in a statement. “With this study, we found that TET enzymes are perhaps related to the regulation of these structures, which could in turn explain one mechanism for acquisition of genomic instability in the absence of TET enzymes.”
The researchers used CRISPR-Cas9 to deplete enzymes that regulate G-quadruplexes and R-loops. Doing so led to an increase in programmed death in the TET-deficient B cells but not in control B cells.
Shukla and colleagues also observed an upregulation of the DNMT1 enzyme in the TET-deficient B cells. DNMT and TET are opposing enzymes in the DNA methylation process, with the former adding the methyl groups to the DNA molecule and the latter removing them. Without TET, the normal balance was disrupted.
Given the two enzymes’ relationship, the researchers wanted to see the effect of removing DNMT1 in TET-deficient B cells. Turns out, mice without TET2 and TET3 quickly died from B-cell lymphoma, with a median survival of 20 weeks. In contrast, mice with additional DNMT1 deletion lived much longer at a median 98 weeks.
What’s more, TET-deficient B cells from mice that had additional DNMT1 alteration displayed a notable decrease in the levels of G-quadruplex and R-loop almost to the levels of normal B cells, the team found.
The findings suggest that G-quadruplexes and R-loops could be therapeutically targeted in cancers with TET loss of function, the researchers said in the study. Further, several DNMT inhibitors such as Bristol Myers Squibb’s Vidaza (azacitidine) are already available for blood cancer.
“Follow-up studies in preclinical models could test whether a combination of G-quadruplex-stabilizing agents and [DNMT] inhibitors might synergize to delay the onset and/or progression of B-cell lymphomas and other malignancies with TET loss of function,” the team wrote in the study.