PLCG2 Exon-Skipped Variants: Insights into Their Potential Role in Chronic Lymphocytic Leukemia

Bruton tyrosine kinase inhibitors (BTKi) are successful in treatment of chronic lymphocytic leukemia (CLL), yet acquired resistance remains a challenge. In our study, REC-1 cells that acquired resistance to BTKi tirabrutinib by long-term treatment gained a splice-site mutation (SSM) c.2236-1G>T in PLCG2. Notably, a deletion spanning c.2236-23_2238 at the same splice site (SSD) was identified in CLL from a patient who had disease progression during ibrutinib therapy. These alterations resulted in the exon-skipped variant, ∆21, which exhibited enhanced Phospholipase activity. PLCG2 SSM REC-1 exhibited increased anti-IgM-mediated calcium flux and resistance to Btk inhibition but retained sensitivity to PLCγ inhibitors. By analyzing additional primary CLL samples, we identified widespread expression of two additional exon-skipped variants, ∆22 and ∆20-22, independent of PLCG2 genetic alterations. The active ∆20-22, which is also described in PLCG2-associated immune dysregulation (PLAID), was predominantly present in CLL compared to healthy donor samples. Expression analysis in CLL cells isolated from patients before and after targeted treatments revealed a decrease in expression of ∆20-22 post-venetoclax (p = 0.02)while no change in expression was observed after ibrutinib treatment, indicating that these variants may persist in the presence of ibrutinib but might be lost in the absence of drug selection pressure. Our study reveals novel PLCG2 splice-site alterations and exon-skipped variants beyond the classical Btk or PLCG2 mutations, to have a potential role in BTKi resistance and inform treatment selection.