Effects of ibrutinib and venetoclax on the expression of immune checkpoint molecules in leukemic blasts of patients with acute lymphoblastic leukemia

Abstract

A major challenge in cancer treatment is the ability of tumor cells to evade immune responses, reducing therapy effectiveness and leading to disease recurrence. Small molecule inhibitors (SMIs) have shown promise in treating malignancies due to their targeted action, affordability, and accessibility. Acute lymphoblastic leukemia (ALL), the most common pediatric malignancy, involves the uncontrolled growth of immature B and T lymphocytes. Key molecules such as BTK and Bcl-2 contribute to disease progression by promoting cell survival and preventing apoptosis. However, mechanisms of immune evasion in ALL remain unclear. This study evaluates the effects of ibrutinib and venetoclax on immune checkpoint molecules (Gal-9, PD-L1, CD200, CD155, CD47) and TGF-β in ALL. Methods: Leukemic cells were isolated from 20 ALL patients using magnetic-activated cell sorting (MACS), achieving >98% purity (flow cytometry). Cells were treated with serial concentrations of ibrutinib and venetoclax, and cytotoxicity was measured by MTT. Apoptosis and cell viability were assessed through flow cytometry and MTT, while mRNA levels of immune checkpoint molecules and TGF-β were analyzed using Real-Time PCR. Results: Both SMIs significantly reduced cell proliferation and increased apoptosis, with venetoclax showing superior effects. Treatments also altered the mRNA expression of immune checkpoint ligands and TGF-β.