Anti-CD20 Immunotherapy Augments the Chemotherapy-Induced Senescence Response In Human Lymphoma Cells

Targeting the B-cell antigen CD20 in addition to conventional DNA damaging chemotherapy has dramatically improved patient prognosis in a variety of B-cell malignancies without overtly increasing treatment toxicity. The therapeutic benefit of rituximab as the prototypic anti-CD20 compound has often been attributed to classical antibody effector functions of the immune system, and, to some extent, to induction of apoptosis as well. Interestingly, there is no consensus regarding the intracellular pathways triggered by CD20 signaling. Cellular senescence, a terminal growth arrest condition, complements apoptosis as another cell-cycle exit program that is acutely inducible in malignant cells in response to DNA damaging chemotherapy, and is associated with favorable treatment outcome in preclinical cancer models. Senescent cells are characterized by retained viability, distinct morphology, and a cytokine secretion pattern that mediates an intense cross-talk to the microenvironment and cells of the immune system.

Here, we provide evidence for induction of cellular senescence by rituximab in cell lines derived from aggressive and indolent human B-cell lymphomas. Mimicking the in vivo presence of Fc-receptor-bearing cells by addition of a F(ab’)₂-fragment directed against the constant part of human IgG, we observed diminished growth, altered cellular morphology and senescence-associated ß-galactosidase reactivity after anti-CD20 immunotherapy in vitro. Strikingly, when applied together with a moderate concentration of the anthracycline adriamycin, anti-CD20 treatment strongly augmented the fraction of lymphoma cells entering a senescent cell-cycle arrest. Expression and secretion of cytokines typically associated with cellular senescence (such as IL-6 and IL-8) were increased afterin vitro immunochemotherapy (analysed by RQ-PCR and fluorescent bead immunoassays of media supernatants). Unexpectedly, we detected higher levels of DNA damage signaling markers (phosphorylated H2AX [serine 139] and phosphorylated p53 [serine 15]) with anti-CD20 co-treatment compared to single agent adriamycin application.

Our data demonstrate the induction of cellular senescence, which has a profound impact on tumor biology and tumor/host interactions in preclinical tumor models, as a consequence of standard lymphoma immunotherapy in humans. The anti-CD20-mediated augmentation of the anthracycline-induced DNA damage response provides a mechanistic explanation for the stronger induction of the senescent phenotype after immunochemotherapy and may contribute to the clinical superiority of treatment regimens that include rituximab. Further elucidation of the synergy between antineoplastic agents and CD20-directed compounds with respect to induction of cellular senescence as a favorable treatment outcome opens the clinical perspective of specifically engaging the senescence program by therapeutic approaches.

No relevant conflicts of interest to declare.