Functional Implication of p53-Mediated DNA Damage Response on the B-Cell Lymphoma Microenvironment Interaction in Chemoimmunotherapy

Resistance against chemotherapy remains a challenge in the current treatment of B cell malignancies. In previous work we could show that macrophages are an essential element in the synergistic response to the administration of the DNA damaging agent Cyclophosphamide and monoclonal antibodies. The combination treatment strongly increases tumor clearance overall survival in the treatment refractory humanized BCL2-MYC-dependent double hit lymphoma hMB mouse model.

Here we observed that the alkylating agent Cyclophosphamide is inducing the secretion of distinct cytokines like VEGF-A, CCL4, IL8, IL10 and TNFα by the leukemic cells. This in turn leads to a repolarization of tumor associated macrophages from a suppressive to an activated phenotypic state, which is designated as acute secretory activating phenotype (ASAP). We analyzed the functional implications of the DNA damage response pathway for the generation of the ASAP and synergy in chemoimmunotherapy. When disrupting DNA damage response pathway by knock-down of key elements like ATM, DNA-PK or p53 in the leukemic cells we could prevent the formation of the stimulatory ASAP effect on macrophage phagocytic capacity. Particularly p53 status seems to possess a central regulating role on macrophage mediated malignant cell depletion. TP53 activation via Nutlin-3A treatment of lymphoma cell enhances ADCP in in p53 wildtype cells, while not displaying enhancement in p53-deficient lymphoma cells. Utilizing the hMB model for modeling in vivo treatment of lymphoma bearing mice we could demonstrate diminished ASAP and ADCP for p53-deficient lymphoma treated with cyclophosphamide in vivo . Using primary human CLL patient cells comparing both wildtype and p53-deficient status, the p53-deficient CLL cells failed to induce the stimulatory, cytokine-mediated effect on macrophage phagocytosis in response to combination treatment as seen with the p53 proficient CLL cells. This observation is underlined by in vivo modeling of CLL therapy by treating Eµ-TCL1/p53^wt/wt as well as Eµ-TCL1p53^-/- mice. Here, low-dose cyclophosphamide treated Eµ-TCL1p53^-/- mice failed to induce an antibody mediated stimulatory effect on macrophage phagocytosis capacity as seen with Eµ-TCL1/p53^wt/wt mice.

In this work, we are able to recapitulate the effect of ASAP mediated increase of macrophage mediated antibody dependent cellular phagocytosis in the Eµ-Tcl1 mouse model and therefore demonstrate that the ASAP is not only restricted to double hit lymphoma, but also shows functional relevance in low-grade lymphoma such as CLL. Moreover, we indicate that p53 functional status influences the generation of the ASAP in independent models in vitro and in vivo as in primary CLL patient cells. Mediated by this process p53 regulates the interraction with tumor asociated macrophages and the tumor microenvironment.Finally our ongoing research offers possibility to reveal and tailor new treatment approaches for chemorefractory patients.