Non-Genotoxic Targeting of p53-Dependent and p53-Independent Signaling Pathways Efficiently Kills Hodgkin/Reed-Sternberg Cells.

Although the majority of patients with classical Hodgkin lymphoma (cHL) can be cured by conventional chemotherapy, a substantial proportion of patients will finally develop treatment-induced secondary malignancies. Therefore, non-genotoxic targeting of major cellular survival pathways could be an interesting novel treatment strategy for patients with cHL. In this study, we focused on the analysis of p53-dependent and p53-independent signaling pathways in Hodgkin/Reed-Sternberg (HRS) cells. To test whether p53 signaling is functional in cHL and whether activation of the p53 pathway is sufficient to kill HRS cells, we employed a recently developed small-molecule antagonist of MDM2, designated nutlin-3a, that disrupts the p53-MDM2 interaction. Nutlin-3a efficiently increased the level of p53 and induced expression of p53 downstream targets in Hodgkin cell lines with wild-type p53, whereas no effects were observed in Hodgkin cell lines that harbor p53 mutations. Activation of the p53 pathway led to strong induction of apoptosis in p53 wild-type Hodgkin cell lines. Knock-down of p53 by RNA interference protected cells from nutlin-induced apoptosis, demonstrating that nutlin-3a exerts its effects strictly through p53. In addition, MDM2 inhibition strongly sensitized HRS cells to cytotoxic drugs, such as doxorubicin, etoposide, or vincristine. In view of the fact that HRS cells are characterized by high constitutive NF-κB activity, we also analyzed the effects of a second non-genotoxic agent, geldanamycin, which is an inhibitor of the HSP90/NF-κB pathway. Titration experiments showed that the pro-apoptotic effects of geldanamycin correlate with the mutation status of IκB proteins, demonstrating strong induction of apoptosis in cell lines with wild-type IκB. Furthermore, Hodgkin cell lines that contain wild-type IκB but lack functional p53 (through mutation or siRNA knock-down) are resistant to nutlin treatment, but still respond to treatment with geldanamycin. This indicates that inhibitors of HSP90 induce apoptosis in HRS cells in a p53-independent manner. Therefore, combined targeting of p53-dependent and p53-independent pathways could be a promising approach to develop highly effective and less genotoxic treatment strategies for patients with cHL.