MDM2 phenotypic and genotypic profiling, respective to TP53 genetic status, in diffuse large B-cell lymphoma patients treated with rituximab-CHOP immunochemotherapy: a report from the International DLBCL Rituximab-CHOP Consortium Program

Abstract

MDM2 is a key negative regulator of the tumor suppressor p53, however, the prognostic significance of MDM2 overexpression in diffuse large B-cell lymphoma (DLBCL) has not been defined convincingly. In a p53 genetically-defined large cohort of de novo DLBCL patients treated with rituximab, cyclophosphamide, hydroxydaunorubicin, vincristine, and prednisone (R-CHOP) chemotherapy, we assessed MDM2 and p53 expression by immunohistochemistry (n = 478), MDM2 gene amplification by fluorescence in situ hybridization (n = 364), and a single nucleotide polymorphism in the MDM2 promoter, SNP309, by SNP genotyping assay (n = 108). Our results show that MDM2 overexpression, unlike p53 overexpression, is not a significant prognostic factor in overall DLBCL. Both MDM2 and p53 overexpression do not predict for an adverse clinical outcome in patients with wild-type p53 but predicts for significantly poorer survival in patients with mutated p53. Variable p53 activities may ultimately determine the survival differences, as suggested by the gene expression profiling analysis. MDM2 amplification was observed in 3 of 364 (0.8%) patients with high MDM2 expression. The presence of SNP309 did not correlate with MDM2 expression and survival. This study indicates that evaluation of MDM2 and p53 expression correlating with TP53 genetic status is essential to assess their prognostic significance and is important for designing therapeutic strategies that target the MDM2-p53 interaction.

Figure 1

MDM2 and p53 expression in DLBCL patients treated with R-CHOP. (A-B) Histogram showing the distribution of MDM2 and p53 expression levels in the DLBCL cohort. X-axis, percentage of immunopositive cells in tumors; Y-axis, numbers of DLBCL patients. (C) Illustration of p53 and MDM2 kinetic pulses in tumor cells under stress conditions. In a single cell, because of the fluctuating p53/MDM2 levels in oscillatory pulses, the IHC pattern is (1) p53^–/MDM2^– at zero time point A or E after cellular stress is removed; (2) p53⁺/MDM2⁺ at time point B; (3) p53^–/MDM2⁺ at time point C; and (4) p53⁺/MDM2^– at time point D. Same patterns over cell population may be generated by evaluation of the percentage of MDM2⁺ cells and defining overexpression by certain cutoffs. (D) Representative MDM2 immunohistochemical staining patterns.

Figure 2

Impact of MDM2 and p53 expression on OS in de novo DLBCL patients with WT- or MUT-p53.

Figure 3

Impact of MDM2 and p53 expression on PFS in de novo DLBCL patients with WT- or MUT-p53.

Figure 4

Concurrent evaluation of p53 and MDM2 overexpression in DLBCL.

Figure 5

Comparison and characterization of differentially expressed genes in subgroups of DLBCL patients treated with R-CHOP. (A) DEGs in patients with MDM2⁺ and MDM2^– DLBCL with WT-p53. (B) DEGs in patients with MDM2⁺ and MDM2^– DLBCL with MUT-p53. (C) DEGs in patients with p53⁺ and p53^– DLBCL with WT-p53. (D) MDM2 amplification shown by FISH. In every single cell, the orange signals (MDM2), by at least two times, outnumber the green signals (centromere 12). (E-F) Impact of MDM2 SNP309 on OS and PFS in DLBCL patients.