MYC Translocations and Expression Are Clinically Important in R-CHOP Treated Patients with De Novo DLBCL.

MYC, an oncogene associated with cellular proliferation, is deregulated as a result of chromosomal translocation in Burkitt lymphoma (BL), and in 8-12% of diffuse large B cell lymphomas (DLBCL). In 2006, 2 studies defined the molecular features of BL and DLBCL by gene expression profiling (GEP) (Dave, NEJM 2006; Hummel, NEJM 2006) and identified a subset of cases that resembled DLBCL by morphology, but by GEP, expressed MYC and MYC target genes similar to classical BL, i.e. molecular BL (mBL) signature. The clinical outcome of these cases is poorly defined. More recently, MYC expression and MYC translocations (MYC tr+) have been associated with an inferior overall survival (OS) in de novo DLBCL patients (pts) treated with R-CHOP (Rimsza, Blood 2008; Savage, Blood 2009) but the prognostic impact of BCL2 protein expression and concurrent BCL2 translocations (BCL2 tr+) is poorly understood. We investigated the prognostic impact of the presence of a mBL signature by GEP, high MYC mRNA expression, and the presence of a MYC tr+ with or without a concurrent BCL2 tr+ in DLBCL pts treated uniformly with R-CHOP.

315 samples were reviewed by a panel of expert hematopathologists and classified according to the WHO 2008 criteria. Pts with high grade B cell lymphoma otherwise unclassifiable, BL, primary mediastinal B cell lymphoma (PMBCL), T-cell-rich B cell lymphoma and pts that were not treated with R-CHOP were excluded from this analysis. The remaining 259 DLBCL samples were subjected to GEP as previously described (Lenz, NEJM 2008). Tissue microarrays (TMA) were constructed in cases with available paraffin material. 184 had successful GEP arrays, 186 were included on a TMA and 151 cases were assessed on both platforms. Presence of a mBL signature was determined according to the method described by Dave (NEJM 2006). MYC expression was determined using log normalized expression values from Affymetrix U133 Plus 2.0 probe set id 202431_s_at and dichotomized into high versus low expression using a cut off threshold determined by the statistical software X-Tile (http://www.tissuearray.org/rimmlab/). The presence of MYC tr+ or BCL2 tr+ was determined by fluorescence in situ hybridization (FISH) using MYC and BCL2 breakapart probes (Abbott) on TMAs. BCL2 protein expression was determined by immunohistochemistry (IHC) using clone 124 (Dako). Correlation between variables and association with OS was performed by Pearson Chi-Square, Kaplan-Meier and Cox regression analysis using SPSS statistical software.

A mBL signature was found in 4/184 samples (2%). One case was MYC tr+, one was MYC tr-, and the MYC translocation status was unknown in the remaining 2 cases. All 4 pts with a mBL had a complete response to R-CHOP lasting >2 years after diagnosis. MYC tr+, BCL2 tr+ or concurrent MYC tr+/ BCL2 tr+ were present in 12%, 20% and 4% of 186 DLBCL cases, respectively. BCL2 tr+ were predominately found in the germinal center B cell (GCB) molecular subtype (36%) compared to the activated B cell (ABC) or unclassifiable (U) subtypes (4% and 19%, p=0.0001) but were not associated with an inferior OS. In contrast, MYC tr+ were not associated with a specific molecular subtype (GCB 15%, ABC 8%, U 19%, p=0.2) but were associated with an inferior OS (p=0.0078). When dichotomizing patients with MYC tr+ according to the BCL2 status, pts with concurrent MYC tr+/ BCL2 tr+ (4%) and pts with MYC tr+ and BCL2 protein-positive biopsies (7%) had a markedly inferior OS compared to pts with MYC tr+ and BCL2 protein-negative biopsies or pts with no MYC tr (median OS 7 months vs. not reached, both p < 0.00001). The presence of MYC tr+ correlated with high MYC expression in 6/16 (38%) MYC tr+ cases whereas high MYC expression was present in 5/111 (5%) MYC tr- cases (p=0.0001). High MYC expression alone was also associated with an inferior OS (p<0.00001). In multivariate analysis, high MYC mRNA expression, concurrent MYC tr+/ BCL2 tr+, and the IPI were independent predictors of OS (p=0.04, p=0.05, p=0.007, respectively).

MYC expression, as prognostic marker in DLBCL, should be investigated in routine clinical practice. Cytogenetic analysis to determine MYC and BCL2 translocation status by FISH and/or karyotype of de novo DLBCL samples, and BCL2 protein expression by IHC are clinically indicated to identify a group of high-risk pts that may benefit from up-front intensified therapy.

Connors:Roche Canada: Research Funding. Gascoyne:Roche Canada: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.