Gene Expression Profiling by a Quantitative Nuclease Protection Assay (qNPA) but Not Immunohistochemical Algorithms Predicts Clinical Outcome in Diffuse Large B-Cell Lymphoma.
Ahmet Dogan, Nazan Ozsan, Ming Mai, Julie A Vrana, Matthew J Maurer, Thomas M Habermann, Bollette L Caron, James R Cerhan, Rebecca F McClure. Mayo Clinic, Rochester, MN
Background: Gene expression profiling (GEP) on frozen tumor tissue has identified biological subgroups (GCB and ABC) of diffuse large B-cell lymphoma (DLBCL) with distinct clinical outcomes. However, immunohistochemistry (IHC) algorithms developed to translate the GEP findings into routine clinical practice using fixed paraffin embedded (PE) specimens have had mixed results. Recently, a quantitative nuclease protection assay (qNPA) that could reproducibly measure the expression levels of multiple genes in routine PE specimens has been reported. In this study we used the qNPA to measure expression of genes implicated in DLBCL outcome on PE specimens and compared our results wih IHC algorithms.
Design: We studied 72 cases of newly diagnosed DLBCL treated with rituximab and anthracycline based chemotherapy. IHC for CD10, BCL6, IRF4, GCET1, MUM1, FOXP1 and LMO2 was performed on PE sections and DLBCL biological subgroups GCB and ABC were assigned according to previously published criteria. qNPA was performed on the diagnostic PE specimens of 57 samples to determine gene expression of 36 genes according to previously established method (Blood 2008;112:3425). Cox proportional hazards models were used to assess the association of IHC and qNPA data with event-free and overall survival. Wilcoxon rank sum tests were used to compare the distributions of qNPA and IHC results.
Results: None of the individual IHC markers or algorithms (CD10/BCL6/IRF4, GCET1/CD10/IRF4/BCL6/FOXp1 and LMO2) predicted clinical outcome (all p > 0.25). The qNPA assay identified several individual genes with significant association with overall survival (FAM38A: (HR=0.41), PLAU (HR=0.48), CCL3 (HR=0.55), NR4A3 (HR=0.50), all p<0.05. Several additional genes trended towards significant association in the direction of previously reported studies: ACTN1 (HR=0.51), GCET1 (HR=0.62), SOD2 (HR=0.59), all p <0.10. A ratio score based on the qNPA expression of genes from the six-gene model of Lossos (NEJM 2004;350:1828; BCL6, FN1, BCL2, CCL3, CCND2, LMO2) was associated with overall survival (p=0.09). Including the next two most prognostic genes available from the Lossos analysis (MYC and PLAU) further increased the utility of the model (p=0.004). In genes with both IHC and qNPA results (BCL6, GCET1, LMO2), IHC positivity was associated with significantly higher gene expression on qNPA (all p < 0.003).
Conclusions: The qNPA technology offers a robust, multiplexed, quantitative platform for translation of GEP findings in DLBCL into routine clinical practice.
Monday, February 28, 2011 11:45 AM
Platform Session: Section B, Monday Morning