Smeland, Erlend B.

Abstract Primary mediastinal large B-cell lymphoma (PMBCL) is recognized as a distinct entity in the World Health Organization classification. Currently, the diagnosis relies on consensus of histopathology, clinical variables, and presentation, giving rise to diagnostic inaccuracy in routine practice. Previous studies have demonstrated that PMBCL can be distinguished from subtypes of diffuse large B-cell lymphoma (DLBCL) based on gene expression signatures. However, requirement of fresh-frozen biopsy material has precluded the transfer of gene expression–based assays to the clinic. Here, we developed a robust and accurate molecular classification assay (Lymph3Cx) for the distinction of PMBCL from DLBCL subtypes based on gene expression measurements in formalin-fixed, paraffin-embedded tissue. A probabilistic model accounting for classification error, comprising 58 gene features, was trained on 68 cases of PMBCL and DLBCL. Performance of the model was subsequently evaluated in an independent validation cohort of 158 cases and showed high agreement of the Lymph3Cx molecular classification with the clinicopathological diagnosis of an expert panel (frank misclassification rate, 3.8%). Furthermore, we demonstrate reproducibility of the assay with 100% concordance of subtype assignments at 2 independent laboratories. Future studies will determine Lymph3Cx’s utility for routine diagnostic purposes and therapeutic decision making.

SummaryPatients with mantle cell lymphoma (MCL) generally have a dismal prognosis. Intensified induction treatment with rituximab and high dose cytarabine (R_HDAC), and consolidation with high‐dose therapy with autologous stem cell support has resulted in 10‐year overall survival (OS) higher than 60%. However, the clinical course varies. Diagnostic tools capable of stratifying patients include the MCL International Prognostic Index (MIPI), gene expression‐based proliferation signature, Ki‐67 proliferation index or tumour cell morphology. Here, we tested the performance of a newly developed Nanostring‐based RNA expression‐based proliferation assay (MCL35) on formalin‐fixed paraffin‐embedded tumour tissue from younger patients recruited in or treated according to Nordic MCL protocols compared to the prognosticators listed above. Seventy‐four patients were included and the assay performed well in all cases except four, which had inadequate RNA quality. The patients were evenly distributed in the MCL35 low‐, intermediate‐ and high‐risk categories. MCL35 low‐ and intermediate‐ risk groups had overlapping progression‐free survival (PFS), while patients in the high‐risk category had significantly inferior PFS. Combining MCL35 with MIPI or the MIPI‐C (MIPI with the addition of binary Ki67 score +/−30%) showed a better discrimination than either assessment alone. In conclusion, the MCL35 assay alone or combined with MIPI or MIPI‐C scores can identify patients who still have a dismal outcome despite intensified treatment.