Describing Treatment of Primary Mediastinal Large B Cell Lymphoma Using Rigorously Defined Molecular Classification: A Retrospective Analysis
Authors: Casulo Carla , Strawderman Myla , Steiner Raphael , Delage Carolyne , Faugh Tina , Maurer Matthew J. , Link Brian K. , Rosenthal Allison C. , Ramsower Colleen , Burack Richard , Syrbu Sergei , Feldman Andrew L. , Rock Philip J , Cerhan James R. , Flowers Christopher , Rimsza Lisa M. , Friedberg Jonathan W. 
Published: November 2020

Introduction
Primary mediastinal large B cell lymphoma (PMBCL) is a rare non-Hodgkin lymphoma (NHL) with a female predominance; often presenting with a large anterior mediastinal mass. Though PMBCL has clinical and molecular features overlapping with Hodgkin lymphoma, it is a distinct entity defined by the World Health Organization classification. PMBCL is heterogeneously treated, and most patients receive front line therapy with either rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone (R-CHOP) with radiotherapy (RT), or the more intensive etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin with rituximab (EPOCH-R) regimen. Diagnosis of PMBCL is made using clinicopathologic criteria and radiographic imaging, however gene expression profiling (GEP) studies reveal a characteristic genotypic signature distinct from diffuse large B cell lymphoma (DLBCL). Molecular classification of PMBCL using the Lymph3Cx assay from formalin-fixed paraffin-embedded tissue (FFPE) is feasible, reproducible, and highly concordant in a training and validation cohort (Mottok et al. Blood 2018). Using a multicenter cohort of patients, we sought to estimate the rate of mis-match among patients with a clinical diagnosis of PMBCL using Lymph3Cx, and describe treatment selections and outcomes for each group.
Methods
Patients were identified from a cohort of patients with newly diagnosed NHL from the University of Iowa and Mayo Clinic Molecular Epidemiology Resource, and the Lymphoma Epidemiology of Outcomes cohort. Patients were enrolled between 2002-2019, and included if they had clinically defined PMBCL. FFPE was retrieved from hematopathology archives of participating academic centers. All diagnoses of PMBCL were based on expert hematopathology review at the time of therapy, and all cases underwent classification by GEP using the Lymph3Cx assay.
Lymph3Cx was performed in the clinical lab at the Mayo Clinic in Arizona: Contiguous unstained sections were deparaffinized and macrodissected to enrich for tumor content before RNA isolation;100-200 ng of total RNA was used in an nCounter Elements XT, hybridized, and processed the following day using the nCounter FLEX system. Raw counts were processed through the Lymph3Cx algorithm and results reported as probability of PMBCL (≥0.90 as PMBCL, ≤0.10 as DLBCL all other results "Unclear PMBCL/DLBCL") (A. Mottok et al, Blood, 2018). For cases classified as DLBCL, the Lymph2Cx cell-of-origin classifier results was reported (Scott et al, JCO, 2016).
Time to event endpoints were described with Kaplan-Meier plots by groups defined by mismatch status and compared with a logrank test. Binary outcomes will be presented with 90% exact confidence intervals.
Results
Fifty patients were identified. Median age was 35 years (range 19-70). Sixty four percent were women. Median follow up was 47 months. Treatments included R-CHOP (44%), EPOCH-R (44%), and MACOP-B [methotrexate with leucovorin rescue, doxorubicin, cyclophosphamide, vincristine, prednisone, and bleomycin] (6%), other (4%). Ten patients (20%) had events (defined as progression or death). Three patients in the entire cohort (6%) died. The Kaplan-Meier estimated survival at 47 months (median follow-up) is 92%.
The Lymph3Cx assay yielded gene expression data of sufficient quality in 47/50 cases (94%, 90% CI=85.2, 98.3%). Of 47 cases clinically identified as PMBCL, 5 unclear were DLBCL/PMBCL and 1 was Germinal Center B cell subtype of DLBCL. Among these 6 patients, 4 received R-EPOCH (66%), 1 received R-CHOP (16.6%). One patient had missing treatment data. One patient had an event requiring subsequent therapy; all patients remain alive.
Conclusions
Using 47 patients with PMBCL defined by histology, clinical and radiographic findings, and molecular features, we demonstrate high concordance between clinical phenotype and molecular genotype of PMBCL by Lymph3Cx. Among the 6 patients not classified as PMBCL, most received R-EPOCH. Differences in outcome by mis-match status await longer follow-up and further accrual of subjects to our data base. Our data suggest molecular genotyping may have a role in mediastinal presentations of large cell lymphoma to optimize treatment decision making.

Disclosures
Maurer: Nanostring: Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees; Celgene / BMS: Research Funding; Morphosys: Membership on an entity's Board of Directors or advisory committees; Kite: Membership on an entity's Board of Directors or advisory committees. Cerhan:BMS/Celgene: Research Funding; NanoString: Research Funding. Flowers:AbbVie: Consultancy, Research Funding; Kite: Research Funding; Burroughs Wellcome Fund: Research Funding; Genentech, Inc./F. Hoffmann-La Roche Ltd: Consultancy, Research Funding; Denovo Biopharma: Consultancy; Celgene: Consultancy, Research Funding; Cancer Prevention and Research Institute of Texas: Research Funding; TG Therapeutics: Research Funding; Eastern Cooperative Oncology Group: Research Funding; V Foundation: Research Funding; Bayer: Consultancy; National Cancer Institute: Research Funding; Millennium/Takeda: Consultancy, Research Funding; Gilead: Consultancy, Research Funding; Acerta: Research Funding; Spectrum: Consultancy; Pharmacyclics/Janssen: Consultancy; Karyopharm: Consultancy; OptumRx: Consultancy; Leukemia and Lymphoma Society: Membership on an entity's Board of Directors or advisory committees; BeiGene: Consultancy. Friedberg:Acerta Pharma - A member of the AstraZeneca Group, Bayer HealthCare Pharmaceuticals.: Other; Astellas: Consultancy; Bayer: Consultancy; Kite Pharmaceuticals: Research Funding; Portola Pharmaceuticals: Consultancy; Roche: Other: Travel expenses; Seattle Genetics: Research Funding.