Abstract
Gene expression profiling (GEP) has stratified diffuse large B-cell lymphoma (DLBCL) into molecular subgroups that correspond to different stages of lymphocyte development–namely germinal center B-cell like and activated B-cell like. This classification has prognostic significance, but GEP is expensive and not readily applicable into daily practice, which has lead to immunohistochemical algorithms proposed as a surrogate for GEP analysis. We assembled tissue microarrays from 475 de novo DLBCL patients who were treated with rituximab-CHOP chemotherapy. All cases were successfully profiled by GEP on formalin-fixed, paraffin-embedded tissue samples. Sections were stained with antibodies reactive with CD10, GCET1, FOXP1, MUM1 and BCL6 and cases were classified following a rationale of sequential steps of differentiation of B cells. Cutoffs for each marker were obtained using receiver-operating characteristic curves, obviating the need for any arbitrary method. An algorithm based on the expression of CD10, FOXP1 and BCL6 was developed that had a simpler structure than other recently proposed algorithms and 92.6% concordance with GEP. In multivariate analysis, both the International Prognostic Index and our proposed algorithm were significant independent predictors of progression-free and overall survival. In conclusion, this algorithm effectively predicts prognosis of DLBCL patients matching GEP subgroups in the era of rituximab therapy.
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References
Jaffe ES, Harris NL, Stein H, Isaacson PG . Classification of lymphoid neoplasms: the microscope as a tool for disease discovery. Blood 2008; 112: 4384–4399.
Coiffier B, Thieblemont C, Van Den Neste E, Lepeu G, Plantier I, Castaigne S et al. Long-term outcome of patients in the LNH-98.5 trial, the first randomized study comparing rituximab-CHOP to standard CHOP chemotherapy in DLBCL patients: a study by the Groupe d'Etudes des Lymphomes de l'Adulte. Blood 2010; 116: 2040–2045.
Alizadeh AA, Eisen MB, Davis RE, Ma C, Lossos IS, Rosenwald A et al. Distinct types of diffuse large B-cell lymphoma identified by gene expression profiling. Nature 2000; 403: 503–511.
Rosenwald A, Wright G, Chan WC, Connors JM, Campo E, Fisher RI et al. The use of molecular profiling to predict survival after chemotherapy for diffuse large-B-cell lymphoma. N Engl J Med 2002; 346: 1937–1947.
Shaffer AL, Rosenwald A, Staudt LM . Lymphoid malignancies: the dark side of B-cell differentiation. Nat Rev Immunol 2002; 2: 920–932.
Lenz G, Staudt LM. . Aggressive lymphomas. N Engl J Med 2010; 362: 1417–1429.
Fu K, Weisenburger DD, Choi WW, Perry KD, Smith LM, Shi X et al. Addition of rituximab to standard chemotherapy improves the survival of both the germinal center B-cell-like and non-germinal center B-cell-like subtypes of diffuse large B-cell lymphoma. J Clin Oncol 2008; 26: 4587–4594.
A predictive model for aggressive non-Hodgkin's lymphoma The International Non-Hodgkin's Lymphoma Prognostic Factors Project. N Engl J Med 1993; 329: 987–994.
Hans CP, Weisenburger DD, Greiner TC, Gascoyne RD, Delabie J, Ott G et al. Confirmation of the molecular classification of diffuse large B-cell lymphoma by immunohistochemistry using a tissue microarray. Blood 2004; 103: 275–282.
Nyman H, Adde M, Karjalainen-Lindsberg ML, Taskinen M, Berglund M, Amini RM et al. Prognostic impact of immunohistochemically defined germinal center phenotype in diffuse large B-cell lymphoma patients treated with immunochemotherapy. Blood 2007; 109: 4930–4935.
Natkunam Y, Farinha P, Hsi ED, Hans CP, Tibshirani R, Sehn LH et al. LMO2 protein expression predicts survival in patients with diffuse large B-cell lymphoma treated with anthracycline-based chemotherapy with and without rituximab. J Clin Oncol 2008; 26: 447–454.
Choi WW, Weisenburger DD, Greiner TC, Piris MA, Banham AH, Delabie J et al. A new immunostain algorithm classifies diffuse large B-cell lymphoma into molecular subtypes with high accuracy. Clin Cancer Res 2009; 15: 5494–5502.
Xia ZG, Xu ZZ, Zhao WL, Zhao SQ, Ding F, Chen Y et al. The prognostic value of immunohistochemical subtyping in Chinese patients with de novo diffuse large B-cell lymphoma undergoing CHOP or R-CHOP treatment. Ann Hematol 2010; 89: 171–177.
Seki R, Ohshima K, Fujisaki T, Uike N, Kawano F, Gondo H et al. Prognostic impact of immunohistochemical biomarkers in diffuse large B-cell lymphoma in the rituximab era. Cancer Sci 2009; 100: 1842–1847.
Nyman H, Jerkeman M, Karjalainen-Lindsberg ML, Banham AH, Leppa S . Prognostic impact of activated B-cell focused classification in diffuse large B-cell lymphoma patients treated with R-CHOP. Mod Pathol 2009; 22: 1094–1101.
Meyer PN, Fu K, Greiner TC, Smith LM, Delabie J, Gascoyne RD et al. Immunohistochemical methods for predicting cell of origin and survival in patients with diffuse large B-cell lymphoma treated with rituximab. J Clin Oncol 2011; 29: 200–207.
Montes-Moreno S, Roncador G, Maestre L, Martinez N, Sanchez-Verde L, Camacho FI et al. Gcet1 (centerin), a highly restricted marker for a subset of germinal center-derived lymphomas. Blood 2008; 111: 351–358.
Barrans SL, Fenton JA, Banham A, Owen RG, Jack AS . Strong expression of FOXP1 identifies a distinct subset of diffuse large B-cell lymphoma (DLBCL) patients with poor outcome. Blood 2004; 104: 2933–2935.
Brown PJ, Ashe SL, Leich E, Burek C, Barrans S, Fenton JA et al. Potentially oncogenic B-cell activation-induced smaller isoforms of FOXP1 are highly expressed in the activated B cell-like subtype of DLBCL. Blood 2008; 111: 2816–2824.
Gutiérrez-García G, Cardesa-Salzmann T, Climent F, Gonzalez-Barca E, Mercadal S, Mate JL et al. Gene-expression profiling and not immunophenotypic algorithms predicts prognosis in patients with diffuse large B-cell lymphoma treated with immunochemotherapy. Blood 2011; 117: 4836–4843.
Marko NF, Frank B, Quackenbush J, Lee NH . A robust method for the amplification of RNA in the sense orientation. BMC Genomics 2005; 6: 27.
Liu WM, Li R, Sun JZ, Wang J, Tsai J, Wen W et al. PQN and DQN: algorithms for expression microarrays. J Theoretical Biol 2006; 243: 273–278.
Wright G, Tan B, Rosenwald A, Hurt EH, Wiestner A, Staudt LM . A gene expression-based method to diagnose clinically distinct subgroups of diffuse large B cell lymphoma. Proc Natl Acad Sci USA 2003; 100: 9991–9996.
Williams PM, Li R, Johnson NA, Wright G, Heath J-D, Gascoyne RD . A novel method of amplification of FFPET-derived RNA enables accurate disease classification with microarrays. J Mol Diagn 2010; 12: 680–686.
Lenz G, Wright G, Dave SS, Xiao W, Powell J, Zhao H et al. Stromal gene signatures in large-B-cell lymphomas. N Engl J Med 2008; 359: 2313–2323.
Tzankov A, Zlobec I, Went P, Robl H, Hoeller S, Dirnhofer S . Prognostic immunophenotypic biomarker studies in diffuse large B cell lymphoma with special emphasis on rational determination of cut-off scores. Leuk Lymphoma 2010; 51: 199–212.
de JD, Xie W, Rosenwald A, Chhanabhai M, Gaulard P, Klapper W et al. Immunohistochemical prognostic markers in diffuse large B-cell lymphoma: validation of tissue microarray as a prerequisite for broad clinical applications (a study from the Lunenburg Lymphoma Biomarker Consortium). J Clin Pathol 2009; 62: 128–138.
Banham AH, Beasley N, Campo E, Fernandez PL, Fidler C, Gatter K et al. The FOXP1 winged helix transcription factor is a novel candidate tumor suppressor gene on chromosome 3 p. Cancer Res 2001; 61: 8820–8829.
Youden WJ . Index for rating diagnostic tests. Cancer 1950; 3: 32–35.
Cheson BD, Horning SJ, Coiffier B, Shipp MA, Fisher RI, Connors JM et al. Report of an international workshop to standardize response criteria for non-Hodgkin's lymphomas. NCI Sponsored International Working Group. J Clin Oncol 1999; 17: 1244–1253.
Kaplan E, Meier P . Nonparametric estimation from incomplete observations. J Am Stat Assoc 1958; 53: 457–481.
Cox DR . Regression models and life-tables. J R Stat Soc 1972; 34: 187–220.
Rimsza LM, Leblanc ML, Unger JM, Miller TP, Grogan TM, Persky DO et al. Gene expression predicts overall survival in paraffin-embedded tissues of diffuse large B-cell lymphoma treated with R-CHOP. Blood 2008; 112: 3425–3433.
Dunleavy K, Pittaluga S, Czuczman MS, Dave SS, Wright G, Grant N et al. Differential efficacy of bortezomib plus chemotherapy within molecular subtypes of diffuse large B-cell lymphoma. Blood 2009; 113: 6069–6076.
Hernandez-Ilizaliturri FJ, Deeb G, Zinzani PL, Pileri SA, Malik F, Macon WR et al. Higher response to lenalidomide in relapsed/refractory diffuse large b-cell lymphoma in nongerminal center b-cell-like than in germinal center b-cell-like phenotype. Cancer 2011; 117: 5058–5066.
Uddin S, Hussain AR, Ahmed M, Al-Dayel F, Bu R, Bavi P et al. Inhibition of c-MET is a potential therapeutic strategy for treatment of diffuse large B-cell lymphoma. Lab Invest 2010; 90: 1346–1356.
Milhollen MA, Traore T, Adams-Duffy J, Thomas MP, Berger AJ, Dang L et al. MLN4924, a NEDD8-activating enzyme inhibitor, is active in diffuse large B-cell lymphoma models: rationale for treatment of NF-{kappa}B-dependent lymphoma. Blood 2010; 116: 1515–1523.
Hailfinger S, Lenz G, Ngo V, Posvitz-Fejfar A, Rebeaud F, Guzzardi M et al. Essential role of MALT1 protease activity in activated B cell-like diffuse large B-cell lymphoma. Proc Natl Acad Sci USA 2009; 106: 19946–19951.
Schwickert TA, Lindquist RL, Shakhar G, Livshits G, Skokos D, Kosco-Vilbois MH et al. In vivo imaging of germinal centres reveals a dynamic open structure. Nature 2007; 446: 83–87.
Frazer JK, Jackson DG, Gaillard JP, Lutter M, Liu YJ, Banchereau J et al. Identification of centerin: a novel human germinal center B cell-restricted serpin. Eur J Immunol 2000; 30: 3039–3048.
Falini B, Mason DY . Proteins encoded by genes involved in chromosomal alterations in lymphoma and leukemia: clinical value of their detection by immunocytochemistry. Blood 2002; 99: 409–426.
Hu H, Wang B, Borde M, Nardone J, Maika S, Allred L et al. Foxp1 is an essential transcriptional regulator of B cell development. Nat Immunol 2006; 7: 819–826.
Lenz G, Wright GW, Emre NC, Kohlhammer H, Dave SS, Davis RE et al. Molecular subtypes of diffuse large B-cell lymphoma arise by distinct genetic pathways. Proc Natl Acad Sci USA 2008; 105: 13520–13525.
Klein U, Casola S, Cattoretti G, Shen Q, Lia M, Mo T et al. Transcription factor IRF4 controls plasma cell differentiation and class-switch recombination. Nat Immunol 2006; 7: 773–782.
Lossos IS, Alizadeh AA, Rajapaksa R, Tibshirani R, Levy R . HGAL is a novel interleukin-4-inducible gene that strongly predicts survival in diffuse large B-cell lymphoma. Blood 2003; 101: 433–440.
Lossos IS, Czerwinski DK, Alizadeh AA, Wechser MA, Tibshirani R, Botstein D et al. Prediction of survival in diffuse large-B-cell lymphoma based on the expression of six genes. N Engl J Med 2004; 350: 1828–1837.
Natkunam Y, Lossos IS, Taidi B, Zhao S, Lu X, Ding F et al. Expression of the human germinal center-associated lymphoma (HGAL) protein, a new marker of germinal center B-cell derivation. Blood 2005; 105: 3979–3986.
Lu X, Chen J, Malumbres R, Cubedo Gil E, Helfman DM, Lossos IS . HGAL a lymphoma prognostic biomarker, interacts with the cytoskeleton and mediates the effects of IL-6 on cell migration. Blood 2007; 110: 4268–4277.
Natkunam Y, Zhao S, Mason DY, Chen J, Taidi B, Jones M et al. The oncoprotein LMO2 is expressed in normal germinal-center B cells and in human B-cell lymphomas. Blood 2007; 109: 1636–1642.
Pan Z, Shen Y, Du C, Zhou G, Rosenwald A, Staudt LM et al. Two newly characterized germinal center B-cell-associated genes, GCET1 and GCET2, have differential expression in normal and neoplastic B cells. Am J Pathol 2003; 163: 135–144.
Amen F, Horncastle D, Elderfield K, Banham AH, Bower M, Macdonald D et al. Absence of cyclin-D2 and Bcl-2 expression within the germinal centre type of diffuse large B-cell lymphoma identifies a very good prognostic subgroup of patients. Histopathology 2007; 51: 70–79.
Shaffer AL, Shapiro-Shelef M, Iwakoshi NN, Lee AH, Qian SB, Zhao H et al. XBP1, downstream of Blimp-1, expands the secretory apparatus and other organelles, and increases protein synthesis in plasma cell differentiation. Immunity 2004; 21: 81–93.
Harris NL, Jaffe ES, Stein H, Banks PM, Chan JK, Cleary ML et al. A revised European-American classification of lymphoid neoplasms: a proposal from the International Lymphoma Study Group [see comments]. Blood 1994; 84: 1361–1392.
Gatter KC, Warnke RA . Diffuse large B-cell lymphoma. In: Jaffe ES, Harris NL, Stein H, Vardiman JW eds Pathology and Genetics: Tumours of Haematopoietic and Lymphoid Tissues. World Health Organization Classification of Tumours. IARC Press Lyon, France, 2001, 171–174.
Stein H, Warnke RA, Chan WC, Jaffe ES, Chan JKC, Gatter KC et al. Diffuse large B-cell lymphoma, not otherwise specified. In: Swerdlow SH, Campo E, Harris NL eds WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues 4th edn. IARC Lyon, France, 2008, 233–237.
Acknowledgements
We thank our consortium program team of pathologists, hematologists and clinicians, and each of the contributing center principal physicians for their support in selection, evaluation and contribution of the cases. We thank our patients, former and current hematopathology and hematology/oncology fellows, and research scientists (Chih-Jian Lih, Paul M. Williams, Lynn Trinh and Yuchaun Tai) for their support. Technical and publication editing supports from Maitrayee Goswami and Virginia Mohlere from the Department of Scientific Publications are greatly appreciated. The abstract was presented as oral communication at the Lugano ICML Conference on June 16, 2011. CV is a honorable visiting hematologist supported by San Bortolo Hospital, Vicenza, Italy and The University of Texas MD Anderson Cancer Center. KHY is supported by The University of Texas MD Anderson Cancer Center Institutional R and D Fund, Institutional Research Grant Award, MD Anderson Cancer Center SPORE Research Development Program Award, Gundersen Lutheran Medical Foundation Award and Forward Lymphoma Fund. This study is also partially supported by the Zurich Stiftung zur Krebsbekaempfung and NCI/NIH (R01CA138688 and 1RC1CA146299). Publicly available data sets: All primary sequencing data will be made publicly available through the GEO archive through accession GSE#31312.
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Designed research: CV and KHY. Performed research: CV, YL, ZYXM, WL, SMM, MAP, MBM, LW and KHY. Contributed vital new reagents, resource, and analytical tools under approved IRB and MTA: CV, YL, RNM, AT, WW, WL, ESGD, SMM, KD, AC, WT, AO, YZ, GB, JNW, SON, CD, EDH, XFZ, RSG, WWLC, FZ, JT, XYZ, JHVK, QH, MAP, MBM, CEBR, LJM, LW and KHY. Collected data and follow-up under approved IRB and MTA: CV, ZYXM, RNM, TMG, AT, ESGD, SMM, KD, AC, WT, AO, YZ, GB, JNW, HYW, SON, CD, EDH, XFZ, RSG, WWLC, FZ, MC, JT, XYZ, JHVK, QH, WA, JE, MP, AJMF, MAP, MBM, CEBR, LJM and KHY. Contributed vital strategies, participated in discussions and provided scientific input: CV, YL, ZYXM, RNM, TMG, YL, AT, WW, WL, BSK, ESGD, SMM, KD, AC, WT, AO, YZ, GB, JNW, HYW, SON, CD, EDH, XFZ, RSG, WWLC, FZ, MC, JT, XYZ, JHVK, QH, WA, JE, MP, AJMF, MAP, MBM, CEBR, LJM, LW and KHY. Analyzed data: CV and KHY. Performed and supported statistical analysis: CV, AT and KHY. Wrote the paper: CV and KHY.
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Visco, C., Li, Y., Xu-Monette, Z. et al. Comprehensive gene expression profiling and immunohistochemical studies support application of immunophenotypic algorithm for molecular subtype classification in diffuse large B-cell lymphoma: a report from the International DLBCL Rituximab-CHOP Consortium Program Study. Leukemia 26, 2103–2113 (2012). https://doi.org/10.1038/leu.2012.83
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DOI: https://doi.org/10.1038/leu.2012.83
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