ASBMT. Controversies in BMT for Lymphoma. Randy D. Gascoyne 1, Craig Moskowitz 2, Thomas C. Shea 3, * Section VI: BMT for Lymphoma

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1 Biol Blood Marrow Transplant 19 (2013) S26eS32 Section VI: BMT for Lymphoma Controversies in BMT for Lymphoma Randy D. Gascoyne 1, Craig Moskowitz 2, Thomas C. Shea 3, * ASBMT American Society for Blood and Marrow Transplantation 1 British Columbia Cancer Agency, Vancouver, British Columbia, Canada 2 Division of Hematologic Oncology, Lymphoma and Adult BMT Services (Attending Physician), Memorial Sloan Kettering Cancer Center, Department of Medicine, Weill Medical College of Cornell University, Ithaca, New York 3 Bone Marrow and Stem Cell Transplant Program, UNC Lineberger Comprehensive Cancer Center, UNC, Chapel Hill, North Carolina INTRODUCTION The following educational supplement on Controversies in Diffuse Large B Cell Lymphoma addresses three primary areas. The first section, Biology of Diffuse Large B Cell Lymphomas, focuses on recent advances in understanding the pathogenesis and biology of this disorder with a spotlight on new classification methods that should lead to improved understanding and treatment of this disease. The second section, State of Salvage Therapy for Relapsed and Primary Refractory Diffuse Large B Cell Lymphoma, addresses recent trials that have provided a new benchmark for outcomes of patients undergoing autologous hematopoietic stem cell transplantation (HSCT) after failing initial therapy with rituximab-containing induction regimens. This section reviews some of the new prognostic data that will help categorize patients into those with a good or poor prognosis after autologous HSCT and how novel salvage regimens are needed to improve these results. The third section, Hematopoietic Stem Cell Transplantation Conditioning Regimens and Post-Transplant Maintenance Therapy for Diffuse Large B-Cell Lymphoma, assesses outcomes achieved with a variety of novel and conventional conditioning regimens, including the use of radioimmunotherapy and allogeneic HSCT for poor-prognosis patients. This section also describes potential uses for new agents as maintenance therapy after autologous HSCT. BIOLOGY OF DIFFUSE LARGE B CELL LYMPHOMAS Randy D. Gascoyne Diffuse, aggressive, large B cell lymphomas represent the most frequent histological subtype of non-hodgkin lymphoma encountered in clinical practice in North America. This category includes a spectrum of lymphoma entities, the most common of which is diffuse large B cell lymphoma (DLBCL), not otherwise specified. Also included are distinct entities and subtypes such as primary mediastinal large B cell lymphoma (PMBCL), plasmablastic lymphoma, primary central nervous system DLBCL, and T cell/histiocyte-rich large B cell lymphoma, to name but a few. Normal Germinal Center Normal B cells arise in the bone marrow and undergo primary rearrangement of their immunoglobulin genes before antigen exposure. They exit the bone marrow bearing Financial disclosure: See Acknowledgments on page S32. * Correspondence and reprint requests: Dr. Thomas C. Shea, UNC Lineberger Comprehensive Cancer Center, Chapel Hill, NC. address: sheat@med.unc.edu (T.C. Shea) /$ e see front matter Ó 2013 American Society for Blood and Marrow Transplantation. surface immunoglobulin receptors and seed peripheral lymphoid organs where they encounter antigen. If the antigen induces a T-dependent antibody response, then germinal center formation occurs and represents the anatomical site for the final steps in the generation of antibody diversity. Two important genetic mechanisms occur within the germinal center, including somatic hypermutation and class switch recombination. Both processes require double-stranded DNA breaks and the presence of activation-induced cytidine deaminase. Differentiation through the germinal center is orchestrated by a number of key transcription factors, including BCL6, PRDM1, IRF4, and XBP1. B cells that survive re-express BCL2 and are selected because they possess high-affinity antibody on their surface. These cells exit the germinal center and further differentiate into either mature plasma cells or long-lived memory B cells. Thus, the germinal center is largely responsible for generating both components of long-lived humoral immunity. This complex biology is required to optimize the diversity of our immune response but is equally fraught with errors, some of which lead to the development of B cell non-hodgkin lymphomas [1]. Diffuse Large B Cell Lymphomas Gene Expression Profiling (GEP) has established the presence of at least 3 main molecular subtypes of DLBCL, including the germinal center B cell type (GCB), the activated B cell type (ABC), and PMBCL based on the stage of B cell differentiation and presumed cell-of-origin [2]. GCB and ABC together comprise 80% of all DLBCL cases and are associated with differences in survival, genetics, and mutational profiles highlighting distinct biology (Figure 1). These cell-of-origin distinctions can have a differential impact on the response to novel therapies that are beginning to make their way into the upfront treatment regimens used for this group of malignancies. Lessons learned from a thorough understanding of the biology inform on rational treatment approaches and a path forward for more targeted therapies. Immunohistochemical algorithms have been developed as surrogate testing strategies for determining cell-of-origin, because Affymetrix GEP (Santa Clara, CA) requires frozen tissue that is generally not available in routine clinical practice. The most popular, the Hans algorithm, is based on 3 routine immunohistochemical stains, CD10, BCL6, and MUM1, and is variably associated with outcome, in part because it is a 2-class predictor (GCB versus non-gcb) that tries to recapitulate a three-class GEP predictor (GCB versus ABC versus unclassifiable) but is also fraught with technical

2 R.D. Gascoyne et al. / Biol Blood Marrow Transplant 19 (2013) S26eS32 S27 Figure 1. Critical events in the pathogenesis of DLBCL are depicted. For each of the 4 major molecular entities, the more frequent copy number alterations are shown as well as recurrent mutations. The ABC subtype is thought to arise from a cell just before germinal center exit, having some features of plasmablasts. Both copy number changes and recurrent mutations drive the 3 major themes in ABC pathogenesis, namely constitutive NF-kB signaling, chronic B cell receptor (BCR) signaling, and a block in terminal B cell differentiation to mature plasma cells. The pathogenic events that underlie both PMBCL and nodular sclerosis Hodgkin lymphoma (NSHL) are similar and include up-regulated JAK-STAT signaling and molecular events that foster immune privilege. difficulties and issues related to reproducibility [3]. The development of a robust testing strategy for distinguishing cell-of-origin subtypes is now of paramount importance. The GCB subtype of DLBCL accounts for roughly 50% of all cases, shows a favorable outcome, and reveals a gene expression program reminiscent of normal germinal center B cells. Cases harboring the t(14;18) are only found within the GCB subtype of DLBCL. Recurrent mutations of EZH2, TNFRSF14, MEF2B, GNA13, and SGK1 are all found to be highly enriched within this category [4]. Next-generation sequencing approaches have been used to uncover an infrequent, but novel, gene fusion in GCB. Cases with t(3;3)(q26.32;q28) fuses TBL1XR1 to TP63 and, interestingly, the presence of this translocation was associated with GCB cases that were primarily refractory to R-CHOP therapy [5]. The ABC subtype is recognized to have constitutive NF-kB activation on the basis of gene expression and the pattern of recurrent, somatic mutations found. Recurrent mutations in CARD11, MYD88, and CD79B are found within the ABC subtype, suggesting that constitutive signaling downstream of the B cell receptor is important for ABC biology and a logical target for more specific therapies [2]. Phase II data and emerging phase III trials have shown that new agents such as the proteosome inhibitor bortezomib, ibrutinib targeting Bruton s tyrosine kinase, fostamatinib targeting SYK, CAL-101 targeting phosphatidylinositol 3-kinase, and lenalidomide all seem to be most effective against ABC cases [2]. Base paireresolution next-generation sequencing data have also shown that histone modification and epigenetic chromatin remodeling are critical new aspects of DLBCL biology not previously appreciated. The discovery of recurrent mutations in EZH2, a member of the polycomb repressor complex-2 responsible for laying down repressive chromatin marks that silence gene expression, was found to be recurrently mutated in 22% of GCB DLBCL cases [6]. Mutations are also found in follicular lymphoma, whereas loss-of-function alterations characterize myeloid neoplasms. In follicular lymphoma and DLBCL they were clearly established as gainof-function mutations. All EZH2 mutations in DLBCL are heterozygous, requiring the presence of the wild-type allele to produce the altered phenotype. The mutant allele accelerates the addition of the second and third methyl groups to H3K27 lysine residues of core histones, but only if the wildtype enzyme adds the first methyl group [7]. Recurrent mutations were also discovered to involve additional genes involved in histone modification, including MLL2, CREBBP, EP300, MEF2B, and SETD2. In aggregate, these findings clearly implicate an unprecedented role for histone modification and chromatin remodeling in the pathogenesis of DLBCL and, importantly, highlight novel targets for therapy in selected patients harboring these alterations. Recurrent mutation in another group of genes has served to refocus attention on the importance of immune escape as an oncogenic driver in DLBCL pathobiology. Based on GEP studies, the Leukemia Lymphoma Molecular Profiling Project (LLMPP) consortium showed that non-neoplastic immune cells in DLBCL were important contributors to survival after R-CHOP therapy [8]. A gene signature implicating macrophages, connective tissue elements, and myeloid precursors (stromal-1) was associated with favorable outcome, whereas a gene signature of angiogenesis and adipocytes (stromal-2) was associated with inferior survival. The LLMPP had earlier established that tumors with decreased expression of human leukocyte antigen (HLA)-DR (major histocompatibility

3 S28 R.D. Gascoyne et al. / Biol Blood Marrow Transplant 19 (2013) S26eS32 complex class II antigen) were also associated with markedly inferior outcomes. Recurrent mutations and other genetic alterations of a number of genes, including CD58, B2M, TNFRSF14, CD273, CD274, CIITA, TNFSF9, and several HLA genes, clearly now implicate that escape from immunosurveillance is an important mechanism underlying the pathogenesis of these tumors [4,9]. Translocation of the MYC oncogene can be seen in 6% to 14% of de novo DLBCL cases and is also associated with inferior survival [10,11]. Importantly, expression of MYC protein occurs in many more cases and when seen together with BCL2 protein expression is associated with diminished survival [12,13]. Finally, next-generation sequencing techniques were recently used to identify a recurrent gene fusion in PMBCL, accounting for nearly 40% of all cases and associated with inferior survival [14]. CIITA is common to all fusions, is located on chromosome 16p, and is the master transcriptional controller of major histocompatibility complex class II expression (ie, HLA-DR). The recurrent fusions implicate CIITA as a promiscuous gene, but in 50% of the cases the fusion partner involves the ligands of PD-1 located on chromosome 9p24, PD-L1 (CD274), and PD-L2 (CD273) that produce T cell exhaustion for the infiltrating T cells in the tumor microenvironment. This novel mechanism produces an immunological double hit, whereby both partners in a gene fusion promote immune privilege by allowing the tumor cells to hide from immune attack on one hand (loss of CIITA leading to down-regulation of HLA-DR) while upregulating the ligands of PD-1 that antagonize the T cell immune response to the tumor on the other. The finding of these novel genetic mechanisms now identifies obvious candidate tumors and specific patients likely to benefit from targeted approaches using antibody-based therapies that disrupt PD-1 signaling pathways [15]. REFERENCES 1. Victora GD, Nussenzweig MC. Germinal centers. Annu Rev Immunol. 2012;30: Lenz G, Staudt LM. Aggressive lymphomas. N Engl J Med. 2010;362: Hans CP, Weisenburger DD, Greiner TC, et al. Confirmation of the molecular classification of diffuse large B-cell lymphoma by immunohistochemistry using a tissue microarray. Blood. 2004;103: Morin RD, Mendez-Lago M, Mungall AJ, et al. Frequent mutation of histone-modifying genes in non-hodgkin lymphoma. Nature. 2011; 476: Scott DW, Mungall KL, Ben-Neriah S, et al. TBL1XR1/TP63: a novel recurrent gene fusion in B-cell non-hodgkin lymphoma. Blood. 2012; 119: Morin RD, Johnson NA, Severson TM, et al. Somatic mutations altering EZH2 (Tyr641) in follicular and diffuse large B-cell lymphomas of germinal-center origin. Nat Genet. 2010;42: Yap DB, Chu J, Berg T, et al. Somatic mutations at EZH2 Y641 act dominantly through a mechanism of selectively altered PRC2 catalytic activity, to increase H3K27 trimethylation. Blood. 2011;117: Lenz G, Wright G, Dave SS, et al. Stromal gene signatures in large-b-cell lymphomas. N Engl J Med. 2008;359: Challa-Malladi M, Lieu YK, Califano O, et al. Combined genetic inactivation of beta2-microglobulin and CD58 reveals frequent escape from immune recognition in diffuse large B cell lymphoma. Cancer Cell. 2011;20: Savage KJ, Johnson NA, Ben-Neriah S, et al. MYC gene rearrangements are associated with a poor prognosis in diffuse large B-cell lymphoma patients treated with R-CHOP chemotherapy. Blood. 2009;114: Barrans S, Crouch S, Smith A, et al. Rearrangement of MYC is associated with poor prognosis in patients with diffuse large B-cell lymphoma treated in the era of rituximab. J Clin Oncol. 2010;28: Johnson NA, Slack GW, Savage KJ, et al. Concurrent expression of MYC and BCL2 in diffuse large B-cell lymphoma treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone. J Clin Oncol. 2012;30: Green TM, Young KH, Visco C, et al. Immunohistochemical double-hit score is a strong predictor of outcome in patients with diffuse large B-cell lymphoma treated with rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone. J Clin Oncol. 2012;30: Steidl C, Shah SP, Woolcock BW, et al. MHC class II transactivator CIITA is a recurrent gene fusion partner in lymphoid cancers. Nature. 2011; 471: Steidl C, Gascoyne RD. The molecular pathogenesis of primary mediastinal large B-cell lymphoma. Blood. 2011;118: STATE OF SALVAGE THERAPY FOR RELAPSED AND PRIMARY REFRACTORY DIFFUSE LARGE B CELL LYMPHOMA Craig Moskowitz The CORAL (Collaborative Trial in Relapsed Aggressive Lymphoma) intergroup trial compared rituximab, ifosfamide, etoposide, and carboplatin (R-ICE) with rituximab, dexamethasone, cytarabine, and cisplatin (R-DHAP) for patients with relapsed and primary refractory diffuse large B cell lymphoma (DLBCL). The overall response rate was 63%; 38% of patients achieved complete response as determined by CT criteria. There was no difference between the response rates in the R-ICE (63.5%; confidence interval: 56% to 70%) and R-DHAP (62.8%; confidence interval: 55% to 69%) groups or in the mobilization-adjusted response rates (52% versus 54%, respectively) [1,2]. Prior exposure to rituximab predicted for response rate (51% in rituximab treated versus 83% in rituximab naive patients) in the CORAL study. It is interesting that a response rate of 83% confirms the original report of R-ICE in rituximabnaive patients [3]. However, because all patients in the United States with DLBCL receive rituximab as part of primary therapy, a benchmark of 51% should be considered the starting point for any new salvage program; however, other prognostic factors can improve or more likely worsen these numbers. Remission duration of <1 year and a second-line ageadjusted international prognostic index (IPI) >1 also had a negative impact on response rate, 46% versus 88% for remission duration and 52% versus 71% for second-line IPI [4]. One can imagine a number of scenarios where salvage or second-line therapy followed by High-Dose Therapy/Autologous Stem Cell Transplant may not be the correct treatment for these patients. A simple example is a 56-year-old man, 7 months post-therapy with R-CHOP-21, who develops adenopathy above and below the diaphragm with an elevated lactate dehydrogenase. If outcome is analyzed by intent to treat, the patient has at best a 20% cure rate. Response rate to RICE or R-DHAP is approximately 50% and, if chemosensitive, a 40% chance of 3-year progression-free survival. Recently, Cuccuini et al. reported a secondary analysis from the CORAL study that can further reduce the likelihood of curability for this patient population [5]. Up to 10% of patients with DLBCL have a MYC/8q24 oncogene rearrangement; 150 patients had their pathology specimen analyzed with fluorescein in situ hybridization probes for MYC/8q24, BCL2/18q21, and BCL6/3q27. These results were then correlated to clinical characteristics and immunohistochemistry for CD10, BCL6, MUM1, FOXP1, and BCL2 expression. Twentyfour cases (16%) had a MYC rearrangement associated with either a BCL2/18q21 rearrangement (n ¼ 16) or a BCL6/3q27 rearrangement (n ¼ 4), so-called double-hit DLBCL [6].MYC þ rearrangement was associated with a germinal center B cell type (GCB) phenotype and high-risk second-line ageadjusted IPI. Outcome of patients with MYC þ DLBCL was significantly worse than MYC DLBCL, with a 4-year

4 R.D. Gascoyne et al. / Biol Blood Marrow Transplant 19 (2013) S26eS32 S29 progression-free survival of 22% versus 45% (P ¼.01) and a 4-yeay overall survival at 33% versus 62% (P ¼.02). The outcome was not influenced by type of salvage therapy. The cell of origin has also been evaluated by Thieblemont et al. in patients on the CORAL trial and GCB patients had an improved outcome with R-DHAP as long as there was no evidence of overexpression of MYC [7]. To summarize, any patient with an activated B cell type subtype or those with a GCB subtype and overexpression of MYC is considered to have pathologically unfavorable disease. At the time of workup of the relapsed/refractory DLBCL, it is evident from the CORAL trial that patients with remission duration of <1 year, second-line age-adjusted IPI of 2 or 3, activated B cell type subtype DLBCL, or MYC þ DLBCL with a GCB phenotype are unlikely to do well with standard salvage therapy and an autotransplant with BEAM (BCNU, etoposide, ara-c, melphalan)-type conditioning regimens. This is approximately 70% of patients who are transplant eligible for relapsed and refractory DLBCL. Studies evaluating immunotherapy post-transplantation, including allotransplantation, new conditioning regimens with radioimmunotherapy, and other combinations of chemotherapy for salvage treatment, are needed. A number of centers use and are studying gemcitabine or oxaliplatin-based salvage regimens; these regimens can be administered as an outpatient, which is certainly appealing. The National Cancer Institute of Canada has evaluated gemcitabine, cisplatin, and rituximab (R-GDP). The response rates are similar to R-DHAP, but it is better tolerated [8]. A large phase III trial comparing R-GDP and R-DHAP will be presented at the American Society of Hematology meetings in Encouraging results have been reported with the combination of rituximab, gemcitabine, and oxaliplatin (R-GEMOX). The French lymphoma study groups treated 46 patients with R-GEMOX and after four cycles the response rate was 83%, of which 50% of patients achieved a complete response [9]. The treatment was generally well tolerated; whether these results will hold up in a patient population that failed rituximab needs to be studied. Another interesting approach is a variation of DHAP, which substitutes oxaliplatin for cisplatin, referred to as the R-DHAX regimen. Lignon et al. treated 91 patients with the regimen of which 20% had baseline renal dysfunction and 42 patients had DLBCL. The response rate was 77% regardless of prior rituximab therapy; this program can also be administered as an outpatient [10]. Our current approach is somewhat different in that we are using an alternative anti-cd20, ofatumomab, with standard salvage chemotherapy and have recently led a multicenter phase II trial combining ofatumumab and DHAP or ICE [11] that showed promising activity with an ORR of 61%. Treatment was fairly well tolerated, with no unexpected toxicity, and stem cell mobilization was adequate. The results were promising in the subset of patients with primary refractory or early-relapsing disease, a group that did less well on the CORAL study. To see whether ofatumumab is superior to rituximab in the salvage setting, a phase III randomized trial of ofatumumab and DHAP and rituximab and DHAP is ongoing. Several novel targeted agents are currently under study, many of which are downstream of the B cell receptor, including spleen tyrosine kinase (syk) [12], which modulates and enhances the signal from the receptor. Additional targets downstream from syk are also being studied, the most exciting of which are inhibitors of Bruton s tyrosine kinase, such as ibrutinib and phosphatidylinositol 3-kinase inhibitors [13,14]. These oral agents have little single-agent toxicity, and phase II studies are being initiated to combine these inhibitors with standard salvage therapy. Finally, a number of antibody-drug or toxin conjugates are in various stages of testing. For example, one trial is combining inotuzumab ozogamicin, a humanized anti-cd22 antibody conjugated to the toxin, and calicheamicin with rituximab, gemcitabine, and oxaliplatin (R-Gem Ox) [15]. In summary, current second-line or salvage therapy is inadequate in the R-CHOP era. Newer regimens such as those described above are urgently needed. REFERENCES 1. Gisselbrecht C, Glass B, Mounier N, et al. Salvage regimens with autologous transplantation for relapsed large B-cell lymphoma in the rituximab era. J Clin Oncol. 2010;28: Gisselbrecht CSN, Mounier N, Singh Gill D, et al. Rituximab maintenance after autologous stem cell transplantation in relapsed patients with CD20þ diffuse large B-cell lymphoma (DLBCL): CORAL final analysis. J Clin Oncol. 2012;30: Kewalramani T, Zelenetz AD, Nimer SD, et al. Rituximab and ICE as second-line therapy before autologous stem cell transplantation for relapsed or primary refractory diffuse large B-cell lymphoma. Blood. 2004;103: Hamlin PA, Zelenetz AD, Kewalramani T, et al. Age-adjusted international prognostic index predicts autologous stem cell transplantation outcome for patients with relapsed or primary refractory diffuse large B-cell lymphoma. Blood. 2003;102: Cuccuini W, Briere J, Mounier N, et al. MYCþ diffuse large B-cell lymphoma is not salvaged by classical R-ICE or R-DHAP followed by BEAM plus autologous stem cell transplantation. Blood. 2012;119: Aukema SM, Siebert R, Schuuring E, et al. Double-hit B-cell lymphomas. Blood. 2011;117: Thieblemont C, Briere J, Mounier N, et al. The germinal center/activated B-cell subclassification has a prognostic impact for response to salvage therapy in relapsed/refractory diffuse large B-cell lymphoma: a bio- CORAL study. J Clin Oncol. 2011;29: Crump M, Baetz T, Couban S, et al. Gemcitabine, dexamethasone, and cisplatin in patients with recurrent or refractory aggressive histology B-cell non-hodgkin lymphoma: a phase II study by the National Cancer Institute of Canada Clinical Trials Group (NCIC-CTG). Cancer. 2004;101: El Gnaoui T, Dupuis J, Belhadj K, et al. Rituximab, gemcitabine and oxaliplatin: an effective salvage regimen for patients with relapsed or refractory B-cell lymphoma not candidates for high-dose therapy. Ann Oncol. 2007;18: Lignon J, Sibon D, Madelaine I, et al. Rituximab, dexamethasone, cytarabine, and oxaliplatin (R-DHAX) is an effective and safe salvage regimen in relapsed/refractory B-cell non-hodgkin lymphoma. Clin Lymph Myel Leuk. 2010;10: Matasar MJ, Rodriguez MA, Fennessy M, et al. A phase II study of ofatumumab in combination with ICE or DHAP chemotherapy in relapsed or refractory aggressive B-cell lymphoma prior to autologous stem cell transplantation (ASCT). Blood. 2011;118:957 [abstract]. 12. Friedberg JW, Sharman J, Sweetenham J, et al. Inhibition of Syk with fostamatinib disodium has significant clinical activity in non-hodgkin lymphoma and chronic lymphocytic leukemia. Blood. 2010;115: Lannutti BJ, Meadows SA, Herman SE, et al. CAL-101, a p110delta selective phosphatidylinositol-3-kinase inhibitor for the treatment of B-cell malignancies, inhibits PI3K signaling and cellular viability. Blood. 2011;117: Yang Y, Shaffer AL 3rd, Emre NC, et al. Exploiting synthetic lethality for the therapy of ABC diffuse large B cell lymphoma. Cancer Cell. 2012;21: Advani A, Coiffier B, Czuczman MS, et al. Safety, pharmacokinetics, and preliminary clinical activity of inotuzumab ozogamicin, a novel immunoconjugate for the treatment of B-cell non-hodgkin s lymphoma: results of a phase I study. J Clin Oncol. 2010;28: HEMATOPOIETIC STEM CELL TRANSPLANTATION CONDITIONING REGIMENS AND POST-TRANSPLANT MAINTENANCE THERAPY FOR DIFFUSE LARGE B-CELL LYMPHOMA Thomas C. Shea Since 1995, when the Parma trial reported a 46% 5-year event-free survival for patients who received salvage

5 S30 R.D. Gascoyne et al. / Biol Blood Marrow Transplant 19 (2013) S26eS32 autologous hematopoietic stem cell transplantation (HSCT) versus 12% for those receiving chemotherapy alone, the standard of care for patients with relapsed diffuse large B cell lymphoma (DLBCL) has been salvage therapy followed by autologous HSCT [1]. Since that landmark report, the addition of the anti-cd20 antibody, rituximab, has led to an approximately 20% improvement in event-free survival after initial anthracycline-based chemotherapy for these patients [2]. Despite these advances, nearly 50% of all patients with DLBCL will eventually relapse with their disease, at which point salvage therapy with autologous stem cell transplantation remains the best option for cure in this population. Hamlin and colleagues at Memorial Sloan Kettering reported that the secondary age-adjusted international prognostic index (saaipi) using stage of disease, performance status, and lactate dehydrogenase at the time of relapse were important predictors for eventual outcome [3]. More recent data have identified the value of a negative positron emission tomography scan before transplant [4], and the CORAL (Collaborative Trial in Relapsed Aggressive Lymphoma) study has shown that failure after the use of rituximab in the initial treatment, primary refractory disease, relapse within a year of initial diagnosis, or saaipi >1 were also associated with a poor outcome [5]. Three-year event-free survival rates in this trial were 21%, 20%, and 18% for those who had been treated with previous rituximab, had relapsed within 12 months of diagnosis, or those with saaipi scores of 2 to 3, respectively, versus 47%, 45%, and 40% for those without these characteristics. Only about 50% of these high-risk patients were able to achieve a complete or partial remission to salvage therapy. Forty percent of this chemotherapyresponsive population was able to achieve durable remission with transplantation, but this constituted only 20% of the entire high-risk group. Other biological factors such as the activated B cell type versus germinal center B cell type phenotypes and the presence of MYC translocations are outlined in the articles Biology of Diffuse Large B Cell Lymphomas and State of Salvage Therapy for Relapsed and Primary Refractory Diffuse Large B Cell above [6]. Clearly, new treatment strategies that include better salvage and more effective transplant regimens are needed. Autologous HSCT Conditioning Regimens Although the most common transplant regimen in use is the combination of BCNU, etoposide, cytarabine, and melphalan, or BEAM, there have been many other efforts to improve on the efficacy of this treatment program [7], but no large comparative studies have documented a superiority of one regimen over another (Table 1). A number of investigators have evaluated the busulfanecyclophosphamideevp-16 regimen, which has shown promise in phase II trials but has not been compared in a head to head fashion with the generally less toxic BEAM regimen [8]. 131 I Tositumomab radioimmunotherapy was added to the BEAM regimen in a recent prospective, randomized Clinical Trials Network trial and showed no significant improvement in overall outcome compared with BEAM plus rituximab [9]. The addition of 90 Y ibritumomab tiuxetan has shown interesting phase II data. One small phase III trial of 43 patients comparing 90 Y ibritumomab tiuxetan plus BEAM with BEAM indicated a significant overall survival advantage (91% versus 62% overall survival at 2 years) for 90 Y ibritumomab tiuxetan plus BEAM [10]. In another report from the City of Hope, 90 Y ibritumomab tiuxetan was administered along with BEAM Table 1 Methods to Improve Hematopoietic Stem Cell Transplant Outcomes for DLBCL Interventions Outcomes Autologous conditioning regimens BEAM Standard : low morbidity and 3-5% mortality [5,6] 131 I Tositumomab þ BEAM Large randomized negative study [9] 90 Y ibritumomab tiuxetan þ Phase II and a small positive BEAM randomized study [10,11] Busulfan þ cyclophosphamide þ More toxicity, no definite benefit VP-16 compared with BEAM [8] TBI-containing regimens Increased early and late toxicities; no survival benefit [12,13] Allogeneic transplant regimens Fully ablative regimens Reduced-intensity regimens Tandem auto/allo HSCT Increased transplant-related mortality outweighs benefit as initial therapy but may be effective salvage therapy [15-17] Effective as salvage after autologous HSCT [17] Promising phase I/II data for high-risk disease [14] Maintenance regimens Effective phase II salvage Ibrutinib, inotuzumab ozogamicin regimens [20,21] Phase II maintenance regimens Anti-PD1 antibody, CT-011 [22] Rituximab Overall ineffective in phase III trial [18] and compared with a total body irradiation (TBI)eVP- 16ecyclophosphamide regimen used at that institution [11]. The 90 Y ibritumomab tiuxetan-beam regimen had substantially less toxicity and superior overall outcomes compared with the TBI regimen. Although this trial suggested the potential value of adding radioimmunotherapy, the trial also demonstrated the increased early toxicities such as mucositis as well as the late toxicities including myelodysplasia, cataracts, and dry mouth observed in the TBI-treated patients [12,13]. These side effects have been noted in other large series and, in conjunction with a general lack of additional long-term benefit after TBI-based regimens, have resulted in a general reduction in the use of radiation containing regimens for autologous HSCTs for lymphoma in recent years. Allogeneic HSCT It is tempting to look at the outcomes of allogeneic HSCT regimens for patients with high-risk disease such as those identified in the CORAL study (Table 1). Although phase II studies have shown that reduced-intensity allogeneic HSCTs are generally well tolerated after failure of autologous HSCTs and can be used in conjunction with autologous HSCTs as a tandem auto/allo approach, registry data indicate that outcomes are comparable or better with a single autologous as compared with an initial allogeneic HSCT [14-17]. An alternative strategy that uses an autologous HSCT and then a non-ablative or reduced-intensity transplant at the time of subsequent relapse may be more effective. A large report from the Center for International Blood and Marrow Transplant Research (CIBMTR) compared 877 patients who had undergone an initial autologous HSCT with 79 patients who underwent allogeneic HSCT between 1995 and 2003 [15]. The allogeneic patients had higher risk features, including more advanced stage and more prior therapy, but also demonstrated a significant increase in treatment-related

6 R.D. Gascoyne et al. / Biol Blood Marrow Transplant 19 (2013) S26eS32 S31 mortality, treatment failure, overall mortality, and a comparable risk of disease progression compared with the autologous HSCT patients. Although they concluded that the results with allogeneic HSCT appeared better than might have been expected with a comparable population of patients undergoing autologous HSCT, the high treatment-related mortality and lack of clear benefit has led to the conclusion that survival after autologous HSCTs was generally better than with allogeneic HSCTs even with follow-up out to 7 years or more. The CIBMTR [16] and the European Group for Blood and Marrow Transplantation described outcomes with allogeneic HSCT for DLBCL patients who had failed a previous autologous HSCT [17]. In the European Group for Blood and Marrow Transplantation report, 101 patients were evaluated and found to have a 41.7% relapse-free and 53.8% overall survival at 3 years. Treatment-related mortality was higher in patients over the age of 45 and in those who had recurred less than 12 months after their initial transplant or in those with refractory disease. Reduced-intensity transplants were associated with a trend toward a lower mortality and a higher relapse rate but without any significant differences in progression-free or overall survival between these approaches regardless of whether this was with a related or unrelated donor. Perhaps the identification of high-risk patients as defined in the CORAL study or other biological features such as MYC translocation will identify patients who might benefit from early allogeneic HSCT as part of welldesigned clinical trials [7]. Maintenance Therapy after Autologous HSCT Despite some favorable phase II reports, the use of maintenance therapy has not been explored in a systematic fashion after autologous stem cell transplantation. In an effort to address this issue, the CORAL trial randomized patients to rituximab maintenance or no maintenance after their initial transplant [18]. Across the entire group of patients there was no advantage for the use of rituximab, but there was an improvement for women who received rituximab versus those who did not. It has been reported that this may in part be due to enhanced clearance of rituximab in men, resulting in decreased efficacy because of its shorter half-life [19]. Although other agents that have activity and modest toxicity are being considered, such as the Btk inhibitor, ibrutinib [20], the anti-cd22 immunoconjugate inotuzumab ozogamicin [21], or the anti PD-1 antibody [22] CT-011, there are no standard approaches to maintenance therapy in this setting. The Alliance for Clinical Trials in Oncology and the Clinical Trials Network are currently developing a trial to explore this question with an anticipated goal of being open in Summary A number of transplant approaches have been tried in patients with DLBCL following relapse after initial induction therapy, and there are data that a single autologous HSCT can cure 20% to 40% or of these patients, depending on their chemosensitivity and clinical and biological prognostic factors [5-11,15]. There are registry data indicating that although allografts are potentially curative after failure of an autologous HSCT, there are no data indicating that outside of a clinical trial, allogeneic HSCTs should be preferred over autologus transplants as initial transplant therapy after induction failure. Fianlly, whereas maintenance therapies are attractive after transplant for high-risk DLBCL patients, it is not yet clear that this approach will demonstrate the benefit that has been seen in other settings such as for multiple myeloma patients after treatment with lenalidomide [23]. REFERENCES 1. Philip T, Guglielmi C, Hagenbeek A, et al. Autologous bone marrow transplantation as compared with salvage chemotherapy in relapses of chemotherapy-sensitive non-hodgkin s lymphoma. N Engl J Med. 1995; 333: Coiffier B, Lepage E, Briere J, et al. CHOP chemotherapy plus rituximab compared with CHOP alone in elderly patients with diffuse large-b-cell lymphoma. N Engl J Med. 2002;346: Hamlin PA, Zelenetz AD, Kewalramani T, et al. Age-adjusted international prognostic index predicts autologous stem cell transplantation outcome for patients with relapsed or primary refractory diffuse large B-cell lymphoma. Blood. 2003;102: Alousi AM, Saliba RM, Okoroji GJ, et al. Disease staging with positron emission tomography or gallium scanning and use of rituximab predict outcome for patients with diffuse large B-cell lymphoma treated with autologous stem cell transplantation. Br J Haematol. 2008; 142: Gisselbrecht C, Glass B, Mounier N, et al. Salvage regimens with AHSCT for relapsed large B-cell lymphoma in the rituximab era. J Clin Oncol. 2010;28: Caballero MD, Rubio V, Rifon J, et al. BEAM chemotherapy followed by autologous stem cell support in lymphoma patients: analysis of efficacy, toxicity and prognostic factors. Bone Marrow Transplant. 1997;20: Cuccuini W, Josette Briere J, Mounier N, et al. MYCþ diffuse large B-cell lymphoma is not salvaged by classical R-ICE or R-DHAP followed by BEAM plus autologous stem cell transplantation. Blood. 2012;119: Kim JE, Lee DH, Yoo C, et al. 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7 S32 R.D. Gascoyne et al. / Biol Blood Marrow Transplant 19 (2013) S26eS32 Blood and Marrow Transplantation Registry. J Clin Oncol. 2011;29: Gisselbrecht C, Glass B, Laurent G, et al. Maintenance with rituximab after autologous stem cell transplantation in relapsed patients with CD20þ diffuse large B-cell lymphoma (DLBCL): CORAL final analysis. J Clin Oncol. 2011;29: abstr Carella A, de Souza C, Luminari S, et al. The prognostic role of gender in diffuse large B-cell lymphoma treated with rituximab containing regimens; a FIL/GEMOH retrospective study. Leuk Lymph Staudt L, Dunleavy K, Buggy L, et al. The Bruton s tyrosine kinase (Btk) inhibitor PCI modulates chronic active BCR signaling and induces tumor regression in relapsed/refractory ABC DLBCL. Blood. 2011;118: abstr Advani A, Coiffier B, Czuczman M, et al. Safety, pharmacokinetics, and preliminary clinical activity of inotuzumab ozogamicin, a novel immunoconjugate for the treatment of B-cell non-hodgkin - s lymphoma: results of a phase I study. J Clin Oncol. 2010;28: Gordon L, Weller E, Armand P, et al. A phase II study of CT-011, an anti- PD-1 antibody, after AUSCT in recurrent/refractory DLBCL: first analysis of progression-free-survival (PFS), overall survival (Os) and toxicity. Ann Oncol. 2011;22(Suppl: 4):63 [abstract]. 23. McCarthy P, Owzar K, Hofmeister C, et al. Lenalidomide after stem-cell transplantation for multiple myeloma. N Engl J Med. 2012;366: SUMMARY In this supplement the authors have outlined some of the important recent advances and areas for future investigation that will shape our current and future treatments of DLBCL. With a better understanding of the biology of this distinct yet heterogeneous group of diseases, we will more precisely classify the disease that each patient has and develop specific treatments to improve the results of both initial and salvage therapy. Improved classification schemas and therapeutic interventions based on specific disease biology should then lead to more effective application of both autologous and allogeneic HSCTs for these patients. It is hoped that these approaches, coupled with maintenance therapies such as those that have prolonged survival after autologous HSCT for multiple myeloma, will enhance the curability of DLBCL as well. ACKNOWLEDGMENTS Financial disclosure: The authors have nothing to disclose.