ALBERT

Description: Journal of Applied Meteorology and Climatology, Ahead of Print. 〈br/〉

Description: Atmosphere, Vol. 8, Pages 197: Evaluation of Surface Fluxes in the WRF Model: Case Study for Farmland in Rolling Terrain Atmosphere doi: 10.3390/atmos8100197 Authors: Xia Sun Heather Holmes Olabosipo Osibanjo Yun Sun Cesunica Ivey The partitioning of available energy into surface sensible and latent heat fluxes impacts the accuracy of simulated near surface temperature and humidity in numerical weather prediction models. This case study evaluates the performance of the Weather Research and Forecasting (WRF) model on the simulation of surface heat fluxes using field observations collected from a surface flux tower in Oregon, USA. Further, WRF-modeled heat flux sensitivities to North American Mesoscale (NAM) and North American Regional Reanalysis (NARR) large-scale input forcing datasets; U.S. Geological Survey (USGS) and the Moderate Resolution Imaging Spectroradiometer (MODIS) land use datasets; Pleim-Xiu (PX) and Noah land surface models (LSM); Yonsei University (YSU) and Mellor-Yamada-Janjic (MYJ) planetary boundary layer (PBL) schemes using the Noah LSM; and Asymmetric Convective Model version 2 (ACM2) PBL scheme using PX LSM are investigated. The errors for simulating 2-m temperature, 2-m humidity, and 10-m wind speed were reduced on average when using NAM compared with NARR. Simulated friction velocity had a positive bias on average, with the YSU PBL scheme producing the largest overestimation in the innermost domain (0.5 km horizontal grid resolution). The simulated surface sensible heat flux had a similar temporal behavior as the observations but with a larger magnitude. The PX LSM produced lower and more reliable sensible heat fluxes compared with the Noah LSM. However, Noah latent heat fluxes were improved with a lower RMSE compared to PX, when NARR forcing data was used. Overall, these results suggest that there is not one WRF configuration that performs best for all the simulated variables (surface heat fluxes and meteorological variables) and situations (day and night).

Description: Key Points Immune stimulation of cyclin D1 transgenic mice bearing Bim-deficient B cells induces an MCL phenotype. The induced lymphoma of EμCycD1CD19CREBimfl/fl mice highlights the collaborative roles of Bim deletion and cyclin D1 expression in MCL.

Description: Diffuse large B-cell lymphoma (DLBCL) exhibits significant biological and transcriptional heterogeneity which is conferred, in part, by pathologic modulation of lineage-specific and growth-associated master regulatory transcription factors (TF). Chromatin associated with TF binding sites is markedly enriched in histone proteins that are post-translationally modified by lysine side-chain acetylation. This mark facilitates the opening of chromatin and recruits a class of co-activators which recognize ε-acetyl lysine through a bromodomain. The sub-family of bromodomain and extra-terminal domain (BET) co-activators (BRD2, BRD3 and BRD4) are appealing, in part, because transgenic expression of BRD2 caused a DLBCL-like neoplasm in mice. We recently developed the first BET inhibitor, JQ1, and now explore the role of BET bromodomains in oncogenic transcription and assess BET family members as therapeutic targets in DLBCL. Nanomolar doses of JQ1 and 3 structurally dissimilar BET bromodomain inhibitors decreased the cellular proliferation of a broad panel of DLBCL cell lines of all transcriptionally defined types whereas the inactive enantiomer, JQ1R, had no effect. BRD2 and BRD4 depletion similarly decreased the proliferation of multiple DLBCL cell lines. We next explored the therapeutic potential of BET inhibition in two independent DLBCL xenotransplantation models, Ly1 and Toledo. In the first xenograft model, JQ1-treated mice had a prolongation of overall survival (p = 0.003). In the second model, JQ1-treated animals had significantly delayed tumor progression and decreased lymphomatous infiltration of spleen and bone marrow. To define the transcriptional pathways regulated by BET bromodomain proteins, we performed transcriptional profiling of multiple vehicle and JQ1-treated DLBCL cell lines. Following JQ1 treatment, we observed downregulation of multiple MYD88/TLR and BCR signaling pathway components and functionally validated MYC and E2F target gene sets. BET inhibition decreased MYC transcripts and protein in the DLBCL cell line panel suggesting that BET bromodomains directly modulate MYC transcription. In contrast, JQ1 treatment did not measurably alter E2F1 transcript or protein abundance suggesting a co-activator role of the BET bromodomains for E2F1. To explore the role of BET bromodomains in oncogenic E2F1 transcriptional signaling, we performed ChIPSeq experiments in Ly1 cells, using a chemical genetic approach. Rank-ordering of all transcriptionally active promoters based on H3K4me3 enrichment and RNA Pol II occupancy identifies pervasive binding and spatial colocalization of BRD4 and E2F1 to active promoter elements. We identified a JQ1-mediated transcriptional elongation defect across E2F1-bound promoters, responsible for the downregulation of E2F1 targets. As oncogenic TFs may signal to RNA Pol II through distal enhancer elements, we also characterized the genome-wide localization of BRD4 to enhancers in the Ly1 DLBCL cell line. Rank-ordering of enhancer regions by H3K27ac enrichment reveals that BRD4 binds to the vast majority of active enhancers in the Ly1 genome. Strikingly, the BRD4 load is asymmetrically distributed throughout the genome at enhancer sites with only a small subset of BRD-loaded “super enhancers (SE)”, 285/18330 (1.6%), accounting for 32% of all BRD4 enhancer binding in the cell. The POU2AF1 locus emerged as the most BRD4-overloaded enhancer in Ly1. BET inhibition reduced RNA Pol II elongation of POU2AF1, with a concomitant increase in promoter-paused RNA Pol II near the transcriptional start site. Accordingly, JQ1 treatment decreased POU2AF1 transcript abundance and protein expression and reduced the expression of a POU2AF1 target gene set. POU2AF1 depletion with independent shRNAs significantly decreased the proliferation of Ly1 and enforced POU2AF1 expression decreased the sensitivity of Ly1 cells to JQ1 treatment. Additional super enhancer-driven genes that were sensitive to JQ1 treatment include ones which promote and maintain the B-cell gene expression program and limit plasma cell differentiation. Our data suggest that BET inhibition limits the growth of DLBCLs by at least two complementary activities: a specific effect on genes that define a given cell type by high BRD4 loading at enhancers and the selective suppression of transcription at E2F- and MYC- driven target genes. + Contributed equally Disclosures: Qi: Patent for JQ1: holds patent for JQ1, holds patent for JQ1 Patents & Royalties. Young:Syros Pharmaceuticals: Consultancy, Equity Ownership, Membership on an entity’s Board of Directors or advisory committees; Enzon Pharmaceuticals: Equity Ownership, Membership on an entity’s Board of Directors or advisory committees. Bradner:Tensha Therapeutics: Equity Ownership, Scientific founder of Tensha which is translating drug-like derivatives of the JQ1 chemical probe of BET bromodomains used in this study, as cancer therpeutics. As such, the Dana-Farber Cancer Institute and Dr. Bradner have been granted minority equity. Other; Syros Pharmaceuticals: Equity Ownership, Scientific founder of Syros which is discovering Super Enhancers as a new class of gene control elements. As such, the Dana-Farber Cancer Institute and Dr. Bradner have been granted minority equity., Scientific founder of Syros which is discovering Super Enhancers as a new class of gene control elements. As such, the Dana-Farber Cancer Institute and Dr. Bradner have been granted minority equity. Other.

Description: Introduction. Classical Hodgkin Lymphomas (cHL) include small numbers of malignant Reed-Sternberg (RS) cells within an extensive but ineffective inflammatory/immune cell infiltrate. In cHL, chromosome 9p24.1 alterations increase the abundance of the PD-1 ligands, PD-L1 and PD-L2, and their further induction via JAK2-STAT signaling. PD-1 ligands engage the PD-1 receptor on T-cells and induce PD-1 signaling and T-cell exhaustion. Tumor cells expressing PD-1 ligands on their surface utilize the PD-1 pathway to evade an effective immune response. In recent pilot studies, PD-1 blockade was associated with high response rates and durable remissions in relapsed/refractory cHL. The unique composition of cHL limits its analysis with high throughput genomic assays. Therefore, the precise incidence, nature and prognostic significance of PD-L1 and PD-L2 alterations in cHL remain undefined. Herein, we utilize a recently developed fluorescence in situ hybridization (FISH) assay to characterize 9p24.1/PD-L1/PD-L2 alterations in a cohort of 108 newly diagnosed cHL patients (pts) who were uniformly treated with StanfordV (a combined modality therapy regimen) and have longterm followup. Methods. Pts were characterized as Ann Arbor early stage I/II favorable risk (ES-F), early stage unfavorable risk (bulk ≥ 10cm or ≥ .33 mediastinal dimension and/or B symptoms) (ES-U) or advanced stage III/IV (AS). ES-F pts received 8 weeks of Stanford V and 30 Gy involved field radiation (IFR); ES-U and AS pts received 12 weeks of Stanford V and 36 Gy IFR to initial sites 〉 5 cm. FISH was performed on formalin-fixed paraffin-embedded diagnostic biopsy specimens using bacterial artificial chromosome probes which covered CD274/PD-L1 (labeled with spectrum orange) and PDCD1LG2/PD-L2 (labeled with spectrum green) and a control centromeric probe (spectrum aqua-labeled CEP9, from 9p11-q11). Malignant RS cells were identified by their nuclear morphologic features and 50 RS cells/case were analyzed. Nuclei with a target:control probe ratio of at least 3:1 were defined as amplified (amp), those with a probe ratio of more than 1:1 but less than 3:1 were classified as relative copy gain, and those with a probe ratio of 1:1 but more than 2 copies of each probe were defined as polysomic for chromosome 9p. In each case, the percent and magnitude of disomy, polysomy, copy gain and amp were noted. In accordance with clinically approved diagnostic criteria, cases were classified by the highest observed level of 9p24.1 alteration. Specifically, cases with polysomy lacked copy gain or amp and cases with copy gain lacked amp. Immunohistochemical staining for PD-L1/PAX5 was performed as previously described and PD-L1 expression in PAX5 dim+ malignant RS cells and PAX5- infiltrating normal cells was assessed separately. Results. Almost all newly diagnosed cHL pts in this series had concordant alterations of the PD-L1 and PD-L2 loci; disomy was found in only 1% (1/108), polysomy in 5% (5/108), copy gain in 56% (61/108) and amp in 36% (39/108) of study pts. There was a correlation between intensity of PD-L1 protein expression and relative genetic alterations in this series. Two additional pts had translocations of PD-L1 or PD-L2 (2%, 2/108). We next assessed the association between specific types of PD-L1/PD-L2 alterations, clinical risk factors and outcome. Overall, the progression-free survival (PFS) was significantly lower for AS pts compared to ES-F/U pts (p=0.017). A model of PFS for the cHL pts by genetic alteration indicated that PFS was also significantly lower for pts with amp (p=0.02). Consistent with these findings, the incidence of 9p24.1 amp increased by clinical risk group: ES-F, 24%; ES-U, 34%; and AS, 50% (p=0.024, Kruskal-Wallis test). Therefore, we fit a full model of clinical and genetic factors including B-symptoms, bulk, stage and amp. Despite the association of amp with increased clinical risk groups, the genetic alteration further delineated PFS in the multivariate model (p=0.075). Conclusions. PD-L1/PD-L2 alterations are a defining feature of cHL with rare polysomy and more frequent copy gain and amp. There is an increased incidence of amp in pts with AS disease and a highly significant association of PD-L1/PD-L2 amp with PFS. These findings underscore the importance of genetically defined PD-1 mediated immune evasion in cHL and provide a rationale for the efficacy of PD-1 blockade in this disease. Disclosures Rodig: Perkin Elmer: Membership on an entity's Board of Directors or advisory committees; BMS: Research Funding. Shipp:BMS: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bayer: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Research Funding; Gilead: Consultancy; Merck: Membership on an entity's Board of Directors or advisory committees.

Description: Introduction. Primary testicular lymphoma (PTL) and primary central nervous system lymphoma (PCNSL) are large B-cell lymphomas (LBCL) that occur in immune privileged (IP) sites and share certain clinical and molecular features. To date, the treatment of these IP lymphomas is largely empiric and more effective targeted therapies are needed. Methods. To define actionable genetic features of IP lymphomas, we performed comprehensive genomic analyses of 21 PCNSLs and 7 PTLs and validated specific alterations in an independent cohort of 43 additional PTLs. Recurrent copy number alterations (CNAs) were detected using high-density single nucleotide polymorphism (SNP) arrays and the GISTIC algorithm and integrated with transcriptional profiles to identify candidate driver genes. Recurrent somatic mutations were identified using a combination of whole exome sequencing (WES) of paired tumor/normal samples and whole transcriptome sequencing (RNA-Seq) of the additional tumors without paired normal samples. Results. In systemic diffuse large B-cell lymphomas (DLBCLs), multiple low-frequency CNAs and associated target genes decrease p53 activity and perturb cell cycle regulation; infrequent somatic mutations of TP53 also deregulate these pathways (Cancer Cell, 2012; 22:359-372). In contrast, PCNSLs and PTLs primarily exhibit bi-allelic deletion of the upstream regulator of p53 activity and cell cycle, CDKN2A (~70% PCNSLs and ~80% of PTLs) and rarely have copy loss or somatic mutations of TP53 or CNAs of additional pathway components. The most commonly mutated genes in PCNSL and PTL, CD79B and MYD88, are also perturbed in a subset of systemic DLBCLs. However, mutations of these two genes are much more frequent in IP lymphomas (70% MYD88 and 61% CD79B of analyzed PCNSLs and PTLs) and these alterations are commonly found in the same cases (57% of cases in this series). These data indicate that concurrent oncogenic activation of the B-cell receptor (BCR) and the Toll-like receptor (TLR) signaling pathways is a characteristic feature of IP lymphomas with implications for targeted therapies. Among the IP lymphomas, ~20% of PCNSLs and ~40% PTLs exhibit 3q12.3/NFKBIZ copy gain and increased expression of the NFKBIZ protein product, IκB-ζ, an atypical IκB family member induced by TLR signaling. In our PTL series, MYD88 wild-type tumors had the highest 3q12.3/NFKBIZ copy gains, and ~90% of all analyzed PTLs had structural bases for NFκB activation via the TLR pathway. Lentiviral-mediated IκB-ζ knockdown decreased expression of the IκB-ζ target genes, IκB-α and BCL-xL, and induced apoptosis of LBCL cell lines with MYD88 L265P mutations, NFKBIZ gain or both alterations. In addition, enforced expression of NFKBIZ enhanced the growth of LBCLs with normal NFKBIZ copy numbers. Taken together, these data suggest that many IP lymphomas depend upon oncogenic MYD88/NFKBIZ signaling. Although the majority of CNAs and somatic mutations were shared by PCNSLs and PTLs, certain alterations were primarily observed in PTL. In both the initial and independent validation series, 〉 40% of PTLs exhibited copy gain of chromosome 9p24.1/CD274 (PD-L1) / PDCD1LG2 (PD-L2) and associated overexpression of the PD-1 ligands. These observations were of particular interest because 9p24.1 copy gain is a characteristic abnormality in two additional lymphoid malignancies, primary mediastinal LBCL and classical Hodgkin lymphoma, PD-1 signaling promotes tumor immune evasion and the PD-1 pathway is targetable. We also identified one PTL in which a novel translocation juxtaposed the regulatory elements of TBL1XR1 (chromosome 3) to the start codon-bearing exon 2 of PDCD1LG2 (PD-L2) (chromosome 9). This translocation, which was detected by RNA-Seq and confirmed by 5’ RACE and a newly developed split-apart FISH assay, resulted in dramatic overexpression of the PD-L2 protein. These data suggest that PTLs utilize several genetic mechanisms to deregulate the PD-1 ligands and limit anti-tumor immunity. Conclusions. Integrative and comparative genomic studies define PCNSL and PTL as related but unique lymphoid malignancies with targetable genetic alterations, and associated p53 deficiency and cell cycle deregulation, concurrent oncogenic BCR and TLR signaling and PD-1 dependent immune evasion that warrant further clinical investigation. Note: B.C. and M.G.M.R have made equal contributions to this research. Disclosures Feuerhake: Roche Pharma Research and Early Development (pRED) from 2008-2012: Employment. Freeman:Merck: on the PD-1 pathway Patents & Royalties; EMD-Serrono: on the PD-1 pathway Patents & Royalties; Boehringer-Ingelheim: on the PD-1 pathway Patents & Royalties; Amplimmune: on the PD-1 pathway Patents & Royalties; Roche: on the PD-1 pathway Patents & Royalties; Bristol-Myers-Squibb: on the PD-1 pathway Patents & Royalties; Novatis: on the PD-1 pathway, on the PD-1 pathway Patents & Royalties. Shipp:Sanofi: Research Funding; Bayer: Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers-Squibb: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Membership on an entity's Board of Directors or advisory committees; Merck: Membership on an entity's Board of Directors or advisory committees; Janssen R&D: Membership on an entity's Board of Directors or advisory committees.

Description: Introduction: High dose chemotherapy followed by autologous stem cell transplantation (ASCT) cures a subset of patients with chemosensitive relapsed or refractory (rel/ref) diffuse large B-cell lymphoma (DLBCL). Several factors associated with post-ASCT outcome have been identified, including pre-ASCT PET status, but better biomarkers are needed in order to optimally select candidates for the procedure. In other lymphoma subtypes with defining chromosomal translocations, PCR detection of pre- and post-ASCT minimal residual disease (MRD) in peripheral blood and of tumor contamination in the stem cell product is associated with inferior outcome. Until recently, MRD detection in DLBCL was limited by the rarity of detectable circulating disease using conventional techniques. The immunosequencing platform (Adaptive Biotechnologies, Corp.) is a next-generation-sequencing (NGS)-based MRD assay that detects small amounts of circulating tumor DNA (CTD) in patients with lymphoid malignancies. The assay detects CTD at diagnosis in most DLBCL patients and CTD levels track with response to induction therapy (Armand, 2013). Persistence of CTD or recurrence of CTD after completion of therapy is highly associated with DLBCL relapse (Kurtz, 2015; Roschewski, 2015). We evaluated whether CTD in autologous stem cell grafts was predictive of outcome in patients with rel/ref DLBCL undergoing ASCT. Methods: We retrospectively studied patients with rel/ref DLBCL, including transformed indolent lymphoma (TIL), who had paired archival tumor and autologous stem cell specimens and underwent ASCT at Brigham and Women's Hospital/Dana-Farber Cancer Institute from 2003-2013. Genomic tumor DNA was extracted from archival formalin-fixed paraffin-embedded (FFPE) tissue and analyzed using the NGS-based MRD assay. PCR amplification of IGH-VDJ, IGH-DJ,and IGK regions using universal consensus primers was performed followed by NGS to determine the tumor clonotype(s), defined as having a frequency 〉 5% in the tumor specimen. DNA from all available autologous peripheral blood or bone marrow stem cell specimens from each patient was amplified using universal consensus primers and sequenced to determine the level of CTD, defined as the number of lymphoma molecules per diploid genome. Results: We identified 98 eligible patients with rel/ref DLBCL/TIL. The median age was 60 (range 22-77) years; 63% were male; 65% had DLBCL, 29% had TIL, and 5% had primary mediastinal DLBCL; the median number of prior lines of therapy was 2 (range 2-5); all had received prior rituximab; 38% had primary refractory disease; 60% were in complete remission at ASCT; 96% received CBV conditioning. Median follow-up was 56 (range 19-123) months. The 4y progression-free survival (PFS) and overall survival (OS) in the entire cohort were 46% and 64%, respectively. Among 83 patients (85%) with sufficient DNA for clonotype determination, a clonotype was identified in 59 (71%). CTD data was complete in 53 patients (52 received peripheral blood stem cells (PBSC) and 1 received bone marrow). Eight patients (15%) had detectable CTD (CTD+) in the stem cell autografts (all PBSC) and 6/8 relapsed after ASCT. One CTD+ patient had early non-relapse mortality less than 1 month after ASCT and was never restaged. Seven of 8 CTD+ patients had TIL histology, 5 of whom relapsed (4 with aggressive lymphoma). The 4y PFS and OS in CTD+ v CTD- patients were 13% v 48% (p=0.01), and 38% v 67% (p=0.013), respectively [Figure 1]. In multivariable models including CTD status and pre-ASCT characteristics, CTD+ was the only factor associated with OS (HR 3.1, p=0.018), but was not significantly associated with PFS. Discussion: In patients with rel/ref DLBCL undergoing ASCT, the presence of CTD in the autologous stem cell graft is associated with inferior survival. CTD detection in the autograft may be more common in patients with TIL. In studies evaluating CTD detection in DLBCL, the plasma compartment has been more sensitive for detecting CTD than mononuclear cells. The use of concentrated cell specimens in this study may have decreased the sensitivity of the assay. Nevertheless, if the present findings are confirmed in a larger population, CTD detection may permit the identification of a subgroup of patients with a particularly poor outcome after ASCT, for whom alternative approaches could be considered. Figure 1. Overall (A) and Progression-Free (B) Survival in CTD+ vs CTD- Patients Figure 1. Overall (A) and Progression-Free (B) Survival in CTD+ vs CTD- Patients Figure 2. Figure 2. Disclosures Herrera: Sequenta, Inc.: Research Funding; Pharmacyclics: Research Funding; Genentech: Research Funding. Kong:Adaptive Biotechnologies, Corp.: Employment, Other: Stockholder. Davids:Genentech: Other: ad board; Pharmacyclics: Consultancy; Janssen: Consultancy. Rodig:Perkin Elmer: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Research Funding. Faham:Adaptive Biotechnologies Corp.: Employment, Other: Stockholder. Armand:BMS: Research Funding; Infinity: Consultancy, Research Funding; Merck: Consultancy, Research Funding; Sequenta, Inc.: Research Funding.

Description: Key Points PCNSLs and PTLs have a defining genetic signature that differs from other LBCLs and suggests rational targeted therapies. PCNSLs and PTLs frequently exhibit 9p24.1/PD-L1/PD-L2 copy number alterations and translocations, likely genetic bases of immune evasion.

Description: Background: Double-hit lymphomas (DHL) are a subset of diffuse large B-cell lymphoma (DLBCL) with concurrent chromosomal rearrangements involving the MYC and BCL2 or BCL6 genes, and are associated with dismal outcomes with standard upfront therapy. Double expressing lymphomas (DEL) are a subset of DLBCL with co-expression of MYC and BCL2 by immunohistochemistry (IHC), and also have a poor prognosis with standard therapy. While DHL status may be associated with inferior outcomes in patients with relapsed or refractory (rel/ref) disease (Cuccuini, Blood, 2012), little is known about the outcome of DHL patients with rel/ref disease who proceed to autologous stem cell transplantation (ASCT), and no study to date has examined the outcome of patients with DEL following ASCT. We evaluated the prognostic impact of DHL and DEL status in patients with rel/ref DLBCL who underwent ASCT at 2 centers. Methods: We retrospectively studied patients with rel/ref DLBCL, including transformed indolent lymphoma (TIL), who underwent ASCT at Brigham and Women's Hospital/Dana-Farber Cancer Institute (DFCI) and City of Hope (COH) between 1/2000 and 7/2013. The most recent biopsy prior to ASCT was used for testing when possible. IHC for MYC and BCL2 were performed using the Ventana (MYC: DFCI) and Leica BOND III (MYC: COH; BCL2: DFCI, COH) platforms according to standard protocols. For DEL, IHC cutoffs of ≥ 40% MYC-positive and ≥ 50% BCL2-positive cells were used (Johnson, JCO, 2012). Fluorescence in situ hybridization (FISH) for MYC was performed using LSI MYC dual-color break-apart probes (Abbott Molecular, Des Plaines, IL). MYC -rearranged cases had FISH for BCL2 and BCL6 performed using LSI BCL2 and BCL6 dual-color break-apart probes. DHL was defined as 〉 20% nuclei with break-apart signals for MYC and BCL-2 and/or BCL-6. Results: 201 patients with available archival tissue and clinical data were included. The median age was 60 (range 30-77) years; 60% were male; 26% had TIL; the median number of prior lines of therapy was 2 (range 2-5); 99% had prior rituximab; 53% had primary refractory disease or early (〈 6 mo) relapse; 60% were in CR by PET at ASCT; and conditioning regimens were: 66% CBV v 16% BEAM v 10% rituximab and/or ibritumomab tiuxetan-BEAM v 8% other. Overall, the 4y progression-free survival (PFS) and overall survival (OS) were 44% and 61%, respectively. Among 185 patients with complete IHC data, 38% were DEL. The 4y PFS and OS in patients with DEL compared to non-DEL patients were 37% v 52% (p =0.001), and 51% v 69% (p =0.005), respectively [Figure 1]. Results were similar using other reported IHC cutoffs for DEL (e.g. MYC ≥ 40%/BCL2 ≥ 70%, Green, JCO, 2012). Among 93 patients with complete FISH and IHC data available, 13% had MYC rearrangement: 4% were MYC/BCL2 DHL, 3% were MYC/BCL6 DHL, and 2% had rearrangements of all 3 loci. The 4y PFS and OS in DHL v non-DHL were 30% v 42% (p =0.042), and 40% v 57% (p =0.026), respectively. Patients with DEL (excluding DHL) and patients with DHL had similar PFS, which was inferior to non-DEL/non-DHL patients (4y PFS 35% v 30% v 45%, respectively, p =0.026) [Figure 2]. In multivariable models testing pre-ASCT variables, including PET response to salvage, DEL (HR 2.1, p=0.0002), TIL histology (HR 1.8, p=0.009), and SD/PD at ASCT (HR 2.9, p=0.025) were associated with poorer PFS, while DEL (HR 2.0, p=0.004) and SD/PD (HR 3.1, p=0.021) were associated with poorer OS. Neither MYC (≥ 40%) nor BCL2 (≥ 50%) expression alone was independently associated with PFS or OS. When analysis was restricted to the subset of patients with complete IHC and FISH data, DEL (HR 1.9, p=0.023), DHL (HR 2.4, p =0.048), and SD/PD at ASCT (HR 7.6, p =0.009) were associated with inferior PFS. No center effect was observed. Conclusions: DEL and DHL status are both associated with inferior PFS in patients with rel/ref DLBCL who undergo ASCT, regardless of remission status. Although ASCT remains a potentially curative approach, these patients should be targeted for study of pre- or post-ASCT relapse risk reduction strategies. Figure 1. (A) Overall survival and (B) Progression-Free Survival after ASCT in DEL vs non-DEL Patients Figure 1. (A) Overall survival and (B) Progression-Free Survival after ASCT in DEL vs non-DEL Patients Figure 2. Progression-Free Survival after ASCT in Patients with DEL vs DHL vs non-DEL/DHL Figure 2. Progression-Free Survival after ASCT in Patients with DEL vs DHL vs non-DEL/DHL Disclosures Herrera: Genentech: Research Funding; Pharmacyclics: Research Funding; Sequenta, Inc.: Research Funding. Budde:Atara Biotherapeutics: Consultancy; Seattle Genetics, Inc.: Research Funding; Merck: Research Funding; Ikara Inc: Patents & Royalties. Chen:Seattle Genetics, Inc.: Consultancy, Other: Travel expenses, Research Funding, Speakers Bureau; Genentech: Consultancy, Speakers Bureau; Millennium: Consultancy, Research Funding, Speakers Bureau. Davids:Pharmacyclics: Consultancy; Janssen: Consultancy; Genentech: Other: ad board. Nademanee:Seattle Genetics Inc.: Research Funding; Celgene: Consultancy; Gilead: Consultancy; Spectrum: Research Funding. Siddiqi:Pharmacyclics: Research Funding, Speakers Bureau; Janssen: Speakers Bureau. Forman:Mustang: Research Funding; Amgen: Consultancy. Rodig:Perkin Elmer: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Research Funding. Krishnan:Onyx: Speakers Bureau; Janssen: Consultancy; Millenium: Speakers Bureau; BMS: Consultancy; Jazz: Consultancy; Celgene: Consultancy, Speakers Bureau. Armand:Merck: Consultancy, Research Funding; Infinity: Consultancy, Research Funding; BMS: Research Funding; Sequenta, Inc.: Research Funding.

Description: Background: Double-hit lymphomas (DHL) - diffuse large B-cell lymphomas (DLBCL) with concurrent rearrangements of MYC and BCL2 and/or BCL6, and double-expressor lymphomas (DEL) - DLBCL with co-expression of MYC and BCL2 by immunohistochemistry (IHC), are associated with poor outcomes after standard chemoimmunotherapy. We have previously demonstrated that patients with relapsed or refractory (rel/ref) DHL and DEL have inferior outcomes after autologous stem cell transplantation (autoSCT) compared to patients with neither DEL nor DHL [Herrera et al, ASH 2015]. Although patients with DEL and DHL have inferior outcomes after chemotherapy-based treatment modalities, we hypothesized that allogeneic SCT (alloSCT) could potentially abrogate that negative prognostic impact. Data are extremely limited regarding the outcome of patients with DHL who undergo alloSCT, and no study has examined alloSCT outcomes in patients with DEL. We studied alloSCT outcomes in a multicenter cohort of rel/ref DLBCL patients and evaluated the prognostic impact of DEL and DHL status. Methods: We retrospectively studied patients with rel/ref DLBCL, transformed indolent lymphoma (TIL), or high-grade B-cell lymphoma unclassified (BCLU) who had available tumor tissue and underwent alloSCT at Dana-Farber Cancer Institute, Massachusetts General Hospital, or City of Hope between 1/2000 and 5/2014. IHC for MYC, BCL2, and BCL6 were performed. DEL was defined as MYC expression in ≥ 40% tumor cells and BCL2 expression in ≥ 50% tumor cells. FISH for MYC was performed using dual-color break-apart probes. Cases with MYC-rearrangement had FISH performed for BCL2 and BCL6 using break-apart probes. Rearrangement was defined as ≥ 10% nuclei with break-apart signals. DHL was defined as concurrent rearrangement of MYC and BCL2 and/or BCL6. Results: Tumor tissue was available in 103 patients, among whom we could obtain complete IHC and FISH data in 74. In these 74 patients, the median age was 54 years (range 24-69); 69% had DLBCL/BCLU whereas 31% had TIL; the median number of prior therapies was 4 (range 2-9); 58% had prior autoSCT; 73% were in complete or partial remission (CR/PR) at alloSCT; 77% had reduced intensity conditioning (RIC); 78% had a matched related or unrelated donor. 4y progression-free survival (PFS), overall survival (OS), cumulative incidence of relapse (CIR), and non-relapse mortality (NRM) in the overall cohort were 34%, 40%, 44% and 22%, respectively, with a median follow-up of 46 months for survivors. 47% of patients had DEL and 14% had DHL. The proportion of patients with a history of primary refractory disease was higher among DHL (60%) and DEL (52%) patients compared to nonDHL/nonDEL patients (37%), although the difference was not significant (p=0.3). Overall, there were no significant differences in clinical characteristics between patients with DHL, DEL, and nonDHL/nonDEL. Neither DEL nor DHL were significantly associated with outcome (Figure). The 4y PFS in DEL v non-DEL patients was 29% v 39% (p=0.2), 4y OS 30% v 49% (p=0.11), 4y CIR 50% v 40% (p=0.3), and 4y NRM 21% v 22% (p=1.0). The 4y PFS in DHL v non-DHL patients was 40% v 33% (p=0.6), OS 50% v 37% (p=0.4), CIR 40% v 45% (p=0.9), and NRM 20% v 22% (p=0.8). In multivariable Cox models for PFS and OS, age ≥ 55 (PFS: HR 0.4, p=0.002; OS: HR 0.4, p=0.005), refractory disease (not CR/PR) at alloSCT (PFS: HR 2.4, p=0.009; OS HR 2.6, p=0.007), and TIL (PFS HR 0.4, p=0.018; OS HR 0.4, p=0.028) were associated with PFS and OS, but DEL (PFS HR 1.2, p=0.5; OS HR 1.6, p=0.12) and DHL (PFS HR 0.8, p=0.7; OS HR 0.8, p=0.7) were not. We also constructed multivariable competing risk regression models for CIR and NRM. Age, remission status, histology, and conditioning intensity were associated with relapse, while no factor was significantly associated with NRM. Neither DEL (CIR HR 1.2, p=0.7, NRM HR 0.8, p=0.7) nor DHL (CIR HR 1.1, p=0.9, NRM HR 0.8, p=0.8) were associated with either outcome in those models. Conclusions: AlloSCT produced durable remissions in heavily treated rel/ref DLBCL patients, regardless of DEL and DHL status. In our cohort, DEL and DHL status did not have a significant prognostic impact. Although patients with DEL or DHL have poorer outcomes after chemoimmunotherapy and autoSCT, our results suggest that alloSCT may overcome the chemoresistance of double-hit/double-expressor tumors. Figure Progression-Free Survival After AlloSCT in DEL, DHL, and nonDEL/nonDHL Patients Figure. Progression-Free Survival After AlloSCT in DEL, DHL, and nonDEL/nonDHL Patients Disclosures Herrera: Adaptive Biotechnologies: Research Funding; Genentech: Research Funding; Immune Design: Research Funding; Merck: Research Funding; Pharmacyclics: Research Funding; Seattle Genetics: Research Funding. Song:Seattle Genetics: Consultancy. Chen:Genentech: Consultancy, Speakers Bureau; Millenium: Consultancy, Research Funding, Speakers Bureau; Seattle Genetics: Consultancy, Honoraria, Research Funding, Speakers Bureau; Merck: Consultancy, Research Funding. Chen:Incyte Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Research Funding. Koreth:LLS: Research Funding; amgen inc: Consultancy; takeda pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; kadmon corp: Membership on an entity's Board of Directors or advisory committees; prometheus labs inc: Research Funding; millennium pharmaceuticals: Research Funding. Pillai:Trillium Therapeutics: Research Funding. Siddiqi:Janssen Biotech: Research Funding, Speakers Bureau; Seattle Genetics: Speakers Bureau; Juno Therapeutics: Research Funding; Kite Pharma: Research Funding; Acerta Pharma: Research Funding; MedImmune: Research Funding; Genentech: Research Funding; TG Therapeutics: Research Funding. Zain:Seattle Genetics: Consultancy, Speakers Bureau; Celgene: Consultancy, Speakers Bureau. Kwak:XEME BioPharma: Consultancy, Equity Ownership; Antigenics: Equity Ownership; Celltrion: Consultancy; Sella Life Sciences: Consultancy. Nademanee:Celgene: Consultancy; Seattle Genetics: Consultancy, Research Funding. Weinstock:Novartis: Consultancy, Research Funding. Soiffer:Kiadis: Membership on an entity's Board of Directors or advisory committees; Juno: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees. Rodig:Bristol-Myers Squibb: Honoraria, Research Funding; Perkin Elmer: Membership on an entity's Board of Directors or advisory committees. Armand:Roche: Research Funding; Pfizer: Research Funding; Sequenta Inc: Research Funding; Merck: Consultancy, Research Funding; Infinity Pharmaceuticals: Consultancy; Bristol-Myers Squibb: Consultancy, Research Funding.