ALBERT

Description: Background: Peripheral T-cell lymphoma (PTCL) consists of an uncommon and heterogeneous group of lymphomas that are often challenging to diagnose and classify. Since most patients also have a poor survival with standard multiagent chemotherapy, more effective therapeutic approaches are needed to improve patient outcome. Table1: Pathological diagnosis Number of cases profiled AITL 36 ALK(+)ALCL 19 ALK (−)ALCL 08 ATLL 12 T/NK 14 PTCLU 44 Other rare entities 10 Methods: A mRNA profiling study using Affymetrix HGU133+2 arrays on 143 cases of PTCL and NK-cell lymphoma (NKCL) from the International Peripheral T-cell Lymphoma Project, was conducted on pre-treatment biopsies. These included the following pathologically classified cases (Table 1). In addition, we also profiled nine NK cell lines, seven T cell lines, normal resting and activated CD4+ and CD8+ T cells and resting and IL2- activated NK cells from healthy individuals. BRB-ArrayTools was used to develop gene classifiers for the major PTCL entities and survival predictors for AITL based on gene expression data. Results: We have identified key molecular signatures for PTCL and NKCL that have allowed us to construct a robust classifier for AITL (207 transcripts), ALK+ ALCL (94), ATLL (225) and NKCL (127). PTCL-U group may have 3 or 4 molecular subgroups and additional studies with more cases, are necessary to further define this group. Misclassified cases were identified and re-assigned to the molecularly defined entities, including re-assigning of 9/44 PTCL-U to AITL. We have confirmed the enriched expression of genes identified in follicular helper T-cells in AITL, suggesting that AITL is derived from this T-cell subset. A number of oncogenic pathways (e.g. NF-κB, HIF-a,VEGF, IL6) and tumor/host interactions that contributed to local tumor-induced immunosuppression (e.g. TGF-b), were identified in AITL. A molecular predictor of outcome was developed for AITL and validated by leave one-out-cross validation. Since PTCL is an uncommon disease, future studies will require the collaboration of multiple large clinical groups with tissue resources for both discovery and validation. Conclusion: This study has demonstrated that GEP will allow the construction of robust and biologically-meaningful classifiers for PTCL, and prognosticators can be derived for well-defined entities with a sufficient number of cases. GEP will also allow us to identify therapeutically-relevant oncogenic pathways and tumor/host interactions that may lead to improvement in the therapy and outcome of patients with PTCL and NKCL. (This study is a part of the International T-cell Lymphoma Project)

Description: Abstract 2648 Follicular lymphoma (FL) is an indolent lymphoma and the second most common type of non-Hodgkin lymphoma in the Western world. It is characterized by the t(14;18) chromosomal translocation, which is present in up to 90% of cases. About 40% of FL cases eventually transform into a more aggressive lymphoma (tFL), most commonly diffuse large B-cell lymphoma (DLBCL). To identify the secondary chromosomal abnormalities that contribute to the development of FL, and to its transformation, we undertook a large study using the Affymetrix 250k NspI SNP array to identify copy number abnormalities (CNAs) in 198 FL and 79 tFL samples, 75% of which have concurrent gene expression profiling studies using Affymetrix U133+2 or A+B arrays for correlative analysis. There were 22 recurrent chromosomal abnormalities that were present in over 10% of FL cases, including gains of 7, 12, 18, 21, X, 1q, 2q, 5p, 6p, 8q, 17q and loss of 6q. We also identified 20 smaller CNAs that occurred in over 5% of FL cases, the most frequent being loss of chromosome 1p36.33-p36.31 including TNFRSF14, loss of chromosome 10q23.1-q25.1 encompassing several possible cancer-related genes such as PTEN, gain of chromosome 2p16.1-p15 including REL, and gain of chromosome 8q24.13-q24.3 including MYC. Univariate Cox regression models were used to analyze the CNA regions that occurred in at least 10 FL cases as predictors of overall survival. Four recurrent CNAs were predictive of survival in univariate analysis below the p=0.05 significance level, and two were found to be borderline significant. A gain of X or the p arm of X was predictive of poor survival. Additionally, two losses on 6q (6q13–15 and 6q23.3–24.1) were associated with poor survival. The 6q23.3–24.1 loss contains TNFAIP3, which encodes a negative regulator of the NF-kB pathway, and is a frequent site of homozygous loss. Additionally, a gain of chromosome 8 that includes the MYC gene, and a loss of chromosome 9 that includes CDKN2A, were borderline predictors of poor survival. Patients with FLs that have 7 or more abnormalities had worse survival than those with fewer abnormalities. We also compared the CNAs found in tFL samples to FL samples and identified 26 abnormalities that were at least 5 times more frequent in tFL and present in at least 5% of tFLs. A gain of 3q27.3-q28 containing 5 genes including BCL6 and LPP, for example, was found in 11% of tFL case, but only 2% of FL cases. We also found differences in the deletion of Beta-2 Microglobulin (B2M) between FL and tFL. The B2M locus is deleted in 8% of FLs, but in 21% of tFLs. B2M, a subunit of the MHC class I molecule, is known to be repressed by mechanisms such as mutation and deletion in de novo DLBCL, as a way for the tumor to evade immune surveillance. HLA-A- B, and/or -C were deleted in 5% of FLs and almost 9% of tFLs. CD58, which plays a role in T- and NK-cell immune responses, was deleted in 3% of FLs and 11% of tFLs. Overall, 19% of FLs and 37% of tFLs had an abnormality in CD58, B2M, and/or HLA class I, indicating that evasion of immune surveillance is important in transformation to a more aggressive disease. We also compared CNAs from tFL cases to those found in de novo GCB-DLBCL cases and identified several that differed markedly between the 2 diseases, such as a gain of chromosome 21 which was present in 21% of tFL cases but only 3% of DLBCL cases. In conclusion, FL, tFL, and de novo GCB-DLBCL share common CNAs, but the prevalence of the individual lesions differ among the 3 entities. Functional validation of potential candidate genes will determine important pathways in the development and progression of FL, and identify possible targets for therapeutic intervention. Disclosures: No relevant conflicts of interest to declare.

Description: Abstract 1568 Introduction Non-Hodgkin lymphoma (NHL) represents 60% of lymphoma diagnoses in children, and NHL subtypes change considerably from childhood to adulthood.[1] Recent studies have implicated age-associated biological differences in certain NHL subtypes.[2] AYAs with cancer fall between pediatric and adult patients clinically and potentially biologically. Although environmental stressors and the psychosocial transition of adolescence likely impacts these outcomes, age-related biological differences need to be better understood. The majority of pediatric NHLs are high-grade tumors, whereas low- and intermediate-grade tumors are more common among adults.[1] Recent studies have found that young adults with NHL have poorer survival compared to children.[2] Although the incidence of AYA lymphoma has increased over the past 20 years, survival has not significantly improved, demanding a better understanding of the epidemiology and biology of lymphoma among this patient population.[3] Methods Cases were identified using the Nebraska Lymphoma Study Group database and chart review. All patients with DLBCL from 1983–2010 were identified (n = 1328), and 36 (2.7%) cases are included in this study of AYA DLBCL (age 13–30 years). The Kaplan-Meier method was used to estimate overall survival (OS) and event-free survival (EFS) distributions, and the log-rank test was used to compare survival distributions between groups. OS is defined as the time from the beginning of therapy to death or last follow-up. EFS is defined as the time from the beginning of therapy to progression, death, or last follow-up. P-values less than 0.05 are considered to be statistically significant. SAS software V 9.2 (SAS Institute Inc., Cary, NC) was used for all data analysis. The study was approved by the Institutional Review Board. Results The median age of the 36 AYA DLBCL patients was 24.2 years (range, 14.5–29.8) with a female to male ratio of 1.8:1, and 53% were primary mediastinal B-cell lymphoma. Patient characteristics are shown in table 1. Of the 36 patients, 18 have died and 18 are alive at last follow-up. Fifteen deaths were due to lymphoma, 1 treatment related, 1 unrelated to disease, and 1 of unknown cause. The 5-year EFS is 52% (95% CI 34–67%, figure 1) and OS is 58% (95% CI 49–72%). The median follow-up of patients alive at last follow-up is 8.8 years (range, 1.8 – 29). OS was not affected by gender (p=0.32), stage (p=0.43), LDH (p=0.11), B symptoms (p=0.98), or size of the largest mass at diagnosis (0.93). Nearly all patients (97%) were treated on adult chemotherapy protocols. Discussion This study presents data on 36 AYA patients with DLBCL. The 5-year EFS was 52% and OS was 58%. Pediatric patients report a 5-year EFS of 87–96% [2, 4] compared to adults which have a 5-year EFS of 44–80%.[4] In contrast to other reports, in this study, gender, elevated LDH, and advanced stage (III-IV) did not impact EFS or OS, although the comparisons may be under-powered due to the small sample size. Although pediatric patients have better outcomes compared to adults, AYA patients have worse EFS than both pediatric and adult DLBCL patients, demanding further understanding of the biology of AYA DLBCL and improved treatment strategies. Disclosures: No relevant conflicts of interest to declare.

Description: Background: Angioimmunoblastic T-cell lymphoma (AITL) is a common subtype of peripheral T-cell lymphoma (PTCL) with distinct pathological features and poor prognosis. Currently used chemotherapy is mostly unsuccessful with a 3-year overall survival of less than 30%. We and others have identified frequent mutations affecting IDH2 at arginine-172 (R172),TET2, DNMT3A and RHOA in AITL. The biochemical and functional consequences of IDH2R172 mutations in T cells have not been demonstrated. In this study, we performed targeted re-sequencing of epigenetic regulators, including IDH2, TET2 and DNMT3A in molecularly defined PTCL cases and analyzed the biochemical changes associated with IDH2R172 mutations as well as alterations in gene expression profiling (GEP), DNA methylation and histone modification that may improve our understanding of the pathogenetic mechanisms in AITL. Methods: We performed targeted re-sequencing of epigenetic regulators IDH2, TET2 and DNMT3A in AITL (n = 39) and PTCL subtypes (n = 53) with corresponding GEP. Due to lack of appropriate cell lines derived from AITL, we chose to use Jurkat T cells, a T-ALL cell line frequently used for T-cell functional studies and normal CD4+T cell to study the biochemical consequences of IDH2R172 mutations in T-cells. Liquid chromatography- tandem mass spectrometry was utilized for the detection of intracellular level of the 2-hydroxyglutarate and levels of 5-methycytosine and 5-hydroxymethycytosine in genomic DNA. Alterations of histone lysine tri-methylation were assessed by immunohistochemistry in AITL specimens and western blotting in vitro. Results: TET2 mutations appear to be the founder mutations in AITL with 82.1% (32/39) mutated cases and present as the major clone in the majority of mutant cases (75%; 24/32). TET2 mutations were also observed at a lower frequency in PTCL-NOS molecular subgroups (TBX21 (46%; 10/18); GATA3 (41%; 5/12)) and ALK negative-ALCL (33.3%, 4/12). The mutations in DNMT3A were observed at similar frequency in AITL (38.5%, 15/39) and PTCL-NOS subgroups [TBX21 (33%; 6/18); GATA3 (25%; 3/12)). However, IDH2R172 mutations were found predominantly in AITL (33.3%, 13/39), but rarely in PTCL-NOS subgroups (6.7%, 1/15 in PTCL-NOS). Remarkably, IDH2R172 mutant cases formed a unique cluster in unsupervised hierarchical clustering, and IDH2R172mutation defined a unique subset within AITL with a distinct gene expression signature. We observed that ectopic expression of IDH2R172K in the Jurkat T cell line led to a markedly increased level of intracellular 2-HG and up-regulation of the repressive histone methylation mark H3K27me3. Furthermore, a significant increase in 5-methylcytosine and corresponding decrease in 5-hydroxymethycytosine in genomic DNA was also observed in IDH2R172K transduced Jurkat and primary CD4+ T cells. Consistent with these findings, significant increase in aglobal DNA hypermethylation in proximal promoter regions and a global increase of the repressive histone mark H3K27me3 was observed in AITL harboring IDH2R172 mutants. Integrative analysis of GEP and promoter methylation identified several recurrently hypermethylated genes including negative regulators of the NF-kB pathway. Conclusion: IDH2R172 mutations define a unique subgroup of patients in angioimmunoblastic T-cell lymphoma with a distinct gene expression profile. IDH2R172 mutations are associated with global promoter hypermethylation in genomic DNA and trimethylation of H3K27 in AITL specimens. The current findings suggest that abnormal methylation associated with IDH2R172 mutations contribute to lymphomagenesis in AITL. Disclosures Fu: Nanostring: The author is a potential inventor on a patent application using Nanostring technology for the Lymph2Cx assay, which has been licensed from the NIH by Nanostring Patents & Royalties. Greiner:Nanostring: The author is a potential inventor on a patent applicaiton using Nanostring technology for a different assay, which has been licensed from the NIH by Nanostring Patents & Royalties.

Description: Although there are many recognized health benefits for the consumption of omega-3 (n-3) long-chain polyunsaturated fatty acids (LCPUFA), intake in the United States remains below recommended amounts. This analysis was designed to provide an updated assessment of fish and n-3 LCPUFA intake (eicosapentaenoic (EPA), docosahexaenoic acid (DHA), and EPA+DHA) in the United States adult population, based on education, income, and race/ethnicity, using data from the 2003-2014 National Health and Nutrition Examination Survey (NHANES) (n = 44,585). Over this survey period, participants with less education and lower income had significantly lower n-3 LCPUFA intakes and fish intakes (p 〈 0.001 for all between group comparisons). N-3 LCPUFA intake differed significantly according to ethnicity (p 〈 0.001), with the highest intake of n-3 LCPUFA and fish in individuals in the “Other” category (including Asian Americans). Supplement use increased EPA + DHA intake, but only 7.4% of individuals consistently took supplements. Overall, n-3 LCPUFA intake in this study population was low, but our findings indicate that individuals with lower educational attainment and income are at even higher risk of lower n-3 LCPUFA and fish intake.

Description: Purpose: To assess the efficacy of vincristine-laden platelets in patients with refractory thrombocytopenia. Patients and methods: Patients who received vincristine-laden platelets for refractory thrombocytopenia between 1991 and 2003 were included in this retrospective study. Vincristine (1 mg) was added to the platelets and the bag was incubated for 1 hour prior to transfusion. The following data were collected: age and gender of the patient, nature of the underlying disease causing thrombocytopenia, baseline renal and hepatic function, previous therapy for thrombocytopenia, platelet counts on days 0, 1, 3, 7, 15, 30 and 90 following vincristine-laden platelet transfusion, number of units of platelets transfused in the week prior to and the week after transfusion of vincristine-laden platelets. The rate of change in platelet count was calculated for each patient as a slope. Results: 20 patients were included in the study. The underlying disease causing thrombocytopenia was lung cancer (n=4), breast cancer following autologous hematopoietic stem cell transplantation and acute myeloid leukemia (n=3 each), myelodysplastic syndrome (n=2), acute lymphoid leukemia, chronic lymphoid leukemia, chronic myeloid leukemia, multiple myeloma, ovarian cancer, aspergillosis, cytomegalovirus infection and systemic lupus erythematosus (n=1 each). The median rate of change of platelet count was 550/μL/day (range, −1000 to 12,800/μL/day) and was significantly greater than 0 (p=0.003, Wilcoxon rank sum test). There was a significant decrease in the number of units of platelets transfused in the week after vincristine-laden platelet transfusion (median, 6; range, 0–16) as compared to the week prior to the transfusion (median, 6.5; range, 0–17). The median change in the number of units of platelets transfused was −1.5 (p=0.031, Wilcoxon rank sum test). Patients were also compared based on their underlying diagnosis to evaluate if patients who did not have bone marrow involvement by their primary disorder did better than those who did have marrow involvement. Patients who had a hematological malignancy with bone marrow involvement had a median rate of change in platelet counts post-vincristine, of 15/μL/day (range, −32 to 2900/μL/day). Patients who had other disorders had a median rate of change of 1230/μL/day (range, −100 to 12,800/μL/day). These rates were not statistically significantly different (p=0.27, Wilcoxon rank sum test). There was no difference in the change in the number of units of platelets transfused between the two groups. The change in the number of units of platelets transfused in the hematological malignancy group was −1.0 (range, −7 to 6) as compared to −4 (range, −7 to 4) in the non-hematological malignancy group (p=0.18, Wilcoxon rank sum test). Conclusions: Vincristine-laden platelet transfusion significantly increased the platelet counts and decreased the need for platelet transfusion. Thus transfusion of vincristine-laden platelets is an easy, inexpensive method to manage refractory thrombocytopenia. Figure Figure