Novak, Anne J.

Abstract Purpose: Transformation of follicular lymphoma is a critical event associated with a poor prognosis. The role of the tumor microenvironment in previous transformation studies has yielded conflicting results. Experimental Design: To define cell subtypes associated with transformation, we examined tissue specimens at diagnosis from patients with follicular lymphoma that later transformed and, using immunohistochemistry (IHC), stained for CD68, CD11c, CD21, CXCL13, FOXP3, PD1, and CD14. Cell content and the pattern of expression were evaluated. Those identified as significantly associated with time to transformation (TTT) and overall survival (OS) were further characterized by flow cytometry and multicolor IHC. Results: Of note, 58 patients were analyzed with median TTT of 4.7 years. The pattern of PD1+ and CD14+ cells rather than the quantity of cells was predictive of clinical outcomes. On multivariate analysis, including the follicular lymphoma international prognostic index score, CD14+ cells localized in the follicle were associated with a shorter TTT (HR, 3.0; P = 0.004). PD1+ cells with diffuse staining were associated with a shorter TTT (HR, 1.9; P = 0.045) and inferior OS (HR, 2.5; P = 0.012). Multicolor IHC and flow cytometry identified CD14+ cells as follicular dendritic cells (FDC), whereas PD1+ cells represented two separate populations, TFH and exhausted T cells. Conclusion: These results identify the presence of PD1+ T cells and CD14+ FDC as independent predictors of transformation in follicular lymphoma. Clin Cancer Res; 20(11); 2862–72. ©2014 AACR.

Key Points STAT5 is constitutively phosphorylated in malignant B cells obtained from patients with Waldenström's macroglobulinemia. Inhibition of STAT5 signaling significantly decreases IgM production and may be useful therapeutically for patients with high IgM levels.

Abstract Purpose: Although the International Prognostic Score (IPS) is the gold standard for risk-stratifying patients with classical Hodgkin lymphoma (cHL), these criteria do not accurately predict outcome. As cytokines are critically involved in driving cHL, we tested whether pretreatment serum cytokine levels could provide additional prognostic information. Experimental Design: Thirty cytokines were measured in pretreatment serum from 140 patients with cHL and compared with 50 nonlymphoma controls. Patients were followed for event-free survival (EFS) and overall survival (OS), and Cox proportional hazards regression models were used to assess the association of individual cytokines and the cytokine profiles with outcome via unadjusted and IPS-adjusted HR. Results: Twelve cytokines (EGF, bFGF, G-CSF, HGF, IL-6, IL-8, IL-12, IL-2R, IP-10, MIG, TNF-α, and VEGF) were significantly (P < 0.05) higher in patients with cHL than controls; elevated levels of HGF, IL-6, IL-2R, IP-10, and MIG were all associated with poorer EFS. Only interleukin-2 receptor (IL-2R; P = 0.002) and interleukin (IL)-6 (P < 0.001) were independently prognostic. Patients with increased IL-6 and IL-2R had a significantly higher risk of early relapse and death, a finding that remained significant even after IPS-based risk stratification. Although elevated IL-6 and IL-2R correlated with the IPS, soluble CD30 (sCD30), and thymus and activation-related chemokine (TARC) levels, the two-cytokine model remained independently predictive of prognosis. Conclusions: Elevated pretreatment serum cytokines are associated with increased disease relapse and inferior survival in cHL. Thus, the pretreatment cytokine profile, particularly serum levels of IL-6 and IL-2R, may be used to identify patients with cHL at high risk for early-disease relapse. Clin Cancer Res; 19(24); 6812–9. ©2013 AACR.

Abstract Background: Our genome-wide association study (GWAS) of chronic lymphocytic leukemia (CLL) identified 4 highly correlated intronic variants within the IRF8 gene that were associated with CLL. These results were further supported by a recent meta-analysis of our GWAS with two other GWAS of CLL, supporting the IRF8 gene as a strong candidate for CLL risk. Methods: To refine the genetic association of CLL risk, we conducted Sanger sequencing of IRF8 in 94 CLL cases and 96 controls. We then conducted fine mapping by genotyping 39 variants (of which 10 were identified from sequencing) in 745 CLL cases and 1,521 controls. We also assessed these associations with risk of other non-Hodgkin lymphoma (NHL) subtypes. Results: The strongest association with CLL risk was observed with a common single-nucleotide polymorphism (SNP) located within the 3′ untranslated region (UTR) of IRF8 (rs1044873, log additive OR = 0.7, P = 1.81 × 10−6). This SNP was not associated with the other NHL subtypes (all P > 0.05). Conclusions: We provide evidence that rs1044873 in the IRF8 gene accounts for the initial GWAS signal for CLL risk. This association appears to be unique to CLL with little support for association with other common NHL subtypes. Future work is needed to assess functional role of IRF8 in CLL etiology. Impact: These data provide support that a functional variant within the 3′UTR of IRF8 may be driving the GWAS signal seen on 16q24.1 for CLL risk. Cancer Epidemiol Biomarkers Prev; 22(3); 461–6. ©2013 AACR.

AbstractMany common genetic variants have been associated with non-Hodgkin lymphoma (NHL), but individual study results are often conflicting. To confirm the role of putative risk alleles in B-cell NHL etiology, we performed a validation genotyping study of 67 candidate single nucleotide polymorphisms within InterLymph, a large international consortium of NHL case-control studies. A meta-analysis was performed on data from 5633 B-cell NHL cases and 7034 controls from 8 InterLymph studies. rs3789068 in the proapoptotic BCL2L11 gene was associated with an increased risk for B-cell NHL (odds ratio = 1.21, P random = 2.21 × 10−11), with similar risk estimates for common B-cell subtypes. PRRC2A rs3132453 in the HLA complex class III region conferred a reduced risk of B-cell NHL (odds ratio = 0.68, P random = 1.07 × 10−9) and was likewise evident for common B-cell subtypes. These results are consistent with the known biology of NHL and provide insights into shared pathogenic components, including apoptosis and immune regulation, for the major B-cell lymphoma subtypes.

Abstract Background: Non-Hodgkin lymphoma (NHL) is a malignancy of lymphocytes, and there is growing evidence for a role of germline genetic variation in immune genes in NHL etiology. Methods: To identify susceptibility immune genes, we conducted a 2-stage analysis of single-nucleotide polymorphisms (SNP) from 1,253 genes using the Immune and Inflammation Panel. In Stage 1, we genotyped 7,670 SNPs in 425 NHL cases and 465 controls, and in Stage 2 we genotyped the top 768 SNPs on an additional 584 cases and 768 controls. The association of individual SNPs with NHL risk from a log-additive model was assessed using the OR and 95% confidence intervals (CI). Results: In the pooled analysis, only the TAP2 coding SNP rs241447 (minor allele frequency = 0.26; Thr655Ala) at 6p21.3 (OR = 1.34, 95% CI 1.17–1.53) achieved statistical significance after accounting for multiple testing (P = 3.1 × 10−5). The TAP2 SNP was strongly associated with follicular lymphoma (FL, OR = 1.82, 95%CI 1.46–2.26; p = 6.9 × 10−8), and was independent of other known loci (rs10484561 and rs2647012) from this region. The TAP2 SNP was also associated with diffuse large B-cell lymphoma (DLBCL, OR = 1.38, 95% CI 1.08–1.77; P = 0.011), but not chronic lymphocytic leukemia (OR = 1.08; 95% CI 0.88–1.32). Higher TAP2 expression was associated with the risk allele in both FL and DLBCL tumors. Conclusion: Genetic variation in TAP2 was associated with NHL risk overall, and FL risk in particular, and this was independent of other established loci from 6p21.3. Impact: Genetic variation in antigen presentation of HLA class I molecules may play a role in lymphomagenesis. Cancer Epidemiol Biomarkers Prev; 21(10); 1799–806. ©2012 AACR.

Summary A recent meta‐analysis of three genome‐wide association studies of chronic lymphocytic leukaemia ( CLL ) identified two common variants at the 6p21.31 locus that are associated with CLL risk. To verify and further explore the association of these variants with other non‐ H odgkin lymphoma ( NHL ) subtypes, we genotyped 1196 CLL cases, 1699 NHL cases, and 2410 controls. We found significant associations between the 6p21.31 variants and CLL risk (rs210134: P = 0·01; rs210142: P = 6·8 × 10 −3 ). These variants also showed a trend towards association with some of the other NHL subtypes. Our results validate the prior work and support specific genetic pathways for risk among NHL subtypes.

Abstract Cytokines within the tumor microenvironment play an important role in supporting the growth and survival of B-cell malignancies. One such cytokine, IL-21, promotes the growth of myeloma and Hodgkin lymphoma cells while inducing apoptosis in chronic lymphocytic leukemia. However, the biologic significance of IL-21 has not been examined in Waldenstrom macroglobulinemia (WM), a B-cell lymphoma characterized by elevated serum IgM and a lymphoplasmacytic bone marrow infiltrate. We report here on the presence of IL-21 in the bone marrow of patients with WM and have identified activated T cells as the source of this cytokine. We readily detected the IL-21 receptor on malignant WM B cells and show that IL-21 significantly increases both IgM secretion and cellular proliferation of these cells with no effect on viability. IL-21 rapidly induces phosphorylation of STAT3 in WM cells, and treatment of the WM cell line MWCL-1 with a STAT3 inhibitor abolished the IL-21–mediated increases in cellular proliferation and IgM secretion. IL-21 also increased the expression of known STAT3 targets involved in B-cell differentiation, including BLIMP-1, XBP-1, IL-6, and IL-10. Overall, our data indicate that IL-21 in the bone marrow microenvironment significantly affects the biology of WM tumor cells through a STAT3-dependent mechanism.

AbstractGenetic aberrations, including trisomies 3 and 18, and well-defined IGH translocations, have been described in marginal zone lymphomas (MZLs); however, these known genetic events are present in only a subset of cases. Here, we report the cloning of an IGH translocation partner on chromosome X, t(X;14)(p11.4;q32) that deregulates expression of an poorly characterized orphan G-protein–coupled receptor, GPR34. Elevated GPR34 gene expression was detected independent of the translocation in multiple subtypes of non-Hodgkin lymphoma and distinguished a unique molecular subtype of MZL. Increased expression of GPR34 was also detected in tissue from brain tumors and surface expression of GPR34 was detected on human MZL tumor cells and normal immune cells. Overexpression of GPR34 in lymphoma and HeLa cells resulted in phosphorylation of ERK, PKC, and CREB; induced CRE, AP1, and NF-κB–mediated gene transcription; and increased cell proliferation. In summary, these results are the first to identify a role for a GPR34 in lymphoma cell growth, provide insight into GPR34-mediated signaling, identify a genetically unique subset of MZLs that express high levels of GPR34, and suggest that MEK inhibitors may be useful for treatment of GPR34-expressing tumors.