Citations

AbstractSTAT3 regulates cell growth by up-regulating downstream targets, such as Myc. The frequency of phosphorylated STAT3 (pSTAT3) and Myc expression and their prognostic relevance is unknown within diffuse large B-cell lymphoma (DLBCL) germinal center B-cell (GCB) and non-GCB subtypes. pSTAT3 and Myc were studied by immunohistochemistry (IHC) on tumors from 40 DLBCL patients uniformly treated on a clinical trial of epratuzumab/rituximab-CHOP. A total of 35% of cases were pSTAT3-positive, and pSTAT3 positivity was more frequent in the non-GCB (P = .06) type but did not correlate with event-free survival (EFS). Myc expression was observed in 50% of cases and was more frequent in non-GCB type (P = .07). Myc-positive cases had inferior EFS in all patients, including the GCB and pSTAT3-positive cases, were more likely to express Myc (P = .06). Myc translocations involving the major breakpoint regions were found in 10% (3 of 29) of cases, and all 3 cases were GCB and had an inferior EFS (P = .09). pSTAT3, but not Myc expression, was correlated with elevated pretreatment serum cytokines, such as IL-10 (P = .05), G-CSF (P = .03), and TNF-α (P = .04). pSTAT3 IHC in DLBCL tumors has the potential to identify patients for STAT3 pathway–directed therapy; Myc IHC is a potential marker for inferior EFS in GCB patients.

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.

Abstract Diffuse Large B cell lymphoma (DLBCL) is the most common form of non-Hodgkin's lymphoma, and is associated with variable clinical presentation as well as variable response to standard chemotherapy regimens. Most significant advances in the treatment of DLBCL over the last 15 years, has been the addition of rituximab to standard chemotherapy. However, still 40% of patients relapse and succumbing to the disease. To improve clinical outcomes for DLBCL patients, it is urgent to identify novel molecular targets and stratify patients according to the expression of specific biomarkers that predict outcomes for the individual patient. Recently, it has been reported that the Signal transducer and activator of transcription 3 (STAT3) is over expressed in DLBCL human cell lines. However, no study has reported the STAT3 expression in DLBCL patient samples or its molecular subtypes (germinal centre B cell type (GCB) and Non-GCB cell type). The phosphorylation of STAT3 (pSTAT3) at its Tyr-705 residue (pSTAT3Tyr705) leads to its activation, followed by dimerization and translocation to the nucleus. Constitutively active STATs (particularly STAT3 and STAT5) contribute to the malignant phenotype in both human cancer cell lines and primary tumors. We hypothesize that constitutive STAT3 activity might be associated with drug resistance and adverse outcome in DLBCL patients. We determined whether STAT3 is constitutively active in patients with DLBCL, and whether activated STAT3 levels correlate with disease severity. Expression of total (tSTAT3) or tyrosine phosphorylated STAT3 (pSTAT3) was determined by in 40 DLBCL tumors. pSTAT3 was found in 52% (21/40) of cases with variable expression {18% low, (+); 25% moderate, (++); and 10% very high (+++)}. However 80% (32/40) of patients expressed tSTAT3. pSTAT3 was expressed in both germinal centre B cells (GCB) (46%) and Non-GCB (69%) DLBCL cases which were molecular subtyped by the Hans method. Furthermore in-vitro inhibition of STAT3 dephosphorylated STAT3 and caused an inhibitory effect on phospho-STAT3 positive DLBCL cells; there was minimal effect on phospho-STAT3 negative cells. Event free survival was lower in pSTAT3 positive compared to pSTAT3 negative DLBCL patients. In summary, our data provide the first evidence that over-expression of pSTAT3 is common in DLBCL, this expression does not clearly relate only to non-GCB tumors and therefore STAT3 inhibitors should be tested in all DLBCL. Pre-treatment assessment of pSTAT3 tumor status will be important to understand whether this IHC marker can preselect patients for response or lack of response to new JAK/STAT targeted agents such as TG101348 (Sanofi Aventis) and BMS-911543 (Bristol-Myers Squibb). Our findings suggest a potential novel therapeutic approach of targeting the inhibition of STAT3 activity in a majority of DLBCL patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2576. doi:1538-7445.AM2012-2576

Abstract Background. Several recent studies have reported an inverse relationship between sun exposure and non-Hodgkin (NHL) risk. One hypothesis is that vitamin D mediates this effect. We evaluated the association of sun exposure at various ages and common variants in vitamin D related genes with risk of NHL and its major subtypes. Methods. Newly diagnosed NHL cases (N=1023) and frequency matched controls (N=1254) were enrolled in a clinic-based case-control study conducted at the Mayo Clinic. Thirty-eight single nucleotide polymorphisms (SNPs) from 5 candidate genes relevant to vitamin D metabolism and sun sensitivity (IRF4, RXR, VDR, CYP24A1, CYP27B1) were genotyped. Participants also completed a detailed questionnaire, including self-reported sun exposure at ages 13-21, 22-40, and 41+ years. Odds ratios (OR), 95% confidence intervals (CI), and tests for trend and effect modification were estimated using unconditional logistic regression adjusted for age and gender. ORs are reported for the highest vs. lowest exposure category or as the ordinal OR for SNPs. Results. The mean age of diagnosis/enrollment was 61 years for cases and 60 years for controls. The most common subtypes were chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL; 34%), follicular (FL; 24%), and diffuse large B-cell (DLBCL; 18%) lymphomas. There was a significant decrease in NHL risk with increased sun exposure at ages 13-21 years (OR=0.66, 95% CI 0.44-1.00; ptrend=0.04), which attenuated with increasing age at exposure. Exploratory analysis by NHL subtype indicated a trend toward a protective effect of sun exposure at ages 13-21 years for each of the major subtypes, though these estimates did not reach statistical significance. Of the 38 SNPs evaluated, there was a significant association for 3 SNPs in VDR and 1 SNP in CYP24A1: rs3819545 (OR=1.24, 95% CI 1.10-1.40, p=0.0004), rs2239186 (OR=1.22, 95% CI 1.05-1.41, p=0.009), and rs886441 (OR=0.82, 95% CI 0.70-0.96, p=0.02) for VDR and rs2762939 (OR=0.85, 95% CI 0.75-0.98, p=0.02) for CYP24A1. Both the VDR and CYP24A1 associations were observed for CLL/SLL, FL, but were attenuated for DLBCL. There was no evidence of any significant effect modification of sun exposure at ages 13-21 years by genotype in these four SNPs for NHL overall, or for NHL subtypes. Conclusions. We observed an inverse association between the frequency of sun exposure at ages 13-21 years and NHL risk, which attenuated for sun exposure at older ages, suggesting that early life exposure may be more relevant. While genetic variation in VDR and CYP24A1 were associated with NHL risk, there was no evidence for an interaction of sun exposure with these genes, suggesting that a sun association may not act through the vitamin D metabolism pathway. This would be consistent with a lack of association of circulating vitamin D with NHL risk recently published from the Cohort Consortium pooling project. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1894. doi:10.1158/1538-7445.AM2011-1894

Abstract Background: NHL is a malignancy of lymphocytes, and there is growing evidence for a role of immune dysfunction in the etiology of this malignancy. We conducted a 2-stage analysis of single nucleotide polymorphisms (SNPs) from 1,253 genes using the ParAllele (now Affymetrix) Immune and Inflammation Panel with risk of NHL. Methods: In Stage 1, we successfully genotyped 7,670 SNPs in a clinic-based study of 425 NHL cases and 465 frequency matched controls seen at the Mayo Clinic. Genes were tagged with SNPs from HapMap to provide coverage at r2 = 0.8 and minor allele frequency (MAF) >0.05. In Stage 2, we genotyped the top 768 SNPs on an additional 584 cases and 768 controls from the Mayo Clinic Study, and then conducted a pooled analysis of all 1009 cases and 1233 controls. The association of individual SNPs with NHL risk was assessed using the Odds Ratios (ORs) and 95% confidence intervals, adjusted for age and gender. The most prevalent homozygous genotype was used as the reference group, and each SNP was modeled individually as having a log-additive effect. Results: Overall, the mean age at diagnosis was 62.0 years for cases and the mean age at enrollment for controls was 61.1 years. The most common subtypes were chronic lymphocytic leukemia (CLL, N=343), follicular lymphoma (FL, N=245), and diffuse large B-cell lymphoma (DLBCL, N=178). In the pooled analysis, there were 3 SNPs at p<0.0004. The top SNP was rs241447 (MAF=0.26) from TAP2 at 6p21.3 (OR=1.34, 95% CI 1.17-1.53, p=0.00003). This SNP is a cSNP that leads to a missense change (Ala→Thr). The second SNP was rs2857597 (MAF=0.26) from AIF1, also at 6p21.3 (OR=0.77, 95% CI 0.67-0.88, p=0.0002). The region spanning these two SNPs had multiple risk SNPs with p<0.01, including SNPs in BAT2 (rs3132453), BAT3 (rs2242656), C2 (rs7746553), HLA-DRA (rs8084, rs7192), and TAP2 (rs1894408). Of these SNPs, only rs241447 (TAP2) and rs7192 (HLA-DRA) displayed any substantial LD (r2=0.84, all other r2<0.25). The third SNP was rs754505 (MAF=0.07) from NFATC1 at 18q23 (OR=0.63, 95% CI 0.49-0.81, p=0.0003). There were 4 additional SNPs in this gene with a p<0.05 (rs7240932, rs396127, rs177820, rs183374). In subtype analysis, the SNPs from 6p21.3 showed the strongest associations with FL (including the TAP2 SNP rs241447, p=6.9 × 10−8), although there were also associations with CLL (except for the TAP2 and C2 SNPs) and DLBCL (except for the HLA-DRA SNPs). For NFATC1 the associations were similar across the main NHL subtypes. Conclusions: We identified strong associations with NHL risk in the 6p21.3 region, which includes the MHC locus. Our results were particularly striking for FL and supports recent GWAS and candidate gene studies that implicate this region. We also identified NFATC1 as a susceptibility gene. NFATC1 is known to regulate the expression of growth and survival genes including MYC, TNF, CD40L, and BAFF, all of which have also been linked to lymphomagenesis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2761. doi:10.1158/1538-7445.AM2011-2761

Abstract Immunoglobulin light-chain (AL) amyloidosis is a rare, incurable plasma cell disorder. Its therapy has benefited immensely from the expanding drug armamentarium available for multiple myeloma. Pomalidomide in combination with weekly dexamethasone (Pom/dex) is active among patients with relapsed myeloma. In the present study, we explored the Pom/dex combination in patients with previously treated AL. Patients were eligible for this prospective phase 2 trial if they had had at least one prior regimen and if they had reasonably preserved organ function. Patients were treated with oral Pom/dex. Thirty-three patients were enrolled. The median age was 66 years. Median time from diagnosis to on-study was 37 months. Eighty-two percent had cardiac involvement. The confirmed hematologic response rate was 48%, with a median time to response of 1.9 months. Organ improvement was documented in 5 patients. The median overall and progression-free survival rates were 28 and 14 months, respectively; the 1-year overall and progression-free survival rates were 76% and 59%, respectively. There was a discordance between the hematologic response and the N-terminal pro-brain natriuretic peptide response. The most common grade 3-5 adverse events, regardless of attribution, were neutropenia and fatigue. We conclude that pomalidomide appears to be a valuable drug covering an unmet clinical need in patients with previously treated AL. The trial is registered at www.clinicaltrials.gov as NCT00558896.

SummaryThe pathological background in nodular lymphocyte predominant Hodgkin lymphoma (NLPHL) consists of lymphocytes and histocytes. This study analysed the peripheral blood absolute lymphocyte count/absolute monocyte count ratio at diagnosis (ALC/AMC‐DX) on the impact of survival inNLPHL. One hundred and three consecutiveNLPHLpatients that were followed atMayoClinic from 1974 to 2010 were included in the study. Receiver operating characteristic and area under the curve were used forALC/AMC‐DXcut‐off value analysis and proportional‐hazards models were used to compare survival based on theALC/AMC‐DXratio. With a median follow‐up of 8·9 years (range: 0·3–31 years), anALC/AMC‐DX≥2·1 was the best cut‐off value for survival with an area under the curve of 0·82, a sensitivity of 70% and specificity of 84%. After adjusting for theInternationalPrognosticScore (IPS),ALC/AMC‐DXremained an independent prognostic factor for overall survival [HazardRatio (HR), 0·33, 95% confidence interval (CI), 0·15–0·71%,P < 0·004]; lymphoma‐specific survival (HR, 0·05; 95%CI, 0·01–0·68%,P < 0·002); progression‐free survival (HR, 0·30; 95%CI, 0·14–0·60%,P < 0·006), and time to progression (HR, 0·06, 95%CI, 0·04–0·30%,P < 0·004).ALC/AMC‐DXis a low cost, already standarized, biomarker to predict clinical outcomes inNLPHL.

AbstractCytokines are deregulated in cancers and can contribute to tumor growth. In patients with diffuse large-cell lymphoma (DLBCL), we observed higher levels of JAK/STAT pathway-related serum cytokines (ie, IL-6, IL-10, epidermal growth factor, and IL-2) compared with controls. Of these, only IL-10 activated the JAK2 pathway in lymphoma cells in vitro. Patients with high serum IL-10 had shorter event-free survival (EFS) than patients with low levels (P > .01) and high IL-10 was correlated with high lactase dehydrogenase (P = .0085) and higher International Prognostic Index scores (P = .01). To explore the mechanism by which IL-10 may contribute to an inferior EFS, we investigated the effect of IL-10 on the JAK2 pathway and found that the IL-10/IL-10 receptor complex up-regulated JAK2 signaling. Neutralizing Ab to IL-10 inhibited constitutive and IL-10–induced JAK2/STAT3 phosphorylation. JAK2 inhibition dephosphorylated JAK2 and STAT3 and caused an inhibitory effect on phospho-JAK2–positive DLBCL cells; there was a minimal effect on phospho-JAK2–negative cells. Apoptosis induced by JAK2 inhibition was dependent on inhibition of autocrine IL-10 and c-myc expression and independent of Bcl-2 family expression. These results provide the rationale for testing JAK2 inhibitors in DLBCL patients, and indicate that serum IL-10 may be a biomarker to identify patients more likely to respond to JAK2-targeted therapy.

Abstract The mammalian target of rapamycin (mTOR) plays crucial roles in proliferative and antiapoptotic signaling in lymphoid malignancies. Rapamycin analogs, which are allosteric mTOR complex 1 (mTORC1) inhibitors, are active in mantle cell lymphoma and other lymphoid neoplasms, but responses are usually partial and short-lived. In the present study we compared the effects of rapamycin with the dual mTORC1/mTORC2 inhibitor OSI-027 in cell lines and clinical samples representing divers lymphoid malignancies. In contrast to rapamycin, OSI-027 markedly diminished proliferation and induced apoptosis in a variety of lymphoid cell lines and clinical samples, including specimens of B-cell acute lymphocytic leukemia (ALL), mantle cell lymphoma, marginal zone lymphoma and Sezary syndrome. Additional analysis demonstrated that OSI-027–induced apoptosis depended on transcriptional activation of the PUMA and BIM genes. Overexpression of Bcl-2, which neutralizes Puma and Bim, or loss of procaspase 9 diminished OSI-027–induced apoptosis in vitro. Moreover, OSI-027 inhibited phosphorylation of mTORC1 and mTORC2 substrates, up-regulated Puma, and induced regressions in Jeko xenografts. Collectively, these results not only identify a pathway that is critical for the cytotoxicity of dual mTORC1/mTORC2 inhibitors, but also suggest that simultaneously targeting mTORC1 and mTORC2 might be an effective anti-lymphoma strategy in vivo.