Functional coexpression of Interleukin (IL)‐7 and its receptor (IL‐7R) on Hodgkin and Reed‐Sternberg cells: Involvement of IL‐7 in tumor cell growth and microenvironmental interactions of Hodgkin's lymphoma

The clinical and pathological features of classical Hodgkin lymphoma (cHL) mirror an abnormal tissue and systemic immune response due to the production of a variety of cytokines and chemokines by the malignant Hodgkin‐Reed‐Sternberg (H‐RS) cells and/or surrounding reactive cells. Here, we demonstrate that HL‐derived cell lines (L‐428, KM‐H2, HDLM‐2, L‐1236 and L‐540) and primary H‐RS cells from lymph node tissues of HL patients express the IL‐7(R) receptor. IL‐7 appears to be involved in autocrine circuitries of HL because L‐1236, HDLM‐2 and KM‐H2 cells display the constitutive production of IL‐7 and neutralizing anti‐IL‐7 antibodies induces a statistically significant inhibition of their basal proliferation. In addition, IL‐7, either exogenous or fibroblasts‐derived, promotes the clonogenic growth and reduces apoptosis of cultured H‐RS cells, being also able to partially protect these cells from the cytotoxic effects of doxorubicin. We also provide evidence that IL‐7 stimulates IL‐6 secretion from IL‐7R‐expressing fibroblasts from HL‐involved lymph nodes (HLFs), and that a striking increase in IL‐6 secretion can be observed in cocultures of HLFs with L1236 cells. Finally, we show that L‐1236 cells‐derived IL‐7 represents a costimulator for proliferation of purified CD4+CD25+CD127^dim/? regulatory T cells (Tregs). Taken together, our data indicates that the IL‐7/IL‐7R axis constitutes an additional signaling pathway between H‐RS cells and their reactive cellular background, thereby affecting proliferation and survival of tumor cells, acting as a cofactor for Tregs expansion and enhancing the microenviromental production of IL‐6, a cytokine associated with the presence of “B” symptoms and a poor outcome in HL patients. © 2009 UICC     

Interleukin‐13 and its signaling pathway is associated with obesity‐related colorectal tumorigenesis

The incidence of colorectal cancer (CRC) has been on the rise, which is linked to the increasing prevalence of obesity, based on global epidemiological evidence. Although chronic inflammation is implicated in tumor development, the mechanisms underlying obesity‐associated CRC remain unknown. Here, we sought to identify the inflammatory cytokines and their roles in obesity‐related colorectal tumorigenesis using cytokine array analyses in a mouse model. Colorectal tumorigenesis was induced through i.p. injection of azoxymethane once a week for 6 weeks in 6‐week‐old female WT C57Black/6J mice and the obesity diabetes model mouse KK/TaJcl, KK‐Ay/TaJcl. The formation of aberrant crypt foci and colorectal tumors were more frequent in obese mice compared with WT mice, and both serum interleukin (IL)‐13 and IL‐13 receptor (R) expression in the normal intestinal mucosal epithelium were significantly increased in the obese mice. Furthermore, addition of IL‐13 to a human CRC cell line and a human colon organoid culture altered the phenotype of intestinal epithelial cells. Knockdown experiments further revealed that IL‐13Rα1 dominantly induced mucosal proliferation. Collectively, These results suggest an association between anti‐inflammatory cytokines and colorectal carcinogenesis, and provide new research directions for cancer prevention strategies. In particular, inflammation provoked by obesity, notably by increased expression of the cytokine IL‐13, could play an important role in the carcinogenesis of obesity‐related CRC.     

Targeting IL‐13Rα2 in human pancreatic ductal adenocarcinoma with combination therapy of IL‐13‐PE and gemcitabine

Pancreatic cancer is an aggressive disease with only limited therapeutic options available. We have identified that 71% pancreatic ductal adenocarcinoma (PDA) express high levels of IL‐13Rα2, a high‐affinity receptor for IL‐13. To target IL‐13Rα2, we have developed a recombinant immunotoxin, which is a fusion of IL‐13 and Pseudomonas exotoxin (IL‐13‐PE). Since IL‐13‐PE and a commonly used cytotoxic drug gemcitabine act by a different mechanism, we hypothesized that they synergize in mediating antitumor response. Both IL‐13‐PE and gemcitabine‐mediated cytotoxicity to two pancreatic cancer cell lines and when combined synergistic cytotoxicity was observed. This synergism was also demonstrated in vivo in an orthotopic mouse model of human PDA. IL‐13‐PE and gemcitabine showed complete eradiation of tumors as assessed by whole body imaging of GFP‐transfected tumors in 57% of mice in an early cancer model resulting into prolongation of survival. In contrast, monotherapy with either agent did not produce complete eradiation, but tumor volumes were significantly decreased. In advanced PDA model, combination therapy also produced dramatic reduction in tumor growth and enhanced survival compared to animals treated with either agent alone. When IL‐13Rα2 was knocked‐down by RNAi prior to tumor implantation, IL‐13‐PE and gemcitabine did not synergize indicating that IL‐13Rα2 is essential. Mechanistically, gemcitabine increased IL‐13Rα2 expression in vitro and in vivo, which resulted in a synergism of combination therapy. Interestingly, PDA cancer stem cells were resistant to gemcitabine, but not to IL‐13‐PE. These results suggest that combination therapy with IL‐13‐PE and gemcitabine may be a useful strategy for PDA therapy.     

Cardiovascular mortality among patients with non‐Hodgkin lymphoma: Differences according to lymphoma subtype

Survival rates of patients with non‐Hodgkin lymphoma (NHL) have improved over the last decade. However, cardiotoxicities remain important adverse consequences of treatment with chemotherapy and radiation, although the burden of cardiovascular mortality (CVM) in such patients remains unknown. We conducted a retrospective cohort study of patients greater than or equal to 20 years of age diagnosed with diffuse large B‐cell lymphoma (DLBCL), follicular lymphoma (FL), and chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) between 2000 and 2013 using data extracted from the United States Surveillance, Epidemiology, and End Results (SEER) database. Our primary endpoint was CVM. The association between NHL and CVM was evaluated using multivariable Cox regression analysis after adjusting for other patient characteristics. We calculated standardized mortality ratios (SMRs) for CVM, comparing NHL patients with the general population. We identified 153 983 patients who met the inclusion criteria (69 329 with DLBCL, 48 650 with CLL/SLL, and 36 004 with FL). The median follow‐up was 37 months (interquartile range, 10‐78 months); the mean patient age was 66.24 (±14.69) years; 84 924 (55.2%) were men; 134 720 (87.5%) were White, and 131 912 (85.7%) did not receive radiation therapy. Overall, 9017 patients (5.8%) died from cardiovascular disease, and we found that NHL patients had a higher risk of CVM than the general population, after adjusting for age (SMR 15.2, 95% confidence interval: 14.89‐15.52). The rates of CVM were 5.1%, 8%, and 4.4% in patients with DLBCL, CLL/SLL, and FL, respectively. Furthermore, across all NHL subtypes, older age, higher stage at the time of diagnosis (particularly stage 4), male sex, and living in the south were associated with higher risks of CVM. Our data suggest that risk assessment and careful cardiac monitoring are recommended for NHL patients, particularly those with the CLL/SLL subtypes.     

Vasoactive intestinal peptide increases apoptosis of hepatocellular carcinoma by inhibiting the cAMP/Bcl‐xL pathway

Vasoactive intestinal peptide (VIP) is a modulator of inflammatory responses. VIP receptors are expressed in several tumor types, such as colorectal carcinoma. The study described herein was conducted to confirm the presence of VIP and its receptors (VPAC1 and VPAC2) in surgically resected hepatocellular carcinoma (HCC) tissues and in the HCC cell line Huh7. The mechanism responsible for apoptosis of HCC cells was then examined because VIP treatment (10^?10 M) significantly suppressed proliferation of Huh7 cells. In examining apoptosis‐related proteins, we found caspase‐3 to be significantly increased and Bcl‐xL and cyclic AMP (cAMP) response element‐binding protein (CREB) to be significantly decreased in Huh7 cells cultured with VIP. Furthermore, the CREB level and phosphorylation were reduced. These effects were reversed by the addition of VIP receptor antagonist or cAMP antagonist Rp‐cAMPS. Pretreatment with cAMP analogue blocked the increased apoptosis, suggesting that VIP induces apoptosis via a PKA‐independent signaling mechanism. Our data indicate that VIP prevents the progression of HCC by apoptosis through the cAMP/Bcl‐xL pathway.     

阻断STAT3信号通路对恶性淋巴瘤细胞凋亡的影响

目的探讨阻断信号转导与转录激活因子3(STAT3)信号通路对恶性淋巴瘤细胞凋亡的影响。方法用0、200、400、800、1600 nmol/L的STAT3信号通路特异性阻断剂JSI-124作用于恶性淋巴瘤细胞株Raji,噻唑蓝(MTT)法检测细胞增殖水平,计算半数抑制浓度。用半数抑制浓度的JSI-124作用于Raji细胞,流式细胞术检测细胞凋亡和细胞周期,Western blot法检测细胞周期蛋白D1(cyclin D1)、STAT3、磷酸化的STAT3(p-STAT3)、活化的半胱氨酸天冬氨酸蛋白水解酶3(cleaved caspase 3)的蛋白相对表达水平。结果随着JSI-124作用浓度的升高,细胞存活率逐渐下降。计算半数抑制浓度为(615.62±52.98)nmol/L,故后续选用600 nm/L的JSI-124作用于恶性淋巴瘤细胞。600 nm/L作用组细胞凋亡率明显高于0 nm/L作用组(P﹤0.01)。600 nm/L作用组G0/G1期细胞所占比例明显低于0 nm/L作用组,G2/M期细胞所占比例明显高于0 nm/L作用组,差异均有统计学意义(P﹤0.01);两组S期细胞所占比例比较,差异无统计学意义(P﹥0.05)。600 nm/L作用组p-STAT3、cyclin D1蛋白相对表达水平明显低于0 nm/L作用组,cleaved caspase 3蛋白相对表达水平明显高于0 nm/L作用组,差异均有统计学意义(P﹤0.01)。结论阻断STAT3信号通路可促进恶性淋巴瘤细胞凋亡,抑制细胞增殖,将细胞周期阻滞在G2/M期。     

Use of non‐steroidal anti‐inflammatory drugs and risk of non‐Hodgkin lymphoma: a systematic review and meta‐analysis

Epidemiological study findings regarding the association between use of non‐steroidal anti‐inflammatory drugs (NSAIDs) and risk of non‐Hodgkin lymphoma (NHL) have been inconsistent. We aimed to systematically review epidemiological studies of the association and calculate pooled relative risks using meta‐analytic methods. We searched eight electronic literature databases and three clinical trial registers to identify all studies (including observational studies and randomized clinical trials) of the association published prior to October 2013. Identified studies were independently reviewed by two researchers. We used a random effects model to calculate pooled odds ratio (PORs). Heterogeneity amongst studies was examined using Cochran's Q and I‐squared (I²) tests; and sources of heterogeneity were explored using subgroup and meta‐regression analyses. A total of 17 studies (12 case‐control studies and five cohort studies), all adult studies, were included. Use of NSAIDs was not associated with overall risk of NHL [POR = 1.05, and 95% confidence interval (95% CI) 0.90–1.22] or NHL subtypes including B‐cell lymphoma, T‐cell lymphoma, follicular lymphoma, diffuse large B‐cell lymphoma and chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL). Aspirin use was associated with reduced risk of CLL/SLL (POR = 0.70, 95% CI 0.54–0.91) but not with the risk of all NHLs (POR = 1.02, 95% CI 0.89–1.17). Use of non‐aspirin NSAIDs was associated with increased risk of NHL (POR = 1.41, 95% CI 1.01–1.97) amongst females only. The epidemiologic evidence remains inconclusive. Effects of NSAIDs may differ by drug type, NHL subtype, and sex and more studies taking into consideration these differences are needed. Copyright © 2014 John Wiley & Sons, Ltd.     

Down‐regulation of microRNA‐9 leads to activation of IL‐6/Jak/STAT3 pathway through directly targeting IL‐6 in HeLa cell

MicroRNA‐9 (miR‐9) presents to exert distinct and even opposite functions in different kinds of tumors through targeting different cellular genes. However, its role in cervical adenocarcinoma remains uncertain. Here, we report that miR‐9 is down‐regulated in cervical adenocarcinoma due to its frequent promoter‐hypermethylation and exerts its tumor suppressor role through inhibiting several novel target genes, including interleukin‐6 (IL‐6). The promoters of miR‐9 precursors (mir‐9‐1, ‐2, and ‐3) were hypermethylated in cervical adenocarcinoma tissues. Demethylation treatment of HeLa dramatically increased the expression of mature miR‐9. Both in vitro and in vivo functional experiments confirmed that miR‐9 can inhibit the proliferation, migration, and malignant transformation abilities of HeLa cells. Bioinformatics methods and array‐based RNA expression profiles were used to screen the downstream target genes of miR‐9. Dual‐luciferase reporting assay, real‐time qPCR, and ELISA or Western blot confirmed four genes (CKAP2, HSPC159, IL‐6, and TC10) to be novel direct target genes of miR‐9. Pathway annotation analysis of the differently expressed genes (DEGs) induced by ectopic miR‐9 expression revealed the enrichment in Jak/STAT3 pathway, which is one of the downstream pathways of IL‐6. Ectopic expression of miR‐9 in HeLa inhibited Jak/STAT3 signaling activity. Moreover, such effect could be partially reversed by the addition of exogenous IL‐6. In conclusion, our results here present a tumor suppressor potential of miR‐9 in cervical adenocarcinoma for the first time and suggest that miR‐9 could repress tumorigenesis through inhibiting the activity of IL‐6/Jak/STAT3 pathway. © 2015 The Authors. Molecular Carcinogenesis, published by Wiley Periodicals, Inc.     

Periodontal disease and risk of non‐Hodgkin lymphoma in the Health Professionals Follow‐Up Study

Periodontal disease is a chronic inflammatory condition that has been associated with chronic diseases, including cancer. In an earlier prospective cohort analysis within the Health Professionals Follow‐Up Study (HPFS), we observed a 31% higher risk of non‐Hodgkin lymphoma (NHL) among participants with severe periodontal disease at baseline. Here, we extend the study with an additional 8 years of follow‐up, and conduct analyses with updated periodontal disease status and NHL subtypes. The HPFS is an ongoing prospective cohort study of 51,529 men in the USA Between baseline in 1986 and 2012, 875 cases of NHL were diagnosed, including 290 chronic lymphocytic leukemia/small lymphocytic lymphomas (CLL/SLL), 85 diffuse large B‐cell lymphomas and 91 follicular lymphomas. We performed multivariable Cox proportional hazards regression to evaluate associations of interest. History of periodontal disease at baseline was positively associated with risk of NHL overall (hazard ratio (HR) = 1.26, 95% confidence interval (CI): 1.06–1.49) and CLL/SLL (HR = 1.41, 95% CI: 1.04–1.90). With updated periodontal status, HRs were 1.30 (95% CI: 1.11–1.51) for NHL overall and 1.41 (95% CI: 1.08–1.84) for CLL/SLL. In contrast, after adjusting for periodontal disease, tooth loss was inversely associated with NHL, suggesting that other causes or consequences of tooth loss may have different implications for NHL etiology. Our findings suggest that periodontal disease is a risk factor for NHL. Whether periodontal disease is a direct or indirect cause of NHL, or is a marker of underlying systemic inflammation and/or immune dysregulation, warrants further investigation.     

GBV‐C/hepatitis G virus infection and non‐Hodgkin lymphoma: a case control study

We investigated whether there was an association between GBV‐C viremia and the development of non‐Hodgkin lymphoma (NHL) in 553 NHL cases and 438 controls from British Columbia, Canada. Cases were aged 20–79, diagnosed between March 2000 and February 2004, and resident in Greater Vancouver or Victoria. Cases and controls were tested for GBV‐C RNA by RT‐PCR and positive samples were genotyped. Overall, GBV‐C RNA was detected in 4.5% of NHL cases vs. 1.8% of controls [adjusted odds ratio (OR) = 2.72, 95% confidence interval (CI) = 1.22–6.69]. The association between GBV‐C RNA detection and NHL remained even after individuals with a history of prior transfusion, injection drug use and hepatitis C virus sero‐positivity were excluded. GBV‐C viremia showed the strongest association with diffuse large B cell lymphoma (adjusted OR = 5.18, 95% CI = 2.06–13.71). Genotyping was performed on 29/33 GBV‐C RNA positive individuals; genotypes 2a (n = 22); 2b (n = 5) and 3 (n = 2) were identified, consistent with the distribution of genotypes found in North America. This is the largest case‐control study to date associating GBV‐C viremia and NHL risk. As GBV‐C is known to be transmitted through blood products this may have important implications for blood safety.     

Establishment and characterization of a novel Hodgkin lymphoma cell line,AM‐HLH,carrying the Epstein‐Barr virus genome integrated into the host chromosome

We describe the establishment and characterization of a cell line, AM‐HLH, obtained from a patient with Epstein‐Barr virus–positive (EBV⁺) nodular sclerosis–type Hodgkin lymphoma (HL). The cells were positive for CD2 and CD30 and negative for CD15. The immunoglobulin heavy‐ and κ light–chain genes were rearranged. The karyotype was of the triploid range. Southern blotting using the EBV terminal repeat probe detected 3 hybridizing bands that were identical to those of the parental HL material. The cells expressed EBV‐encoded RNAs as well as latent genes (EBNA1, EBNA2, LMP1, and LMP2A) and lytic genes (BZLF1 and BALF2). Fluorescence in situ hybridization (FISH) with the cosmid pJB8 clone containing a fragment of EBV DNA as a probe revealed multiple hybridization signals at a marker chromosome. Additional FISH using whole chromosome painting and centromere probes in combination with multicolor FISH determined that multiple EBV copies were clustered within the chromosome 20 materials of the marker chromosome. Culture supernatants of AM‐HLH contained IL‐10 as measured by the bead‐based immunoassay. It is possible that an integrated EBV genome and cellular genes on chromosome 20 were coamplified, leading to the enhanced expression of genes involved in cell growth control. The AM‐HLH cell line will be useful to clarify the role of cytokines in the development of EBV⁺ HL.     

Lifetime alcohol intake and risk of non‐Hodgkin lymphoma: Findings from the Melbourne Collaborative Cohort Study

Cohort studies have reported inconsistent evidence regarding alcohol intake and risk of non‐Hodgkin lymphoma (NHL), mostly based on alcohol intake assessed close to study enrolment. We examined this association using alcohol intake measured from age 20 onwards. We calculated usual alcohol intake for 10‐year periods from age 20 using recalled frequency and quantity of beverage‐specific consumption for 37,990 participants aged 40–69 years from the Melbourne Collaborative Cohort Study. Cox regression was performed to derive hazard ratios (HRs) and 95% confidence intervals (CIs) for the association between alcohol intake (g/day) and NHL risk. After a mean follow‐up of 19.3 years, 538 NHL cases were diagnosed. Approximately 80% of participants were either lifetime abstainers or consumed below 20 g of ethanol/day. All categories of lifetime alcohol intake were associated with about 20% lower incidence of NHL compared with lifetime abstention, but there was no evidence of a trend by amount consumed (HR = 0.97 per 10 g/day increment in intake, 95% CI: 0.92–1.03; p value = 0.3). HRs for beer, wine and spirits were 0.91 (95% CI: 0.83–1.00; p value = 0.05), 1.03 (95% CI: 0.94–1.12; p value = 0.6), and 1.06 (95% CI: 0.83–1.37; p value = 0.6), respectively, per 10 g/day increment in lifetime intake. There were no significant differences in associations between NHL subtypes. In this low‐drinking cohort, we did not detect a dose‐dependent association between lifetime alcohol intake and NHL risk.