Long‐term prognostic significance of interleukin‐17‐producing T cells in patients with non‐small cell lung cancer

The presence of interleukin (IL)‐17‐producing T cells has recently been reported in non‐small cell lung cancer (NSCLC) patients. However, the long‐term prognostic significance of these populations in NSCLC patients remains unknown. In the present study, we collected peripheral blood from 82 NSCLC patients and 22 normal healthy donors (NC). Percentages of IL‐17‐producing CD4⁺T (Th17), CD8⁺T (Tc17) and γδT cells (γδT17) were measured to determine their association with clinical outcomes and overall survival (OS) in NSCLC. All NSCLC patients were followed up until July 2018. Median follow‐up time was 13.5 months (range 1‐87 months). The 3‐ and 5‐year survival rate was 27% and 19.6%, respectively. We found that Th17 cells and γδT17 cells were significantly increased, whereas Tc17 cells were markedly decreased in patients with NSCLC compared with those in NC. In addition, Th17 cells were significantly positively associated with T helper type 1 cells (Th1), whereas γδT17 cells were significantly negatively associated with γδT ⁺ interferon (IFN)‐γ⁺ cells. High percentages of peripheral Tc17 cells were significantly associated with favorable 5‐year OS (P = .025), especially in patients with early TNM stage (P = .016). Furthermore, high percentages of peripheral Th17 cells were positively associated with favorable 5‐year OS in patients with late TNM stage (P = .002). However, no significant association was observed between γδT17 cells and OS, regardless of the TNM stage. In conclusion, our findings suggest that enhanced Th17 and reduced Tc17 cells in the peripheral blood could be a significant predictor of a favorable prognosis for NSCLC patients.     

IL‐17A‐producing CD30+ Vδ1 T cells drive inflammation‐induced cancer progression

Although it has been suspected that inflammation is associated with increased tumor metastasis, the exact type of immune response required to initiate cancer progression and metastasis remains unknown. In this study, by using an in vivo tumor progression model in which low tumorigenic cancer cells acquire malignant metastatic phenotype after exposure to inflammation, we found that IL‐17A is a critical cue for escalating cancer cell malignancy. We further demonstrated that the length of exposure to an inflammatory microenvironment could be associated with acquiring greater tumorigenicity and that IL‐17A was critical for amplifying such local inflammation, as observed in the production of IL‐1β and neutrophil infiltration following the cross‐talk between cancer and host stromal cells. We further determined that γδT cells expressing Vδ1 semi‐invariant TCR initiate cancer‐promoting inflammation by producing IL‐17A in an MyD88/IL‐23‐dependent manner. Finally, we identified CD30 as a key molecule in the inflammatory function of Vδ1T cells and the blockade of this pathway targeted this cancer immune‐escalation process. Collectively, these results reveal the importance of IL‐17A‐producing CD30⁺ Vδ1T cells in triggering inflammation and orchestrating a microenvironment leading to cancer progression.     

Phenotypic characterization and prognostic impact of circulating γδ and αβ T‐cells in metastatic malignant melanoma

Human T cells carrying γδ T‐cell receptors (TCRs) represent a minor population relative to those with αβ TCRs. There has been much interest recently in the possibility of using these γδ T‐cells in cancer therapy because they can kill tumor cells in vitro in an MHC‐unrestricted manner, and possess potential regulatory capability and antigen‐presenting capacity. The presence of γδ T‐cells in late‐stage melanoma patients and their relationship with survival has not been extensively explored, although relatively lower percentages of total γδ T‐cells and Vδ2+ cells have been reported. Here, we present a detailed analysis of associations of γδ T‐cell subsets and differentiation stages with survival in Stage IV patients, compared with CD4+ and CD8+ αβ T‐cells. We found an increased Vδ1:Vδ2‐ratio and a decreased CD4:CD8‐ratio in patients compared to healthy controls, on the basis both of relative frequencies and absolute cell counts per μL blood. Nonetheless, Kaplan–Meier analyses showed that a higher than median frequency of Vδ1+ cells was negatively associated with survival, whereas there were no positive or negative associations with frequencies of Vδ2+ cells. Correlations of cell differentiation status with survival revealed a negative association of early‐differentiated Vδ1+ T cells with survival, both on the basis of relative frequencies and absolute counts. There was also a positive correlation between the frequencies of early‐differentiated CD8+ αβ T‐cells and survival. Our findings suggest peripheral blood frequencies of Vδ1+ T‐cells as a potential prognostic marker in melanoma. The mechanisms by which higher abundance of Vδ1+ cells are associated with poorer survival require determination.     

Clinicopathological analysis of 17 primary cutaneous T‐cell lymphoma of the γδ phenotype from Japan

Primary cutaneous γδ T‐cell lymphoma (PCGD‐TCL) is an aggressive lymphoma consisting of clonal proliferation of mature activated γδ T‐cells of a cytotoxic phenotype. Because primary cutaneous γδ T‐cell lymphoma is a rare disease, there are few clinicopathological studies. In addition, T‐cell receptor (TCR) γδ cells are typically immunostained in frozen sections or determined by TCRβ negativity. We retrospectively analyzed 17 primary cutaneous T‐cell lymphomas of the γδ phenotype (CTCL‐γδ) in a clinicopathological and molecular study using paraffin‐embedded sections. Among 17 patients, 11 had CTCL‐γδ without subcutaneous panniculitis‐like T‐cell lymphoma (SPTCL) features and six had CTCL‐γδ with SPTCL features. Immunophenotypically, some significant differences were found in CD8 and CD56 positivity between our patient series of CTCL‐γδ patients with SPTCL features and SPTCL‐γδ patients described in the previous literature. A univariate analysis of 17 CTCL‐γδ patients showed that being more than 60 years old, presence of visceral organ involvement, and small‐to‐medium cell size were poor prognostic factors. In addition, the 5‐year overall survival rate was 42.4% for the CTCL‐γδ patients without SPTCL features and 80.0% for those with SPTCL features. Consequently, there was a strikingly significant difference in overall survival among SPTCL, CTCL‐γδ with SPTCL features and CTCL‐γδ without SPTCL features (P = 0.0005). Our data suggests that an indolent subgroup may exist in CTCL‐γδ. Studies on more cases, including those from other countries, are warranted to delineate the clinicopathological features and the significance in these rare lymphomas.     

TGF‐β inactivation and TGF‐α overexpression cooperate in an in vivo mouse model to induce hepatocellular carcinoma that recapitulates molecular features of human liver cancer

Hepatocellular carcinoma (HCC) results from the cumulative effects of deregulated tumor suppressor genes and oncogenes. The tumor suppressor and oncogenes commonly affected include growth factors, receptors and their downstream signaling pathway components. The overexpression of transforming growth factor alpha (TGF‐α) and the inhibition of TGF‐β signaling are especially common in human liver cancer. Thus, we assessed whether TGF‐α overexpression and TGF‐β signaling inactivation cooperate in hepatocarcinogenesis using an in vivo mouse model, MT1/TGFa;AlbCre/Tgfbr2^flx/flx mice (“TGFa;Tgfbr2^hepko”), which overexpresses TGF‐α and lacks a TGF‐β receptor in the liver. TGF‐β signaling inactivation did not alter the frequency or number of cancers in mice with overexpression of TGF‐α. However, the tumors in the TGFa;Tgfbr2^hepko mice displayed increased proliferation and increased cdk2, cyclin E and cyclin A expression as well as decreased Cdkn1a/p21 expression compared to normal liver and compared to the cancers arising in the TGF‐α overexpressing mice with intact TGF‐β receptors. Increased phosphorylated ERK1/2 expression was also present in the tumors from the TGFa;Tgfbr2^hepko mice and correlated with downregulated Raf kinase inhibitor protein expression, which is a common molecular event in human HCC. Thus, TGF‐β signaling inactivation appears to cooperate with TGF‐α in vivo to promote the formation of liver cancer that recapitulates molecular features of human HCC.     

Regulatory mechanisms mediated by peroxisome proliferator‐activated receptor‐β/δ in skin cancer

Considerable progress has been made during the past 20 years towards elucidating the role of peroxisome proliferator‐activated receptor‐β/δ (PPARβ/δ) in skin cancer. In 1999, the original notion that PPARβ/δ was involved with epithelial cell function was postulated based on a correlation between PPARβ/δ expression and the induction of messenger RNAs encoding proteins that mediate terminal differentiation in keratinocytes. Subsequent studies definitively revealed that PPARβ/δ could induce terminal differentiation and inhibit proliferation of keratinocytes. Molecular mechanisms have since been discovered to explain how this nuclear receptor can be targeted for preventing and treating skin cancer. This includes the regulation of terminal differentiation, mitotic signaling, endoplasmic reticulum stress, and cellular senescence. Interestingly, the effects of activating PPARβ/δ can preferentially target keratinocytes with genetic mutations associated with skin cancer. This review provides the history and current understanding of how PPARβ/δ can be targeted for both nonmelanoma skin cancer and melanoma and postulates how future approaches that modulate PPARβ/δ signaling may be developed for the prevention and treatment of these diseases.     

Regulation of peroxisome proliferator‐activated receptor‐β/δ by the APC/β‐CATENIN pathway and nonsteroidal antiinflammatory drugs

Studies indicate that peroxisome proliferator‐activated receptor‐β/δ (PPARβ/δ) can either attenuate or potentiate colon cancer. One hypothesis suggests that PPARβ/δ is upregulated by the adenomatous polyposis coli (APC)/β‐CATENIN pathway and a related hypothesis suggests that PPARβ/δ is downregulated by nonsteroidal antiinflammatory drugs (NSAIDs). The present study examined these possibilities using in vivo and in vitro models. While APC/β‐CATENIN‐dependent expression of CYCLIN D1 was observed in vivo and in vitro, expression of PPARβ/δ was not different in colon or intestinal polyps from wild‐type or Apc^min heterozygous mice or in human colon cancer cell lines with mutations in APC and/or β‐CATENIN. No difference in the level of PPARβ/δ was found in colon from wild‐type or Apc^min heterozygous mice following treatment with NO‐donating aspirin (NO‐ASA). NSAIDs inhibited cell growth in RKO (wild‐type APC) and DLD1 (mutant APC) human colon cancer cell lines but expression of PPARβ/δ was not downregulated in these cell lines in response to a broad concentration range of celecoxib, indomethacin, NS‐398, or nimesulide. However, indomethacin caused an increase in PPARβ/δ mRNA and protein that was accompanied with increased expression of a known PPARβ/δ target gene. Interestingly, expression of PPARα was also increased in the human colon cancer cell lines by several NSAIDs at the highest concentration examined. Results from these studies provide additional evidence indicating that PPARβ/δ is not upregulated by the APC/β‐CATENIN pathway. Further, these studies suggest that increased PPARβ/δ and/or PPARα by NSAIDs in human colon cancer cell lines could contribute to the mechanisms underlying the chemopreventive effects of NSAIDs. Mol. Carcinog. © 2009 Wiley‐Liss, Inc.     

Amino‐terminal fragments of laminin γ2 chain retract vascular endothelial cells and increase vascular permeability

Laminin γ2 (Lmγ2) chain, a subunit of laminin‐332, is a typical molecular marker of invading cancer cells, and its expression correlates with poor prognosis of cancer patients. It was previously found that forced expression of Lmγ2 in cancer cells promotes their invasive growth in nude mice. However, the mechanism of the tumor‐promoting activity of Lmγ2 remains unknown. Here we investigated the interaction between Lmγ2 and vascular endothelial cells. When treated with an N‐terminal proteolytic fragment of γ2 (γ2pf), HUVECs became markedly retracted or shrunken. The overexpression of Lmγ2 or treatment with γ2pf stimulated T‐24 bladder carcinoma cells to invade into the HUVEC monolayer and enhanced their transendothelial migration in vitro. Moreover, γ2pf increased endothelial permeability in vitro and in vivo. As the possible mechanisms, γ2pf activated ERK and p38 MAPK but inactivated Akt in HUVECs. Such effects of γ2pf led to prominent actin stress fiber formation in HUVECs, which was blocked by a ROCK inhibitor. In addition, γ2pf induced delocalization of VE‐cadherin and β‐catenin from the intercellular junction. As possible receptors, γ2pf interacted with heparan sulfate proteoglycans on the surface of HUVECs. Moreover, we localized the active site of γ2pf to the N‐terminal epidermal growth factor‐like repeat. These data suggest that the interaction between γ2pf and heparan sulfate proteoglycans induces cytoskeletal changes of endothelial cells, leading to the loss of endothelial barrier function and the enhanced transendothelial migration of cancer cells. These activities of Lmγ2 seem to support the aberrant growth of cancer cells.     

The Karyopherin proteins,Crm1 and Karyopherin β1, are overexpressed in cervical cancer and are critical for cancer cell survival and proliferation

The Karyopherin proteins are involved in nucleo‐cytoplasmic trafficking and are critical for protein and RNA subcellular localization. Recent studies suggest they are important in nuclear envelope component assembly, mitosis and replication. Since these are all critical cellular functions, alterations in the expression of the Karyopherins may have an impact on the biology of cancer cells. In this study, we examined the expression of the Karyopherins, Crm1, Karyopherin β1 (Kpnβ1) and Karyopherin α2 (Kpnα2), in cervical tissue and cell lines. The functional significance of these proteins to cancer cells was investigated using individual siRNAs to inhibit their expression. Microarrays, quantitative RT‐PCR and immunofluorescence revealed significantly higher expression of Crm1, Kpnβ1 and Kpnα2 in cervical cancer compared to normal tissue. Expression levels were similarly elevated in cervical cancer cell lines compared to normal cells, and in transformed epithelial and fibroblast cells. Inhibition of Crm1 and Kpnβ1 in cancer cells significantly reduced cell proliferation, while Kpnα2 inhibition had no effect. Noncancer cells were unaffected by the inhibition of Crm1 and Kpnβ1. The reduction in proliferation of cancer cells was associated with an increase in a subG1 population by cell cycle analysis and Caspase‐3/7 assays revealed increased apoptosis. Crm1 and Kpnβ1 siRNA‐induced apoptosis was accompanied by an increase in the levels of growth inhibitory proteins, p53, p27, p21 and p18. Our results demonstrate that Crm1, Kpnβ1 and Kpnα2 are overexpressed in cervical cancer and that inhibiting the expression of Crm1 and Kpnβ1, not Kpnα2, induces cancer cell death, making Crm1 and Kpnβ1 promising candidates as both biomarkers and potential anticancer therapeutic targets. © 2008 Wiley‐Liss, Inc.     

Children with endemic Burkitt lymphoma are deficient in EBNA1‐specific IFN‐γ T cell responses

Endemic Burkitt lymphoma (eBL) is the most common childhood cancer in equatorial Africa and is linked to Epstein–Barr virus (EBV) and Plasmodium falciparum coinfections early in life. Epstein–Barr nuclear antigen 1 (EBNA1) is the sole viral latent antigen expressed in BL tumors. Loss of EBNA1‐specific immune surveillance could allow eBL emergence. Therefore, EBNA1‐specific T cell responses were analyzed by IFN‐γ ELISPOT in Kenyan children with eBL and compared to healthy children with divergent malaria exposure. Significantly fewer children with eBL, 16% (7/44) had EBNA1‐specific IFN‐γ responses in contrast to healthy children living in a malaria holoendemic area or in an area with sporadic malaria transmission, 67% (40/60) and 72% (43/60) responders, respectively (p < 0.003). Children with eBL maintained IgG₁ dominated antibody responses to EBNA1 similar to healthy children suggesting a selective loss of IFN‐γ secreting EBNA1‐specific T cells in the presence of intact humoral immunity. CD8⁺ T cell responses to EBV lytic and latent antigens not expressed in the tumors were similarly robust in eBL patients compared to healthy children. In addition, CD4⁺ T cell responses to a malaria protein, merozoite surface protein 1, were present in lymphoma patients. This study demonstrates a selective loss of EBNA1‐specific T cell responses in children with eBL and suggests a potential immunotherapeutic target for this EBV‐associated lymphoma. © 2008 Wiley‐Liss, Inc.     

Th17 cells and interleukin‐17 increase with poor prognosis in patients with acute myeloid leukemia

Although Th17 cells play crucial roles in the pathogenesis of many autoimmune and inflammatory disorders, their roles in malignancies are currently under debate. The role and mechanism of Th17 cells in patients with acute myeloid leukemia (AML) remain poorly understood. Here we demonstrated that the frequency of Th17 cells was significantly increased in peripheral blood mononuclear cells (PBMCs) and bone marrow mononuclear cells from AML patients compared with healthy donors. Plasma levels of interleukin (IL)‐17, IL‐22, IL‐23, IL‐1β, IL‐6, and transforming growth factor (TGF)‐β1 were significantly increased in blood and bone marrow in AML patients compared with healthy donors. The in vitro experiments demonstrated that IL‐1β, IL‐6, IL‐23, but not TGF‐β1 promoted the generation and differentiation of Th17 cells from naive CD4⁺ T cells in humans. IL‐17A, a signature cytokine secreted by Th17 cells, induced the proliferation of IL‐17 receptor (IL‐17R)‐positive AML cells via IL‐17R, in which activation of PI3K/Akt and Jak/Stat3 signaling pathway may play important roles. In addition, combination of IL‐17A and IL‐22 significantly reduced the generation of Th1 cells and the production of interferon (IFN)‐γ from healthy donor or AML patient peripheral blood mononuclear cells. Patients with high Th17 cell frequency had poor prognosis, whereas patients with high Th1 cell frequency had prolonged survival. Combined analysis of Th1 and Th17 cell frequencies improved the ability to predict patient outcomes. In conclusion, Th17 cells play a crucial role in the pathogenesis of AML and may be an important therapeutic target and prognostic predictor.     

bFGF-mediated phosphorylation of δ-catenin increases its protein stability and the ability to induce the nuclear redistribution of β-catenin

Recently, we have shown that δ-catenin strengthened the epidermal growth factor receptor (EGFR)/Erk1/2 signaling pathway through the association between EGFR and δ-catenin. Now, we further analyzed the correlation between basic fibroblast growth factor (bFGF)/fibroblast growth factor receptor 1 (FGFR1) and δ-catenin in prostate cancer and investigated the molecular mechanism underlying the role of bFGF/FGFR1 modulation in CWR22Rv-1 (Rv-1) cells. Here, we demonstrated that bFGF phosphorylated the tyrosine residues of δ-catenin in Rv-1 cells and further proved that the bFGF mediated FGFR1/δ-catenin tyrosine phosphorylation was time dependent. Furthermore, we demonstrated that bFGF stabilized the expression of δ-catenin through weakening its association with GSK3β and enhancing its stability to induce β-catenin into the nuclear by strengthening the processing of E-cadherin. In a word, these results indicated that bFGF/FGFR1 signaling pathway could enhance the tumor progression of prostate cancer via δ-catenin.     

γ‐Glutamyltransferase and cancer risk: The Korean cancer prevention study

Elevated serum γ‐glutamyltransferase (GGT) is a marker of hepatic injury and is associated with risk of chronic disease. However, the value of GGT as a biomarker for cancer risk remains unclear. Therefore, we evaluated the association of serum GGT with cancer incidence among more than 1.6 million Koreans. We included 1,662,087 Koreans (1,108,121 men and 553,966 women aged 20–95 years) who received health insurance from the National Health Insurance Service and had a biennial medical evaluation between 1995 and 1998. Follow‐up was through December 2012. Using Cox proportional hazards models, we adjusted for age, smoking status, alcohol consumption, exercise and body mass index after exclusion of early cases (cancer diagnosis or death within 5 years of starting follow‐up) and estimated hazard ratios (HRs) of overall and organ‐specific cancer incidence by GGT quintiles. During the 17‐year follow‐up, 129,087 new cancer cases occurred among the participants. Across levels of GGT, there was a positive gradient of HR and the highest quintile of GGT (≥60 IU/L) had the highest HR for all cancers in both men and women. By cancer site, the association was strongest for liver cancer, comparing the highest and lowest strata in men [HR, 6.67; 95% confidence interval (95%CI), 5.88–7.57] and in women (HR, 7.57; 95%CI, 6.41–8.94). Significant associations were also observed for cancers of the esophagus, larynx, stomach, colorectal, bile duct and lung in men and of the bile duct in women. Increased serum GGT level is independently associated with risk of cancer.     

Evaluation of the safety,pharmacokinetics and treatment effects of an ανβ3 integrin inhibitor on bone turnover and disease activity in men with hormone‐refractory prostate cancer and bone metastases

Aim: This study aimed to evaluate the safety, pharmacokinetics and treatment effects of an α_νβ₃ integrin inhibitor on bone turnover and disease activity in men with hormone‐refractory prostate cancer (HRPC) and bone metastases. Methods: A total of 21 patients with bone metastases and HRPC were randomized to receive MK‐0429 200 mg b.i.d. or 1600 mg b.i.d. for 4 weeks. Toxicity, pharmacokinetics and markers of bone turnover and tumor activity were examined. Results: Nausea was the most common adverse event: one (200‐mg group) and 11 (1600‐mg group) patients. At 4 weeks, mean AUC_{0–12 h} was 210 mmol*h (200‐mg group) and 673 mmol*h (1600‐mg group); mean C_max values were 42 mmol/L (200‐mg group) and 154 mmol/L (1600‐mg group). Urinary cross‐linked N‐telopeptides of type I collagen to creatinine ratio (uNTx), a bone turnover biomarker, showed a change from baseline of ?43.4 percent (200‐mg group) and ?34.1 percent (1600‐mg group). There was an increase in serum prostate specific antigen (PSA), a marker for disease activity, of 54.1 percent (200‐mg group) and 44.5 percent (1600‐mg group). Conclusion: MK‐0429 was generally well tolerated, with the most common side‐effect being nausea. There was some evidence of an early reduction of bone turnover, indicating a potential for clinical use in the treatment of MBD although serum PSA was unexpectedly increased during the study.     

α-Adrenergic receptors may contribute to the hypertriglyceridemia associated with tumour growth

The inoculation of the Yoshida AH-130 ascites hepatoma to rats resulted in an important loss of adipose tissue associated with a decrease in lipoprotein lipase (LPL) activity. Tumour burden also resulted in an important hyperlipidemia which affected both triglyceride and free fatty acids. Administration of phentolamine (an α-adrenergic antagonist) to tumour-bearing rats did not influence LPL activity, but it reversed the increase in plasma triglycerides associated with tumour burden. It is suggested that the hypertriglyceridemia associated with tumour growth may be, in part, a consequence of the effect of catecholamines on hepatic triglyceride secretion, via α-adrenergic receptors.     

Estrogen receptor β and breast cancer

A second estrogen receptor, estrogen receptor-beta, was identified in 1996 and has led to an intensive re-evaluation of the role of estrogens in normal physiological and disease processes. While much has been learnt about this new receptor, there remain many outstanding questions, particularly regarding its prognostic significance and therapeutic implications.