Elevated pleural effusion IL-17 is a diagnostic marker and outcome predictor in lung cancer patients
© Xu et al.; licensee BioMed Central Ltd. 2014
Received: 20 February 2014
Accepted: 2 April 2014
Published: 8 May 2014
Interleukin 17 (IL-17) is a proinflammatory cytokine produced mainly by CD4+ T-lymphocytes and may be important in tumor cell growth and progression. In this study, we aimed to evaluate the diagnostic and prognostic value of pleural effusion levels of IL-17 in lung cancer patients with malignant pleural effusion (MPE).
Pleural effusion samples were collected from 78 lung cancer patients with MPE and from 45 patients with nonmalignant pleural effusion. Pleural fluid concentrations of IL-17 were measured by using enzyme-linked immunosorbent assays.
Malignant effusion exhibited higher IL-17 levels than nonmalignant effusion (20.49 ± 5.27 pg/ml vs. 13.16 ± 2.25 pg/ml; P < 0.01). Lung cancer patients with pleural fluid IL-17 levels below 15 pg/ml had longer overall survival than those patients with higher levels (10.8 months vs. 4.7 months; P < 0.05). On the basis of multivariate analysis, we found that pleural fluid IL-17 level was an independent prognostic factor in lung cancer patients with MPE.
Measurement of IL-17 levels might be a useful diagnostic and prognostic test for lung cancer patients with MPE.
Lung cancer is the leading cause of cancer-related mortality in the world . Approximately 15% of lung cancer patients have pleural effusion at the time of initial diagnosis, and 50% develop it later in the course of their disease . Differentiating malignant from nonmalignant pleural effusion is a critical problem, and conventional methods have proven inadequate [3–6]. Pleural fluid cytology has traditionally been the analytical method of choice for the detection of tumor cells in pleural fluid. However, sensitivity varies between 30% and 60% , and blindly obtained pleural needle biopsy specimens offer little additional sensitivity . Although the presence of tumor cells in pleural effusion is a diagnostic marker of malignant pleural effusion (MPE), the probability of finding them is low. For negative cytology pleural effusion, some of the currently used indices, such as carcinoembryonic antigen, neuron-specific enolase and cytokeratin 19 fragments (CYFRA 21-1), have a certain extent of differential value; however, their specificity and sensitivity are limited [9, 10]. Therefore, searching for new indices is very important.
In addition to diagnostic issues, patients with MPE have a poor prognosis and are difficult to treat effectively . Despite advances in treatment modalities, the overall survival (OS) is still very short. The present standard treatment is to evacuate the pleural fluid, followed by intravenous chemotherapy or intrapleural chemotherapy . However, it was found that not all patients were benefited from the addition of chemotherapy, especially in patients with short OS. Therefore, prognostic assessment of the patient is essential. Hsu et al. proved that the expression level of angiogenetic biomarkers was significantly correlated with patient survival and pleural effusion control . In addition, researchers in recent molecular and genetic profiling studies identified several markers as diagnostic and prognostic factors of lung cancer.
The improved understanding of pleural effusion immunopathogenesis could lead to the development of immunodiagnostic tools to facilitate its differential diagnosis. Investigators in large-scale studies have reported that lymphocytes play an important role in the pathogenesis of pleural effusion [14–17]. CD4+ T cells can be differentiated into interleukin 17 (IL-17)-producing T helper (Th17) cells. Th17 cells produce unique cytokines, including IL-17A, IL-17F and IL-22. These cytokines play a role in inflammation and cancers [18–21]. In previous studies, researchers have reported that the levels of IL-17 are elevated in pleural effusion [22, 23]. However, whether there is a significant difference in their expression level as well as in the relationship between pleural fluid IL-17 concentrations and the prognosis for patients with lung cancer have not been evaluated to date.
In our present study, we assessed pleural effusion of various etiologies in 123 patients to determine whether IL-17 pleural fluid could be used as a diagnostic indicator of lung cancer and a predictor of survival time. We found that IL-17 is elevated in malignant effusion caused by lung cancer, as well as being potentially predictive of survival outcome.
Patient characteristics a
Mean age (±SD), years
56.3 ± 12.5
55.6 ± 11.7
Squamous cell carcinoma
Small-cell lung carcinoma
This study was approved by the Ethics Committee of Nanjing Chest Hospital, and informed consent was obtained from each patient.
Diagnostic criteria for pleural effusion
The diagnostic criteria for MPE were as follows. Cytological evidence of malignant cells present in pleural effusion or in biopsy specimens. Tuberculous (TB) pleural effusion was diagnosed according to the following findings: identification of acid-fast bacilli in pleural fluid, caseous granulomas in a pleural biopsy specimen and a high level of pleural fluid adenosine deaminase (>40 U/L) with an improvement of the pleurisy after anti-TB therapy. Parapneumonic effusion was characterized by any pleural effusion associated with pneumonia and response to antibiotics. Patients with pleural empyema were also included in this group. Effusions related to heart failure were collected from patients with documented heart failure but without neoplastic or other disease.
Sample collection and determination of IL-17 concentration
Fresh pleural effusion specimens were collected before treatment and centrifuged at 1,500 × g for 10 minutes at -4°C. The supernatants were immediately stored at -80°C until use. The IL-17 concentrations were determined by enzyme-linked immunosorbent assay (ELISA) with the commercial human IL-17 Ready-SET-Go! ELISA Kit (eBioscience, San Diego, CA, USA). All assays were run in duplicate, with dilutions as appropriate, and the technicians were blinded to clinical data.
All statistical analyses of differences between malignant effusions and nonmalignant pleural effusion were performed using the Mann–Whitney U test. The diagnostic accuracy of IL-17 in discriminating between lung cancer with malignant and nonmalignant pleural effusion was compared by constructing receiver operating characteristic (ROC) curves. The optimum cutoff point from the ROC analysis was established by selecting the value that provided the greatest sum of sensitivity and specificity. Survival analyses were performed using the Kaplan–Meier method, and significant differences in survival rates were compared using the logrank test. The Cox proportional hazards regression model was used to compare the relative influence of different prognostic factors. P < 0.05 was considered to indicate statistical significance.
Levels of IL-17 in pleural effusion
Diagnostic value of IL-17 in malignant pleural effusion
Relationship between IL-17 concentration and clinicopathological factors in lung cancer patients with malignant pleural effusion
Interleukin 17 levels in pleural effusion of lung cancer patients a
Patients ( n)
IL-17 (pg/ml) Mean ± SD
27.08 ± 4.87
19.92 ± 5.28
21.23 ± 5.30
19.87 ± 5.23
20.33 ± 6.12
21.52 ± 5.05
0 or 1
19.99 ± 5.37
2 to 4
21.41 ± 5.21
20.46 ± 5.18
20.62 ± 6.12
20.34 ± 4.58
20.83 ± 6.01
20.34 ± 5.36
20.15 ± 4.97
Prognostic significance of pleural fluid IL-17 for lung cancer patients with malignant pleural effusion
Univariate analysis for overall survival in lung cancer patients with malignant pleural effusion a
Patients ( n)
0 or 1
2 to 4
Multivariate Cox proportional hazards analysis for overall survival in lung cancer patients with malignant pleural effusion a
0.593 to 2.689
0.146 to 0.742
0 or 1
2 to 4
0.778 to 4.745
0.083 to 2.479
0.449 to 3.112
To the best of our knowledge, this study is the first on pleural effusion in which IL-17 levels were investigated for their diagnostic and prognostic power simultaneously in lung cancer patients with MPE. Our findings suggest that profoundly elevated pleural fluid concentrations of IL-17 (≥15 pg/ml) measured at the onset of effusion correlated with shorter survival in lung cancer patients. Furthermore, multivariate analysis of prognostic factors identified pleural fluid IL-17 concentration as an independent prognostic factor for OS. These results suggest that IL-17 concentration measured in pleural effusion is an indicator of not only the presence of lung cancer but also the patient’s survival outcome.
IL-17 is a proinflammatory cytokine produced mainly by CD4+ T lymphocytes. It may be important in tumor cell growth and may contribute to the aggressiveness of human tumors . Recently, accumulating evidence has shown that IL-17-positive cells are frequently present in multiple cancers, including prostate cancer , colorectal cancer , hepatocellular carcinoma , breast cancer , ovarian cancer  and non-small-cell lung cancer . Researchers in some studies have reported that IL-17 cell expression in MPE is elevated [15, 23]. Most of the reports indicate that IL-17 may protect against tumors by promoting immune system–mediated tumor rejection . These results suggest the IL-17 plays an important role in tumor immune system evasion. One inference derived from this hypothesis is that IL-17 concentration in pleural effusion could be a good indicator in the follow-up of tumor patients, which is supported by our presently reported results.
Differentiating malignant from nonmalignant pleural effusion is a clinical problem, and conventional methods have proven inadequate [3–6]. A reliable marker for rapid and accurate diagnosis of pleural effusion is greatly needed. In this study, the IL-17 concentrations in malignant pleural effusion were higher than those in nonmalignant and TB pleural effusion. Our results suggest that IL-17 should be a tumor marker for the diagnosis of MPE. No statistically significant correlation was observed between IL-17 concentration and histological type of lung cancer. We assigned 15 pg/ml IL-17 in MPE as the diagnostic cutoff value, which had sensitivity of 79.5% and specificity of 91.1%. This result shows that IL-17 concentration could be a valuable marker in the differential diagnosis of malignant and nonmalignant pleural effusion. Further studies of the potential efficacy of this marker are needed.
IL-17 concentrations in MPE associated with lung cancer are significantly higher than those in nonmalignant pleural effusion. Determination of IL-17 concentration in pleural effusion is diagnostically informative and IL-17 concentration is an independent prognostic factor that shows promise in the follow-up of lung cancer patients who develop pleural effusion.
- Kasahara K, Shibata K, Shintani H, Iwasa K, Sone T, Kimura H, Nobata K, Hirose T, Yoshimi Y, Katayama N, Ishiura Y, Kita T, Nishi K, Nakatsumi Y, Ryoma Y, Fujimura M, Nakao S: Randomized phase II trial of OK-432 in patients with malignant pleural effusion due to non-small cell lung cancer. Anticancer Res 2006, 26: 1495–1499.PubMedGoogle Scholar
- Cheng D, Liang B, Li YH: Application of MMP-7 and MMP-10 in assisting the diagnosis of malignant pleural effusion. Asian Pac J Cancer Prev 2012, 13: 505–509. 10.7314/APJCP.2012.13.2.505PubMedView ArticleGoogle Scholar
- Aoe K, Hiraki A, Murakami T, Eda R, Maeda T, Sugi K, Takeyama H: Diagnostic significance of interferon-γ in tuberculous pleural effusions. Chest 2003, 123: 740–744. 10.1378/chest.123.3.740PubMedView ArticleGoogle Scholar
- Aoe K, Hiraki A, Murakami T, Murakami K, Makihata K, Takao K, Eda R, Maeda T, Sugi K, Darzynkiewicz Z, Takeyama H: Relative abundance and patterns of correlation among six cytokines in pleural fluid measured by cytometric bead array. Int J Mol Med 2003, 12: 193–198.PubMedGoogle Scholar
- Hiraki A, Aoe K, Matsuo K, Murakami K, Murakami T, Onoda T, Sugi K, Takeyama H, Eda R: Simultaneous measurement of T-helper 1 cytokines in tuberculous pleural effusion. Int J Tuberc Lung Dis 2003, 7: 1172–1177.PubMedGoogle Scholar
- Hiraki A, Aoe K, Eda R, Maeda T, Murakami T, Sugi K, Takeyama H: Comparison of six biological markers for the diagnosis of tuberculous pleuritis. Chest 2004, 125: 987–989. 10.1378/chest.125.3.987PubMedView ArticleGoogle Scholar
- Sriram KB, Relan V, Clarke BE, Duhig EE, Yang IA, Bowman RV, Lee YC, Fong KM: Diagnostic molecular biomarkers for malignant pleural effusions. Future Oncol 2011, 7: 737–752. 10.2217/fon.11.45PubMedView ArticleGoogle Scholar
- Heffner JE: Diagnosis and management of malignant pleural effusions. Respirology 2008, 13: 5–20.PubMedView ArticleGoogle Scholar
- Villena V, López-Encuentra A, Echave-Sustaeta J, Martín-Escribano P, Ortuño-de-Solo B, Estenoz-Alfaro J: Diagnostic value of CA 549 in pleural fluid: comparison with CEA, CA 15.3 and CA 72.4. Lung Cancer 2003, 40: 289–294. 10.1016/S0169-5002(03)00120-XPubMedView ArticleGoogle Scholar
- Porcel JM, Vives M, Esquerda A, Salud A, Pérez B, Rodríguez-Panadero F: Use of a panel of tumor markers (carcinoembryonic antigen, cancer antigen 125, carbohydrate antigen 15–3, and cytokeratin 19 fragments) in pleural fluid for the differential diagnosis of benign and malignant effusions. Chest 2004, 126: 1757–1763. 10.1378/chest.126.6.1757PubMedView ArticleGoogle Scholar
- Aoe K, Hiraki A, Yamazaki K, Nakamura Y, Murakami T, Maeda T, Nishimura M, Sugi K, Ueoka H: Elevated pleural fluid RCAS1 is a diagnostic marker and outcome predictor in lung cancer patients. Int J Oncol 2006, 29: 65–72.PubMedGoogle Scholar
- Su WC, Lai WW, Chen HH, Hsiue TR, Chen CW, Huang WT, Chen TY, Tsao CJ, Wang NS: Combined intrapleural and intravenous chemotherapy, and pulmonary irradiation, for treatment of patients with lung cancer presenting with malignant pleural effusion: a pilot study. Oncology 2003, 64: 18–24. 10.1159/000066516PubMedView ArticleGoogle Scholar
- Hsu IL, Su WC, Yan JJ, Chang JM, Lai WW: Angiogenetic biomarkers in non-small cell lung cancer with malignant pleural effusion: correlations with patient survival and pleural effusion control. Lung Cancer 2009, 65: 371–376. 10.1016/j.lungcan.2008.12.007PubMedView ArticleGoogle Scholar
- Ibrahim L, Salah M, Abd El Rahman A, Zeidan A, Ragb M: Crucial role of CD4 + CD25 + FOXP3+ T regulatory cell, interferon-γ and interleukin-16 in malignant and tuberculous pleural effusions. Immunol Invest 2013, 42: 122–136. 10.3109/08820139.2012.736116PubMedView ArticleGoogle Scholar
- Ye ZJ, Zhou Q, Gu YY, Qin SM, Ma WL, Xin JB, Tao XN, Shi HZ: Generation and differentiation of IL-17-producing CD4+ T cells in malignant pleural effusion. J Immunol 2010, 185: 6348–6354. 10.4049/jimmunol.1001728PubMedView ArticleGoogle Scholar
- Yang HB, Shi HZ: T lymphocytes in pleural effusion. Chin Med J (Engl) 2008, 121: 579–580.Google Scholar
- Qin XJ, Shi HZ, Liang QL, Huang LY, Yang HB: CD4+CD25+ regulatory T lymphocytes in tuberculous pleural effusion. Chin Med J (Engl) 2008, 121: 581–586.Google Scholar
- Wang L, Yi T, Kortylewski M, Pardoll DM, Zeng D, Yu H: IL-17 can promote tumor growth through an IL-6-Stat3 signaling pathway. J Exp Med 2009, 206: 1457–1464. 10.1084/jem.20090207PubMedPubMed CentralView ArticleGoogle Scholar
- Sfanos KS, Bruno TC, Maris CH, Xu L, Thoburn CJ, DeMarzo AM, Meeker AK, Isaacs WB, Drake CG: Phenotypic analysis of prostate-infiltrating lymphocytes reveals TH17 and Treg skewing. Clin Cancer Res 2008, 14: 3254–3261. 10.1158/1078-0432.CCR-07-5164PubMedPubMed CentralView ArticleGoogle Scholar
- Miyahara Y, Odunsi K, Chen W, Peng G, Matsuzaki J, Wang RF: Generation and regulation of human CD4+ IL-17-producing T cells in ovarian cancer. Proc Natl Acad Sci U S A 2008, 105: 15505–15510. 10.1073/pnas.0710686105PubMedPubMed CentralView ArticleGoogle Scholar
- Kryczek I, Banerjee M, Cheng P, Vatan L, Szeliga W, Wei S, Huang E, Finlayson E, Simeone D, Welling TH, Chang A, Coukos G, Liu R, Zou W: Phenotype, distribution, generation, and functional and clinical relevance of Th17 cells in the human tumor environments. Blood 2009, 114: 1141–1149. 10.1182/blood-2009-03-208249PubMedPubMed CentralView ArticleGoogle Scholar
- Wang T, Lv M, Qian Q, Nie Y, Yu L, Hou Y: Increased frequencies of T helper type 17 cells in tuberculous pleural effusion. Tuberculosis (Edinb) 2011, 91: 231–237. 10.1016/j.tube.2011.02.002View ArticleGoogle Scholar
- Ye ZJ, Zhou Q, Zhang JC, Li X, Wu C, Qin SM, Xin JB, Shi HZ: CD39+ regulatory T cells suppress generation and differentiation of Th17 cells in human malignant pleural effusion via a LAP-dependent mechanism. Respir Res 2011, 12: 77. 10.1186/1465-9921-12-77PubMedPubMed CentralView ArticleGoogle Scholar
- Dong C: TH17 cells in development: an updated view of their molecular identity and genetic programming. Nat Rev Immunol 2008, 8: 337–348. 10.1038/nri2295PubMedView ArticleGoogle Scholar
- Steiner GE, Newman ME, Paikl D, Stix U, Memaran-Dagda N, Lee C, Marberger MJ: Expression and function of pro-inflammatory interleukin IL-17 and IL-17 receptor in normal, benign hyperplastic, and malignant prostate. Prostate 2003, 56: 171–182. 10.1002/pros.10238PubMedView ArticleGoogle Scholar
- Le Gouvello S, Bastuji-Garin S, Aloulou N, Mansour H, Chaumette MT, Berrehar F, Seikour A, Charachon A, Karoui M, Leroy K, Farcet JP, Sobhani I: High prevalence of Foxp3 and IL17 in MMR-proficient colorectal carcinomas. Gut 2008, 57: 772–779. 10.1136/gut.2007.123794PubMedView ArticleGoogle Scholar
- Zhang JP, Yan J, Xu J, Pang XH, Chen MS, Li L, Wu C, Li SP, Zheng L: Increased intratumoral IL-17-producing cells correlate with poor survival in hepatocellular carcinoma patients. J Hepatol 2009, 50: 980–989. 10.1016/j.jhep.2008.12.033PubMedView ArticleGoogle Scholar
- Zhu X, Mulcahy LA, Mohammed RA, Lee AH, Franks HA, Kilpatrick L, Yilmazer A, Paish EC, Ellis IO, Patel PM, Jackson AM: IL-17 expression by breast-cancer-associated macrophages: IL-17 promotes invasiveness of breast cancer cell lines. Breast Cancer Res 2008, 10: R95. 10.1186/bcr2195PubMedPubMed CentralView ArticleGoogle Scholar
- Chen X, Wan J, Liu J, Xie W, Diao X, Xu J, Zhu B, Chen Z: Increased IL-17-producing cells correlate with poor survival and lymphangiogenesis in NSCLC patients. Lung Cancer 2010, 69: 348–354. 10.1016/j.lungcan.2009.11.013PubMedView ArticleGoogle Scholar
- Benchetrit F, Ciree A, Vives V, Warnier G, Gey A, Sautès-Fridman C, Fossiez F, Haicheur N, Fridman WH, Tartour E: Interleukin-17 inhibits tumor cell growth by means of a T-cell-dependent mechanism. Blood 2002, 99: 2114–2121. 10.1182/blood.V99.6.2114PubMedView ArticleGoogle Scholar
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