國立成功大學「邁向頂尖大學計畫」
延攬優秀人才工作報告表
NCKU’s “Aim for the Top University Project” Work Report Form for Distinguished Scholars
□續聘continuation of employment ■離職resignation
100 年 7 月 13 日更新
受聘者姓名
Name of the Employee 徐宗溢 Male Female■男
聘 期 Period of Employment from 103 年(y) 01 月(m) 01 日(d) to 103 年(y) 06 月(m) 30 日(d) 研究或教學或科技研發與 管理計畫名稱 The project title of research,
teaching, technology development and management
Sp1 在肺癌形成所扮演之角色及其 新穎抑制劑的藥效評估之探討 計畫主持人 （申請單位主管） Project Investigator (Head of Department/Center)
洪建中
(吳俊忠)
補助延聘編號Grant Number HUA 104 - 3 - 22 -063
一、 研究、教學、科技研發與管理工作全程經過概述。（由受聘人填寫）
Please summarize the entire research, teaching, or science and technology R&D and management work process (To be
completed by the employee)
Positive Feedback Regulation between IL10 and EGFR Promotes Lung Cancer Formation Abstract
Purpose: The role of interleukin (IL)10, a well-known tumor-promoting cytokine, in the tumorigenesis of
various cancer types is controversial. The objective is to clarify whether IL10 is required for lung tumorigenesis.
Experimental design: IL10 levels were measured in lung tumor tissues and serum, both of which were from
human and lung cancer transgenic mice (EGFRL858R and Kras4bG12D). EGFRL858R/IL10-/- and Kras4bG12D/IL10
-/-were used to confirm the essential role of IL10 in initiating lung cancer by histological analysis. The gene expression profile in EGFRL858R/IL10-/- was uncovered by microarray analysis. Effect of IL10 on oncogenic
signaling was studied in vitro and in vivo. Furthermore, we studied the role of Src in IL10-mediated signaling for lung cancer development.
Results: Increased IL10 levels in lung tumors and serum of patients correlated with poor prognosis. In
EGFRL858R- and Kras4bG12D-induced lung cancer mice, which spontaneously develop lung tumors, IL10 levels
were significantly increased in lungs and serum. EGFRL858R/IL10-/- and Kras4bG12D/IL10-/- mice were generated,
and it was found that IL10 knockout inhibited lung tumor development and decreased levels of infiltrating M2
macrophages and tumor-promoting Treg lymphocytes. To elucidate the molecular mechanism, we showed that
EGF increased IL10 expression by enhancing nucleolin-mediated mRNA stability of IL10, thus activated IL10-dependent tumor-promotes JAK1/STAT3, Src, PI3K/Akt, and Erk signaling. Interestingly, IL10-induced
Figure 1. IL10 is over expressed in lung cancer. A. Upper panel: the representative image of IL10 expression from 6 independent groups. The human specimens were homogenized for Western blotting targeting IL10. Lower panel: The relevance between survival rate and IL10 level from 27 and 32 lung cancer patients with higher and unaltered IL10, respectively. B. Upper panel: IL10 level in human serum was measured by ELISA. Data were expressed as mean±s.e.m. Lower panel: The average serum concentration of IL10, 38.16 pg/ml, was set as the cutting edge to evaluate the correlation with prognosis. C, E. IL10 level in Kras4bG12D- and EGFRL858R-induced lung cancer mice. Lungs from mice
were prepared for RT-PCR, Western blotting and HE staining. D, F. IL10 levels in the serum from Kras4bG12D and EGFRL858R mice with or without lung cancer were measured by ELISA.
Figure 2. EGF induces IL10 expression, and IL10 increases proliferation. A(a). After treatment with EGF for 24h in serum-free media, mouse lung primary cells were harvested for RT-PCR and Western blotting targeting IL10. (b). The medium was collected for detecting IL10 by ELISA. B(a). IL10 secretion in EGF-treated H1299 cells (b). H1299 cells-expressed pGL2-IL10 promoter were harvested for luciferase reporter assay after EGF treatment. PGE2 and LPS treatments are positive
controls. Data were expressed as mean±s.e.m. C. After LY294002 treatment for 24h, H1299 cells were harvested for Western blotting to detect IL10. The result was quantified as lower panel. D. GFP-nucleolin (NCL)-overexpressed cells were treated with LY294002 for 24h, and harvested for Western blotting. Lower panel is the quantified result. Data were expressed as mean±s.e.m. (*p<0.05, **p<0.01). E. After treatment with LY294002 or GFP-NCL overexpression for 24h, cells were treated with actinomycin D for indicated time interval. Total RNA was extracted for RT-qPCR targeting IL10.
Figure 3. IL10-deficent mice failed to develop lung cancer induced by Kras4bG12D and
EGFRL858R. A. The mRNA level of IL10 in indicated transgenic mice. B. The protein level of
IL10 was detected by IHC. C. After doxycycline administration for 5 months, lungs of Kras4bG12D
mice were excised for counting surface tumor nodules. D. Lungs excised from
doxycycline-treated Kras4bG12D (5 months) and EGFRL858R (1 month) were
immunohistochemically stained by HE and the anti-CD163 antibody, and immunofluorescently stained by anti-CD4 and anti-FoxP3 antibodies. Brown and grey in slides of Kras4bG12D and
Figure 4. IL10 regulates EGFR expression through Src activation. A. The proliferation-related gene expression profile from microarray analysis in EGFRL858R and EGFRL858R/IL10-/-. B(a). Lung extracts
were homogenized for detecting EGFR expression by Western blotting, and the signal of EGFR was quantified. Data was expressed as mean±s.e.m. (***p<0.001). (c). The histological slides of lungs were stained for EGFR by IHC. C(a). Four scores to define IL10 expression detected by IHC in 192 lung cancer patients. (b). The IL10 expression in stages I&II and III&IV. (c). Negative (scores 0 and 1) and positive (scores 2 and 3) IL10 expression was correlated with survival period of patients. D. EGFR expression was detected in normal and tumor parts in 27 IL10-positive patients. (a). Upper panel is the
Figure 5. IL10 induces oncogenic signaling through inducing the interaction of IL10RA with phospho-Src. A. Effect of IL10 knockout on EGFRL858R and Kras4bG12D-induced signaling. Lungs
were homogenized for Western Blotting using the indicated antibody. Lower panel is the quantitative result. Data were expressed as mean±s.e.m. (*p<0.05, **p<0.01, ***p<0.001). B,C,D. After treatment with IL10 for 24 h, cell harvests were prepared for Western blotting with indicated antibodies. The lower panel of C and D is quantitative result. E. The immune-complex immunoprecipitated by the anti-Src antibody was analyzed by Western blotting using the indicated antibody. F. After transfection with wild-type HA-Src or kinase-dead HA-Src (aa 1-249), for 24h, H1299 cells with or without IL10 treatment were harvested for immunoprecipitation by the anti-HA antibody, and analyzed by Western blotting. Upper panel is the domain illustration of HA-Src. G. After transfection of pHA-Src and pHA-Src(1-249), and IL10 treatment, cellular extracts were prepared for Western blotting using indicated antibodies. Lower panel is the quantitative result for phospho-JAK1 and phospho-Akt. Data
were expressed as mean±s.e.m. *p<0.05. H,I. After transfection with indicated plasmids and IL10 treatment, cellular extracts were prepared for immunoprecipitation using the anti-HA or anti-GFP antibody. The immunoprecipitated complex was analyzed by Western blotting.
Please evaluate the performance of research, teaching or science and technology R&D and management Work: (To be completed
by Project Investigator or Head of Department/Center)
(1)是否達到延攬預期目標？
Has the expected goal of recruitment been achieved?
是。
徐宗溢博士在 101/8-104/6 發表了三篇 SCI 科學論文
1. Mol Pharmacol. 2012 Dec;82(6):1115-28. PMID: 22956772 2. Oncogene. 2015 Feb 12;34(7):826-37.PMID: 24632608
3. Oncotarget. 2015 Mar 30. [Epub ahead of print] PMID: 25909174
(2)研究或教學或科技研發與管理的方法、專業知識及進度如何？
What are the methods, professional knowledge, and progress of the research, teaching, or R&D and management work? 除了上述的成果, 本篇研究報告的內容也正在投稿到 SCI 國際期刊。
(3)受延攬人之研究或教學或科技研發與管理成果對該計畫(或貴單位)助益如何？
How have the research, teaching, or R&D and management results of the employed person given benefit to the project (or your unit)?
協助科技部計畫穩定的進行以及經費使用的管理。
(4)受延攬人於補助期間對貴單位或國內相關學術科技領域助益如何？
How has the employed person, during his or her term of employment, benefited your unit or the relevant domestic academic field?
找到新穎的肺癌發生機制與新的肺癌生物標記，有助於往後治療肺癌與預防肺癌研究策略的 發展。
(5)具體工作績效或研究或教學或科技研發與管理成果：
Please describe the specific work performance, or the results of research, teaching, or R&D and management work:
有效發展本實驗室肺癌轉譯醫學研究策略(細胞、動物、人體)。
(6)是否續聘受聘人? Will you continue hiring the employed person? □續聘Yes ■不續聘No ※ 此報告表篇幅以三～四頁為原則。This report form should be limited to 3-4 pages in principle.
※ 此表格可上延攬優秀人才成果報告繳交說明網頁下載。
