臺灣博碩士論文加值系統

靈芝蛋白 (GMI) 是一重組小孢子靈芝的真菌免疫調節蛋白，它具有調節免疫、調節血糖、抗發炎、抗氧化和預防癌症功效是廣為人知的事，不過其防癌機制尚不是很明確。本研究中，我們發現GMI可抑制異種移植動物的腫瘤生長。由前人的研究可以得知，表皮生長因子受體 (EGFR)在非小細胞肺癌(NSCLC)中經常過度地表現並擁有很大比例的突變進而導致病人的存活率降低。在這篇研究中，我們進一步地去探討GMI在EGFR上的調控與肺癌細胞存活率中所扮演的角色。我們發現GMI能夠透過調控EGFR表現量降低和抑制EGFR自體磷酸化，有效地消除EGFR活性並誘導細胞凋亡，在機制的探討上，我們發現GMI依賴EGFR酪胺酸激酶之活性以誘導蛋白酶體相關的EGFR降解作用。綜合以上所述，我們的研究結果顯示GMI可以促進EGFR降解作用並誘導細胞凋亡，因此GMI具有成為對抗肺癌之蛋白質藥物的潛力。

GMI is a recombinant fungal immunomodulatory protein cloned from Ganoderma microsporum. It exerts well-known modulating immune function, regulation of blood sugar, anti-inflammatory, anti-oxidation and cancer efficacy. However, the mechanisms of anti-cancer is not defined yet. Here, we demonstrated that GMI inhibits tumor growth in a xenograft animal model. Epidermal growth factor receptor (EGFR) is frequently overexpression in non-small cell lung cancer (NSCLC), reducing survival of patients and NSCLC genome conceals a large proportion of EGFR mutation. We investigated the role of GMI in EGFR regulation and viability of lung cancer cells. We found that GMI effectively induced apoptosis as well as abolished EGFR activity and its downstream signaling pathways via downregulation of EGFR expression and suppression of EGFR autophosphorylation. Mechanistically, we demonstrated that GMI-induced proteasome-mediated EGFR degradation depended on the tyrosine kinase activity of EGFR. In summary, our results suggest that GMI promotes EGFR degradation and induces apoptosis, indicating that GMI may be a potential protein drug against lung cancer.

Abbreviations I
Abstract II
中文摘要 III
Content IV
Introduction - 1 -
Materials and methods - 4 -
Cell lines - 4 -
Preparation of the recombinant GMI protein - 4 -
Reagents and antibodies - 5 -
Tumor xenograft animal model - 5 -
Cell viability assay - 6 -
Cell cycle analysis - 6 -
Apoptosis analysis - 7 -
Western blot analysis - 7 -
Immunoprecipitation - 7 -
In vitro ubiquitination assay - 8 -
Statistical analysis - 8 -
Results - 9 -
1. GMI suppresses tumorigenesis and reduces EGFR expression in LLC1-bearing mouse model - 9 -
2. GMI inhibits NSCLC cells proliferation - 9 -
3. GMI induces apoptosis in EGFR mutated H1975 and PC9 cells - 10 -
4. GMI suppresses downstream signaling and downregulates expression of EGFR - 10 -
5. GMI modulates EGFR expression by accelerating ubiquitin-dependent proteasomal degradation - 10 -
6. GMI-induced EGFR degradation depends on kinase activity - 11 -
7. GMI is effective in gefitinib-resistant cells - 12 -
Discussion - 13 -
References - 16 -
Figure legends - 27 -
Figure 1. GMI inhibits tumor progression and reduces EGFR protein levels in LLC1 cell-bearing male C57BL6 mice. - 27 -
Figure 2. GMI inhibits cell viability and colony formation. - 30 -
Figure 3. GMI induces cell apoptosis. - 32 -
Figure 4. GMI blocks EGFR signaling. - 34 -
Figure 5. GMI downregulates EGFR expression - 36 -
Figure 6. GMI induces EGFR degradation. - 38 -
Figure 7. GMI-induced EGFR degradation depends on kinase activity. - 41 -
Figure 8. GMI affects gefitinib-resistant cells. - 43 -

Amann, J., S. Kalyankrishna, P. P. Massion, J. E. Ohm, L. Girard, H. Shigematsu, M. Peyton, D. Juroske, Y. Huang, J. Stuart Salmon, Y. H. Kim, J. R. Pollack, K. Yanagisawa, A. Gazdar, J. D. Minna, J. M. Kurie and D. P. Carbone (2005). "Aberrant epidermal growth factor receptor signaling and enhanced sensitivity to EGFR inhibitors in lung cancer." Cancer Res 65(1): 226-235.
Arfaoui, A., L. Kriaa, N. Znaidi, S. Gritli, H. Bouacha, R. Zermani and S. Rammeh (2014). "Over-expression of EGFR is closely correlated to poor prognosis in Tunisian patients with non-small cell lung adenocarcinoma." J Immunoassay Immunochem 35(3): 256-268.
Bethune, G., D. Bethune, N. Ridgway and Z. Xu (2010). "Epidermal growth factor receptor (EGFR) in lung cancer: an overview and update." J Thorac Dis 2(1): 48-51.
Boh, B., M. Berovic, J. Zhang and L. Zhi-Bin (2007). "Ganoderma lucidum and its pharmaceutically active compounds." Biotechnol Annu Rev 13: 265-301.
Castanon, E., C. Rolfo, D. Vinal, I. Lopez, J. P. Fusco, M. Santisteban, P. Martin, L. Zubiri, J. I. Echeveste and I. Gil-Bazo (2015). "Impact of epidermal growth factor receptor (EGFR) activating mutations and their targeted treatment in the prognosis of stage IV non-small cell lung cancer (NSCLC) patients harboring liver metastasis." J Transl Med 13: 257.
Chiu, L. Y., M. E. Hu, T. Y. Yang, I. L. Hsin, J. L. Ko, K. J. Tsai and G. T. Sheu (2015). "Immunomodulatory Protein from Ganoderma microsporum Induces Pro-Death Autophagy through Akt-mTOR-p70S6K Pathway Inhibition in Multidrug Resistant Lung Cancer Cells." PLoS One 10(5): e0125774.
Guo, L., C. J. Kozlosky, L. H. Ericsson, T. O. Daniel, D. P. Cerretti and R. S. Johnson (2003). "Studies of ligand-induced site-specific phosphorylation of epidermal growth factor receptor." J Am Soc Mass Spectrom 14(9): 1022-1031.
Hirsch, F. R., M. Varella-Garcia, P. A. Bunn, Jr., M. V. Di Maria, R. Veve, R. M. Bremmes, A. E. Baron, C. Zeng and W. A. Franklin (2003). "Epidermal growth factor receptor in non-small-cell lung carcinomas: correlation between gene copy number and protein expression and impact on prognosis." J Clin Oncol 21(20): 3798-3807.
Hsiao, A. J., L. H. Chen and T. H. Lu (2015). "Ten leading causes of death in Taiwan: A comparison of two grouping lists." J Formos Med Assoc 114(8): 679-680.
Hsin, I. L., C. C. Ou, M. F. Wu, M. S. Jan, Y. M. Hsiao, C. H. Lin and J. L. Ko (2015). "GMI, an Immunomodulatory Protein from Ganoderma microsporum, Potentiates Cisplatin-Induced Apoptosis via Autophagy in Lung Cancer Cells." Mol Pharm 12(5): 1534-1543.
Hsin, I. L., C. C. Ou, T. C. Wu, M. S. Jan, M. F. Wu, L. Y. Chiu, K. H. Lue and J. L. Ko (2011). "GMI, an immunomodulatory protein from Ganoderma microsporum, induces autophagy in non-small cell lung cancer cells." Autophagy 7(8): 873-882.
Hsu, H. C., C. I. Hsu, R. H. Lin, C. L. Kao and J. Y. Lin (1997). "Fip-vvo, a new fungal immunomodulatory protein isolated from Volvariella volvacea." Biochem J 323 ( Pt 2): 557-565.
Huang, K. Y., S. H. Kao, W. L. Wang, C. Y. Chen, T. H. Hsiao, S. B. Salunke, J. J. Chen, K. Y. Su, S. C. Yang, T. M. Hong, C. S. Chen and P. C. Yang (2016). "Small Molecule T315 Promotes Casitas B-Lineage Lymphoma-Dependent Degradation of Epidermal Growth Factor Receptor via Y1045 Autophosphorylation." Am J Respir Crit Care Med 193(7): 753-766.
Inamura, K., H. Ninomiya, Y. Ishikawa and O. Matsubara (2010). "Is the epidermal growth factor receptor status in lung cancers reflected in clinicopathologic features?" Arch Pathol Lab Med 134(1): 66-72.
Jackman, D. M., B. Y. Yeap, L. V. Sequist, N. Lindeman, A. J. Holmes, V. A. Joshi, D. W. Bell, M. S. Huberman, B. Halmos, M. S. Rabin, D. A. Haber, T. J. Lynch, M. Meyerson, B. E. Johnson and P. A. Janne (2006). "Exon 19 deletion mutations of epidermal growth factor receptor are associated with prolonged survival in non-small cell lung cancer patients treated with gefitinib or erlotinib." Clin Cancer Res 12(13): 3908-3914.
Jeurink, P. V., C. L. Noguera, H. F. Savelkoul and H. J. Wichers (2008). "Immunomodulatory capacity of fungal proteins on the cytokine production of human peripheral blood mononuclear cells." Int Immunopharmacol 8(8): 1124-1133.
Jinn, T. R., C. M. Wu, W. C. Tu, J. L. Ko and J. T. Tzen (2006). "Functional expression of FIP-gts, a fungal immunomodulatory protein from Ganoderma tsugae in Sf21 insect cells." Biosci Biotechnol Biochem 70(11): 2627-2634.
Khan, E. M., R. Lanir, A. R. Danielson and T. Goldkorn (2008). "Epidermal growth factor receptor exposed to cigarette smoke is aberrantly activated and undergoes perinuclear trafficking." FASEB J 22(3): 910-917.
Ko, J. L., C. I. Hsu, R. H. Lin, C. L. Kao and J. Y. Lin (1995). "A new fungal immunomodulatory protein, FIP-fve isolated from the edible mushroom, Flammulina velutipes and its complete amino acid sequence." Eur J Biochem 228(2): 244-249.
Kris, M. G., R. B. Natale, R. S. Herbst, T. J. Lynch, Jr., D. Prager, C. P. Belani, J. H. Schiller, K. Kelly, H. Spiridonidis, A. Sandler, K. S. Albain, D. Cella, M. K. Wolf, S. D. Averbuch, J. J. Ochs and A. C. Kay (2003). "Efficacy of gefitinib, an inhibitor of the epidermal growth factor receptor tyrosine kinase, in symptomatic patients with non-small cell lung cancer: a randomized trial." JAMA 290(16): 2149-2158.
Levkowitz, G., H. Waterman, S. A. Ettenberg, M. Katz, A. Y. Tsygankov, I. Alroy, S. Lavi, K. Iwai, Y. Reiss, A. Ciechanover, S. Lipkowitz and Y. Yarden (1999). "Ubiquitin ligase activity and tyrosine phosphorylation underlie suppression of growth factor signaling by c-Cbl/Sli-1." Mol Cell 4(6): 1029-1040.
Liao, C. H., Y. M. Hsiao, C. P. Hsu, M. Y. Lin, J. C. Wang, Y. L. Huang and J. L. Ko (2006). "Transcriptionally mediated inhibition of telomerase of fungal immunomodulatory protein from Ganoderma tsugae in A549 human lung adenocarcinoma cell line." Mol Carcinog 45(4): 220-229.
Mok, T. S., Y. L. Wu, S. Thongprasert, C. H. Yang, D. T. Chu, N. Saijo, P. Sunpaweravong, B. Han, B. Margono, Y. Ichinose, Y. Nishiwaki, Y. Ohe, J. J. Yang, B. Chewaskulyong, H. Jiang, E. L. Duffield, C. L. Watkins, A. A. Armour and M. Fukuoka (2009). "Gefitinib or carboplatin-paclitaxel in pulmonary adenocarcinoma." N Engl J Med 361(10): 947-957.
NICE (2011). The Diagnosis and Treatment of Lung Cancer (Update). Cardiff (UK).
Amann, J., S. Kalyankrishna, P. P. Massion, J. E. Ohm, L. Girard, H. Shigematsu, M. Peyton, D. Juroske, Y. Huang, J. Stuart Salmon, Y. H. Kim, J. R. Pollack, K. Yanagisawa, A. Gazdar, J. D. Minna, J. M. Kurie and D. P. Carbone (2005). "Aberrant epidermal growth factor receptor signaling and enhanced sensitivity to EGFR inhibitors in lung cancer." Cancer Res 65(1): 226-235.
Arfaoui, A., L. Kriaa, N. Znaidi, S. Gritli, H. Bouacha, R. Zermani and S. Rammeh (2014). "Over-expression of EGFR is closely correlated to poor prognosis in Tunisian patients with non-small cell lung adenocarcinoma." J Immunoassay Immunochem 35(3): 256-268.
Bethune, G., D. Bethune, N. Ridgway and Z. Xu (2010). "Epidermal growth factor receptor (EGFR) in lung cancer: an overview and update." J Thorac Dis 2(1): 48-51.
Boh, B., M. Berovic, J. Zhang and L. Zhi-Bin (2007). "Ganoderma lucidum and its pharmaceutically active compounds." Biotechnol Annu Rev 13: 265-301.
Castanon, E., C. Rolfo, D. Vinal, I. Lopez, J. P. Fusco, M. Santisteban, P. Martin, L. Zubiri, J. I. Echeveste and I. Gil-Bazo (2015). "Impact of epidermal growth factor receptor (EGFR) activating mutations and their targeted treatment in the prognosis of stage IV non-small cell lung cancer (NSCLC) patients harboring liver metastasis." J Transl Med 13: 257.
Chen, G., P. Kronenberger, E. Teugels, I. A. Umelo and J. De Greve (2012). "Targeting the epidermal growth factor receptor in non-small cell lung cancer cells: the effect of combining RNA interference with tyrosine kinase inhibitors or cetuximab." BMC Med 10: 28.
Chiu, L. Y., M. E. Hu, T. Y. Yang, I. L. Hsin, J. L. Ko, K. J. Tsai and G. T. Sheu (2015).
"Immunomodulatory Protein from Ganoderma microsporum Induces Pro-Death Autophagy through Akt-mTOR-p70S6K Pathway Inhibition in Multidrug Resistant Lung Cancer Cells." PLoS One 10(5): e0125774.
Guo, L., C. J. Kozlosky, L. H. Ericsson, T. O. Daniel, D. P. Cerretti and R. S. Johnson (2003). "Studies of ligand-induced site-specific phosphorylation of epidermal growth factor receptor." J Am Soc Mass Spectrom 14(9): 1022-1031.
Hirsch, F. R., M. Varella-Garcia, P. A. Bunn, Jr., M. V. Di Maria, R. Veve, R. M. Bremmes, A. E. Baron, C. Zeng and W. A. Franklin (2003). "Epidermal growth factor receptor in non-small-cell lung carcinomas: correlation between gene copy number and protein expression and impact on prognosis." J Clin Oncol 21(20): 3798-3807.
Hsiao, A. J., L. H. Chen and T. H. Lu (2015). "Ten leading causes of death in Taiwan: A comparison of two grouping lists." J Formos Med Assoc 114(8): 679-680.
Hsin, I. L., C. C. Ou, M. F. Wu, M. S. Jan, Y. M. Hsiao, C. H. Lin and J. L. Ko (2015). "GMI, an Immunomodulatory Protein from Ganoderma microsporum, Potentiates Cisplatin-Induced Apoptosis via Autophagy in Lung Cancer Cells." Mol Pharm 12(5): 1534-1543.
Hsin, I. L., C. C. Ou, T. C. Wu, M. S. Jan, M. F. Wu, L. Y. Chiu, K. H. Lue and J. L. Ko (2011). "GMI, an immunomodulatory protein from Ganoderma microsporum, induces autophagy in non-small cell lung cancer cells." Autophagy 7(8): 873-882.
Hsu, H. C., C. I. Hsu, R. H. Lin, C. L. Kao and J. Y. Lin (1997). "Fip-vvo, a new fungal immunomodulatory protein isolated from Volvariella volvacea." Biochem J 323 ( Pt 2): 557-565.
Huang, K. Y., S. H. Kao, W. L. Wang, C. Y. Chen, T. H. Hsiao, S. B. Salunke, J. J. Chen, K. Y. Su, S. C. Yang, T. M. Hong, C. S. Chen and P. C. Yang (2016). "Small Molecule T315 Promotes Casitas B-Lineage Lymphoma-Dependent Degradation of Epidermal Growth Factor Receptor via Y1045 Autophosphorylation." Am J Respir Crit Care Med 193(7): 753-766.
Inamura, K., H. Ninomiya, Y. Ishikawa and O. Matsubara (2010). "Is the epidermal growth factor receptor status in lung cancers reflected in clinicopathologic features?" Arch Pathol Lab Med 134(1): 66-72.
Jackman, D. M., B. Y. Yeap, L. V. Sequist, N. Lindeman, A. J. Holmes, V. A. Joshi, D. W. Bell, M. S. Huberman, B. Halmos, M. S. Rabin, D. A. Haber, T. J. Lynch, M. Meyerson, B. E. Johnson and P. A. Janne (2006). "Exon 19 deletion mutations of epidermal growth factor receptor are associated with prolonged survival in non-small cell lung cancer patients treated with gefitinib or erlotinib." Clin Cancer Res 12(13): 3908-3914.
Jeurink, P. V., C. L. Noguera, H. F. Savelkoul and H. J. Wichers (2008). "Immunomodulatory capacity of fungal proteins on the cytokine production of human peripheral blood mononuclear cells." Int Immunopharmacol 8(8): 1124-1133.
Jinn, T. R., C. M. Wu, W. C. Tu, J. L. Ko and J. T. Tzen (2006). "Functional expression of FIP-gts, a fungal immunomodulatory protein from Ganoderma tsugae in Sf21 insect cells." Biosci Biotechnol Biochem 70(11): 2627-2634.
Khan, E. M., R. Lanir, A. R. Danielson and T. Goldkorn (2008). "Epidermal growth factor receptor exposed to cigarette smoke is aberrantly activated and undergoes perinuclear trafficking." FASEB J 22(3): 910-917.
Ko, J. L., C. I. Hsu, R. H. Lin, C. L. Kao and J. Y. Lin (1995). "A new fungal immunomodulatory protein, FIP-fve isolated from the edible mushroom, Flammulina velutipes and its complete amino acid sequence." Eur J Biochem 228(2): 244-249.
Kris, M. G., R. B. Natale, R. S. Herbst, T. J. Lynch, Jr., D. Prager, C. P. Belani, J. H. Schiller, K. Kelly, H. Spiridonidis, A. Sandler, K. S. Albain, D. Cella, M. K. Wolf, S. D. Averbuch, J. J. Ochs and A. C. Kay (2003). "Efficacy of gefitinib, an inhibitor of the epidermal growth factor receptor tyrosine kinase, in symptomatic patients with non-small cell lung cancer: a randomized trial." JAMA 290(16): 2149-2158.
Levkowitz, G., H. Waterman, S. A. Ettenberg, M. Katz, A. Y. Tsygankov, I. Alroy, S. Lavi, K. Iwai, Y. Reiss, A. Ciechanover, S. Lipkowitz and Y. Yarden (1999). "Ubiquitin ligase activity and tyrosine phosphorylation underlie suppression of growth factor signaling by c-Cbl/Sli-1." Mol Cell 4(6): 1029-1040.
Liao, C. H., Y. M. Hsiao, C. P. Hsu, M. Y. Lin, J. C. Wang, Y. L. Huang and J. L. Ko (2006). "Transcriptionally mediated inhibition of telomerase of fungal immunomodulatory protein from Ganoderma tsugae in A549 human lung adenocarcinoma cell line." Mol Carcinog 45(4): 220-229.
Lin, T. Y. and H. Y. Hsu (2016). "Ling Zhi-8 reduces lung cancer mobility and metastasis through disruption of focal adhesion and induction of MDM2-mediated Slug degradation." Cancer Lett 375(2): 340-348.
Mok, T. S., Y. L. Wu, S. Thongprasert, C. H. Yang, D. T. Chu, N. Saijo, P. Sunpaweravong, B. Han, B. Margono, Y. Ichinose, Y. Nishiwaki, Y. Ohe, J. J. Yang, B. Chewaskulyong, H. Jiang, E. L. Duffield, C. L. Watkins, A. A. Armour and M. Fukuoka (2009). "Gefitinib or carboplatin-paclitaxel in pulmonary adenocarcinoma." N Engl J Med 361(10): 947-957.
Ohsaki, Y., S. Tanno, Y. Fujita, E. Toyoshima, S. Fujiuchi, Y. Nishigaki, S. Ishida, A. Nagase, N. Miyokawa, S. Hirata and K. Kikuchi (2000). "Epidermal growth factor receptor expression correlates with poor prognosis in non-small cell lung cancer patients with p53 overexpression." Oncol Rep 7(3): 603-607.
Paez, J. G., P. A. Janne, J. C. Lee, S. Tracy, H. Greulich, S. Gabriel, P. Herman, F. J. Kaye, N. Lindeman, T. J. Boggon, K. Naoki, H. Sasaki, Y. Fujii, M. J. Eck, W. R. Sellers, B. E. Johnson and M. Meyerson (2004). "EGFR mutations in lung cancer: correlation with clinical response to gefitinib therapy." Science 304(5676): 1497-1500.
Pao, W. and N. Girard (2011). "New driver mutations in non-small-cell lung cancer." Lancet Oncol 12(2): 175-180.
Pao, W. and K. E. Hutchinson (2012). "Chipping away at the lung cancer genome." Nat Med 18(3): 349-351.
Pao, W., V. A. Miller, K. A. Politi, G. J. Riely, R. Somwar, M. F. Zakowski, M. G. Kris and H. Varmus (2005). "Acquired resistance of lung adenocarcinomas to gefitinib or erlotinib is associated with a second mutation in the EGFR kinase domain." PLoS Med 2(3): e73.
Pirazzoli, V., D. Ayeni, C. B. Meador, B. G. Sanganahalli, F. Hyder, E. de Stanchina, S. B. Goldberg, W. Pao and K. Politi (2016). "Afatinib plus Cetuximab Delays Resistance Compared to Single-Agent Erlotinib or Afatinib in Mouse Models of TKI-Naive EGFR L858R-Induced Lung Adenocarcinoma." Clin Cancer Res 22(2): 426-435.
Riely, G. J., W. Pao, D. Pham, A. R. Li, N. Rizvi, E. S. Venkatraman, M. F. Zakowski, M. G. Kris, M. Ladanyi and V. A. Miller (2006). "Clinical course of patients with non-small cell lung cancer and epidermal growth factor receptor exon 19 and exon 21 mutations treated with gefitinib or erlotinib." Clin Cancer Res 12(3 Pt 1): 839-844.
Saito, T., S. Okada, K. Ohshima, E. Yamada, M. Sato, Y. Uehara, H. Shimizu, J. E. Pessin and M. Mori (2004). "Differential activation of epidermal growth factor (EGF) receptor downstream signaling pathways by betacellulin and EGF." Endocrinology 145(9): 4232-4243.
Schlessinger, J. (2002). "Ligand-induced, receptor-mediated dimerization and activation of EGF receptor." Cell 110(6): 669-672.
Sharma, S. V., D. W. Bell, J. Settleman and D. A. Haber (2007). "Epidermal growth factor receptor mutations in lung cancer." Nat Rev Cancer 7(3): 169-181.
Shepherd, F. A., J. Rodrigues Pereira, T. Ciuleanu, E. H. Tan, V. Hirsh, S. Thongprasert, D. Campos, S. Maoleekoonpiroj, M. Smylie, R. Martins, M. van Kooten, M. Dediu, B. Findlay, D. Tu, D. Johnston, A. Bezjak, G. Clark, P. Santabarbara, L. Seymour and G. National Cancer Institute of Canada Clinical Trials (2005). "Erlotinib in previously treated non-small-cell lung cancer." N Engl J Med 353(2): 123-132.
Siegel, R. L., K. D. Miller and A. Jemal (2015). "Cancer statistics, 2015." CA Cancer J Clin 65(1): 5-29.
Sigismund, S., V. Algisi, G. Nappo, A. Conte, R. Pascolutti, A. Cuomo, T. Bonaldi, E. Argenzio, L. G. Verhoef, E. Maspero, F. Bianchi, F. Capuani, A. Ciliberto, S. Polo and P. P. Di Fiore (2013). "Threshold-controlled ubiquitination of the EGFR directs receptor fate." EMBO J 32(15): 2140-2157.
Veale, D., N. Kerr, G. J. Gibson, P. J. Kelly and A. L. Harris (1993). "The relationship of quantitative epidermal growth factor receptor expression in non-small cell lung cancer to long term survival." Br J Cancer 68(1): 162-165.
Volm, M., W. Rittgen and P. Drings (1998). "Prognostic value of ERBB-1, VEGF, cyclin A, FOS, JUN and MYC in patients with squamous cell lung carcinomas." Br J Cancer 77(4): 663-669.
Wang, P. H., C. I. Hsu, S. C. Tang, Y. L. Huang, J. Y. Lin and J. L. Ko (2004). "Fungal immunomodulatory protein from Flammulina velutipes induces interferon-gamma production through p38 mitogen-activated protein kinase signaling pathway." J Agric Food Chem 52(9): 2721-2725.
Wu, C. T., T. Y. Lin, H. Y. Hsu, F. Sheu, C. M. Ho and E. I. Chen (2011). "Ling Zhi-8 mediates p53-dependent growth arrest of lung cancer cells proliferation via the ribosomal protein S7-MDM2-p53 pathway." Carcinogenesis 32(12): 1890-1896.
Ohsaki, Y., S. Tanno, Y. Fujita, E. Toyoshima, S. Fujiuchi, Y. Nishigaki, S. Ishida, A. Nagase, N. Miyokawa, S. Hirata and K. Kikuchi (2000). "Epidermal growth factor receptor expression correlates with poor prognosis in non-small cell lung cancer patients with p53 overexpression." Oncol Rep 7(3): 603-607.
Paez, J. G., P. A. Janne, J. C. Lee, S. Tracy, H. Greulich, S. Gabriel, P. Herman, F. J. Kaye, N. Lindeman, T. J. Boggon, K. Naoki, H. Sasaki, Y. Fujii, M. J. Eck, W. R. Sellers, B. E. Johnson and M. Meyerson (2004). "EGFR mutations in lung cancer: correlation with clinical response to gefitinib therapy." Science 304(5676): 1497-1500.
Pao, W. and N. Girard (2011). "New driver mutations in non-small-cell lung cancer." Lancet Oncol 12(2): 175-180.
Pao, W. and K. E. Hutchinson (2012). "Chipping away at the lung cancer genome." Nat Med 18(3): 349-351.
Pao, W., V. A. Miller, K. A. Politi, G. J. Riely, R. Somwar, M. F. Zakowski, M. G. Kris and H. Varmus (2005). "Acquired resistance of lung adenocarcinomas to gefitinib or erlotinib is associated with a second mutation in the EGFR kinase domain." PLoS Med 2(3): e73.
Pirazzoli, V., D. Ayeni, C. B. Meador, B. G. Sanganahalli, F. Hyder, E. de Stanchina, S. B. Goldberg, W. Pao and K. Politi (2016). "Afatinib plus Cetuximab Delays Resistance Compared to Single-Agent Erlotinib or Afatinib in Mouse Models of TKI-Naive EGFR L858R-Induced Lung Adenocarcinoma." Clin Cancer Res 22(2): 426-435.
Riely, G. J., W. Pao, D. Pham, A. R. Li, N. Rizvi, E. S. Venkatraman, M. F. Zakowski, M. G. Kris, M. Ladanyi and V. A. Miller (2006). "Clinical course of patients with non-small cell lung cancer and epidermal growth factor receptor exon 19 and exon 21 mutations treated with gefitinib or erlotinib." Clin Cancer Res 12(3 Pt 1): 839-844.
Saito, T., S. Okada, K. Ohshima, E. Yamada, M. Sato, Y. Uehara, H. Shimizu, J. E. Pessin and M. Mori (2004). "Differential activation of epidermal growth factor (EGF) receptor downstream signaling pathways by betacellulin and EGF." Endocrinology 145(9): 4232-4243.
Schlessinger, J. (2002). "Ligand-induced, receptor-mediated dimerization and activation of EGF receptor." Cell 110(6): 669-672.
Sharma, S. V., D. W. Bell, J. Settleman and D. A. Haber (2007). "Epidermal growth factor receptor mutations in lung cancer." Nat Rev Cancer 7(3): 169-181.
Shepherd, F. A., J. Rodrigues Pereira, T. Ciuleanu, E. H. Tan, V. Hirsh, S. Thongprasert, D. Campos, S. Maoleekoonpiroj, M. Smylie, R. Martins, M. van Kooten, M. Dediu, B. Findlay, D. Tu, D. Johnston, A. Bezjak, G. Clark, P. Santabarbara, L. Seymour and G. National Cancer Institute of Canada Clinical Trials (2005). "Erlotinib in previously treated non-small-cell lung cancer." N Engl J Med 353(2): 123-132.
Siegel, R. L., K. D. Miller and A. Jemal (2015). "Cancer statistics, 2015." CA Cancer J Clin 65(1): 5-29.
Sigismund, S., V. Algisi, G. Nappo, A. Conte, R. Pascolutti, A. Cuomo, T. Bonaldi, E. Argenzio, L. G. Verhoef, E. Maspero, F. Bianchi, F. Capuani, A. Ciliberto, S. Polo and P. P. Di Fiore (2013). "Threshold-controlled ubiquitination of the EGFR directs receptor fate." EMBO J 32(15): 2140-2157.
Veale, D., N. Kerr, G. J. Gibson, P. J. Kelly and A. L. Harris (1993). "The relationship of quantitative epidermal growth factor receptor expression in non-small cell lung cancer to long term survival." Br J Cancer 68(1): 162-165.
Volm, M., W. Rittgen and P. Drings (1998). "Prognostic value of ERBB-1, VEGF, cyclin A, FOS, JUN and MYC in patients with squamous cell lung carcinomas." Br J Cancer 77(4): 663-669.
Wang, P. H., C. I. Hsu, S. C. Tang, Y. L. Huang, J. Y. Lin and J. L. Ko (2004). "Fungal immunomodulatory protein from Flammulina velutipes induces interferon-gamma production through p38 mitogen-activated protein kinase signaling pathway." J Agric Food Chem 52(9): 2721-2725.
Wasser S.P., Coates P., Blackman M., Cragg G., Levine M., Moss J., White J. (2005) "Encyclopedia of Dietary Supplements.New York: Marcel Dekker." Reishi or Lingzhi (Ganoderma lucidum) pp. 680–90.
Wu, C. T., T. Y. Lin, H. Y. Hsu, F. Sheu, C. M. Ho and E. I. Chen (2011). "Ling Zhi-8 mediates p53-dependent growth arrest of lung cancer cells proliferation via the ribosomal protein S7-MDM2-p53 pathway." Carcinogenesis 32(12): 1890-1896.