I.
多型性神經膠質母細胞瘤（glioblastoma multiforme, GBM）是成人最常見且最具侵略性的原發性惡性腦腫瘤。它的侵略性和對傳統療法的抵抗性使其成為一種預後不良的高復發率惡性疾病。GBM中的膠質母細胞瘤幹細胞（glioblastoma stem cells, GSC）被認為與該病的抗性與復發有關。甲狀腺素受器作用蛋白質-6（Thyroid Hormone Receptor Interactor 6, TRIP6）是一種焦點附著分子（focal adhesion molecule），主要功能包括細胞運動、抗凋亡信號、轉錄控制和其他細胞反應。此外，我們還發現TRIP6是維持神經幹細胞的重要蛋白質。我們已知TRIP6在GBM細胞中高量表達，並且TRIP6的表現量高者其患者的預後愈差。然而在於腫瘤生成的過程中，TRIP6調控GBM的分子機制仍有進一步探討的空間。Rac三磷酸鳥苷水解酶（Rac GTPases）是Rho GTPases家族的成員。其主要功能是調節肌動蛋白（actin）細胞骨架的重組、細胞遷移、存活和凋亡。此外，我們亦發現Rac可調控神經膠質母細胞瘤類癌幹細胞的維持。前述所提到的兩個蛋白都與肌動蛋白及細胞的移動有關，因此TRIP6和Rac是否能交互調控維持腫瘤幹細胞的特性，以及TRIP6-Rac信號通路能否作為一種新的神經膠質瘤藥物靶點有待進一步研究。我們建立以干擾RNA將TRIP6表現量削弱後再額外表現Rac GTPase家族中Rac2的細胞株，檢測其衍生之類癌幹細胞增殖的能力發現，Rac2 應該作用於TRIP6下游，協助TRIP6調控神經膠質母細胞瘤類癌幹細胞的生長。

II.
大腸直腸癌亦簡稱為大腸癌，據2017年的報導，為世界上第三大被診斷出的惡性腫瘤，並為全球第四大的致死癌症。在台灣也是罹病人數最高的癌症，目前主要的治療方法為手術治療，再來是化學療法，但是大腸直腸癌在晚期的治癒率非常低，且經常在治療後復發與轉移。近年，許多癌症研究學者認為，癌症的預後不良與高機率的復發，是因為在腫瘤當中有一小群細胞具有幹細胞特性，稱為癌幹細胞。它們具有著自我更新、分裂與分化成癌細胞的能力，並認為這些細胞是造成癌症復發及癌細胞對藥物抗藥性的原因。為了篩選出有效抑制大腸癌細胞，尤其是大腸癌幹細胞增生的化學合成藥物，我們針對一系列吲哚合成化合物做大腸癌幹細胞的存活試驗。其中Gavin 05、Garvin14兩支藥物對大腸癌細胞DLD-1所衍生之類癌幹細胞中有抑制效果。為了進一步確認該藥物對於脊椎動物的毒性，我們利用了斑馬魚進行了毒性測試，結果發現隨著給藥時間越長，幼魚的心搏與控制組相比有明顯的下降，但其外表並無明顯畸形。

I.
Glioblastoma is the most common and aggressive malignant primary brain tumor in adults. Its invasiveness and resistance to traditional therapies make it a highly recurrent malignant disease with poor prognosis. Glioblastoma stem cells （GSC） in GBM are considered to be associated with the resistance and the recurrence of the disease. TRIP6 is a focal adhesion molecule which main functions include cell movement, anti-apoptotic signaling, transcriptional control, and other cellular responses. In addition, we also found that TRIP6 is an important protein in maintaining neural stem cells. TRIP6 is also highly expressed in GBM cells. Moreover, the prognosis of GBM patients are poorer with the higher TRIP6 expression levels. However, the molecular mechanism of TRIP6 in regulating GBM progression needs to be explored more. Rac GTPases is a member of Rho GTPases family. They regulate the rearrangement of actin cytoskeleton, cell migration, survival and apoptosis. Furthermore, we found that Rac GTPases positively regulate GSC maintenance. Therefore, whether TRIP6 and Rac coordinately maintain cancer stem cell properties, and whether TRIP6-Rac signaling pathway can serve as a new drug target for GBM need to be further studied . To investigate this possibility, we established TRIP6 shRNA expressing cell lines with overexpressing Rac2, and examined the proliferation ability of these cells-derived tumorspheres. We found that the Rac2 may act downstream of TRIP6 and mediate TRIP6-regulated proliferation of GBM tumorsphere.

II.
Colorectal cancer, also known as colon cancer, is the third largest diagnosed cancer and the fourth leading cause of cancer death in the world according to the report in 2017. In Taiwan, it is also the most common malignant disease. At present, the main treatment is surgery followed by chemotherapy. However, the cure rate of colorectal cancer in the late stage is very low, and the late stage disease often accompanies with relapse and metastasis after treatment. In recent years, many cancer researchers believe that the poor prognosis and high recurrence of cancer are due to a small group of cells with stem cell characteristics, called cancer stem cells. They have the ability of self-renewal, division and differentiation into cancer cells. These cells may be the cause of cancer recurrence and drug resistance. In order to screen the chemosynthetic drugs that can effectively inhibit the proliferation of colorectal cancer cells, especially the colorectal cancer stem cells, we performed the drug screening of indole compounds for inhibition of colorectal cancer cell, DLD-1. The results showed that Gavin 05 and 14 can inhibit DLD-1 cells and its derived tumorspheres. We further examine the toxicity effects of these two compounds by zebrafish. These two compounds caused mild decreation of heart beat rate but no clear abnormaty of appearance.

1. Young, R.M., et al., Current trends in the surgical management and treatment of adult glioblastoma. Ann Transl Med, 2015. 3(9): p. 121.
2. Louis, D.N., et al., The 2007 WHO classification of tumours of the central nervous system. Acta Neuropathol, 2007. 114(2): p. 97-109.
3. Gallego, O., Nonsurgical treatment of recurrent glioblastoma. Curr Oncol, 2015. 22(4): p. e273-81.
4. Mao, H., et al., Deregulated signaling pathways in glioblastoma multiforme: molecular mechanisms and therapeutic targets. Cancer Invest, 2012. 30(1): p. 48-56.
5. Bao, S., et al., Glioma stem cells promote radioresistance by preferential activation of the DNA damage response. Nature, 2006. 444(7120): p. 756-60.
6. Johannessen, T.C., et al., Highly infiltrative brain tumours show reduced chemosensitivity associated with a stem cell-like phenotype. Neuropathol Appl Neurobiol, 2009. 35(4): p. 380-93.
7. Vlashi, E. and F. Pajonk, Cancer stem cells, cancer cell plasticity and radiation therapy. Semin Cancer Biol, 2015. 31: p. 28-35.
8. Vinogradov, S. and X. Wei, Cancer stem cells and drug resistance: the potential of nanomedicine. Nanomedicine (Lond), 2012. 7(4): p. 597-615.
9. Bonnet, D. and J.E. Dick, Human acute myeloid leukemia is organized as a hierarchy that originates from a primitive hematopoietic cell. Nat Med, 1997. 3(7): p. 730-7.
10. Al-Hajj, M., et al., Prospective identification of tumorigenic breast cancer cells. Proc Natl Acad Sci U S A, 2003. 100(7): p. 3983-8.
11. Kim, Y.S., et al., [Molecular markers of cancer stem cells verified in vivo]. Biomed Khim, 2016. 62(3): p. 228-38.
12. Scharenberg, C.W., M.A. Harkey, and B. Torok-Storb, The ABCG2 transporter is an efficient Hoechst 33342 efflux pump and is preferentially expressed by immature human hematopoietic progenitors. Blood, 2002. 99(2): p. 507-12.
13. Wang, Y. and T.D. Gilmore, Zyxin and paxillin proteins: focal adhesion plaque LIM domain proteins go nuclear. Biochim Biophys Acta, 2003. 1593(2-3): p. 115-20.
14. Lin, V.T. and F.T. Lin, TRIP6: an adaptor protein that regulates cell motility, antiapoptotic signaling and transcriptional activity. Cell Signal, 2011. 23(11): p. 1691-7.
15. Bach, I., The LIM domain: regulation by association. Mech Dev, 2000. 91(1-2): p. 5-17.
16. Lai, Y.J., et al., TRIP6 regulates neural stem cell maintenance in the postnatal mammalian subventricular zone. Dev Dyn, 2014. 243(9): p. 1130-42.
17. Zhao, W., et al., TRIP6 promotes cell proliferation in hepatocellular carcinoma via suppression of FOXO3a. Biochem Biophys Res Commun, 2017. 494(3-4): p. 594-601.
18. Gu, Y., et al., Hematopoietic cell regulation by Rac1 and Rac2 guanosine triphosphatases. Science, 2003. 302(5644): p. 445-9.
19. Troeger, A. and D.A. Williams, Hematopoietic-specific Rho GTPases Rac2 and RhoH and human blood disorders. Exp Cell Res, 2013. 319(15): p. 2375-83.
20. Jaffe, A.B. and A. Hall, Rho GTPases: biochemistry and biology. Annu Rev Cell Dev Biol, 2005. 21: p. 247-69.
21. Roberts, A.W., et al., Deficiency of the hematopoietic cell-specific Rho family GTPase Rac2 is characterized by abnormalities in neutrophil function and host defense. Immunity, 1999. 10(2): p. 183-96.
22. Itokowa, T., et al., Osteoclasts lacking Rac2 have defective chemotaxis and resorptive activity. Calcif Tissue Int, 2011. 88(1): p. 75-86.
23. Yang, F.C., et al., Rac2 stimulates Akt activation affecting BAD/Bcl-XL expression while mediating survival and actin function in primary mast cells. Immunity, 2000. 12(5): p. 557-68.
24. Yang, F.C., et al., Rac and Cdc42 GTPases control hematopoietic stem cell shape, adhesion, migration, and mobilization. Proc Natl Acad Sci U S A, 2001. 98(10): p. 5614-8.
25. Kalfa, T.A., et al., Rac GTPases regulate the morphology and deformability of the erythrocyte cytoskeleton. Blood, 2006. 108(12): p. 3637-45.
26. Lai, Y.J., et al., Small G protein Rac GTPases regulate the maintenance of glioblastoma stem-like cells in vitro and in vivo. Oncotarget, 2017. 8(11): p. 18031-18049.
27. Liu, Y., et al., RAC2 acts as a prognostic biomarker and promotes the progression of clear cell renal cell carcinoma. Int J Oncol, 2019. 55(3): p. 645-656.
28. Xia, P., et al., Down-regulation of RAC2 by small interfering RNA restrains the progression of osteosarcoma by suppressing the Wnt signaling pathway. Int J Biol Macromol, 2019. 137: p. 1221-1231.
29. Arnold, M., et al., Global patterns and trends in colorectal cancer incidence and mortality. Gut, 2017. 66(4): p. 683-691.
30. Kekelidze, M., et al., Colorectal cancer: current imaging methods and future perspectives for the diagnosis, staging and therapeutic response evaluation. World J Gastroenterol, 2013. 19(46): p. 8502-14.
31. Phua, L.C., et al., Non-invasive fecal metabonomic detection of colorectal cancer. Cancer Biol Ther, 2014. 15(4): p. 389-97.
32. Yeh, Y.-S., et al., Prognostic and Molecular Factors in Stage II Colorectal Cancer. Genomic Medicine, Biomarkers, and Health Sciences, 2011. 3(1): p. 2-8.
33. Mallard, B.W. and J. Tiralongo, Cancer stem cell marker glycosylation: Nature, function and significance. Glycoconj J, 2017. 34(4): p. 441-452.
34. Kazama, S., et al., Expression of the stem cell marker CD133 is related to tumor development in colorectal carcinogenesis. Asian J Surg, 2018. 41(3): p. 274-278.
35. Pop, M.G., Stem Cell Markers in Colon Cancer, in Basic Principles and Practice in Surgery. 2019, IntechOpen.
36. Sahlberg, S.H., et al., Evaluation of cancer stem cell markers CD133, CD44, CD24: association with AKT isoforms and radiation resistance in colon cancer cells. PLoS One, 2014. 9(4): p. e94621.
37. Senbanjo, L.T. and M.A. Chellaiah, CD44: A Multifunctional Cell Surface Adhesion Receptor Is a Regulator of Progression and Metastasis of Cancer Cells. Front Cell Dev Biol, 2017. 5: p. 18.
38. Spizzo, G., et al., EpCAM expression in primary tumour tissues and metastases: an immunohistochemical analysis. J Clin Pathol, 2011. 64(5): p. 415-20.
39. Simon, M., et al., Epithelial cell adhesion molecule-targeted drug delivery for cancer therapy. Expert Opin Drug Deliv, 2013. 10(4): p. 451-68.
40. Schnell, U., V. Cirulli, and B.N. Giepmans, EpCAM: structure and function in health and disease. Biochim Biophys Acta, 2013. 1828(8): p. 1989-2001.
41. Dana, H., CD166 as a Stem Cell Marker? A Potential Target for Therapy Colorectal Cancer? Journal of Stem Cell Research & Therapeutics, 2016. 1(6).
42. Ribeiro, K.B., et al., KRAS mutation associated with CD44/CD166 immunoexpression as predictors of worse outcome in metastatic colon cancer. Cancer Biomark, 2016. 16(4): p. 513-21.
43. Coppedè, F., et al., Genetic and epigenetic biomarkers for diagnosis, prognosis and treatment of colorectal cancer. World J Gastroenterol, 2014. 20(4): p. 943-56.
44. Nosrati, A., et al., Cancer stem cells CD133 and CD24 in colorectal cancers in Northern Iran. Gastroenterol Hepatol Bed Bench, 2016. 9(2): p. 132-9.
45. Aggarwal, B.B. and H. Ichikawa, Molecular targets and anticancer potential of indole-3-carbinol and its derivatives. Cell Cycle, 2005. 4(9): p. 1201-15.
46. Frydoonfar, H.R., D.R. McGrath, and A.D. Spigelman, Inhibition of proliferation of a colon cancer cell line by indole-3-carbinol. Colorectal Dis, 2002. 4(3): p. 205-207.