Abstract ▼

● Research objectives: The goal of this project is to elucidate the molecular mechanism of cancer progression and cancer drug resistance of refractory ovarian and cervical cancers regulated by DRAK1/2 kinase proteins and to examine the DRAK1 protein expression in patient cancer tissue samples and or

● Research objectives: The goal of this project is to elucidate the molecular mechanism of cancer progression and cancer drug resistance of refractory ovarian and cervical cancers regulated by DRAK1/2 kinase proteins and to examine the DRAK1 protein expression in patient cancer tissue samples and organoid originated from patient cancer cells. In addition, we verify the efficacy of DRAK1 peptide and DRAK1/2 kinase inhibitor in ovarian and cervical cancers in order to develop th personalized diagnostics and therapeutics.

● Experimental Methods

▷ Examine the molecular mechanism of DRAK1 related to regulation of cancer metastasis and drug resistance using cervical cancer cell lines
▷ Examine the biological function of DRAK1 in regulation of cancer metastasis and drug resistance using lung metastasis and xenograft in vivo animal model
▷ Investigate in vitro and in vivo mechanism of cancer metastasis and drug resistance by targeting TRAF6 protein
▷ Determine the efficacy of DRAK1 kinase inhibitor in prevention of tumorigenesis of ovarian cancer cells through in vitro and in vivo study
▷ Analyze DRAK1 protein expression and explore clinical significance and relevance by performing immunohistochemisty of ovarian cancer patient tissue samples
▷ Establish in vitro tumor microenvironment model by isolating cellular components from malignant ascites of patient-derived ovarian cancer spheroids
▷ Establish organoid originated from patients and perform single cell RNA sequencing to evaluate DRAK1 and discover new molecular targets as a noble moleuclar biomarker in ovarian cancer

● Result

Part I: Investigate the role of DRAK1 in regulation of tumorigenesis and metastasis of cervical cancer cells and the molecular mechanism by targeting TRAF6 E3 ligase
▷ DRAK1 interacts with TRAF6 and destabilize TRAF6 protein through blocking its autoubiquitination and downregulates TAK1/p38/MAPK/NF-κB immune signaling pathway
▷ We found that cervical cancer tumorigenesis and lung metastasis was regulated by DRAK1 protein expression level using xenograft animal model
▷ We determined that DRAK1 protein was highly expressed whereas TRAF6 protein expression was downregulated in cervical cancer patient primary tumor tissues. However, TRAF6 expression was upregulated as DRAK1 expression was low in metatstitic cervical cancer patient tumor tissues.
Part II: Investigate the role of DRAK1 in regulation of paclitaxel resistance and molecular mechanism of DRAK1 protein expression regulation in paclitaxel-resistant cervical cancer cells
▷ DRAK1 protein expression was suppressed in paclitaxel-resistant cervical cancer cells and resulted in upregulation of TRAF6 protein expression and TRAF6-mediated TAK1/p38/MAPK/NF-κB signaling cascades.
▷ DRAK1 protein was downregulated by CUL3/SPOP E3 ligase protein through ubiquitination in paclitaxel-resistant cervical cancer cells.
▷ DRAK1 protein was higly expressed in chemoresistnat cervical cancer tissues in comparison with chemosensitive cervical cancer tissues and inversely correlated with TRAF6 protein expression.
Part III: Analyze DRAK1 protein expression and clinical relevance in ovarian cancer patient tissue samples
▷ DRAK1 protein expression showed clinical correleation in mucinous and endometrioid ovarian cancer tissues.
▷ DRAK1 and DRAK2 protein expression were elevated in ovarian primary and metastatic tumor tissues compared to normal tissues.
Part IV: Determine the efficacy of DRAK1 kinase inhibitor in prevention of ovarian tumorigenesis in vitro and in vivo
▷ DRAK1 kinase inhibitor treatment significantly decreased the cell viability of ovarian cancer cell lines where DRAK1 was highly expressed.
▷ DRAK1 kinase inhibitor treatement prevented foci-forming ability of ovarian cancer cells
▷ We examined that DRAK1 kinase inhibitor injection dramatically decreased the tumorigenesis of ovarian cancer cells using xenograft model. Part V: Discover novel biomarker by performing single cell RNA sequencing isolated from malignant ascites of ovarian cancer patient-derived spheroids
▷ Dispase treated in tissue for 1 hour, after thawing the frozen tissue. A number of 1,000,000 cells and a viability of 90% were confirmed, which were conditions that allowed single-cell sequencing to pass through QC. This succeeded in optimizing the experimental conditions.
▷ Through the expression of marker, the presence of various types of cells in ascites was confirmed such as macrophages, cancer cells, monocytes, fibroblasts, B cells. A total of 1082 cells were analyzed. Among them, 520 cells expressed macrophage markers, and 233 cancer cells, which were epithelial cells, were observed.
▷ Nine major clusters were revealed in cancer cells. The heterogeneity of cancer cells showed epithelial cell type and mesenchymal cell type according to the expression of EMT marker.
▷ Through single-cell sequencing, cell type and population heterogeneity and DRAK1/2 expression were confirmed using ascites obtained from the same chemo-resistance patient to determine the difference between before/after in treatment.
▷ Migration and invasion were decreased by DRAKs inhibitors in ovarian cancer cells that induced by acellular fraction of ascites in ovarian cancer.
▷ SIRT2 increased sensitivity of cisplatin response in ovarian cancer. These results suggested that ROS generation was an important inducer.
▷ Using single-cell RNA sequencing (scRNA-seq), CCL5-SCD4 interaction can suppress the fluid accumulation in the abdominal cavity by regulating the immune microenvironment in ovarian cancer.

(source : Project Summary 6p)

목차 Contents

• 표지 ... 1
• 제 출 문 ... 2
• 목차 ... 3
• 요약문 ... 4
• Project Summary ... 6
• 총괄연구과제 연구결과 ... 8
• 1. 총괄연구과제의 최종 연구개발 목표 ... 8
• 2. 총괄연구과제의 최종 연구개발 내용 및 결과 ... 13
• 3. 총괄연구과제의 연구결과 고찰 및 결론 ... 61
• 4. 총괄연구과제의 연구성과 및 목표달성도 ... 63
• 5. 총괄연구과제의 활용계획 ... 77
• 6. 첨부서류 ... 78
• 제1세부연구과제 연구결과 ... 92
• 1. 제1세부연구과제의 최종 연구개발 목표 ... 93
• 2. 제1세부연구과제의 연구대상 및 방법 ... 102
• 3. 제1세부연구과제의 최종 연구개발결과 ... 102
• 4. 제1세부연구과제의 연구결과 고찰 및 결론 ... 138
• 5. 제1세부연구과제의 연구성과 및 목표달성도 ... 140
• 6. 제1세부연구과제의 활용계획 ... 141
• 7. 참고문헌 ... 142
• 제2세부연구과제 연구결과 ... 143
• 1. 제2세부연구과제의 최종 연구개발 목표 ... 144
• 2. 제2세부연구과제의 연구대상 및 방법 ... 149
• 3. 제2세부연구과제의 최종 연구개발결과 ... 152
• 4. 제2세부연구과제의 연구결과 고찰 및 결론 ... 180
• 5. 제2세부연구과제의 연구성과 및 목표달성도 ... 182
• 6. 제2세부연구과제의 활용계획 ... 183
• 7. 참고문헌 ... 184
• 끝페이지 ... 185