Effects of 2-D08 (2′,3′,4′-trihydroxy-flavone) on Proliferation and Apoptosis in Uterine Leiomyosarcoma Cells : Effect of 2-D08 on Uterine Leiomyosarcoma Cells = 2-D08 (2′,3′,4′-trihydroxy-flavone)이 자궁 평활근 육종 세포의 증식과 세포사멸에 미치는 영향에 대한 연구

국문 초록 (Abstract)

목 적: SUMO E2 억제제인 2-D08은 항암 효과를 비롯한 여러 생물학적 기능을 가지고 있다. 그러나 자궁 평활근 육종에서 2-D08의 효과는 알려져 있지 않았다. 본 연구는 인간 유래 자궁 평활근 육종 세포에서 2-D08의 항암 활성을 조사했다.

방 법: SK-LMS-1 (인간 유래 자궁 평활근 육종 세포주) 세포를 본 연구에 사용했다. 세포 생존성 (MTT 분석), 콜로니 형성, Ki67 염색 및 BrdU 증식 분석을 통해 세포 증식에 대한 2-D08의 효과를 평가했다. 카스파제-3 활성도 분석을 통해 2-D08에 의한 세포사멸을 측정하였다. 세포 신호전달에 대한 2-D08의 영향을 평가하기 위해 웨스턴 블롯 분석 및 면역침강법을 사용하였다.

결 과: 2-D08은 SK-LMS-1 세포에서 세포 생존성을 유의하게 억제하였다. 2-D08을 처리하면 SK-LMS-1 세포의 콜로니 형성 능력이 유의하게 억제되었다. Ki67 염색 및 BrdU 분석에서 2-D08이 처리가 된 SK-LMS-1 세포는 항증식 효과가 있음을 발견했다. 또한, 2-D08은 SK-LMS-1 세포에서 p21 단백질을 증가하는 것으로 나타났지만, 카스파제-3 활성화 분석에서 세포자멸을 유도하지 않았다. 그리고 프로테아좀 억제는 SK-LMS-1 세포에서 p21 단백질의 안정 수준을 증가시켰고, 2-D08은 이러한 신호 전달 경로를 조절한다. 게다가 SK-LMS-1 세포에서 α-SM-액틴, TAGLN 및 칼포닌 1의 발현 변화를 확인한 결과 2-D08은 평활근 표현형 전환을 직접 유도하지 않았다.

결 론: 이러한 결과는 SK-LMS-1 세포에서 2-D08이 p21의 안정화를 조절하여 세포 증식을 억제한다는 것을 나타낸다. 따라서 2-D08은 자궁 평활근 육종의 후보 치료 물질이 될 수 있다. 그러나 추가적인 연구를 통해 이 화합물의 적절한 용량을 신중하게 선택해야 한다.

다국어 초록 (Multilingual Abstract)

Objective: 2-D08, a SUMO E2 inhibitor, has several biological functions, including anti-cancer effects. However, the effects of 2-D08 in uterine leiomyosarcoma (Ut-LMS) are unknown. This study explored the anti-cancer activity of 2-D08 against human Ut-LMS cells.

Methods: We used SK-LMS-1 (human Ut-LMS cells) as an in vitro model. Cell viability (MTT assay), colony formation, Ki67 staining, and BrdU proliferation assays were used to evaluate the effect of 2-D08 on cellular proliferation. Caspase-3 activity was assessed by measuring 2-D08-induced apoptosis. The impact of 2-D08 on cellular signaling was evaluated through western blotting and immunoprecipitation techniques.

Results: 2-D08 significantly inhibited cell viability in the SK-LMS-1 cells. 2-D08 treatment markedly suppressed the potential of SK-LMS-1 cells to form colonies. Using Ki67 staining and BrdU assay, we found that 2-D08 treatment had anti-inhibitory effects on the proliferation of SK-LMS-1 cells. Moreover, 2-D08 was also shown to upregulate p21 proteins in SK-LMS-1 cells, though 2-D08 did not further promote Caspase-3 activity and apoptosis. In SK-LMS-1 cells, the inhibition of proteasomes led to elevated steady state levels of p21, and 2-D08 was observed to modulate these signaling pathways. Moreover, the evaluation of α-SM-actin, TAGLN, and calponin 1 expression indicated that 2-D08 did not directly initiate smooth muscle phenotypic switching in SK-LMS-1 cells.

Conclusion: These results indicate that in SK-LMS-1 cells, 2-D08 inhibits cell proliferation by regulating the stabilization of p21. Therefore, 2-D08 show potential as a promising candidate for human Ut-LMS. However, it is crucial to meticulously choose the proper dosage for this compound.

목차 (Table of Contents)

• Introduction 1
• Materials and Methods 4
• Results 9
• A. Effects of 2-D08 on cell viability of Ut-LMS cells 9
• B. 2-D08 inhibits colony formation of Ut-LMS cells 9
• C. 2-D08 suppresses the proliferation of Ut-LMS cells in vitro 10
• D. Effect of 2-D08 on the protein expression related to cell proliferation and apoptosis in Ut-LMS cells 10
• E. 2-D08 treatment inhibits the degradation of p21 by Ubproteasome pathway in Ut-LMS cells 11
• F. The impact of 2-D08 on the expression of differentiation marker proteins in Ut-LMS cells 12
• Discussion 13
• Figures 17
• Reference 29

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