상단 PDF Vinpocetine inhibits the proliferation and induces apoptosis in human colon cancer cells

4. Vinpocetine에 의해 변화되는 세포주기 조절 및 세포자멸사 관련 인자 발현 분석 다음으로 vinpocetine을 처리한 HCT116 세포에서 세포주기 조 절 및 세포자멸사 관련 인자들의 발현 변화를 조사해보았다. 먼저 HCT116 세포를 vinpocetine 함유 배지에서 다양한 시간대로 배양하 여 세포주기 조절에 관련된 유전자의 발현 정도를 탐색해보았다. RT- PCR을 시행해본 결과, Fig. 5A와 같이 cyclin B1, cyclin D1, cyclin E2 및 CDK6의 발현에는 영향을 미치지 않았다. 다른 세포주기 조절 인 자들에 대한 추가적인 연구가 수행되어야 할 것으로 보인다. 세포자멸 사를 조절하는 인자에 대한 분석은 Western blot assay을 시행하여 단백질 수준에서 발현 변화를 확인하였다. 대조군과 비교했을 때 vin- pocetine을 처리한 실험군에서 세포자멸을 억제하는 XIAP와 Bcl-2 단백질의 발현이 감소한 반면, 세포자멸을 유도할 수 있는 활성 형태의 잘려진 PARP 단백질 발현이 vinpocetine 처리의 48시간 배양에서 증 가하였다. 이러한 결과들을 통해 vinpocetine이 대장암 세포 HCT116 의 세포자멸사 관련 유전자들의 발현을 조절함으로써 세포자멸사를 유 도함을 확인할 수 있었다(Fig. 5B).

. DISCUSSION Ⅳ Despite recent advances in understanding the carcinogenesis of colorectal cancer, the increasing incidence and relatively low remission rate of chemotherapy have spurred researchers to develop more effective and safer treatment regimens by adopting novel and innovative approaches. The discovery and use of active medicinal compounds from natural sources has provided alternative treatment choices. Guggulsterone has been reported to have potent anti-tumor activities in some cancer cells, including prostate cancer, 22, 23 lung cancer, acute myeloid leukemia, and breast cancer cells. 24, 25 In addition, recent studies have revealed that guggulsterone inhibits angiogenesis in vitro and in vivo in human prostate cancer cells 26 and suppresses NF-κB activation in leukemia, breast, and multiple myeloma cells. 16,27 However, it was not previously determined whether guggulsterone modulates the apoptotic pathway in colon cancer cells. Moreover, it was not known whether guggulsterone has in vivo anti-cancer effects regardless of cancer type, including colorectal cancer. In the present study, we demonstrated the anti-cancer effect of guggulsterone via the induction of apoptosis in colorectal cancer cells both in vitro and in vivo. Given the central role of apoptosis in cancer treatment, the ability of guggulsterone to inhibit colorectal cancer supports the clinical utility of this agent as an anti-cancer drug for colorectal cancer. To the authors’ knowledge, this is the first report that clearly characterizes the anti-tumor properties of guggulsterone in colon cancer cells and tumor xenografts.

단백질 발현 세포 성장, 세포 사멸과 관련된 단백질 발현을 알아보기 위 해 western blot을 실시하였다. MDA-MB-231 cell을 5 × 10 5 cells/mL의 농도로 100 mm dish에 분주하고 48시간 후에 SFM 으로 교환하여 세포를 starvation하였다. 24시간 후 SFM에 delphinidin을 0, 5, 10, 20 μmol/L로 농도로 첨가하여 treat - ment하였다. 48시간 후에 새로운 treatment 용액으로 교환하 고, 24시간 후에 차가운 rinse buffer (PBS, 1 mM PMSF, 1 mM sodium orthovanadate)를 이용하여 세척하고 cell을 모 아 1,000 rpm에서 5분간 원심분리하였다. 차가운 lysis buffer (137 mM NaCl, 20 mM Tris-Cl, 1% triton X-100, 10% glyc- erol, 1 mM sodium orthovanadate, 1 mM PMSF, 20 μg/mL aprotinin, 10 μg/mL antipain, 10 μg/mL leupeptin, 80 μg/

i) Compound K induces phosphorylation of PERK and eIF-2 α ; ii) Compound K induces phosphorylation of IRE-1 and the spliced XBP transcription factor; iii) Compound K induces cleavage of ATF-6, and subsequent GRP-78 and CHOP expres- sion; and iv) Compound K induces caspase-12 cleavage. Among ER-associated apoptotic molecules, CHOP and caspase-12 are major proapoptotic factors that are closely associated with ER stress (11). Taken together, these observations demonstrate that Compound K induces ER-mediated apoptosis. ER stress response pathways are normally activated as a protective mechanism to ensure cell survival (41). However, during severe ER stress, activation of these pathways leads to increased CHOP expression, which is a crucial element that switches ER stress signaling from pro-survival to proapoptosis (42). The CHOP protein is a member of the CCAAT/enhancer-binding proteins and functions as a dominant-negative inhibitor of gene transcription (38). Expression of CHOP is mainly regulated at the transcriptional level through the PERK/eIF-2 α /ATF-6 pathway (38). CHOP knockout mice show reduced apoptosis in response to ER stress (43). Therefore, CHOP is one of the components of the ER stress-mediated apoptosis pathway. In the present study, suppression of CHOP using CHOP siRNA attenuated Compound K-induced apoptosis.

Compound K is active in biological systems and inhibits glucose uptake in tumor cells. Compound K also possesses chemopreventive activity against chemical carcinogens, impairs metastasis in vivo, and constrains tumor growth through the inhibition of 12-O-tetradecanoylphorbol- 13-acetate-induced cyclooxygenase-2 expression. 10–13 We recently reported that compound K exhibits cytotoxicity against tumor cells by the induction of apoptosis and cell cycle arrest at G 1 phase. These actions occur by a caspase- dependent pathway via mitochondrial disruption and inhibition of telomerase activity. 14,15 Furthermore, combi- nation treatment with compound K and radiation enhances human lung cancer cell death 16 and compound K induces apoptosis in MCF-7 human breast cancer cells by the generation of ROS and modulation of AMP-activated protein kinase signaling. 17 Finally, compound K-mediated gene- ration of ROS leads to apoptosis in HT-29 human colon cancer cells through the modulation of a mitochondria- dependent apoptotic pathway and the mitogen-activated protein kinase pathway. 18

50. Sahu, R.P.; Zhang, R.; Batra, S.; Shi, Y.; Srivastava, S.K. Benzyl isothiocyanate-mediated generation of reactive oxygen species causes cell cycle arrest and induces apoptosis via activation of MAPK in human pancreatic cancer cells. Carcinogenesis 2009, 30, 1744–1753. 51. El-Najjar, N.; Chatila, M.; Moukadem, H.; Vuorela, H.; Ocker, M.; Gandesiri, M.; Schneider- Stock, R.; Gali-Muhtasib, H. Reactive oxygen species mediate thymoquinone-induced apoptosis and activate ERK and JNK signaling. Apoptosis 2010, 15, 183–195.

and subsequent attenuation of proliferation. Overall, the ERBB3 blockade resulted in anti- tumorigenic effects without altering the PI3K/AKT/mTOR pathways in HCT116 cells by inhibiting cell proliferation or elevating Bak- and Bax-dependent apoptosis. Moreover, ERBB3 inhibition can provide significant anti-tumorigenic actions against colon cancer cells, despite the KRAS, PI3KCA and TP53 mutations. Similar to the finding of TBK1 as a potent target for HER2-positive breast cancer [65], an identification of molecular targets regulated by ERBB3 signal pathways will be beneficial to design a new regime for ERBB3-targeted therapy or a combination therapy for treating colorectal cancers.

Here, we show that a haploinsufficient mutation of Ino80, the catalytic ATPase of the INO80 complex, decreased intestinal adenomatous polyps and increased survival in an Apc min/+ mouse model of colon cancer. Experiments using tumors obtained from Apc min/+ mice and cells from human colon cancers showed that this Ino80 defect induced stalled replication forks, the concomitant activation of ATR-Chk1 signaling and an increase in apoptosis, suggesting that Ino80 haploinsufficiency inhibited colon cancer tumorigenesis by activating replication stress-induced ATR-Chk1 signaling to increase apoptosis. Importantly, in human colon cancer, we observed that the INO80 subunits were frequently present in high copy numbers and exhibited a high rate of amplification and increased protein expression. These results show that in contrast to our original prediction that INO80 acts as a tumor suppressor, INO80 actually functions oncogenically to promote colon tumorigenesis. INO80 therefore represents a novel therapeutic target in colon cancer. The results of this study also reinforce the emerging notion that while genomic instability can promote tumorigenesis, in certain genetic contexts, it can also act as a tumor suppressor.

“EFLL”, n-butanol fraction of lemon leaves “BFLL” and water fraction of lemon leaves “WFLL”), several cancer cells were treated with various concentrations of each fraction. MTT cell viability assay was performed after treating human breast cancer cell line MDA-MB-231 (Fig. 1A), human non-small lung cancer cell line A549 (Fig. 1B), human gastric cell lines AGS (Fig. 1C), and SNU-1 (Fig. 1D) for 48 h with 12.5, 25, 50, and 100 μg/mL of each fraction and DMSO was used as negative control. Our data demonstrate that among the tested fractions CFLL had the highest antiproliferative activity. Moreover, our data also show that among the tested cells, human gastric cancer cell line AGS was the most affected after the 48 h treatment in all concentrations treated.

MEK and ERK protein were detected by Western blot analysis. We provide evidence that A. capillaris induces cell cycle arrest, apoptosis, and potently inhibits the mitogen-activated protein kinase pathway. A. capillaris inhibited cell proliferation of LX2 cells in a dose- and time-dependent manner, increased the apoptosis fraction at cell cycle analysis with an accompanying DNA fragmentation, and resulted in a significant decrease in Bcl-2 mRNA levels and an increase in Bax expression. Exposure of LX2 cells to A. capillaris induced caspase-3 activation, but co-treatment of A. capillaris with the pan-caspase inhibitor Z-VAD-FMK, and the caspase-3 inhibitor Z-DEVE-FMK, blocked apoptosis. A. capillaris down-regulated Mcl-1 protein levels and inhibited phosphorylation of MEK/ERK, suggesting that it mediates cell death in LX2 cells through the down-regulation of Mcl-1 protein via a MEK/ERK-independent pathway.

７ 3. Results 3.1. JCX inhibits cell viability in human pancreatic cancer cell lines. The picture of J. chinensis leaf is shown in Fig. 1. The WST assay was conducted to examine whether the JCX mediates the inhibition effects on the cell viability of PANC-1, SNU-213, and 293T. Cells were treated with different concentrations (0, 2.5, 5, 7.5, 10, 15 μg/mL) of JCX for 72 h. The results demonstrated the JCX inhibited the cell viability in a dose-dependent manner. (Fig. 2) We found JCX has higher inhibition in PANC-1 and SNU- 213 human pancreatic cancer cells than 293T, derived from human embryonic kidney cells.

5 CDKN2A, TP53, and SMAD4, are the most important events associated with malignant transformation of pancreatic cancer. 9 KRAS is mutated in almost 90% of pancreatic cancer patients and the mutation is occurred in early stage of the pancreatic cancer malignant transformation. KRAS plays a key role in mitogenic signaling. When growth factor like EGF binds to receptor tyrosine kinase (RTK), KRAS-guanosine diphosphate (GDP) changes to KRAS-guanosine triphosphate (GTP) leading to activation of downstream signaling related to cell proliferation and apoptosis. 10 CDKN2A has a role in cell cycle regulation, which encodes p16 and p14. If there are mutations in CDKN2A, they can not do their role resulting in the inability to arrest the cell cycle even though there is DNA damage or oncogenic activation signal. TP53 plays a key role in cellular stress response by regulating gene expression related to cell cycle arrest and apoptosis resulting in suppression of uncontrolled cell proliferation. 11 Accumulation of these genetic alterations in pancreatic cancer totally leads to dysregulation of cell cycle and apoptosis resulting in uncontrolled proliferation of cancer cells.

Bee venom significantly induces apoptosis in HPV-positive cervical-cancer cells To further investigate whether BV affects cell fate, we carried out PI staining analysis after treating Caski cells with BV. We found that, starting after 12 h treatment, the number of cells entering the sub-G1 phase had increased (Fig. 2A). We then measured the induction of apoptosis in BV-treated Caski cells using annexin V staining followed by FACS analysis, and observed an increase of annexin V positive cells in Caski cells.

In conclusion, our study suggest that the anti-cancer effect of methanol extract of A. sieboldii inhibits the cell growth and induced cell apoptosis in FaDu cells through extrinsic death receptor and intrinsic mitochondrial-dependent apoptotic sig- naling pathway. Although these advantage have been found, in order to clarify the anticancer efficacy of A. sieboldii, the mechanism analysis should be performed in various cancer cells and experimental animal under the same extraction con- ditions.

Here, we report that Juniperus chinensis extract (JCX) inhibits cell viability and migration in human pancreatic cancer cells. JCX increases intracellular ROS levels and regulates the FAK/ERK pathway and apoptosis-related proteins, suggesting that it could be used to develop substances for the treatment of pancreatic cancer.

Key words : 18α-Glycyrrhetinic acid, Human gastric adenocarcinoma cell, AGS, Anti-cancer. Ⅰ. 서론 a) 암은 전 세계에서 가장 생명을 위협하는 질병 중 하나이며 그 중에서 위암은 세계에서 두번째로 암과 관련된 사망 원인이다 1,2) . 위암은 매년 거의 100만 명 의 새로운 환자에게서 진단되고 있으며, 동아시아, 동유럽, 중남미 일부 지역에서 가장 높은 발병률을 보이고 있다 3) . 위암 치료법은 수술만이 유일한 치료 법이라고 알려져 있지만 최근에는 위암 수술 후 각종 화학요법으로 위암 치료 결과를 좋게 하고 있다 4) . 따 라서, 새롭고 더 효과적인 항암제를 연구하고 개발해 야 한다.

과학과 의학기술의 발전으로 암(癌)은 완치율이 점차 높아 지고 있지만, 여전히 한국인의 대표적인 사망원인중 하나이다. 2014년 발표된 중앙암등록본부 자료에 따르면 2011년 우리 나라에서 발생한 전체 암 중에서 구강암은 약 0.2%의 비율을 차지하고 있는 희귀 암으로, 그 예후가 좋지 못하며 5년 동안 생존율도 50%에 불과하다고 알려져 있다 14) . 치료를 위해 외 과수술, 방사선 치료, 화학요법 등이 사용되고 있으나 치료과 정에서 나타날 수 있는 부작용과 낮은 완치율이 문제가 되고 있다. 기존 항암제가 암세포뿐만 아니라 정상세포의 기능도 억제하기 때문에 생길 수 있는 많은 부작용을 극복하기 위해 다양한 표적치료 항암제 개발이 시도되고 있다. 표적치료제 개발이란 암세포에서 특이적으로 확인되는 분자를 표적으로 하여 효과를 나타내는 물질들로부터 치료제를 개발하는 것으로 세포주기조절(cell cycle regulation), 혈관신생(angiogenesis), 세포자멸사(apoptosis) 등과 같은 암 발생과 관련된 신호전달 체계에 대해 이해가 우선되어야 한다.

http://dx.doi.org/10.5468/ogs.2014.57.6.501 pISSN 2287-8572 · eISSN 2287-8580 Introduction A uterine myoma (myoma uteri) is a solid tumor made of fibrous tissue, often called a ‘fibroid’ tumor. Myomas vary in size and number, and are mostly slow-growing with no symptoms. Approximately 25% of myomas will cause symp- toms and need medical treatment [1]. Myomas may grow as a single nodule or in clusters and range in size from 1 mm to over 20 cm in diameter. Myomas are the most frequently diagnosed tumor of the female pelvis and the most common reason for a hysterectomy. Although they are often referred to as tumors, they are not cancerous [2]. Treatment of leio- myoma has mainly been surgical resection, although some medical therapies are available for women who want to pre- serve uterus. Currently, management of leiomyoma relies on reducing the circulating levels of ovarian hormones by using gonadotropin-releasing hormone analogues, such as goserelin

in human uterine leiomyoma cells Youngsun Park, Seunghee Baek. Dept.[r]

Cancer cells usually exhibit increased levels on intracel- lular ROS, which in turn can initiate various cycles leading to further metabolic malfunction and ROS generation (44,45). ROS cause oxidative damage to DNA, proteins, lipids and other cellular components and therefore also significant cellular stress (45). A proposed therapeutic strategy against cancer is to treat cancer cells with pharmacological agents that have pro-oxidant properties which increase the intracellular ROS generation to a toxic threshold that triggers cell death in the cancer cells without harming normal cells (44). Vuvicevic et al showed that BML-275 induces ROS generation in glioma cell line, but AMPK α -siRNA treatment fails to induce ROS generation and apoptosis (22). In this study, an increased generation of ROS upon either BML-275 or AMPK α -siRNA treatment was observed and the intracellular accumulation of ROS seems to be one of critical factors in BML-275-induced apoptosis. To verify this speculation, NAC, scavenger of oxygen-free radicals, was challenged with BML-275. NAC relieved BML-275 or AMPK-siRNA mediated ROS production and improved cell viability based on the clonogenic assay, which suggested that both chemical and genetic inhibitor regulate cell viability via repressing AMPK activity.