Skip to main content

본문내용

종목정보

대출한도

종목토론카테고리

게시판버튼

게시글 제목

코미녹스 교모세포 종 논문 일부 번역

작성자 정보

세자

게시글 정보

조회 1,103 2021/06/27 00:56
수정 2021/06/27 01:02

게시글 내용

       

Anti-Tumor Effects of Sodium Meta-Arsenite in Glioblastoma Cells with Higher Akt Activities

Eun Jeong Lee et al. Int J Mol Sci. .
Free PMC article

Abstract

Glioblastoma is a type of aggressive brain tumor that grows very fast and evades surrounding normal brain, lead to treatment failure. Glioblastomas are associated with Akt activation due to somatic alterations in PI3 kinase/Akt pathway and/or PTEN tumor suppressor. Sodium meta-arsenite, KML001 is an orally bioavailable, water-soluble, and trivalent arsenical and it shows antitumoral effects in several solid tumor cells via inhibiting oncogenic signaling, including Akt and MAPK. Here, we evaluated the effect of sodium meta-arsenite, KML001, on the growth of human glioblastoma cell lines with different PTEN expression status and Akt activation, including PTEN-deficient cells (U87-MG and U251) and PTEN-positive cells (LN229). The growth-inhibitory effect of KML001 was stronger in U87-MG and U251 cells, which exhibited higher Akt activity than LN229 cells. KML001 deactivated Akt and decreased its protein levels via proteasomal degradation in U87-MG cells. KML001 upregulated mutant PTEN levels via inhibition of its proteasomal degradation. KML001 inhibited cell growth more effectively in active Akt-overexpressing LN229 cells than in mock-expressing LN229 cells. Consistent with these results, KML001 sensitized PTEN-deficient cells more strongly to growth inhibition than it did PTEN-positive cells in prostate and breast cancer cell lines. Finally, we illustrated in vivo anti-tumor effects of KML001 using an intracranial xenograft mouse model. These results suggest that KML001 could be an effective chemotherapeutic drug for the treatment of glioblastoma cancer patients with higher Akt activity and PTEN loss.

Keywords: Akt; PTEN; glioblastoma; sodium meta-arsenite.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Expression levels of PTEN and Akt and cell growth in human glioblastoma cell lines. ( A) Total RNA was extracted from glioma cell lines as indicated and subjected to RT-PCR to measure PTEN mRNA levels. ( B) Cells were lysed and 30 μg of total cellular proteins was analyzed by immunoblotting using anti-PTEN, -pAkt, -Akt, and β-actin antibodies. β-actin was used as a loading control. In the right panel, film was exposed a little longer for PTEN immunoblot. The levels of proteins were quantified by a densitometry and normalized to the loading control β-actin (lower panel). ( C) 1 × 10 5 cells were seeded and were grown in the culture media condition for 4 days. The cell numbers of each cell line were counted after trypan blue-staining at indicated times. Statistical analysis was conducted using parametric one-way ANOVA test and post hoc test (Bonferroni correction). Error bars represent standard deviations of the mean of three measurements (* p < 0.05, ** p < 0.01). These experiments were performed three independent times with comparable results.
Figure 2
Figure 2
Effects of KML001 on cell growth in human glioblastoma cell lines. ( A, B) Cells were treated with 0–30 μM KML001 in the culture media without or with FBS for 24 h. Cell growth was measured by MTS assay as described in Section 4. ( C) Cells were treated with 0–30 μM KML001 in the culture media for 24 h and cells were collected, mix an equal volume of trypan blue dye and manually counted by countess TM automated cell counter. ( D) Cells were treated with 10 μM KML001 for 24 h and cell lysates were analyzed by immunoblotting using indicated antibodies. ( E) The levels of proteins were quantified by a densitometry and normalized to β-actin. Data are presented as mean ± SE of three independent experiments (* p < 0.01, ** p < 0.05). These experiments were performed three independent times with comparable results.
Figure 3
Figure 3
Induction of apoptosis with KML001 in U87-MG cells. ( A) U87-MG cells were treated with 0–30 μM KML001 for 24 h and cell morphology was observed using an optical microscope. Scale bar, 100 μm. ( B) U87-MG cells were treated with 0–30 μM KML001 for 24 h, stained with annexin V-FITC and PI, and subjected to flow cytometry analysis. Fluorescence dot blots of annexin V-positive (horizontal axis) and PI-positive (vertical axis) cells are shown. Cells that were positively stained by annexin V-FITC only (early apoptosis) and positive for both annexin V-FITC and PI (late apoptosis) were quantitated, and both subpopulations were considered as overall apoptotic cells. ( C) U87-MG cells were treated with 0–30 μM KML001 for 24 h, and then ΔΨ m was measured using the JC-1 kit by flow cytometer. The loss of ΔΨ m was equated with decreased red fluorescence. ( D) U87-MG cells were treated with 0–30 μM KML001 for 24 h and cell lysates were subjected to immunoblotting analysis using the indicated antibodies. The levels of proteins were quantified by a densitometry and normalized to β-actin (Supplementary Figure S2) Similar results were observed in three independent experiments.
Figure 4
Figure 4
KML001-induced Akt downregulation and PTEN upregulation in U87-MG cells. ( A) Cells were treated with 0–30 μM KML001 for 24 h and cell lysates were subjected to immunoblotting analysis using indicated antibodies. The histogram shows the densitometric measuring of pAkt/Akt ratio relative to control. pAkt/Akt ratio = [(pAkt/β-actin)/(Akt/β-actin)] (right panel). Data are presented as mean ± SE of three independent experiments. ** p < 0.01 indicate significant difference from the control. ( B) U87-MG cells were treated with 0–20 μM KML001 for 24 h and total RNA was extracted and subjected to RT-PCR to assess Akt and PTEN mRNA levels. The levels of RNA were normalized with β-actin. ( C– E) U87-MG cells were treated without or with 10 μM KML001 for 24 h and some KML001-treated cells were incubated with 5 μM or 10 μM MG132 ( C), chloroquine (Chlo) ( D), and 10 μM cycloheximide (CHX) ( E) for indicated times before harvest. Cell lysates were subjected to immunoblotting analysis using indicated antibodies. ( F) KML001-treated U87-MG cells were incubated with MG132 for 8 h and with 10 μM cycloheximide for 1 h before harvest. The levels of proteins were quantified by a densitometry and normalized to β-actin (Supplementary Figure S5) Cell lysates were subjected to immunoblotting analysis using indicated antibodies. These experiments were performed three independent times with comparable results.
Figure 5
Figure 5
KML001-induced cell growth inhibition is correlated with Akt activity in glioblastoma cell lines. ( A, B) U87-MG cells were transfected with lentivirus either control virus only (LacZ) or PTEN and cells were treated with 0–30 μM KML001 for 24 h followed by immunoblotting analysis using indicated antibodies ( A), the levels of proteins were quantified by a densitometry and normalized to β-actin (Supplementary Figure S6) and MTS assay ( B). ( C) U87-MG cells were treated either with 0–30 μM KML001, 0–50 μM Akt inhibitor II, or 0–100 μM LY294002 (PI3K inhibitor) and cell growth was measured by MTS assay. ( D, E) LN229 cells were infected with lentivirus encoding either vector only (Mock), wild type Akt (WT-Akt), or constitutively-active Akt (Active-Akt) and stable cell lines were established as described in the “Materials and Methods”. Cells were treated with 0–30 μM KML001 for 24 h followed by immunoblotting analysis using indicated antibodies ( D), the levels of proteins were quantified by a densitometry and normalized to β-actin (Supplementary Figure S6) and MTS assay ( E). Error bars represent standard deviations. * p < 0.05, ** p < 0.01 versus KML001-treated Mock-LN229 cells. These experiments were performed three independent times with comparable results.
Figure 6
Figure 6
Association of KML001-induced cell growth inhibition with Akt activity and PTEN expression. ( A, B) PC-3 and Du145 cells were treated with 0–30 μM KML001 for 24 h, followed by immunoblotting analysis using the indicated antibodies ( A), the levels of proteins were quantified by a densitometry and normalized to β-actin (Supplementary Figure S7) and LDH release assay for cytotoxicity ( B). ( C, D) MDA-MB231 and BT-549, human breast cancer cells were treated with 0–30 μM KML001 for 24 h. Cells were processed for immunoblotting analysis using the indicated antibodies ( C), The levels of proteins were quantified by a densitometry and normalized to β-actin (Supplementary Figure S7) or cell viability assay by counting of trypan blue-stained cells using countess TM automated cell counter ( D). Error bars represent standard deviations (* p < 0.05, ** p < 0.01). Similar results were observed in three independent experiments.
Figure 7
Figure 7
Anti-tumor effects of KML001 in orthotropic xenograft models. ( A) Schematic procedure of in vivo experiments. On Day 1, U87-MG cells were injected into the mouse brain as described in the “Materials and Methods”. On day 7, the mice (n = 5/group) were treated with KML001 or with PBS (control) orally with 2-day intervals for two weeks and on day 21 the mice were sacrificed. On day 7 and day 21 MRI images of brains were captured. Similar results were observed in two independent experiments. ( B) Brain MRI images of mice were captures on day 21. Representative pictures indicate mouse brain regions of corpus clausum and its surrounding structures. ( C) Tumor volumes were measured as described in the “Materials and Methods”. Statistical analysis was conducted using one-way ANOVA test and post hoc test (Bonferroni correction), ** p < 0.01. ( D) Tumor tissues were fixed and stained with anti-pAkt antibodies. Scale bars = 100 μm. White arrows indicate tumors. ( E) The integrated optical density (IOD) was analyzed for pAkt quantification per histological field (×200 magnification). Error bars represent standard deviations (** p < 0.05 vs. control). Similar results were observed in three independent experiments.
See this image and copyright information in PMC

Similar articles

See all similar articles

References

    1. Le Rhun E., Preusser M., Roth P., Reardon D.A., van den Bent M., Wen P., Reifenberger G., Weller M. Molecular targeted therapy of glioblastoma. Cancer Treat. Rev. 2019;80:101896. doi: 10.1016/j.ctrv.2019.101896. - DOI - PubMed
    1. Rajaratnam V., Islam M.M., Yang M., Slaby R., Ramirez H.M., Mirza S.P. Glioblastoma: Pathogenesis and Current Status of Chemotherapy and Other Novel Treatments. Cancers. 2020;12:937. doi: 10.3390/cancers12040937. - DOI - PMC - PubMed
    1. Gimple R.C., Bhargava S., Dixit D., Rich J.N. Glioblastoma stem cells: Lessons from the tumor hierarchy in a lethal cancer. Genes Dev. 2019;33:591–609. doi: 10.1101/gad.324301.119. - DOI - PMC - PubMed
    1. Li Q.J., Cai J.Q., Liu C.Y. Evolving Molecular Genetics of Glioblastoma. Chin. Med. J. 2016;129:464–471. doi: 10.4103/0366-6999.176065. - DOI - PMC - PubMed
    1. Brennan C.W., Verhaak R.G., McKenna A., Campos B., Noushmehr H., Salama S.R., Zheng S., Chakravarty D., Sanborn J.Z., Berman S.H., et al. The somatic genomic landscape of glioblastoma. Cell. 2013;155:462–477. doi: 10.1016/j.cell.2013.09.034. - DOI - PMC - PubMed

MeSH terms

LinkOut - more resources

  • Full Text Sources

  • Medical

  • Research Materials

    • NCI CPTC Antibody Characterization Program 
    • 교모세포종은 매우 빠르게 자라고 정상적인 뇌를 피해서 치료 실패를 초래하는 공격적인 뇌종양의 일종이다. 교모세포종은 PI3 키나제/Akt 경로 및/또는 PTEN 종양 억제제의 체적 변화로 인해 Akt 활성화와 관련이 있다. 메타아세나이트 나트륨, KML001은 경구 생체 이용, 수용성 및 삼원 비소이며 Akt 및 MAPK를 포함한 종양 신호 억제를 통해 여러 고형 종양 세포에서 항정신성 효과를 나타낸다. PTEN 결핍세포(U87-MG, U251)와 PTEN 양성세포(LN229)를 포함하여 PTEN 발현여부와 Akt 활성화가 다른 인간 교모세포주 성장에 미치는 영향을 평가하였다. KML001의 성장 억제 효과는 LN229 세포보다 높은 Akt 활성을 보인 U87-MG와 U251 세포에서 더 강했다. KML001은 U87-MG 세포에서 단백질 분해를 통해 Akt를 비활성화하고 단백질 수치를 감소시켰다. KML001 단백질 분해 억제를 통해 돌연변이 PTEN 수준을 상향 조정했다. KML001은 LN229 셀의 모의 표현보다 LN229 셀을 과도하게 발현하는 활성 Akt에서 더 효과적으로 세포 성장을 억제하였습니다. 이러한 결과와 일관되게, KML001은 전립선 및 유방암 세포 라인에서 PTEN 양성 세포를 감작한 것보다 PTEN 결핍 세포를 성장 억제에 더 강하게 감작시켰다. 마지막으로, 두개골 내 이종 이식 마우스 모델을 사용하여 KML001의 생체 내 종양 방지 효과를 설명했다. 이러한 결과는 KML001이 높은 Akt 활성과 PTEN 손실을 가진 교모세포종 환자의 치료에 효과적인 화학 치료제가 될 수 있음을 시사한다.

      키워드: Akt; PTEN; 교모세포종; 메타아세나이트 나트륨.
    • 
    • 
    • 유사 기사
      비소 화합물, 메타아세나이트 나트륨(KML001)의 비호지킨 림프종 항종양 효과: 체외 및 생체내 연구
      윤JS, 황DW, 김ES, 김J, 김S, 정HJ, 이SK, 이JH, 엄J, 원YW, 박B, 최JH, 이Y.윤제스 외 연구진 신약 투자. 2016년 2월 34일:1-14.doi: 10.1007/s10637-015-0301-z. 2015년 11월 18일.2016년 신약 투자 PMID: 26581399

      커큐민은 p-AKT/mTOR 경로에 의한 교모세포 증식을 억제하고 PTEN 발현을 증가시킨다.
      왕즈, 류에프, 랴오W, 율, 휴즈, 림, 시아에이치.왕즈 외 연구진 아치 생화학 생물. 2020년 8월 15일; 689:108412. doi: 10.1016/j.abb.2020.108412. Epub 2020 5월 21일.아치 생화학 생물. 2020. PMID: 32445778

      Xihuang 알약은 Akt/m를 통한 Temozolomide 교모세포종 외래 이식물의 항종양 효과를 강화합니다.TOR 의존 경로.
      FuJ, ZhuSH, XuHB, Xu YQ, Wang X, Wang J, Kong PS.푸제이 외 연구진입니다 J 에스노파마콜. 2020년 10월 28일, 261:113071. doi: 10.1016/j.jep.2020.113071. 2020년 6월 27일.J 에스노파마콜. 2020. PMID: 32603676

      교모세포종의 AKT 경로를 노리고 있어
      맥도웰 KA, 리긴스 GJ, 갈리아 GL맥도웰 KA 외 연구진입니다 Curr Pharm Des. 2011;17(23):2411-20. doi:10.2174/138161211797249224Curr Pharm Des. 2011. PMID: 21827416 검토.

      mTORC1과 mTORC2의 이산 신호 메커니즘: 세포와 분자 측면에서 아직 분리되어 있다.
      잔와 유니얄 M, 아민 AG, 쿠퍼 JB, 다스 K, 슈미트 MH, 무라리 R.얀와 유니얄 M 외 Adv Biol Regul. 2017년 5월;64:39-48.doi:10.1016/j.jbior.2016.12.001. 에펍 2017 1월 4일.Adv Biol Regul. 2017. PMID: 28189457 검토.


      유사한 모든 기사 참조


    • https://pubmed.ncbi.nlm.nih.gov/33256086/
    • 

게시글 찬성/반대

  • 7추천
  • 0반대
내 아이디와 비밀번호가 유출되었다? 자세히보기 →

운영배심원의견

운영배심원 의견?
운영배심원의견이란
운영배심원 의견이란?
게시판 활동 내용에 따라 매월 새롭게 선정되는
운영배심원(10인 이하)이 의견을 행사할 수 있습니다.

운영배심원 4인이 글 내리기에 의견을 행사하게 되면
해당 글의 추천수와 반대수를 비교하여 반대수가
추천수를 넘어서는 경우에는 해당 글이 블라인드 처리
됩니다.

댓글목록

댓글 작성하기

댓글쓰기 0 / 1000

게시판버튼

최신글리스트

광고영역

하단영역

약관및지원메뉴

관련사이트이동

씽크풀 사이트에서 제공되는 정보는 오류 및 지연이 있을 수 있으며, 이를 기반으로 한 투자에는 손실이 발생할 수 있습니다.
그 이용에 따르는 책임은 이용자 본인에게 있습니다. 또한 이용자는 본 정보를 무단 복사, 전재 할 수 없습니다.

씽크풀은 정식 금융투자업자가 아닌 유사투자자문업자로 개별적인 투자상담과 자금운용이 불가합니다.
본서비스에서 제공되는 모든 정보는 투자판단의 참고자료로 원금 손실이 발생될 수 있으며, 그 손실은 투자자에게 귀속됩니다.

㈜씽크풀 서울시 영등포구 국제금융로 70, 15층 (여의도동, 미원빌딩)

고객센터 1666-6300 사업자 등록번호 116-81-54775 대표 : 김동진

Copyright since 1999 © ThinkPool Co.,Ltd. All Rights Reserved