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Isoindolone derivative QSN-10c induces leukemic cell apoptosis and suppresses angiogenesis via PI3K/AKT signaling pathway inhibition.

Lv WW, Qin SN, Chen CQ, Zhang JJ, Ren TS, Xu YN, Zhao QC - Acta Pharmacol. Sin. (2014)

Bottom Line: Furthermore, QSN-10c dose-dependently decreased the Δψm in K562 cells, increased the release of cytochrome c and the level of Bax, and decreased the level of Bcl-2, suggesting that QSN-10c-induced apoptosis of K562 cells was mediated via the mitochondrial apoptotic pathway.QSN-10c treatment did not alter the Δψm in HUVECs, but dose-dependently inhibited the expression of VEGF, inhibited the tube formation and cell migration in vitro, and significantly suppressed the number of ISVs in zebrafish embryos in vivo.QSN-10c is a novel antitumor compound that exerts both antitumor and anti-angiogenic effects via inhibiting the PI3K/AKT/GSK3β signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Pharmacy, General Hospital of Shenyang Military Area Command, Shenyang 110840, China [2] Shenyang Pharmaceutical University, Shenyang 110016, China.

ABSTRACT

Aim: 2-(4,6-Dimethoxy-1,3-dioxoisoindolin-2-yl) ethyl 2-chloroacetate (QSN-10c) is one of isoindolone derivatives with antiproliferative activity against human umbilical vein endothelial cells (HUVECs). The aim of this study was to investigate its antitumor activity in vitro and anti-angiogenic effects in vitro and in vivo.

Methods: K562 leukemic cells and HUVECs were used for in vitro studies. Cell viability was examined using MTT assay. Cell apoptosis and mitochondrial transmembrane potential (Δψm) were detected with flow cytometry. Tube formation and migration of HUVECs were studied using two-dimensional Matrigel assay and wound-healing migration assay, respectively. VEGF levels were analyzed with RT-PCR and Western blotting. A zebrafish embryo model was used for in vivo anti-angiogenic studies. The molecular mechanisms for apoptosis in K562 cells and antiangiogenesis were measured with Western blotting.

Results: In antitumor activity studies, QSN-10c suppressed the viability of K562 cells and induced apoptosis in dose- and time-dependent manners. Furthermore, QSN-10c dose-dependently decreased the Δψm in K562 cells, increased the release of cytochrome c and the level of Bax, and decreased the level of Bcl-2, suggesting that QSN-10c-induced apoptosis of K562 cells was mediated via the mitochondrial apoptotic pathway. In anti-angiogenic activity studies, QSN-10c suppressed the viability of HUVECs and induced apoptosis in dose dependent manners. QSN-10c treatment did not alter the Δψm in HUVECs, but dose-dependently inhibited the expression of VEGF, inhibited the tube formation and cell migration in vitro, and significantly suppressed the number of ISVs in zebrafish embryos in vivo. Furthermore, QSN-10c dose-dependently suppressed the phosphorylation of AKT and GSK3β in both HUVECs and K562 cells.

Conclusion: QSN-10c is a novel antitumor compound that exerts both antitumor and anti-angiogenic effects via inhibiting the PI3K/AKT/GSK3β signaling pathway.

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QSN-10c increased apoptosis in K562 cells and HUVECs. PI staining (A–B) and FITC-conjugated Annexin V/PI staining (C-D) were conducted to evaluate the cell apoptosis induced by QSN-10c. (A) Representative patterns of flowcytometric distribution of K562 cells treated with QSN-10c for different time. M1 marks subG1 apoptosic cells. (B) Quantitative comparison of the numbers of subG1 phase in different groups. (C) Representative patterns of flowcytometric distribution of K562 cells. Dot plot was divided into a quadrant: upper left, necrotic cells; upper right, late apoptotic cells; lower left, living cells; and lower right, early apoptotic cells. (D) Graphs demonstrate the sum of cells in early and late apoptosis of K562 cells. (E) Representative patterns of flowcytometric distribution of HUVEC. Cells receiving only DMSO (0.1%) served as a vehicle control. Data were expressed as mean±SD. n=3. cP<0.01 vs control.
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fig3: QSN-10c increased apoptosis in K562 cells and HUVECs. PI staining (A–B) and FITC-conjugated Annexin V/PI staining (C-D) were conducted to evaluate the cell apoptosis induced by QSN-10c. (A) Representative patterns of flowcytometric distribution of K562 cells treated with QSN-10c for different time. M1 marks subG1 apoptosic cells. (B) Quantitative comparison of the numbers of subG1 phase in different groups. (C) Representative patterns of flowcytometric distribution of K562 cells. Dot plot was divided into a quadrant: upper left, necrotic cells; upper right, late apoptotic cells; lower left, living cells; and lower right, early apoptotic cells. (D) Graphs demonstrate the sum of cells in early and late apoptosis of K562 cells. (E) Representative patterns of flowcytometric distribution of HUVEC. Cells receiving only DMSO (0.1%) served as a vehicle control. Data were expressed as mean±SD. n=3. cP<0.01 vs control.

Mentions: The sub-diploid (sub-G1) population after PI staining was quantified to determine the extent of apoptosis induced by QSN-10c. A time-dependent increase in sub-G1 cells was visible after treatment with 100 μmol/L QSN-10c (Figure 3A, 3B, 6, 12, 24, and 48 h after treatment resulted in 3.47%±0.11%, 4.31%±0.46%, 5.38%±0.51%, 11.27%±0.73% sub-G1 cells, respectively, vs 1.75%±0.17%, 2.10%±0.28%, 2.16%±0.29%, 2.25%±0.21% in the respective control groups). However, no significant apoptotic induction was observed in HUVEC cells treated with the same QSN-10c concentration (data not shown).


Isoindolone derivative QSN-10c induces leukemic cell apoptosis and suppresses angiogenesis via PI3K/AKT signaling pathway inhibition.

Lv WW, Qin SN, Chen CQ, Zhang JJ, Ren TS, Xu YN, Zhao QC - Acta Pharmacol. Sin. (2014)

QSN-10c increased apoptosis in K562 cells and HUVECs. PI staining (A–B) and FITC-conjugated Annexin V/PI staining (C-D) were conducted to evaluate the cell apoptosis induced by QSN-10c. (A) Representative patterns of flowcytometric distribution of K562 cells treated with QSN-10c for different time. M1 marks subG1 apoptosic cells. (B) Quantitative comparison of the numbers of subG1 phase in different groups. (C) Representative patterns of flowcytometric distribution of K562 cells. Dot plot was divided into a quadrant: upper left, necrotic cells; upper right, late apoptotic cells; lower left, living cells; and lower right, early apoptotic cells. (D) Graphs demonstrate the sum of cells in early and late apoptosis of K562 cells. (E) Representative patterns of flowcytometric distribution of HUVEC. Cells receiving only DMSO (0.1%) served as a vehicle control. Data were expressed as mean±SD. n=3. cP<0.01 vs control.
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Related In: Results  -  Collection

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fig3: QSN-10c increased apoptosis in K562 cells and HUVECs. PI staining (A–B) and FITC-conjugated Annexin V/PI staining (C-D) were conducted to evaluate the cell apoptosis induced by QSN-10c. (A) Representative patterns of flowcytometric distribution of K562 cells treated with QSN-10c for different time. M1 marks subG1 apoptosic cells. (B) Quantitative comparison of the numbers of subG1 phase in different groups. (C) Representative patterns of flowcytometric distribution of K562 cells. Dot plot was divided into a quadrant: upper left, necrotic cells; upper right, late apoptotic cells; lower left, living cells; and lower right, early apoptotic cells. (D) Graphs demonstrate the sum of cells in early and late apoptosis of K562 cells. (E) Representative patterns of flowcytometric distribution of HUVEC. Cells receiving only DMSO (0.1%) served as a vehicle control. Data were expressed as mean±SD. n=3. cP<0.01 vs control.
Mentions: The sub-diploid (sub-G1) population after PI staining was quantified to determine the extent of apoptosis induced by QSN-10c. A time-dependent increase in sub-G1 cells was visible after treatment with 100 μmol/L QSN-10c (Figure 3A, 3B, 6, 12, 24, and 48 h after treatment resulted in 3.47%±0.11%, 4.31%±0.46%, 5.38%±0.51%, 11.27%±0.73% sub-G1 cells, respectively, vs 1.75%±0.17%, 2.10%±0.28%, 2.16%±0.29%, 2.25%±0.21% in the respective control groups). However, no significant apoptotic induction was observed in HUVEC cells treated with the same QSN-10c concentration (data not shown).

Bottom Line: Furthermore, QSN-10c dose-dependently decreased the Δψm in K562 cells, increased the release of cytochrome c and the level of Bax, and decreased the level of Bcl-2, suggesting that QSN-10c-induced apoptosis of K562 cells was mediated via the mitochondrial apoptotic pathway.QSN-10c treatment did not alter the Δψm in HUVECs, but dose-dependently inhibited the expression of VEGF, inhibited the tube formation and cell migration in vitro, and significantly suppressed the number of ISVs in zebrafish embryos in vivo.QSN-10c is a novel antitumor compound that exerts both antitumor and anti-angiogenic effects via inhibiting the PI3K/AKT/GSK3β signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Pharmacy, General Hospital of Shenyang Military Area Command, Shenyang 110840, China [2] Shenyang Pharmaceutical University, Shenyang 110016, China.

ABSTRACT

Aim: 2-(4,6-Dimethoxy-1,3-dioxoisoindolin-2-yl) ethyl 2-chloroacetate (QSN-10c) is one of isoindolone derivatives with antiproliferative activity against human umbilical vein endothelial cells (HUVECs). The aim of this study was to investigate its antitumor activity in vitro and anti-angiogenic effects in vitro and in vivo.

Methods: K562 leukemic cells and HUVECs were used for in vitro studies. Cell viability was examined using MTT assay. Cell apoptosis and mitochondrial transmembrane potential (Δψm) were detected with flow cytometry. Tube formation and migration of HUVECs were studied using two-dimensional Matrigel assay and wound-healing migration assay, respectively. VEGF levels were analyzed with RT-PCR and Western blotting. A zebrafish embryo model was used for in vivo anti-angiogenic studies. The molecular mechanisms for apoptosis in K562 cells and antiangiogenesis were measured with Western blotting.

Results: In antitumor activity studies, QSN-10c suppressed the viability of K562 cells and induced apoptosis in dose- and time-dependent manners. Furthermore, QSN-10c dose-dependently decreased the Δψm in K562 cells, increased the release of cytochrome c and the level of Bax, and decreased the level of Bcl-2, suggesting that QSN-10c-induced apoptosis of K562 cells was mediated via the mitochondrial apoptotic pathway. In anti-angiogenic activity studies, QSN-10c suppressed the viability of HUVECs and induced apoptosis in dose dependent manners. QSN-10c treatment did not alter the Δψm in HUVECs, but dose-dependently inhibited the expression of VEGF, inhibited the tube formation and cell migration in vitro, and significantly suppressed the number of ISVs in zebrafish embryos in vivo. Furthermore, QSN-10c dose-dependently suppressed the phosphorylation of AKT and GSK3β in both HUVECs and K562 cells.

Conclusion: QSN-10c is a novel antitumor compound that exerts both antitumor and anti-angiogenic effects via inhibiting the PI3K/AKT/GSK3β signaling pathway.

Show MeSH