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Impaired angiogenesis and tumor development by inhibition of the mitotic kinesin Eg5.

Exertier P, Javerzat S, Wang B, Franco M, Herbert J, Platonova N, Winandy M, Pujol N, Nivelles O, Ormenese S, Godard V, Becker J, Bicknell R, Pineau R, Wilting J, Bikfalvi A, Hagedorn M - Oncotarget (2013)

Bottom Line: Chemical blockade of Eg5 inhibits endothelial cell proliferation and migration in vitro.Mitosis-independent vascular outgrowth in aortic ring cultures is strongly impaired after Eg5 or Mklp2 protein inhibition.In vivo, interfering with KIF11/Eg5 function causes developmental and vascular defects in zebrafish and chick embryos and potent inhibition of tumor angiogenesis in experimental tumor models.

View Article: PubMed Central - PubMed

Affiliation: University Bordeaux, LAMC, UMR 1029, F-33405 Talence, France.

ABSTRACT
Kinesin motor proteins exert essential cellular functions in all eukaryotes. They control mitosis, migration and intracellular transport through interaction with microtubules. Small molecule inhibitors of the mitotic kinesin KiF11/Eg5 are a promising new class of anti-neoplastic agents currently evaluated in clinical cancer trials for solid tumors and hematological malignancies. Here we report induction of Eg5 and four other mitotic kinesins including KIF20A/Mklp2 upon stimulation of in vivo angiogenesis with vascular endothelial growth factor-A (VEGF-A). Expression analyses indicate up-regulation of several kinesin-encoding genes predominantly in lymphoblasts and endothelial cells. Chemical blockade of Eg5 inhibits endothelial cell proliferation and migration in vitro. Mitosis-independent vascular outgrowth in aortic ring cultures is strongly impaired after Eg5 or Mklp2 protein inhibition. In vivo, interfering with KIF11/Eg5 function causes developmental and vascular defects in zebrafish and chick embryos and potent inhibition of tumor angiogenesis in experimental tumor models. Besides blocking tumor cell proliferation, impairing endothelial function is a novel mechanism of action of kinesin inhibitors.

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Eg5 inhibition reduces tumor angiogenesis in experimental tumor models(a-d) U87-derived gliomas on the CAM were treated with indicated doses of Eg5 inhibitors. Biomicroscopy images were taken at day 4 of tumor development. Note visible induction of necrosis (arrows in b-d) at the tumor surface after Eg5 inhibition. (e-f) Immunohistological examination of experimental glioma. A dense vascular network occurred in control tumors, whereas Eg5 inhibition leads to reduced tumor angiogenesis underneath the tumor surface (asterisk), denoted by the dashed line (f), especially at two treatments of DMN (g) or ISP (h) per day. (i) A significantly higher number of poorly vascularized tumors (+) was within the ISP-treated group, whereas few tumors treated with ISP appeared highly vascularized (+++). (j) Ispinesib also reduced tumor size (representative tumors are shown); (k) weight (P= 0.0037) and (i) vascular density (P=0.0184) in an orthotopic renal cell carcinoma model. (j). Short-term treatment (48h) with ispinesib lead to a significant increase of dilated capillaries (P=0.0284; n, p) whereas overall vascular density remained unchanged (NS; o).
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Figure 6: Eg5 inhibition reduces tumor angiogenesis in experimental tumor models(a-d) U87-derived gliomas on the CAM were treated with indicated doses of Eg5 inhibitors. Biomicroscopy images were taken at day 4 of tumor development. Note visible induction of necrosis (arrows in b-d) at the tumor surface after Eg5 inhibition. (e-f) Immunohistological examination of experimental glioma. A dense vascular network occurred in control tumors, whereas Eg5 inhibition leads to reduced tumor angiogenesis underneath the tumor surface (asterisk), denoted by the dashed line (f), especially at two treatments of DMN (g) or ISP (h) per day. (i) A significantly higher number of poorly vascularized tumors (+) was within the ISP-treated group, whereas few tumors treated with ISP appeared highly vascularized (+++). (j) Ispinesib also reduced tumor size (representative tumors are shown); (k) weight (P= 0.0037) and (i) vascular density (P=0.0184) in an orthotopic renal cell carcinoma model. (j). Short-term treatment (48h) with ispinesib lead to a significant increase of dilated capillaries (P=0.0284; n, p) whereas overall vascular density remained unchanged (NS; o).

Mentions: Experimental gliomas grown on the chicken CAM are accessible to topical treatment with chemical tyrosine kinase inhibitors such as imatinib mesylate or PTK787 [23]. During the three-day anti-Eg5 treatment period, no measurable tumor size reduction occurred. However, biomicroscopic observation of the tumors at day 4 showed increasing whitish areas at the tumor surface (Fig. 6a-d). These areas contain necrotic cells, as confirmed by subsequent histology (Fig. 6f-h). Immunohistological analysis of the tumors showed reduced numbers of angiogenic capillaries after treatment with a single dose of DMN per day (Fig. 6f). This effect was much stronger after two DMN doses; the tumors were almost completely devoid of blood vessels, whereas tumor cells apart from the necrotic areas appeared normal (Fig. 6g). Tumor cell morphology evidenced by vimentin staining was not altered by DMN treatment, even at the highest doses. Two treatments of ispinesib caused widespread destruction of cellular components of the tumor as evidenced by non-specific staining of dead cells for sambucus nigra agglutinin (SNA lectin), clearly visible below the surface (Fig. 6h). A significantly higher number of ispinesib-treated tumors were found in the poorly vascularized group (P<0.005) and accordingly, only a small number of ispinesib-treated tumors were classified in the highly vascularized group (P<0.005; Fig 6i).


Impaired angiogenesis and tumor development by inhibition of the mitotic kinesin Eg5.

Exertier P, Javerzat S, Wang B, Franco M, Herbert J, Platonova N, Winandy M, Pujol N, Nivelles O, Ormenese S, Godard V, Becker J, Bicknell R, Pineau R, Wilting J, Bikfalvi A, Hagedorn M - Oncotarget (2013)

Eg5 inhibition reduces tumor angiogenesis in experimental tumor models(a-d) U87-derived gliomas on the CAM were treated with indicated doses of Eg5 inhibitors. Biomicroscopy images were taken at day 4 of tumor development. Note visible induction of necrosis (arrows in b-d) at the tumor surface after Eg5 inhibition. (e-f) Immunohistological examination of experimental glioma. A dense vascular network occurred in control tumors, whereas Eg5 inhibition leads to reduced tumor angiogenesis underneath the tumor surface (asterisk), denoted by the dashed line (f), especially at two treatments of DMN (g) or ISP (h) per day. (i) A significantly higher number of poorly vascularized tumors (+) was within the ISP-treated group, whereas few tumors treated with ISP appeared highly vascularized (+++). (j) Ispinesib also reduced tumor size (representative tumors are shown); (k) weight (P= 0.0037) and (i) vascular density (P=0.0184) in an orthotopic renal cell carcinoma model. (j). Short-term treatment (48h) with ispinesib lead to a significant increase of dilated capillaries (P=0.0284; n, p) whereas overall vascular density remained unchanged (NS; o).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 6: Eg5 inhibition reduces tumor angiogenesis in experimental tumor models(a-d) U87-derived gliomas on the CAM were treated with indicated doses of Eg5 inhibitors. Biomicroscopy images were taken at day 4 of tumor development. Note visible induction of necrosis (arrows in b-d) at the tumor surface after Eg5 inhibition. (e-f) Immunohistological examination of experimental glioma. A dense vascular network occurred in control tumors, whereas Eg5 inhibition leads to reduced tumor angiogenesis underneath the tumor surface (asterisk), denoted by the dashed line (f), especially at two treatments of DMN (g) or ISP (h) per day. (i) A significantly higher number of poorly vascularized tumors (+) was within the ISP-treated group, whereas few tumors treated with ISP appeared highly vascularized (+++). (j) Ispinesib also reduced tumor size (representative tumors are shown); (k) weight (P= 0.0037) and (i) vascular density (P=0.0184) in an orthotopic renal cell carcinoma model. (j). Short-term treatment (48h) with ispinesib lead to a significant increase of dilated capillaries (P=0.0284; n, p) whereas overall vascular density remained unchanged (NS; o).
Mentions: Experimental gliomas grown on the chicken CAM are accessible to topical treatment with chemical tyrosine kinase inhibitors such as imatinib mesylate or PTK787 [23]. During the three-day anti-Eg5 treatment period, no measurable tumor size reduction occurred. However, biomicroscopic observation of the tumors at day 4 showed increasing whitish areas at the tumor surface (Fig. 6a-d). These areas contain necrotic cells, as confirmed by subsequent histology (Fig. 6f-h). Immunohistological analysis of the tumors showed reduced numbers of angiogenic capillaries after treatment with a single dose of DMN per day (Fig. 6f). This effect was much stronger after two DMN doses; the tumors were almost completely devoid of blood vessels, whereas tumor cells apart from the necrotic areas appeared normal (Fig. 6g). Tumor cell morphology evidenced by vimentin staining was not altered by DMN treatment, even at the highest doses. Two treatments of ispinesib caused widespread destruction of cellular components of the tumor as evidenced by non-specific staining of dead cells for sambucus nigra agglutinin (SNA lectin), clearly visible below the surface (Fig. 6h). A significantly higher number of ispinesib-treated tumors were found in the poorly vascularized group (P<0.005) and accordingly, only a small number of ispinesib-treated tumors were classified in the highly vascularized group (P<0.005; Fig 6i).

Bottom Line: Chemical blockade of Eg5 inhibits endothelial cell proliferation and migration in vitro.Mitosis-independent vascular outgrowth in aortic ring cultures is strongly impaired after Eg5 or Mklp2 protein inhibition.In vivo, interfering with KIF11/Eg5 function causes developmental and vascular defects in zebrafish and chick embryos and potent inhibition of tumor angiogenesis in experimental tumor models.

View Article: PubMed Central - PubMed

Affiliation: University Bordeaux, LAMC, UMR 1029, F-33405 Talence, France.

ABSTRACT
Kinesin motor proteins exert essential cellular functions in all eukaryotes. They control mitosis, migration and intracellular transport through interaction with microtubules. Small molecule inhibitors of the mitotic kinesin KiF11/Eg5 are a promising new class of anti-neoplastic agents currently evaluated in clinical cancer trials for solid tumors and hematological malignancies. Here we report induction of Eg5 and four other mitotic kinesins including KIF20A/Mklp2 upon stimulation of in vivo angiogenesis with vascular endothelial growth factor-A (VEGF-A). Expression analyses indicate up-regulation of several kinesin-encoding genes predominantly in lymphoblasts and endothelial cells. Chemical blockade of Eg5 inhibits endothelial cell proliferation and migration in vitro. Mitosis-independent vascular outgrowth in aortic ring cultures is strongly impaired after Eg5 or Mklp2 protein inhibition. In vivo, interfering with KIF11/Eg5 function causes developmental and vascular defects in zebrafish and chick embryos and potent inhibition of tumor angiogenesis in experimental tumor models. Besides blocking tumor cell proliferation, impairing endothelial function is a novel mechanism of action of kinesin inhibitors.

Show MeSH
Related in: MedlinePlus