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Bevacizumab and intraocular tumors: an intriguing paradox.

el Filali M, Ly LV, Luyten GP, Versluis M, Grossniklaus HE, van der Velden PA, Jager MJ - Mol. Vis. (2012)

Bottom Line: We determined the effect of bevacizumab on the growth of B16F10 cells inside the eye and on B16F10 and UM cells cultured in vitro.In addition, the effect of bevacizumab on in vitro growth of B16F10 and human UM cells and on the expression of VEGF-A, GLUT-1, and HIF-1α was evaluated.Bevacizumab did not affect proliferation of B16F10 cells in vitro, while it inhibited UM cell proliferation.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, LUMC, Leiden, the Netherlands. m.el_filali@lumc.nl

ABSTRACT

Purpose: Bevacizumab, a humanized monoclonal antibody to vascular endothelial growth factor-A (VEGF-A), was originally developed as an anti-tumor treatment. In ocular oncology, it is being used to treat macular edema due to radiation retinopathy, but it may also be useful for the treatment of primary uveal melanoma (UM) or its metastases. We determined the effect of bevacizumab on the growth of B16F10 cells inside the eye and on B16F10 and UM cells cultured in vitro.

Methods: B16F10 melanoma cells were placed into the anterior chamber of the eye of C57Bl/6 mice and tumor growth was monitored after injection of different doses of bevacizumab or mock injection. In addition, the effect of bevacizumab on in vitro growth of B16F10 and human UM cells and on the expression of VEGF-A, GLUT-1, and HIF-1α was evaluated.

Results: Following intraocular injection of bevacizumab into murine B16 tumor-containing eyes, an acceleration of tumor growth was observed, with the occurrence of anterior chamber hemorrhages. Bevacizumab did not affect proliferation of B16F10 cells in vitro, while it inhibited UM cell proliferation. Expression analysis demonstrated that addition of bevacizumab under hypoxic conditions induced VEGF-A, GLUT-1 and HIF-1α in B16F10 cells as well as in UM cell lines and two of four primary UM tumor cultures.

Conclusions: In contrast with expectations, intraocular injection of bevacizumab stimulated B16F10 melanoma growth in murine eyes. In vitro exposure of B16 and human UM cells to bevacizumab led to paradoxical VEGF-A upregulation. The use of VEGF inhibitors for treatment of macular edema (due to radiation retinopathy) after irradiation of UM should be considered carefully, because of the possible adverse effects on residual UM cells.

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VEGF-A expression of B16F10 cells after bevacizumab treatment The effect of bevacizumab on VEGF-A expression in B16F10 cells was determined using qPCR analysis (top) and ELISA (bottom). The amount of VEGF-A mRNA expression was measured with qPCR under normoxic (black) and hypoxic (gray) conditions after 24 h with different doses bevacizumab treatment. Expression is demonstrated in normalized fold. VEGF-A mRNA expression increased in a dose-dependent manner after treatment with bevacizumab, when cells were cultured under hypoxic conditions. An ELISA was used to measure the amount of VEGF-A protein in the supernatant of UM cell lines after hypoxic exposure for 24 h. Shown is the amount of VEGF-A protein (pg/ml). Paradoxically, the amount of produced protein is reduced in the presence of bevacizumab. AVA1=equivalent human dose: 1.25 mg/5.46 ml; AVA10=10 times the equivalent human dose: 12.5 mg/ 5.46 ml; AVA20=20 times the equivalent human dose: 25 mg/5.46 ml; CO=control group: culture medium.
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f4: VEGF-A expression of B16F10 cells after bevacizumab treatment The effect of bevacizumab on VEGF-A expression in B16F10 cells was determined using qPCR analysis (top) and ELISA (bottom). The amount of VEGF-A mRNA expression was measured with qPCR under normoxic (black) and hypoxic (gray) conditions after 24 h with different doses bevacizumab treatment. Expression is demonstrated in normalized fold. VEGF-A mRNA expression increased in a dose-dependent manner after treatment with bevacizumab, when cells were cultured under hypoxic conditions. An ELISA was used to measure the amount of VEGF-A protein in the supernatant of UM cell lines after hypoxic exposure for 24 h. Shown is the amount of VEGF-A protein (pg/ml). Paradoxically, the amount of produced protein is reduced in the presence of bevacizumab. AVA1=equivalent human dose: 1.25 mg/5.46 ml; AVA10=10 times the equivalent human dose: 12.5 mg/ 5.46 ml; AVA20=20 times the equivalent human dose: 25 mg/5.46 ml; CO=control group: culture medium.

Mentions: As intraocular and intratumoral hemorrhages were visible in eyes receiving bevacizumab, we wondered whether the anti-VEGF-A treatment with bevacizumab had a paradoxical stimulatory effect on VEGF-A gene expression. Indeed, bevacizumab induced VEGF-A mRNA expression in a dose-dependent manner when cells were cultured under hypoxic conditions. Under normoxia, baseline VEGF-A expression is relatively high but not affected by bevacizumab treatment (Figure 4).


Bevacizumab and intraocular tumors: an intriguing paradox.

el Filali M, Ly LV, Luyten GP, Versluis M, Grossniklaus HE, van der Velden PA, Jager MJ - Mol. Vis. (2012)

VEGF-A expression of B16F10 cells after bevacizumab treatment The effect of bevacizumab on VEGF-A expression in B16F10 cells was determined using qPCR analysis (top) and ELISA (bottom). The amount of VEGF-A mRNA expression was measured with qPCR under normoxic (black) and hypoxic (gray) conditions after 24 h with different doses bevacizumab treatment. Expression is demonstrated in normalized fold. VEGF-A mRNA expression increased in a dose-dependent manner after treatment with bevacizumab, when cells were cultured under hypoxic conditions. An ELISA was used to measure the amount of VEGF-A protein in the supernatant of UM cell lines after hypoxic exposure for 24 h. Shown is the amount of VEGF-A protein (pg/ml). Paradoxically, the amount of produced protein is reduced in the presence of bevacizumab. AVA1=equivalent human dose: 1.25 mg/5.46 ml; AVA10=10 times the equivalent human dose: 12.5 mg/ 5.46 ml; AVA20=20 times the equivalent human dose: 25 mg/5.46 ml; CO=control group: culture medium.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3472924&req=5

f4: VEGF-A expression of B16F10 cells after bevacizumab treatment The effect of bevacizumab on VEGF-A expression in B16F10 cells was determined using qPCR analysis (top) and ELISA (bottom). The amount of VEGF-A mRNA expression was measured with qPCR under normoxic (black) and hypoxic (gray) conditions after 24 h with different doses bevacizumab treatment. Expression is demonstrated in normalized fold. VEGF-A mRNA expression increased in a dose-dependent manner after treatment with bevacizumab, when cells were cultured under hypoxic conditions. An ELISA was used to measure the amount of VEGF-A protein in the supernatant of UM cell lines after hypoxic exposure for 24 h. Shown is the amount of VEGF-A protein (pg/ml). Paradoxically, the amount of produced protein is reduced in the presence of bevacizumab. AVA1=equivalent human dose: 1.25 mg/5.46 ml; AVA10=10 times the equivalent human dose: 12.5 mg/ 5.46 ml; AVA20=20 times the equivalent human dose: 25 mg/5.46 ml; CO=control group: culture medium.
Mentions: As intraocular and intratumoral hemorrhages were visible in eyes receiving bevacizumab, we wondered whether the anti-VEGF-A treatment with bevacizumab had a paradoxical stimulatory effect on VEGF-A gene expression. Indeed, bevacizumab induced VEGF-A mRNA expression in a dose-dependent manner when cells were cultured under hypoxic conditions. Under normoxia, baseline VEGF-A expression is relatively high but not affected by bevacizumab treatment (Figure 4).

Bottom Line: We determined the effect of bevacizumab on the growth of B16F10 cells inside the eye and on B16F10 and UM cells cultured in vitro.In addition, the effect of bevacizumab on in vitro growth of B16F10 and human UM cells and on the expression of VEGF-A, GLUT-1, and HIF-1α was evaluated.Bevacizumab did not affect proliferation of B16F10 cells in vitro, while it inhibited UM cell proliferation.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, LUMC, Leiden, the Netherlands. m.el_filali@lumc.nl

ABSTRACT

Purpose: Bevacizumab, a humanized monoclonal antibody to vascular endothelial growth factor-A (VEGF-A), was originally developed as an anti-tumor treatment. In ocular oncology, it is being used to treat macular edema due to radiation retinopathy, but it may also be useful for the treatment of primary uveal melanoma (UM) or its metastases. We determined the effect of bevacizumab on the growth of B16F10 cells inside the eye and on B16F10 and UM cells cultured in vitro.

Methods: B16F10 melanoma cells were placed into the anterior chamber of the eye of C57Bl/6 mice and tumor growth was monitored after injection of different doses of bevacizumab or mock injection. In addition, the effect of bevacizumab on in vitro growth of B16F10 and human UM cells and on the expression of VEGF-A, GLUT-1, and HIF-1α was evaluated.

Results: Following intraocular injection of bevacizumab into murine B16 tumor-containing eyes, an acceleration of tumor growth was observed, with the occurrence of anterior chamber hemorrhages. Bevacizumab did not affect proliferation of B16F10 cells in vitro, while it inhibited UM cell proliferation. Expression analysis demonstrated that addition of bevacizumab under hypoxic conditions induced VEGF-A, GLUT-1 and HIF-1α in B16F10 cells as well as in UM cell lines and two of four primary UM tumor cultures.

Conclusions: In contrast with expectations, intraocular injection of bevacizumab stimulated B16F10 melanoma growth in murine eyes. In vitro exposure of B16 and human UM cells to bevacizumab led to paradoxical VEGF-A upregulation. The use of VEGF inhibitors for treatment of macular edema (due to radiation retinopathy) after irradiation of UM should be considered carefully, because of the possible adverse effects on residual UM cells.

Show MeSH
Related in: MedlinePlus