Abstract
Purpose::
Previously, receptor tyrosine kinase inhibitors (RTKi), possibly acting against VEGF signaling, have shown efficacy against ocular angiogenesis in mouse models of oxygen-induced retinopathy and choroidal neovascularization (CNV). We examined the effect of a novel RTKi, AL-39324 (KDR IC50=4nM), via systemic delivery on experimental CNV lesion volume.
Methods::
Experimental CNV was induced in adult C57BL6/J mice (n=40) by application of three laser burns (523 nm diode laser, 150 mW, 100 msec, 50 µm spot size) to rupture Bruch’s membrane in one eye of each animal at day 0 (D0). Animals were then randomly divided into two treatment arms: prevention arm (n=20) were immediately given 0, 1.5, 5 or 10mg/Kg AL-39324 PO BID continuing for the duration of the study; intervention arm (n=20) were started on the same dosing regimen on D7. All animals were euthanized on D14, at which time the eyes were enucleated, immersion fixed and dissected. The posterior cups were then reacted with rhodamine-conjugated R. communis agglutinin I (a specific vascular stain) and finally mounted flat by 4-5 radial cuts. Volumetric assessment of vasculature was accomplished by digital image capture using a confocal microscope, collecting images (2-µm z-steps) throughout the entire depth of each lesion. Computerized density slicing was then used to measure vascular area in each z-slice, and the volume was calculated by summing areas and multiplying by slice depth.
Results::
In the prevention arm, all tested doses of AL-39324 (3, 10 or 20 mg/Kg/d) significantly decreased CNV lesion volume as compared to vehicle alone (down 60%, P=0.03; down 71%, P=0.04; down 83%, P=0.02). In the intervention arm, only the highest tested dose showed a statistically significant reduction in CNV volume (down 77%, P=0.02); however, a dose-dependent trend was supported by the mid dose of 10 mg/Kg/d (down 25%), although it did not achieve statistical significance.
Conclusions::
Volumetric measurements more accurately reflect the efficacy of treatment since a CNV lesion is a complex three-dimensional object. For example, 2 CNV lesions may have similar maximal cross-sectional areas, but they may differ substantially in depth. Therefore, measurement of maximal area alone may yield suboptimal results. In this study, oral delivery of the RTKi, AL-39324, provided a significant, dose-dependent suppression of vascular volume in experimental CNV.
Keywords: choroid: neovascularization • growth factors/growth factor receptors • receptors: pharmacology/physiology