Multiphoton Phosphorescence Quenching Microscopy Reveals Kinetics of Tumor Oxygenation during Antiangiogenesis and Angiotensin Signaling Inhibition

Abstract

Purpose: The abnormal function of tumor blood vessels causes tissue hypoxia, promoting disease progression and treatment resistance. Although tumor microenvironment normalization strategies can alleviate hypoxia globally, how local oxygen levels change is not known because of inability to longitudinally assess vascular and interstitial oxygen in tumors with sufficient resolution. Understanding the spatial and temporal heterogeneity should help improve the outcome of various normalization strategies. Experimental Design: We developed a multiphoton phosphorescence quenching microscopy system using a low-molecular weight palladium porphyrin probe to measure perfused vessels, oxygen tension and their spatial correlations in vivo in mouse skin, bone marrow, and four different tumor models. Further, we measured the temporal and spatial changes in oxygen and vessel perfusion in tumors in response to anti-VEGFR2 antibody (DC101) and an angiotensin-receptor blocker (losartan). Results: We found that vessel function was highly dependent on tumor type. Although some tumors had vessels with greater oxygen carrying ability than those of normal skin, most tumors had inefficient vessels. Further, inter-vessel heterogeneity in tumors associated with heterogeneous response to DC101 and losartan. Using both vascular and stromal normalizing agents, we show that spatial heterogeneity in oxygen levels persist, even with reductions in mean extravascular hypoxia. Conclusions: High-resolution spatial and temporal responses of tumor vessels to two agents known to improve vascular perfusion globally reveal spatially heterogeneous changes in vessel structure and function. These dynamic vascular changes should be considered in optimizing the dose and schedule of vascular and stromal normalizing strategies to improve the therapeutic outcome.