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

Purpose: The abnormal function of tumor blood vessels causes tissue hypoxia, promoting disease progression and treatment resis-tance. Although tumor microenvironment normalization strategies can alleviate hypoxia globally, how local oxygen levels change is not known because of the inability to longitudinally assess vascular and interstitial oxygen in tumors with sufficient resolution. Understand -ing the spatial and temporal heterogeneity should help improve the outcome of various normalization strategies. Experimental Design: We developed a multiphoton phospho-rescence 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 an 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, intervessel heteroge-neity in tumors is associated with heterogeneous response to DC101 and losartan. Using both vascular and stromal normalizing agents, we show that spatial heterogeneity in oxygen levels persists, 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.

Automated summary

This study reveals the spatial and temporal heterogeneity in tumor vessel function and oxygen levels, which can affect the response to therapeutic agents. The findings suggest that optimizing the dose and schedule of vascular and stromal normalizing strategies should take into account the dynamic changes in tumor vasculature.