Two new "protected" oxyphors for biological oximetry: properties and application in tumor imaging

We report the synthesis, calibration, and examples of application of two new phosphorescent probes, Oxyphor R4 and Oxyphor G4, optimized specifically for in vivo oxygen imaging by phosphorescence quenching. These "protected" dendritic probes can operate in either albumin-rich (blood plasma) or albumin-free (interstitial space) environments at all physiological oxygen concentrations, from normoxic to deep hypoxic conditions. Oxyphors R4 and G4 are derived from phosphorescent Pd-meso-tetra-(3,5-dicarboxyphenyl)-porphyrin (PdP) or Pd-meso-tetra-(3,5-dicarboxyphenyl)-tetrabenzoporphyrin (PdTBP), respectively, and possess features common for protected dendritic probes, i.e., hydrophobic dendritic encapsulation of phosphorescent metalloporphyrins and hydrophilic PEGylated periphery. The new Oxyphors are highly soluble in aqueous environments and do not permeate biological membranes. The probes were calibrated under physiological conditions (pH 6.4-7.8) and temperatures (22-38 °C), showing high stability, reproducibility of signals, and lack of interactions with biological solutes. Oxyphor G4 was used to dynamically image intravascular and interstitial oxygenation in murine tumors in vivo. The physiological relevance of the measurements was demonstrated by dynamically recording changes in tissue oxygenation during application of anesthesia (isofluorane). These experiments revealed that changes in isofluorane concentration significantly affect tissue oxygenation.