Misonidazole binding in SCCVII tumors in relation to the tumor blood supply.

Misonidazole (MISO) binding was examined in murine squamous cell carcinomas as a function of distance from the tumor blood supply. C3H mice bearing subcutaneous SCCVII tumors were injected intraperitoneally with 3H-MISO followed at various times later by intravenous injection or infusion of the fluorescent perfusion probe, Hoechst 33342. Tumors were then excised, and single cell suspensions prepared for fluorescence activated cell sorting. Cells sorted on the basis of Hoechst 33342 fluorescence were examined for 3H-MISO content by liquid scintillation counting or autoradiography. MISO content in the 10% of cells most distant from the blood supply was as much as 8 times greater than the amount in the 10% of cells closest to the blood supply. The largest differentials in MISO content between dimly and brightly staining regions were obtained if (a) tumors greater than about 300 mg were used, (b) at least 3 hr were allowed for MISO metabolism and binding prior to analysis, and (c) cell sorting was performed on the basis of concentration of Hoechst (correcting for cell size) rather than on the basis of total fluorescence intensity per cell. As tumors enlarged, MISO content increased even in the cells closest to the blood supply suggesting a decrease in net tumor oxygenation perhaps caused by intermittent hypoxia.