(A) CendR uptake in live tumor slices responds to nutrient availability. Live tumor
tissue slices (PPC1 xenografts; 4T1 mouse mammary tumors) were obtained and incubated
under the indicated conditions (y-axis) for 6 h. The slices were then suspended in the
same kind of media containing 15 pM R-Ag, and incubated for another two hours at
37°C with gentle shaking on a rocker before etching and fixation. The fluorescence
intensity of R-Ag per cell was quantified using ImageJ, normalized to slices cultured in
complete media and shown as relative uptake (x-axis). **P<0.01 and
***P<0.001 (Student's t-test) in comparison to
complete media conditions of the corresponding tumor types. Error bars indicate SEM (4-7
replicates).
(B) Representative images of R-Ag internalization into PPC1 tumor slices. R-Ag, red;
nuclei, blue. Scale bar, 50 μm.
(C) Representative image of glucose uptake into 4T1 tumors after GLUT IV inhibitor
treatment. Mice bearing 4T1 tumors were treated with intratumoral injections of vehicle
only or GLUT IV inhibitor as described in Methods. IRDye 800CW 2-deoxy-D-glucose (2-DG) in
10 nmol in 100 μl PBS was then injected intravenously into a tail vein and 24 h
later, the fluorescence intensity of 2-DG was captured using Xenogen IVIS 200 imager
(PerkinElmer Inc.). The experiment was conducted twice with at least three mice per
group.
(D) iRGD accumulation in tumors responds to glucose deprivation. FAM
(fluorescein)-labeled iRGD (200 μg in 100 μl PBS) was intravenously
injected into mice bearing 4T1 tumors treated with vehicle alone or with the GLUT IV
inhibitor. After 2 h of circulation, the mice were perfused with PBS containing 1%
BSA to remove peptide remaining in the circulation. The tumors were excised, fixed and
sectioned. FAM-iRGD in the tumors was detected with rabbit anti-FITC antibody. The average
signal intensity per cell was normalized to vehicle-treated tumors as relative uptake. The
experiment was conducted twice with at least three mice per group, and error bars indicate
SEM. *P<0.05 (Student's t-test).