Rapamycin Crosses the Blood–Brain Barrier and Blocks mTOR in Tumor Tissue

(A) Rapamycin concentrations in tumor tissue (filled squares) and peripheral blood (empty circles) grouped by rapamycin dose cohorts (2 mg, 5 mg, or 10 mg per os daily). Intratumoral rapamycin concentration for patient 11 could not be determined due to insufficient frozen tumor material. The last preoperative dose of rapamycin was given on the day of craniotomy and peripheral blood was collected within 24 h of surgery.

(B) Quantification of mTOR activity in tumor tissue by immunohistochemistry. The cartoon to the left depicts the S6 kinase 1 branch of the mTOR signaling pathway resulting in phosphorylation of S6 ribosomal protein at serine 235/236 and serine 240/244. The panel to the right shows a comparison between immunoblotting (top) and IHC (bottom) for the determination of S6 phosphorylation in tumor tissue from rapamycin patients 1, 2, and 3. The fold change in serine 235/236 phosphorylation between S2 and S1 for patients 1, 2, and 3 were 0.45, 1.01, and 0.45, respectively (see Figure S2A).

(C) Changes in S6 phosphorylation between S2 and S1 (y-axis: ratio of S6 phosphorylation in S2 sample to S6 phosphorylation in S1 sample) for all patients for whom matched S1 and S2 samples were available (14/15 rapamycin patients and 9/9 patients who did not receive rapamycin). S6 phosphorylation was determined by IHC using phosphosite-specific antibodies against serine 235/236 (left) and serine 240/244 (right). Please see Figures S1 and S2 for details regarding IHC scoring method and results for individual tumors. p-values for the difference in mean S2/S1 ratios for each group were determined using the Kruskal Wallace test.