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We investigated how the dual inhibition of the molecular mechanism of the mammaliantarget of the rapamycin (mTOR) downstreams, P70S6 kinase (P70S6K) and eukaryoticinitiation factor 4E (eIF4E), can lead to a suppression of the proliferation and progression ofurothelial carcinoma (UC) in an orthotopic mouse non-muscle invasive bladder tumor(NMIBT) model. A KU-7-luc cell intravesically instilled orthotopic mouse NMIBC model wasmonitored using bioluminescence imaging (BLI) in vivo by interfering with differentmolecular components using rapamycin and siRNA technology. We then analyzed theeffects on molecular activation status, cell growth, proliferation, and progression. A highconcentration of rapamycin (10 μM) blocked both P70S6K and elF4E phosphorylation andinhibited cell proliferation in the KU-7-luc cells. It also reduced cell viability andproliferation more than the transfection of siRNA against p70S6K or elF4E. The groups withdual p70S6K and elF4E siRNA, and rapamycin reduced tumor volume and lamina propriainvasion more than the groups with p70S6K or elF4E siRNA instillation, although all groupsreduced photon density compared to the control. These findings suggest that both themTOR pathway downstream of eIF4E and p70S6K can be successfully inhibited by highdose rapamycin only, and p70S6K and Elf4E dual inhibition is essential to control bladdertumor growth and progression.

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