Lysosome-oriented, dual-stage pH-responsive polymeric micelles for β-lapachone delivery

beta-Lapachone (beta-lap), a novel anticancer agent, is bioactivated by NADP(H): quinone oxidoreductase 1 (NQO1), an enzyme over-expressed in numerous tumors, including lung, pancreas, breast, and prostate cancers. Fast renal clearance and methemoglobinemia/hemolytic side-effects from the clinical formulation (beta-lap-hydroxyl propyl-beta-cyclodextrin complex) hindered its clinical translation. Here, we investigated dual-model pH-responsive polymers for beta-lap delivery. Three pH-sensitive linkages, including acylhydrazone, ketal and imine bonds for beta-lap prodrug syntheses result in an aryl imine linkage as the most optimal linkage. The conversion to beta-lap was 2.8%, 4.5% and 100% at pH 7.4, 6.5 and 5.0 in 8 h, respectively. The beta-lap aryl imine prodrug conjugated ultra-pH-sensitive (UPS) polymer reached high beta-lap loading density (8.3%) and exhibited dual-stage responsiveness to pH variation. In pHs under pH(t), at stage I, micelles immediately dissociate and subsequently, entering stage II, micelles start to quickly release beta-lap. An in vitro release study showed that the micelles constantly release beta-lap (14.9 +/- 0.1%) at pHs above pHt in 72 h, whereas there was a boosted release of beta-lap (79.4 +/- 1.2%) at pH 5.0. The micelles' intracellular distribution being predominantly in the lysosome organelle guaranteed their pH-responsive dissociation and subsequently beta-lap controlled release. The M-P micelles retained NQO1-dependent cytotoxicity in A549 lung cancer cells, similar to the free drug in both efficacy and mechanism of cell death. The lysosome-oriented dual-stage ultra-pH-responsive beta-lap prodrug micelles potentially offer an alternative nanotherapeutic strategy for lung cancer, as well as other NQO1+ cancer therapies.