Modulating tumor acidity with hydroxyethyl starch-based nanoparticles by targeting CA9 to eliminate cancer stem cells and overcome immunosuppression.

The acidic microenvironment in solid tumors, primarily driven by Warburg effect, promotes tumor progression, immune evasion, and resistance to therapy. Cancer stem cells (CSCs), a critical subset within tumor tissues, exacerbate this acidity through overexpression of pH-regulating proteins such as carbonic anhydrase IX (CA9), which plays a pivotal role in maintaining pH homeostasis, contributes to immune suppression, and sustains CSC stemness and proliferation. In this study, we designed CA9 inhibitor (CAi) coupled hydroxyethyl starch-based nanoparticles (CHHD-Cu NPs) that integrate doxorubicin (DOX) mediated chemotherapy with copper ions (Cu) mediated chemodynamic therapy to target and eliminate CA9-expressing CSCs. Upon administration, CHHD-Cu NPs bind to CA9 and disrupt pH regulation, thereby lowering intracellular pH and raising extracellular pH. This pH modulation enhances intracellular releases of DOX and Cu and alleviates extracellular acidity to boost effector T cells infiltration and activity. Our rationally designed CHHD-Cu NPs eliminate CSCs in two ways: firstly, by robust intracellular DOX- and copper-induced cytotoxicity, and secondly, via pH modulation-mediated activation of anti-tumor immunity. Our strategy offers novel approaches for treatment of immunosuppressive solid tumors.