Engineering a hypoxia-generating hydrogel to enhance the anticancer activity of prodrug Evofosfamide in lung cancer models

Tumor hypoxia is a hallmark of solid tumors that contributes to therapeutic resistance while also creating an opportunity for selective drug activation. Hypoxia-activated prodrugs such as evofosfamide are designed to exploit these low-oxygen regions, but their clinical efficacy has been limited by the uneven and insufficient distribution of hypoxia within tumors. Here, we developed a shear-thinning hydrogel that locally reduces oxygen levels while serving as a sustained drug delivery system. The material incorporates a laccase-gallic acid enzymatic reaction that rapidly consumes oxygen and establishes hypoxic conditions within the gel, even under normoxic environments. Evofosfamide was successfully loaded into the hydrogel and exhibited controlled, near zero-order release over seven days. Over the 7-day incubation period, the hydrogel maintained its structural integrity, with Evo-loaded STHH retaining positive relative swelling, more than 80% of its initial dry mass, and an interconnected porous architecture, as confirmed by scanning electron microscopy (SEM). In an A549 lung cancer model, evofosfamide showed minimal cytotoxicity under normoxia but significantly increased activity under hypoxia. Notably, the hydrogel reproduced this effect under normoxic culture conditions by creating a localized low-oxygen environment, resulting in sustained reductions in cell viability and proliferation in both 2D and cell aggregate models. Gene expression analysis further revealed that HIF1A mRNA expression was significantly increased in the hydrogel condition compared to normoxic and hypoxic groups. Together, these results demonstrate that this platform generates localized low-oxygen conditions that enhance the activity of hypoxia-responsive therapeutics while maintaining controlled drug delivery, offering a strategy to improve the performance of hypoxia-targeted cancer treatments.