Protein drugs are increasingly used as therapeutics for the treatment of cancer. However, their inherent drawbacks, such as poor stability, low cell membrane and tissue permeability, lack of tumor selectivity, and severe side effects, limit their wide applications in cancer therapy. Herein, screening of a thermo‐pH‐sensitive polymer–glucose oxidase conjugate that can controllably self‐assemble into nanoparticles with improved stability is reported. The size, surface charge, and bioactivity of the conjugate can be tuned by adjustment of the solution temperature and pH. The cellular uptake, intracellular hydrogen peroxide generation, and tumor cell spheroid penetration of the conjugate are greatly enhanced under the acidic tumor microenvironment, leading to increased cytotoxicity to tumor cells. Upon a single intratumoural injection, the conjugate penetrates into the whole tumor tissue but hardly diffuses into the normal tissues, resulting in the eradication of the tumors in mice without perceivable side effects. Simultaneously, the conjugate induces a robust antitumor immunity to efficiently inhibit the growth of distant tumors, especially in combination with an immune checkpoint inhibitor. These findings provide a novel and general strategy to make multifunctional protein‐polymer conjugates with responsiveness to the acidic tumor microenvironment for selective tumor therapy.

No abstract

L‐Asparaginase (ASP) is well‐known for its excellent efficacy in treating hematological malignancies. Unfortunately, the intrinsic shortcomings of ASP, namely high immunogenicity, severe toxicity, short half‐life, and poor stability, restrict its clinical usage. Poly(ethylene glycol) conjugation (PEGylation) of ASP is an effective strategy to address these issues, but it is not ideal in clinical applications due to complex chemical synthesis procedures, reduced ASP activity after conjugation, and pre‐existing anti‐PEG antibodies in humans. Herein, the authors genetically engineered an elastin‐like polypeptide (ELP)‐fused ASP (ASP‐ELP), a core‐shell structured tetramer predicted by AlphaFold2, to overcome the limitations of ASP and PEG‐ASP. Notably, the unique thermosensitivity of ASP‐ELP enables the in situ formation of a sustained‐release depot post‐injection with zero‐order release kinetics over a long time. The in vitro and in vivo studies reveal that ASP‐ELP possesses increased activity retention, improved stability, extended half‐life, mitigated immunogenicity, reduced toxicity, and enhanced efficacy compared to ASP and PEG‐ASP. Indeed, ASP‐ELP treatment in leukemia or lymphoma mouse models of cell line‐derived xenograft (CDX) shows potent anti‐cancer effects with significantly prolonged survival. The findings also indicate that artificial intelligence (AI)‐assisted genetic engineering is instructive in designing protein‐polypeptide conjugates and may pave the way to develop next‐generation biologics to enhance cancer treatment.