Innovative methods for engineering cancer cell membranes promise to manipulate cell–cell interactions and boost cell-based cancer therapeutics. Here, we illustrate an in situ approach to selectively modify cancer cell membranes by employing an enzyme-instructed peptide self-assembly (EISA) strategy. Using three phosphopeptides (pY1, pY2, and pY3) targeting the membrane-bound epidermal growth factor receptor (EGFR) and differing in just one phosphorylated tyrosine, we reveal that site-specific phosphorylation patterns in pY1, pY2, and pY3 can distinctly command their preorganization levels, self-assembling kinetics, and spatial distributions of the resultant peptide assemblies in cellulo. Overall, pY1 is the most capable of producing preorganized assemblies and shows the fastest dephosphorylation reaction in the presence of alkaline phosphatase (ALP), as well as the highest binding affinity for EGFR after dephosphorylation. Consequently, pY1 exhibits the greatest capacity to construct stable peptide assemblies on cancer cell membranes with the assistance of both ALP and EGFR. We further use peptide–protein and peptide–peptide co-assembly strategies to apply two types of antigens, namely ovalbumin (OVA) protein and dinitrophenyl (DNP) hapten respectively, on cancer cell membranes. This study demonstrates a very useful technique for the in situ construction of membrane-bound peptide assemblies around cancer cells and implies a versatile strategy to artificially enrich cancer cell membrane components for potential cancer immunotherapy.

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Protein preorganization is ubiquitous in nature and enables subtle structural rearrangements and precise function executions, making it an attractive approach for generating functional peptide materials. Here, a phosphorylation‐dependent pre‐organization approach to optimize the therapeutic outcomes of a self‐assembling peptide targeting programmed cell death ligand 1 (PD‐L1) is reported. Upon incorporating the imaging probe cyanine 5.5 (Cy5.5) and therapeutic drug 10‐hydroxycamptothecin (HCPT), three phosphorylated precursors (1P, 2P, and 3P) containing identical peptide sequences with different tyrosine phosphorylation sites are constructed. The presence of a phosphate group can stabilize the self‐assembling peptide in a well‐defined, preorganized state. Moreover, variations in the phosphorylation sites lead to different preorganized secondary structures, dephosphorylation rates, and PD‐L1 binding affinities. Responding to alkaline phosphatase dephosphorylation, subtle structural transformation to more ordered states, an increase in binding affinity against PD‐L1, and rapid cellular internalization are observed. Compared with the unphosphorylated control (0P), the preorganization of 1P, 2P, and 3P can optimize tumor retention and resultant therapeutic performance, of which 3P maximizes the final therapeutic outcomes, leading to a 70.3% tumor inhibition rate. This study highlights the great potential to control the subtle structural transformation and therapeutic effects of self‐assembling peptides by preparing their preorganized entities.

The development of novel vaccine adjuvants is essential for the production of modern vaccines against infectious agents and cancer. We recently reported a supramolecular hydrogel of a self-assembling D-tetra-peptide named...