A33 aptamer targeting ERBB-3/HER3: a promising strategy to overcome non-genomic drug resistance to EGFR inhibitors

Therapies targeting the epidermal growth factor receptor (EGFR) are widely used to treat several solid tumors, including lung, colorectal, pancreatic, and head and neck cancers. However, although many patients initially respond, most eventually develop resistance. Two major mechanisms drive this process: secondary EGFR mutations and activation of bypass signaling pathways. Among non-genomic resistance mechanisms, HER3 has emerged as a well-characterized compensatory pathway and a promising therapeutic target. In this study, we investigated a novel strategy to inhibit the HER3 axis using aptamers, an innovative class of therapeutic agents. Aptamers are short, single-stranded DNA or RNA oligonucleotides generated in vitro through the multi-step SELEX process. To select the A33 aptamer, a large random DNA library was incubated with IgB-3, a fusion protein composed of the extracellular domain of HER3 linked to the Fc domain of human IgG1. The resulting 40-nucleotide sequence demonstrated high binding specificity to HER3 compared to a control primer across multiple HER3-expressing cell lines. Since HER3 expression increases following anti-EGFR therapies and contributes to resistance, we evaluated A33 in this context. A33 markedly inhibited HER3 phosphorylation both as a single agent and in combination with ErbB-targeted therapies, leading to reduced downstream signaling activation. It also suppressed HER2 phosphorylation, suggesting interference with the formation of active HER2/HER3 heterodimers. Similar inhibitory effects were observed under NRG1 stimulation. Confocal microscopy confirmed that A33 binds HER3 at the cell membrane without inducing receptor internalization or degradation. Preliminary functional assays showed that A33 significantly reduced tumor cell proliferation and migration in both 2D and 3D models, particularly when combined with anti-EGFR treatments. Overall, these findings support the therapeutic potential of DNA aptamers targeting cell surface proteins, identifying A33 as a selective HER3 inhibitor and a promising combinatorial agent in targeted cancer therapy.