EGFRvIII is expressed only in tumor cells and strongly in glioblastoma and is considered a promising target in cancer diagnosis and therapy. Aptamers are synthetic single-stranded oligonucleotides that bind to biochemical target molecules with high binding affinity and specificity. This study examined the potential of the ⁶⁸Ga-NOTA-EGFRvIII aptamer as a nuclear imaging probe for visualizing EGFRvIII-expressing glioblastoma by positron emission tomography (PET). EGFRvIII aptamer was selected using the SELEX technology, and flow cytometry and fluorescence microscopy verified the high binding affinity to EGFRvIII positive U87MG vIII 4.12 glioma cells but not to EGFRvIII negative U87MG cells. The EGFRvIII aptamer was conjugated with a chelator (1,4,7-triazanonane-1,4,7-triyl)triacetic acid (NOTA) for ⁶⁸Ga-labeling. The ⁶⁸Ga-NOTA-EGFRvIII aptamer was prepared using the preconcentration-based labeling method with a high radiolabeling yield at room temperature. Ex vivo biodistribution analyses confirmed the significantly higher tumor uptake of the ⁶⁸Ga-NOTA-EGFRvIII aptamer in EGFRvIII-expressing xenograft tumors than that in EGFRvIII negative tumors, confirming the specific tumor uptake of the ⁶⁸Ga-NOTA-EGFRvIII aptamer in vivo. PET imaging studies revealed a high retention rate of the ⁶⁸Ga-NOTA-EGFRvIII aptamer in U87MG vIII 4.12 tumors but only low uptake levels in U87-MG tumors, suggesting that the ⁶⁸Ga-NOTA-EGFRvIII aptamer may be used as a PET imaging agent for EGFRvIII-expressing glioblastoma.

Background: CD25, the α-chain of the interleukin-2 (IL-2) receptor, is highly expressed on malignant cells and tumor-infiltrating regulatory T-cells (Tregs) in hematologic malignancies, making it an attractive therapeutic target for tumor elimination and immunomodulation. Methods: We developed a CD25-specific aptamer–drug conjugate (CD25-ApDC) by linking a CD25 aptamer to monomethyl auristatin E via a cathepsin B-cleavable Val-Cit linker. Results: The aptamer exhibited high affinity for CD25 (Kd = 16.4 ± 0.29 nM), rapid receptor-mediated uptake (half-time = 9.6 min), and selective inhibition of IL-2 signaling in CD25^high cells, with no activity in CD25^low cells. In vitro, CD25-ApDC induced selective cytotoxicity, confirmed by apoptosis and G2/M arrest in CD25-positive cancer cells while having no effect on CD25-negative cells. Co-culture studies confirmed selective depletion of CD25^high Treg-like cells, suggesting potential to relieve immune suppression within the tumor microenvironment. In vivo, CD25-ApDC achieved complete tumor remission in xenograft and disseminated models with optimized dosing, showing efficacy and tolerability comparable to Brentuximab vedotin. Increasing drug-to-aptamer ratios further enhanced outcomes, supporting flexible dosing strategies. Conclusions: These findings highlight CD25-ApDC as a promising therapeutic modality for hematologic malignancies, offering advantages in specificity, tissue penetration, and manufacturability over conventional antibody-based therapies.