Novel Tumor Pretargeting System Based on Complementary l-Configured Oligonucleotides.

Area of Science:

• Biochemistry
• Radiopharmacology
• Molecular Biology

Background:

• Unnatural mirror image l-configured oligonucleotides (L-ONs) are promising for in vivo recognition systems in pretargeting.
• L-ONs offer high hybridization velocity, metabolic stability, and low non-specific binding compared to d-configured analogs.
• Pretargeting strategies enhance the efficacy of targeted therapies and diagnostics by separating antibody delivery from radiolabeling.

Purpose of the Study:

• To radiopharmacologically evaluate a novel L-ONs-based pretargeting system using cetuximab (C225) for targeting the epidermal growth factor receptor (EGFR).
• To assess the feasibility of using ⁶⁴Cu-labeled L-ONs for positron emission tomography (PET) imaging in EGFR-expressing tumors.
• To determine optimal pretargeting intervals and evaluate the performance of L-ONs in vivo.

Main Methods:

• Conjugation of a PEGylated 17mer-L-DNA with p-SCN-Bn-NOTA (NOTA') for ⁶⁴Cu radiolabeling.
• Modification of cetuximab (C225) with complementary 17mer-L-DNA (c-L-DNA) and NOTA'.
• In vitro characterization (hybridization, binding assays) and in vivo PET/biodistribution studies in FaDu tumor-bearing mice.

Main Results:

• Modified C225 derivatives showed high binding affinity (low nanomolar) to EGFR.
• PET and biodistribution studies indicated that a 24-hour pretargeting interval balances tumor accumulation, internalization, and background levels.
• Adequate hybridization of ⁶⁴Cu-radiolabeled L-DNA to tumor-bound antibody was observed, yielding a tumor-to-muscle ratio of ~11 and clear tumor visualization up to 72 hours.

Conclusions:

• Complementary L-ONs demonstrate high potential for antibody-based pretargeting approaches.
• The developed system is suitable for diagnostic imaging and potentially for therapeutic applications with various radionuclides.
• This L-ONs pretargeting strategy offers a viable method for improving tumor targeting and imaging in EGFR-expressing cancers.