Site-directed antibodies targeting driver mutations of the KRAS protein

Area of Science:

• Oncology
• Molecular Biology
• Immunology

Background:

• Kirsten rat sarcoma viral oncogene homolog (KRAS) is the most frequently mutated oncogene in human cancers, present in about 30% of tumors.
• Mutations in KRAS, particularly at codon G12, are critical drivers of tumorigenesis.
• Development of mutation-specific antibodies for KRAS has lagged behind inhibitor development.

Purpose of the Study:

• To develop novel, site-directed monoclonal antibodies (mAbs) targeting the KRAS G12D oncogenic driver mutation.
• To characterize the affinity and specificity of these newly developed mAbs against KRAS G12D.
• To evaluate the potential of these mAbs for diagnostic and therapeutic applications in KRAS-mutated cancers.

Main Methods:

• Utilized the Epivolve technology for the development of site-directed monoclonal antibodies.
• Assessed antibody binding affinity using equilibrium dissociation constants (K_D).
• Confirmed specificity through Western blot analysis with purified proteins and cell lines, and via immunocytochemistry in cancer cells.

Main Results:

• Developed site-directed mAbs with high binding affinity for KRAS G12D, exhibiting K_D in the nanomolar range.
• Demonstrated over 1,000-fold greater affinity for KRAS G12D compared to wild-type KRAS.
• Confirmed high specificity and validated co-localization with endogenous KRAS G12D in cancer cells.

Conclusions:

• The developed site-directed mAbs possess high affinity and specificity for the KRAS G12D mutation.
• These mAbs represent promising tools for advancing diagnostic applications.
• The findings support the potential utility of these mAbs in the development of targeted therapies for KRAS-driven cancers.