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Isolation of Novel EGFR-Specific VHH Domains

Department of Radiologyy, Duke University Medical Center, Durham, North Carolina, liz.gottlin{at}duke.edu

Xiangrong Guan

Department of Radiologyy, Duke University Medical Center, Durham, North Carolina

Charles Pegram

Department of Pathologyy, Duke University Medical Center, Durham, North Carolina

Allen Cannedy

North Carolina State University College of Veterinary Medicine, Raleigh, North Carolina

Michael J. Campa

Department of Radiologyy, Duke University Medical Center, Durham, North Carolina

Edward F. Patz, Jr.

Department of Radiologyy, Duke University Medical Center, Durham, North Carolina, Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina

Epidermal growth factor receptor (EGFR) is overexpressed or mutated in a high percentage of tumors. EGFR has long been considered a promising target for cancer diagnostic and therapeutic applications. However, monoclonal antibodies and other large antibody constructs diffuse into tumors slowly, limiting their efficacy. To develop lower molecular weight probes for EGFR and other tumor cell receptors, the authors immunized a llama with the extracellular domains (ECDs) of EGFR and an oncogenic mutant receptor, EGFRvIII, and with extracts of tumor cell lines. From the immune repertoire of the llama, the authors constructed a heavy chain variable domain (VHH domain)—phage library. At ~16 kDa, the VHH domain is a tenth of the size of a monoclonal antibody and is the smallest antibody fragment that retains specificity. By affinity selection from this library, the authors isolated many VHH domains with specificity for EGFR. The VHH domains bind to whole cells expressing the receptor but not to control cells lacking the receptor and can immunoprecipitate EGFR from cell lysates. Some VHH domains have cross-specificity with existing anti-EGFR monoclonal antibodies and have reasonably high (nM) affinities. The llama-VHH domain library is also potentially a rich source of targeting agents directed toward other tumor cell receptors. (Journal of Biomolecular Screening 2009:77-85)

Key Words: VHH domains • EGFR • phage display • cancer

Journal of Biomolecular Screening, Vol. 14, No. 1, 77-85 (2009)
DOI: 10.1177/1087057108327064


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