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Recombinant full-length human IgG1s targeting hormone-refractory prostate cancer

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Abstract

We have used a naive human single-chain fragment variable (scFv) library as a source of random shape repertoire to directly probe the altered surface chemistry of tumor cells. We reported previously the identification of more than 90 internalizing phage monoclonal antibodies targeting prostate cancer cells, including those that are hormone refractory. In this report, we describe the conversion of a panel of those scFvs into full-length human immunoglobulins (IgGs) and show that tumor specificity is retained. We have further shown that antibodies isolated from a naive phage display library can nevertheless be of high affinity towards target tumor cells. In addition, full-length IgGs retain the functionality of parental scFvs including the ability to rapidly enter target cells through receptor-mediated endocytosis and thereby to mediate efficient and specific intracellular payload delivery to tumor cells. We have used recombinant IgGs to immunoprecipitate target antigens and analyzed their molecular composition by mass spectrometry. We have identified one target antigen as activated leukocyte cell adhesion molecule (ALCAM)/MEMD/CD166 and have further studied tissue specificity of this internalizing ALCAM epitope by immunohistochemistry. Our study shows that cell type-specific internalizing human antibody can be readily identified from a naive phage antibody display library, characterized with regards to sequence, affinity, tissue specificity, and antigen identity, and modified genetically and chemically to generate various forms of targeted therapeutics.

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Abbreviations

MAb:

Monoclonal antibody

ScFv:

Single-chain variable domain fragment

CaP:

carcinoma of the prostate

AR:

androgen receptor

ILs:

immunoliposomes

MTX:

methotrexate

K D :

dissociation equilibrium constant

LC–MS/MS:

liquid chromatography–tandem mass spectrometry

IHC:

immunohistochemistry

ALCAM:

activated leukocyte cell adhesion molecule

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Acknowledgement

We thank the UCSF Biomolecular Resource Center and the National Mass Spectrometry Facility for analysis of protein samples, Dr. Jianlong Lou and Richard Tsai for help with mammalian cell transfection. This work is supported by grants from the National Cancer Institute Specialized Programs of Research Excellence (SPORE) in Prostate Cancer (P50 CA89520; JDM) and the National Institute of Health (R01 CA118919 and R21 DK066429; BL).

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Authors and Affiliations

  1. Department of Anesthesia, University of California at San Francisco, 1001 Potrero Avenue, 3C-38, San Francisco, CA, 94110, USA

    Bin Liu, Fraser Conrad, Audrey Roth & James D. Marks

  2. UCSF Comprehensive Cancer Center, University of California at San Francisco, 1001 Potrero Avenue, 3C-38, San Francisco, CA, 94110, USA

    Bin Liu, Jeff P. Simko & James D. Marks

  3. Hermes Biosciences, Inc., South San Francisco, CA, 94115, USA

    Daryl C. Drummond

  4. Department of Anatomical Pathology, University of California at San Francisco, San Francisco, CA, 94143, USA

    Jeff P. Simko

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  1. Bin Liu
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  2. Fraser Conrad
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  3. Audrey Roth
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  4. Daryl C. Drummond
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Correspondence to Bin Liu or James D. Marks.

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Supplemental Fig. 1

IHC study of H3 IgG on a panel of tumor tissues. Results are shown for CaP (a and b), breast cancer (c), colon cancer (d), and bladder cancer tissues (e and f). H3 IgG stained breast (5/5) but not colon cancer tissues (4/4), and one out of five bladder tumor tissues (e) but not the remaining four specimens studied (f). Amplifications: a ×200; b–f ×100 (PDF 989 KB)

Supplementary Fig. 2

Normal cellular structures stained by H3 IgG. a Colon ganglion cells (arrow). b Salivary gland (arrow). c Skin eccrine (arrow). d Bronchial epithelium (arrow). a and b ×100; c and d ×200 (PDF 413 KB)

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Liu, B., Conrad, F., Roth, A. et al. Recombinant full-length human IgG1s targeting hormone-refractory prostate cancer. J Mol Med 85, 1113–1123 (2007). https://doi.org/10.1007/s00109-007-0208-z

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  • DOI: https://doi.org/10.1007/s00109-007-0208-z

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