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A Novel Artificially Humanized Anti-Cripto-1 Antibody Suppressing Cancer Cell Growth
Hiroko Ishii1, Maram H Zahra2, Atushi Takayanagi1
1GSP Enterprise, Inc., 1-4-38 12F Minato-machi, Naniwaku, Osaka 556-0017, Japan.
Abstract:
Cripto-1 is a member of the EGF-CFC/FRL1/Cryptic family and is involved in embryonic development and carcinogenesis. We designed a novel anti-Cripto-1 artificial antibody and assessed the recognition to the antigen and the potential to suppress the growth of cancer stem cells. First, single chain antibody clones were isolated by bio-panning with the affinity to recombinant Cripto-1 protein from our original phage-display library. Then, the variable regions of heavy chain VH and light chain VL in each clone were fused to constant regions of heavy chain CH and light chain CL regions respectively. These fused genes were expressed in ExpiCHO-S cells to produce artificial humanized antibodies against Cripto-1. After evaluation of the expression levels, one clone was selected and the anti-Cripto-1 antibody was produced and purified. The purified antibody showed affinity to recombinant Cripto-1 at 1.1 pmol and immunoreactivity to cancer tissues and cell lines. The antibody was available to detect the immunoreactivity in tissue microarrays of malignant tumors as well as in Cripto-1 overexpressing cells. Simultaneously, the antibody exhibited the potential to suppress the growth of human colon cancer derived GEO cells overexpressing Cripto-1 with IC50 at approximately 110 nM. The artificially humanized antibody is proposed to be a good candidate to target cancer cells overexpressing Cripto-1.
Insights
Researchers developed a novel artificial antibody targeting Cripto-1, a protein implicated in cancer. This antibody shows promise in detecting cancer cells and inhibiting the growth of colon cancer stem cells.
Area of Science:
- Biotechnology
- Immunology
- Oncology
Background:
- Cripto-1 is a protein family member involved in embryonic development and cancer.
- Targeting Cripto-1 presents a potential therapeutic strategy for various cancers.
Purpose of the Study:
- To design and characterize a novel artificial humanized antibody against Cripto-1.
- To evaluate the antibody's efficacy in detecting Cripto-1-expressing cancer cells and suppressing cancer stem cell growth.
Main Methods:
- Phage-display technology was used to isolate single-chain antibody fragments with affinity for Cripto-1.
- Variable and constant regions were fused and expressed in ExpiCHO-S cells to produce artificial humanized antibodies.
- Binding affinity, immunoreactivity in cancer tissues/cell lines, and anti-proliferative effects on colon cancer cells were assessed.
Main Results:
- A purified anti-Cripto-1 antibody demonstrated high affinity (1.1 pmol) to recombinant Cripto-1.
- The antibody successfully detected Cripto-1 in cancer tissues and cell lines, including Cripto-1 overexpressing cells.
- The antibody inhibited the growth of Cripto-1-positive human colon cancer cells with an IC50 of approximately 110 nM.
Conclusions:
- The developed artificial humanized antibody is a potent binder of Cripto-1.
- This antibody shows significant potential as a diagnostic and therapeutic agent for targeting Cripto-1-expressing cancers.
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