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In vivo anti-MUC1+ tumor activity and sequences of high-affinity anti-MUC1-SEA antibodies

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Abstract

Cleavage of the MUC1 glycoprotein yields two subunits, an extracellular alpha-subunit bound to a smaller transmembrane beta-subunit. Monoclonal antibodies (mAbs) directed against the MUC1 alpha–beta junction comprising the SEA domain, a stable cell-surface moiety, were generated. Sequencing of all seven anti-SEA domain mAbs showed that they clustered into four groups and sequences of all groups are presented here. mAb DMB5F3 with picomolar affinity for the MUC1 SEA target was selected for further evaluation. Immunohistochemical staining of a series of malignancies with DMB5F3 including lung, prostate, breast, colon, and pancreatic carcinomas revealed qualitative and qualitative differences between MUC1 expression on normal versus malignant cells: DMB5F3 strongly stained malignant cells in a near-circumferential pattern, whereas MUC1 in normal pancreatic and breast tissue showed only weak apical positivity of ductal/acinar cells. Humanized chimeric DMB5F3 linked to ZZ-PE38 (ZZ IgG-binding protein fused to Pseudomonas exotoxin) induced vigorous cytotoxicity of MUC1+ malignant cells in vitro. The intensity of cell killing correlated with the level of MUC1 expression by the target cell, suggesting a MUC1 expression threshold for cell killing. MUC1+ Colo357 pancreatic cancer cells xenotransplanted into nude and SCID mice models were treated with the chDMB5F3:ZZ-PE38 immunocomplex. In both transplant models, chDMB5F3:ZZ-PE38 exhibited significant in vivo anti-tumor activity, suppressing up to 90% of tumor volume in the SCID model compared with concomitant controls. The efficacy of chDMB5F3:ZZ-PE38 immunotoxin in mediating tumor killing both in vitro and in vivo strongly suggests a clinical role for anti-MUC1 SEA antibody in the treatment of MUC1-expressing malignancies.

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Funding

Funded by Israel Cancer Association (Project 20112024) and Israel Science Foundation (Project 1167/10).

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Author notes

  1. Daniel B. Rubinstein and Daniel H. Wreschner have contributed equally to this work.

Authors and Affiliations

  1. BLAVATNIK CENTER for Drug Discovery, Tel Aviv University, 69978, Ramat Aviv, Israel

    Edward Pichinuk & Michael Chalik

  2. School of Molecular Cell Biology and Biotechnology, Tel Aviv University, 69978, Ramat Aviv, Israel

    Itai Benhar, Ravit Ginat-Koton, Ravit Ziv, Nechama I. Smorodinsky & Daniel H. Wreschner

  3. Gynecology-Oncology Division, Mayanei Hayeshua Medical Center, Bnei Brak, Israel

    Gabi Haran

  4. Department of Biopathology, Centre Régional de Lutte Contre le Cancer, Institut Jean-Godinot, 51100, Reims, France

    Christian Garbar

  5. INSERM U976, Sorbonne Paris Cité, UMR-S 976, Université Paris Diderot, 75475, Paris, France

    Armand Bensussan

  6. Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD, 21231, USA

    Alan Meeker

  7. Division of Hematology, Hôtel-Dieu, University Hospital Nantes, Nantes, France

    Thierry Guillaume

  8. Centre National de la Recherche Scientifique (CNRS), Université d’Angers, Université de Nantes, Nantes, France

    Thierry Guillaume

  9. BioModifying, LLC., Silver Spring, MD, 20902, USA

    Daniel B. Rubinstein & Daniel H. Wreschner

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  1. Edward Pichinuk
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Contributions

EP, MC, IB, RGK, RZ, NIS, GH, CG, AB, AM, TG, DBR, and DHW contributed to the study conception and design. Material preparation, data collection, and analysis were performed by authors DHW, EP, MC, DBR, MC, AM, CG, AB, and NIS. The first draft of the manuscript was written by DHW, DBR and EP. EP, MC, IB, NIS, DBR, and DHW commented on the various versions of the manuscript and contributed to the actual writing and preparation of its final version. EP, MC, IB, RGK, RZ, NIS, GH, CG, AB, AM, TG, DBR, and DHW read and approved the final manuscript.

Corresponding author

Correspondence to Daniel H. Wreschner.

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Conflict of interest

Daniel H. Wreschner and Daniel B. Rubinstein have ownership interest in BioModifying, LLC. All other authors declare that they have no conflict of interest.

Ethical Approval

Animal care and use Use of mice was done under supervision of the Tel Aviv University Institutional Animal Care and Use Committee (TAU-IACUC), Study Approval License number L-04-12-003. To ameliorate suffering including methods of killing, animal welfare and steps were all performed in accordance with regulations stipulated by TAU-IACUC. Samples used for immunohistochemical stainings These were procured from the Biomax tissue bank, see http://biomax.us. With all required approvals therein. Cell line authentication Short Tandem Repeat (STR) analysis (PowerPlexW 1.2 System, Promega, Fitchburg, WI), as described [45], was used to validate the human cell lines. STR profiles were matched with the German Collection of Microorganisms and Cell Cultures (DSMZ) database. Cell lines were obtained as gifts from Prof. I. Keydar.

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Pichinuk, E., Chalik, M., Benhar, I. et al. In vivo anti-MUC1+ tumor activity and sequences of high-affinity anti-MUC1-SEA antibodies. Cancer Immunol Immunother 69, 1337–1352 (2020). https://doi.org/10.1007/s00262-020-02547-2

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