HN125: A Novel Immunoadhesin Targeting MUC16 with Potential for Cancer Therapy

Our plain-English summary of HN125: A Novel Immunoadhesin Targeting MUC16 with Potential for Cancer Therapy, by X. Xiang, M. Feng, M. Felder, et al (Journal of Cancer, 2011).

The study

• Authors: X. Xiang, M. Feng, M. Felder, J.P. Connor, Y.G. Man, M.S. Patankar, M. Ho
• Journal: Journal of Cancer
• Published: 2011;2:280-91
• Cited by: 33 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: The mucin MUC16 expresses the repeating peptide epitope CA125 that has been known for decades to be a well-validated cancer marker that is overexpressed on the cell surface of ovarian cancers and other malignant tumors. In spite of recent efforts to make mouse monoclonal antibodies to MUC16 to treat ovarian cancer, a human monoclonal antibody against this mucin has not been described. MUC16 interacts with mesothelin, a protein that mediates heterotypic cancer cell adhesion, indicating that MUC16 and mesothelin play an important role in the peritoneal implantation and metastasis of ovarian tumors. Therefore, a suitable candidate for therapeutic targeting of MUC16 would functionally block the interaction of MUC16 and mesothelin.

Methodology/principal Findings: Here we report the generation of a novel immunoadhesin, HN125, against MUC16 that consists of a functional MUC16 binding domain of mesothelin (IAB) and the Fc portion of a human antibody IgG1. The yield for purified HN125 proteins is over 100 µg/mL of HEK-293 culture supernatant. We show that HN125 has high and specific affinity for MUC16-expressing cancer cells by flow cytometry and immunohistochemistry. HN125 has the ability to disrupt the heterotypic cancer cell adhesion mediated by the MUC16-mesothelin interaction. Moreover, it elicits strong antibody-dependent cell mediated cytotoxicity against MUC16-positive cancer cells in vitro.

Conclusion/significance: This report describes a novel human immunotherapeutic agent highly specific for MUC16 with potential for treating ovarian cancer and other MUC16-expressing tumors. Because of its lower immunogenicity in patients, a fully human protein is the most desirable format for clinical applications. We believe that the methods developed here may apply to the generation of other tumor-targeting immunoadhesins when it is difficult to obtain a human monoclonal antibody to a given antigen for clinical applications. The resultant immunoadhesins can have advantages usually found in monoclonal antibodies such as ease of purification, high binding affinity and effector functions.

In plain English

This laboratory study reported the construction and testing of HN125, an immunoadhesin made by fusing IAB, the MUC16-binding domain of mesothelin, to the Fc portion of human IgG1. The protein was expressed in HEK-293F cells and purified by protein A affinity chromatography; purity was above 95%, with a yield the authors described as 100 to 500 micrograms per mL of culture supernatant. By flow cytometry, HN125 bound MUC16-positive ovarian cancer cells (OVCAR3) and mesothelioma cells (YOU) but not MUC16-negative A431/H9 or NCI-H226 cells; the Kd on OVCAR3 cells was 13 nM. HN125 stained a human micro-papillary ovarian tumor by immunohistochemistry. In a heterotypic adhesion assay, 100 micrograms per mL of HN125 reduced adherence of mesothelin-positive A431/H9 cells to an OVCAR3 monolayer by approximately 80%, and about 50% at 10 micrograms per mL. With donor peripheral blood mononuclear cells, HN125 killed about 35% of OVCAR3 cells by antibody-dependent cellular cytotoxicity. The authors concluded HN125 should be further evaluated as a potential therapeutic agent.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) experimental study: construction, expression and functional testing of a recombinant Fc fusion protein in cancer cell lines and human tumour tissue
• Who was studied: Human cancer cell lines (A431/H9 epithelial carcinoma overexpressing mesothelin, OVCAR3 ovarian, NCI-H226 and YOU mesothelioma, ECC-1 endometrial), formalin-fixed paraffin-embedded ovarian tumour tissue from 15 patients, and peripheral blood mononuclear cells and NK cells from healthy donors
• Exposure: HN125, an immunoadhesin joining the mesothelin IAB domain (residues 296-359) to the human IgG1 Fc (CH2 and CH3) at the hinge region, applied to cells at stated concentrations
• Main results:
  • Purified HN125 ran as a dimer of about 75 kDa non-reduced and a monomer of about 37 kDa reduced; purity above 95%
  • Protein yield from HEK-293 culture supernatant was 100 to 500 micrograms per mL
  • By flow cytometry HN125 bound MUC16-positive OVCAR3 (ovarian) and YOU (mesothelioma) cells but not MUC16-negative A431/H9 or NCI-H226 cells
  • Reported binding affinity of HN125 for OVCAR3 cells
  • HN125 at 100 micrograms per mL reduced adhesion of mesothelin-positive A431/H9 cells to the MUC16-positive OVCAR3 monolayer by approximately 80%; about 50% blockade at 10 micrograms per mL
  • With healthy-donor PBMC, HN125 killed about 35% of OVCAR3 target cells; no ADCC was found against MUC16-negative A431/H9 cells
  • Isolated NK cells from two healthy donors mediated dose-dependent HN125 lysis of OVCAR3 and ECC-1 cells; no significant complement-dependent cytotoxicity was found
  • Pre-incubation with HN125 or full-length mesothelin only marginally reduced OC125 binding to OVCAR3 cells, suggesting a different epitope
• The authors' conclusion: The authors concluded that HN125 is a fully human immunoadhesin specific for MUC16 that should be further evaluated as a potential therapeutic agent for ovarian cancer and other MUC16-expressing cancers
• Limitations the authors state:
  • The anti-tumour assays used cells grown as monolayers, which the authors noted are less resistant to therapy than cells grown in vivo
  • Validation in 3D spheroid and in vivo mouse models is still needed
  • HN125 has not been directly compared with other anti-MUC16 antibodies
  • The precise mesothelin binding site on MUC16 was not determined

Topics

Medical terms in this study, explained in our Lexicon: Ovarian Cancer, Monoclonal Antibody, Mesothelin, Metastasis, Immunohistochemistry.

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