An Anti-TAZ Monoclonal Antibody Recognizing Cell Surface Expressed TAZ Protein in Human Tumor Cells

Mozhan Haji  Ghaffari; Mahsa  Mohammadzadeh; Miganoosh  Simonian; Mehrdad  Hashemi; Niloufar  Sadeghi; Babak  Negahdari; Mohammadali  Mazloomi; Hodjattallah  Rabbani

doi:10.18502/ajmb.v15i1.11420

Mozhan Haji Ghaffari Department of Medical Biotechnology, School of Advanced Science in Medicine, Tehran University of Medical Science, Tehran, Iran
Mahsa Mohammadzadeh Department of Genetics, Islamic Azad University, Tehran Medical Sciences Branch, Tehran, Iran
Miganoosh Simonian Department of Medical Biotechnology, School of Advanced Science in Medicine, Tehran University of Medical Science, Tehran, Iran
Mehrdad Hashemi Department of Genetics, Islamic Azad University, Tehran Medical Sciences Branch, Tehran, Iran
Niloufar Sadeghi Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, Iran
Babak Negahdari Department of Medical Biotechnology, School of Advanced Science in Medicine, Tehran University of Medical Science, Tehran, Iran
Mohammadali Mazloomi Department of Medical Biotechnology, School of Advanced Science in Medicine, Tehran University of Medical Science, Tehran, Iran
Hodjattallah Rabbani Monoclonal Antibody Research Center, Avicenna Research Institute, ACECR, Tehran, Iran

DOI: https://doi.org/10.18502/ajmb.v15i1.11420

Keywords: Carcinoma, Humans, Monoclonal antibody, TAZ (WWTR1)

Abstract

Background: WWTR1 or TAZ is a transcriptional co-activator protein expressed in cytoplasm which functions as the main downstream effector of the Hippo signaling pathway. This pathway is an evolutionally conserved signal cascade, which plays a pivotal role in organ size control and tumorigenesis. Ectopic expression of TAZ has already been observed in many malignancies, while the ectopic localization of TAZ is reported for the first time. The aim of this study was to produce a specific monoclonal antibody (mAb) against a synthetic peptide derived from WWTR1 protein to be used as a research tool in human carcinomas.

Methods: A 21-mer synthetic peptide (derived from human TAZ protein) was used for immunization of BALB/c mice after conjugation with Keyhole Limpet Haemocyanin (KLH) using hybridoma technology. The generated mAb reacted with the immunizing peptide employing ELISA assay. The reactivity of the antibody with native TAZ protein was assessed through Western blot, immunocytochemistry, and flow cytometry using different cancer cell lines.

Results: The produced mAb could recognize the immunizing peptide in ELISA and K_aff was 0.6×10^-9M. The produced anti-TAZ mAb unlike available commercial anti-TAZ antibody, was capable of specifically recognizing cell surface TAZ in human carcinoma cell lines including MCF-7, Raji, and A431 in Western blot, immunocytochemistry, and flow cytometry assays. As expected, no reactivity was observed using normal Peripheral Blood Mononuclear Cell (PBMC) from healthy donors.

Conclusion: Based on the results, TAZ is ectopically expressed on the surface of tumor cell lines which is not the case in normal cells. The generated mAb has a potential to be used as a research tool in studying the expression of TAZ in human carcinomas in different applications.