Assessment of Cell Cytotoxicity and Comet Assay on HER2/neu Positive Cell Line Due to ¹¹¹In Auger Electrons as DNA-Targeting Radioimmunoconjugate

Title:Assessment of Cell Cytotoxicity and Comet Assay on HER2/neu Positive Cell Line Due to ¹¹¹In Auger Electrons as DNA-Targeting Radioimmunoconjugate

Volume: 15 Issue: 2

Author(s): Behnaz Piroozfar, Behrouz Alirezapour*, Farahnaz Motamedi Sedeh, Mohammad Mirzaii, Amir Reza Jalilian, Miad Hashemizadeh and Gholamreza Raisali

Affiliation:

• Radiation Applications Research School, Nuclear Science and Technology Research Institute, North Karegar St. P. O. Box 11365-3486, Tehran, Iran

Keywords: Targeted therapy, Auger electron, 111In, trastuzumab, NLS peptide, comet assay.

Abstract:

Background: Breast cancer Auger electron therapy is a growing field of study in radioimmunotherapy and oncology research. Trastuzumab, a high affinity-binding monoclonal antibody against HER2/neu is which is over-expressed in breast tumors, is used in radiopharmaceutical development.

Objectives: In this work, the lethal effects of ¹¹¹In³⁺, ¹¹¹In-DTPA-trastuzumab and ¹¹¹In-trastuzumab coupled-nuclear localizing sequence peptide (¹¹¹In-DTPA-NLS-trastuzumab) on malignant cells were studied in vitro.

Methods: DTPA-NLS-trastuzumab was prepared using sulfosuccinimidyl-4-(N-maleimidomethyl) cyclohexane-1-carboxylate (sulfo-SMCC) conjugation with NLS peptide in the first step, followed by conjugation with diethylenetriaminepentaacetic acid (DTPA). Both DTPA-trastuzumab and DTPA- NLS-trastuzumab were labeled with ¹¹¹In followed by purification and quality control techniques. Sk-Br-3 (a HER2/neu+ cell line), was used in the cell viability assessment assay for ¹¹¹In, ¹¹¹In-DTPA-trastuzumab and ¹¹¹In-DTPA-NLS-trastuzumab (3.7 MBq) at 37 ºC. The cytotoxicity of the three species was studied using MTT and comet assay was utilized DNA damage detection.

Results: A significant radiochemical purity for ¹¹¹In-DTPA-NLS-trastuzumab (99.36% ± 0.30%, ITLC) at the DTPA:antibody ratio of 6.90 ± 0.34:1, was obtained. Significant cell viability difference was found for ¹¹¹In-DTPA-NLS-trastuzumab compared to the other treatments at two-time points. In addition, comet assay demonstrated significant DNA damage at 144 h using ¹¹¹In-DTPA- NLS-trastuzumab.

Conclusion: The results of cell viability and cell death using MTT assay and comet assay, respectively, demonstrate the NLS-peptide effectively facilitates ¹¹¹In-trastuzumab transport into the HER2/neu positive cancer cell nuclei to impose the radiotherapeutic effects of Auger electrons on DNA leading to cell death.

Cite this article as:

Piroozfar Behnaz , Alirezapour Behrouz*, Sedeh Motamedi Farahnaz , Mirzaii Mohammad , Jalilian Reza Amir , Hashemizadeh Miad and Raisali Gholamreza , Assessment of Cell Cytotoxicity and Comet Assay on HER2/neu Positive Cell Line Due to ¹¹¹In Auger Electrons as DNA-Targeting Radioimmunoconjugate, Current Radiopharmaceuticals 2022; 15 (2) . https://dx.doi.org/10.2174/1874471014666210625115111