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Licensed Unlicensed Requires Authentication Published by De Gruyter June 25, 2021

Inhibition of Ras and STAT3 activity of 4-(tert-butyl)-N-carbamoylbenzamide as antiproliferative agent in HER2-expressing breast cancer cells

Aguslina Kirtishanti, Siswandono Siswodihardjo, I Ketut Sudiana, Desak G. A. Suprabawati and Aristika Dinaryanti

Abstract

Objectives

Human epidermal growth factor receptor type 2 (HER2)-expressing breast cancer patients indicate poor prognosis in disease progression. HER2 overexpression can increase activities of Ras-mitogen activated protein kinase (Ras-MAPK) pathway and Janus Kinase (JAK)-STAT3, increasing breast cancer cell proliferation as demonstrated by marker Ki67. Therapeutic options for HER2-expressing breast cancer are limited and have major side effects, so anticancer development as an antiproliferative is needed. From previous research, synthetic chemical 4-(tert-butyl)-N-carbamoylbenzamide (4TBCB) compound has cytotoxic activity in vitro on HER2-expressing breast cancer cells. This study wanted to determine the mechanism 4TBCB compound in inhibiting HER2 signaling through Rat Sarcoma (Ras) and signal transducer and activator of transcription 3 (STAT3) pathway in HER2-expressing breast cancer cells.

Methods

Breast cancer cells were isolated from the biopsy tissue of breast cancer patients. The isolated cells were cultured and given 4TBCB test compound with three concentrations (0.305, 0.61, and 1.22 mM) and lapatinib 0.05 mM as a comparison compound. Cancer cell cultures were stained with monoclonal antibodies phosphorylated HER2 (pHER2), phosphorylated Ras (pRas), phosphorylated STAT3 (pSTAT3), and Ki67. The expression of pHER2, pRas, pSTAT3, and Ki67 proteins was observed using the immunofluorescence method and the results were compared with control cells, namely cancer cells that were not given 4TBCB and lapatinib but stained with monoclonal antibodies.

Results

4TBCB compounds (0.61 and 1.22 mM) and lapatinib can reduce pHER2, pRas, pSTAT3, and Ki67 expressions compared to control cells.

Conclusions

4TBCB compounds (0.61 and 1.22 mM) can reduce pHER2, pRas, pSTAT3, Ki67 expressions and predicted to inhibit HER2 signaling through the Ras and STAT3 pathways in HER2-expressing breast cancer cells.


Corresponding author: Siswandono Siswodihardjo, Department of Medicinal Chemistry, Faculty of Pharmacy, University of Airlangga, Surabaya, Indonesia, Phone: +62 8123206328, E-mail:

Funding source: Directorate General of Resources for Science, Technology and Higher Education of Ministry of Research, Technology and Higher Education (KEMENRISTEK DIKTI)

Acknowledgments

We acknowledge Eryk Hendrianto and Aida Ariyanti from the Stem Cell Research Center for supporting the current research.

  1. Research funding: Directorate General of Resources for Science, Technology and Higher Education of Ministry of Research, Technology and Higher Education (KEMRISTEK DIKTI).

  2. Author contributions: All authors have accepted responsibility for the entire content of this manuscript and approved its submission.

  3. Competing interests: Authors state no conflict of interest.

  4. Informed consent: Informed consent was obtained from all individuals included in this study.

  5. Ethical approval: Research involving human subjects complied with all relevant national regulations, institutional policies and is in accordance with the tenets of the Helsinki Declaration (as revised in 2013), and has been approved by Ethical Commission of General Hospital in Surabaya, Indonesia.

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Received: 2020-12-14
Accepted: 2021-03-24
Published Online: 2021-06-25

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