Conjugate of PAMAM Dendrimer, Doxorubicin and Monoclonal Antibody-Trastuzumab: The New Approach of a Well-Known Strategy.

Monika Marcinkowska,Ewelina Sobierajska,Anna Janaszewska,Arkadiusz Chworos,Barbara Klajnert-Maculewicz,Maciej Stanczyk

doi:10.3390/polym10020187

Abstract

The strategy utilizing trastuzumab, a humanized monoclonal antibody against human epidermal growth receptor 2 (HER-2), as a therapeutic agent in HER-2 positive breast cancer therapy seems to have advantage over traditional chemotherapy, especially when given in combination with anticancer drugs. However, the effectiveness of single antibody or antibody conjugated with chemotherapeutics is still far from ideal. Antibody–dendrimer conjugates hold the potential to improve the targeting and release of active substance at the tumor site. In the present study, we developed and synthesized PAMAM dendrimer–trastuzumab conjugates carrying doxorubicin (dox) specifically to cells overexpressing HER-2. 1HNMR, FTIR and RP-HPLC were used to characterize the products and analyze their purity. Toxicity of PAMAM–trastuzumab and PAMAM–dox–trastuzumab conjugates compared with free trastuzumab and doxorubicin towards HER-2 positive (SKBR-3) and negative (MCF-7) human breast cancer cell lines was determined using MTT assay. Furthermore, the cellular uptake and cellular localization were studied by flow cytometry and confocal microscopy, respectively. A cytotoxicity profile of above mentioned compounds indicated that conjugate PAMAM–dox–trastuzumab was more effective when compared to free drug or the conjugate PAMAM–trastuzumab. Moreover, these results reveal that trastuzumab can be used as a targeting agent in PAMAM–dox–trastuzumab conjugate. Therefore PAMAM–dox–trastuzumab conjugate might be an interesting proposition which could lead to improvements in the effectiveness of drug delivery systems for tumors that overexpress HER-2.

Highlights

According to World Health Organization (WHO) reports, breast cancer is the most common tumor among every major ethnic group of women, with almost 1.7 million new cases diagnosed in2012 [1]
The prime example of a monoclonal antibody used in this approach is humanized anti-HER2 mAb trastuzumab (Herceptin) that plays a major role in breast cancer treatment because human epidermal growth factor receptor 2 (HER-2/neu/ErbB) gene amplification or protein overexpression occurs in 20% to 25% of breast tumors
Our research model consisted of human epidermal growth receptor 2 (HER-2) positive (SKBR-3) and negative (MCF-7) human breast cancer cell lines

Summary

Introduction

According to World Health Organization (WHO) reports, breast cancer is the most common tumor among every major ethnic group of women, with almost 1.7 million new cases diagnosed in2012 [1]. The prime example of a monoclonal antibody used in this approach is humanized anti-HER2 mAb trastuzumab (Herceptin) that plays a major role in breast cancer treatment because human epidermal growth factor receptor 2 (HER-2/neu/ErbB) gene amplification or protein overexpression occurs in 20% to 25% of breast tumors. It makes it a potential candidate for targeted antibody therapy since trastuzumab blocks the HER-2 by binding to domain. To introduce thiol groups into G4 dendrimer surface, the primary amine groups were reacted with a 10:1 mole excess of Traut’s reagent in 0.1 M PBS buffer, at room temperature under N2 for 1 h pH 8.0. The final stoichiometric ratio for PAMAM–dox–trastuzumab conjugate was 1:1:1

Methods

Results

Conclusion