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Multicomponent Conjugates of Anticancer Drugs and Monoclonal Antibody with PAMAM Dendrimers to Increase Efficacy of HER-2 Positive Breast Cancer Therapy
2019, Marcinkowska, Monika, Stanczyk, Maciej, Janaszewska, Anna, Sobierajska, Ewelina, Chworos, Arkadiusz, Klajnert-Maculewicz, Barbara
Purpose: Conjugation of nanocarriers with antibodies that bind to specific membrane receptors that are overexpressed in cancer cells enables targeted delivery. In the present study, we developed and synthesised two PAMAM dendrimer-trastuzumab conjugates that carried docetaxel or paclitaxel, specifically targeted to cells which overexpressed HER-2. Methods: The 1H NMR, 13C NMR, FTIR and RP-HPLC were used to analyse the characteristics of the products and assess their purity. The toxicity of PAMAM-trastuzumab, PAMAM-doc-trastuzumab and PAMAM-ptx-trastuzumab conjugates was determined using MTT assay and compared with free trastuzumab, docetaxel and paclitaxel toward HER-2-positive (SKBR-3) and negative (MCF-7) human breast cancer cell lines. The cellular uptake and internal localisation were studied using flow cytometry and confocal microscopy, respectively. Results: The PAMAM-drug-trastuzumab conjugates in particular showed extremely high toxicity toward the HER-2-positive SKBR-3 cells and very low toxicity towards to HER-2-negative MCF-7 cells. As expected, the HER-2-positive SKBR-3 cell line accumulated trastuzumab from both conjugates rapidly; but surprisingly, although a large amount of PAMAM-ptx-trastuzumab conjugate was observed in the HER-2-negative MCF-7 cells. Confocal microscopy confirmed the intracellular localisation of analysed compounds. The key result of fluorescent imaging was the identification of strong selective binding of the PAMAM-doc-trastuzumab conjugate with HER-2-positive SKBR-3 cells only. Conclusions: Our results confirm the high selectivity of PAMAM-doc-trastuzumab and PAMAM-ptx-trastuzumab conjugates for HER-2-positive cells, and demonstrate the utility of trastuzumab as a targeting agent. Therefore, the analysed conjugates present an promising approach for the improvement of efficacy of targeted delivery of anticancer drugs such as docetaxel or paclitaxel. © 2019, The Author(s).
Conjugate of PAMAM Dendrimer, Doxorubicin and Monoclonal Antibody—Trastuzumab: The New Approach of a Well-Known Strategy
2018, Marcinkowska, Monika, Sobierajska, Ewelina, Stanczyk, Maciej, Janaszewska, Anna, Chworos, Arkadiusz, Klajnert-Maculewicz, Barbara
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.