Synthesized Anti-HER2 Trastuzumab-MCC-DM1 Conjugate: An Evaluation of Efficacy and Cytotoxicity
Background: Trastuzumab is a humanized monoclonal antibody that targets site-specifically human epidermal growth factor-2 receptor (HER2) cell surface antigen overexpressed in approximately 20% of human breast carcinomas. Despite its positive therapeutic outcomes, a large proportion of individuals a...
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| Format: | Article |
| Language: | English |
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Shiraz University of Medical Sciences
2023-09-01
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| Series: | Iranian Journal of Immunology |
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| Online Access: | https://iji.sums.ac.ir/article_49411_07f4888549d6cfb0b706534f398c5e5b.pdf |
| _version_ | 1797680523899830272 |
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| author | Soodabeh Shafiee Roya Mirzaei Malihe Salehi Neda Jalili Amir Taheri Leila Farahmand |
| author_facet | Soodabeh Shafiee Roya Mirzaei Malihe Salehi Neda Jalili Amir Taheri Leila Farahmand |
| author_sort | Soodabeh Shafiee |
| collection | DOAJ |
| description | Background: Trastuzumab is a humanized monoclonal antibody that targets site-specifically human epidermal growth factor-2 receptor (HER2) cell surface antigen overexpressed in approximately 20% of human breast carcinomas. Despite its positive therapeutic outcomes, a large proportion of individuals are unresponsive to the treatment with trastuzumab or develop resistance to it.Objective: To evaluate a chemically synthesized trastuzumab-based antibody-drug conjugate (ADC) to improve the trastuzumab therapeutic index.Methods: The current study explored the physiochemical characteristics of the trastuzumab conjugated to a cytotoxic chemotherapy agent DM1 via Succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC) linker, created in our earlier study, using SDS-PAGE, UV/VIS, and RP-HPLC analyses. The antitumor effects of the ADCs were analyzed using MDA-MB-231 (HER2-negative) and SK-BR-3 (HER2-positive) cell lines utilizing in vitro cytotoxicity, viability, and binding assays. Three different formats of a HER2-targeting agent: trastuzumab, synthesized trastuzumab-MCC-DM1, and commercially available drug T-DM1 (Kadcyla®) were compared.Results: UV-VIS spectroscopic analysis showed that the trastuzumab-MCC-DM1 conjugates, on average, entailed 2.9 DM1 payloads per trastuzumab. A free drug level of 2.5% was determined by RP-HPLC. The conjugate appeared as two bands on a reducing SDS-PAGE gel. MTT viability assay showed that conjugating trastuzumab with DM1 significantly improved the antiproliferative effects of this antibody in vitro. Importantly, the evaluations using LDH release and cell apoptosis assays confirmed that trastuzumab maintains its ability to induce cell death response while conjugating with the DM1. The binding efficiency of trastuzumab-MCC-DM1 was comparable to that of the naked trastuzumab.Conclusion: Trastuzumab-MCC-DM1 was found effective against HER2+ tumors. The potency of this synthesized conjugate brings it closer to the commercially available T-DM1. |
| first_indexed | 2024-03-11T23:31:16Z |
| format | Article |
| id | doaj.art-ee7bd5531d414c82b59d000ae6696847 |
| institution | Directory Open Access Journal |
| issn | 1735-1383 1735-367X |
| language | English |
| last_indexed | 2024-03-11T23:31:16Z |
| publishDate | 2023-09-01 |
| publisher | Shiraz University of Medical Sciences |
| record_format | Article |
| series | Iranian Journal of Immunology |
| spelling | doaj.art-ee7bd5531d414c82b59d000ae66968472023-09-20T09:13:41ZengShiraz University of Medical SciencesIranian Journal of Immunology1735-13831735-367X2023-09-0120330331510.22034/iji.2023.96229.242049411Synthesized Anti-HER2 Trastuzumab-MCC-DM1 Conjugate: An Evaluation of Efficacy and CytotoxicitySoodabeh Shafiee0Roya Mirzaei1Malihe Salehi2Neda Jalili3Amir Taheri4Leila Farahmand5Department of Recombinant Proteins, Breast Cancer Research Center, Motamed Cancer Institute, ACECR, Tehran, Iran.Department of Recombinant Proteins, Breast Cancer Research Center, Motamed Cancer Institute, ACECR, Tehran, Iran.Department of Recombinant Proteins, Breast Cancer Research Center, Motamed Cancer Institute, ACECR, Tehran, Iran.Department of Recombinant Proteins, Breast Cancer Research Center, Motamed Cancer Institute, ACECR, Tehran, Iran.Department of Recombinant Proteins, Breast Cancer Research Center, Motamed Cancer Institute, ACECR, Tehran, Iran.Department of Recombinant Proteins, Breast Cancer Research Center, Motamed Cancer Institute, ACECR, Tehran, Iran.Background: Trastuzumab is a humanized monoclonal antibody that targets site-specifically human epidermal growth factor-2 receptor (HER2) cell surface antigen overexpressed in approximately 20% of human breast carcinomas. Despite its positive therapeutic outcomes, a large proportion of individuals are unresponsive to the treatment with trastuzumab or develop resistance to it.Objective: To evaluate a chemically synthesized trastuzumab-based antibody-drug conjugate (ADC) to improve the trastuzumab therapeutic index.Methods: The current study explored the physiochemical characteristics of the trastuzumab conjugated to a cytotoxic chemotherapy agent DM1 via Succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC) linker, created in our earlier study, using SDS-PAGE, UV/VIS, and RP-HPLC analyses. The antitumor effects of the ADCs were analyzed using MDA-MB-231 (HER2-negative) and SK-BR-3 (HER2-positive) cell lines utilizing in vitro cytotoxicity, viability, and binding assays. Three different formats of a HER2-targeting agent: trastuzumab, synthesized trastuzumab-MCC-DM1, and commercially available drug T-DM1 (Kadcyla®) were compared.Results: UV-VIS spectroscopic analysis showed that the trastuzumab-MCC-DM1 conjugates, on average, entailed 2.9 DM1 payloads per trastuzumab. A free drug level of 2.5% was determined by RP-HPLC. The conjugate appeared as two bands on a reducing SDS-PAGE gel. MTT viability assay showed that conjugating trastuzumab with DM1 significantly improved the antiproliferative effects of this antibody in vitro. Importantly, the evaluations using LDH release and cell apoptosis assays confirmed that trastuzumab maintains its ability to induce cell death response while conjugating with the DM1. The binding efficiency of trastuzumab-MCC-DM1 was comparable to that of the naked trastuzumab.Conclusion: Trastuzumab-MCC-DM1 was found effective against HER2+ tumors. The potency of this synthesized conjugate brings it closer to the commercially available T-DM1.https://iji.sums.ac.ir/article_49411_07f4888549d6cfb0b706534f398c5e5b.pdfbreast neoplasmsimmunoconjugatestherapeutic indextrastuzumab |
| spellingShingle | Soodabeh Shafiee Roya Mirzaei Malihe Salehi Neda Jalili Amir Taheri Leila Farahmand Synthesized Anti-HER2 Trastuzumab-MCC-DM1 Conjugate: An Evaluation of Efficacy and Cytotoxicity Iranian Journal of Immunology breast neoplasms immunoconjugates therapeutic index trastuzumab |
| title | Synthesized Anti-HER2 Trastuzumab-MCC-DM1 Conjugate: An Evaluation of Efficacy and Cytotoxicity |
| title_full | Synthesized Anti-HER2 Trastuzumab-MCC-DM1 Conjugate: An Evaluation of Efficacy and Cytotoxicity |
| title_fullStr | Synthesized Anti-HER2 Trastuzumab-MCC-DM1 Conjugate: An Evaluation of Efficacy and Cytotoxicity |
| title_full_unstemmed | Synthesized Anti-HER2 Trastuzumab-MCC-DM1 Conjugate: An Evaluation of Efficacy and Cytotoxicity |
| title_short | Synthesized Anti-HER2 Trastuzumab-MCC-DM1 Conjugate: An Evaluation of Efficacy and Cytotoxicity |
| title_sort | synthesized anti her2 trastuzumab mcc dm1 conjugate an evaluation of efficacy and cytotoxicity |
| topic | breast neoplasms immunoconjugates therapeutic index trastuzumab |
| url | https://iji.sums.ac.ir/article_49411_07f4888549d6cfb0b706534f398c5e5b.pdf |
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