Jafar Mahmoudian; Roya Ghods; Mahmood Jeddi-Tehrani; Nassim Ghaffari-Tabrizi-Wizsy; Mohammad Reza Nejadmoghaddam; Ramin Ghahremanzadeh; Seyed Nasser Ostad; Amir-Hassan Zarnani
Abstract
Background: Placenta-specific 1 (PLAC1) is an oncoplacental genes aberrantly expressed in various malignancies. Antibody-drug conjugates (ADC) offer a promising therapeutic approach by enhancing efficacy and reducing toxicity of treatment compared to cytotoxic small-molecule agents.Objective: To evaluate ...
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Background: Placenta-specific 1 (PLAC1) is an oncoplacental genes aberrantly expressed in various malignancies. Antibody-drug conjugates (ADC) offer a promising therapeutic approach by enhancing efficacy and reducing toxicity of treatment compared to cytotoxic small-molecule agents.Objective: To evaluate the efficacy of an SN38-conjugated monoclonal anti-PLAC1 antibody in a mouse model of breast cancer.Methods: Anti-human PLAC1 monoclonal antibodies were generated and characterized. SN38 was conjugated to an anti-PLAC1 antibody (clone: 2H12C12) and conjugation efficacy was determined by UV spectrophotometry. The antigen-binding activity of the conjugated antibody was assessed using ELISA and flow cytometry. In vitro, the cytotoxic profile of 2H12C12-SN38 was evaluated in MDA-MB-231 breast cancer cells using a fluoroimetric viability assay. The impact of 2H12C12-SN38 on MDA-MB-231 tumor growth and angiogenesis ex vivo was examined using chorioallantoic membrane (CAM) assay followed by immunohistochemical analysis. Pharmacokinetics of 2H12C12-SN38 in mice was determined by serial venipuncture following ADC administration. The inhibitory effects of anti-PLAC1 ADC on tumor growth were evaluated in a nude mouse xenograft model of human breast cancer.Results: The anti-PLAC1 ADC exhibited a substantial cytotoxicity against MDA-MB-231 cells, with effects observed at concentration as low as ~33 nM. In the CAM assay, the ADC significantly reduced the growth of MDA-MB-231 tumor but did not produce a significant effect on tumor angiogenesis. Pharmacokinetic analysis in mice demonstrated an average half-life (t1/2) of approximately 80 hours. In a nude mouse xenograft model, treatment with the ADC resulted in a significant reduction in tumor size compared with isotype-matched antibody-SN38 conjugate, or free SN38.Conclusion: This study represents the first therapeutic application of anti-PLAC1 ADC in a xenograft model of human breast cancer. Our findings support the embryonic origin of cancers and highlight the potential therapeutic value of targeting oncofetal antigens in human breast cancer.
Masoud Hassanzadeh Makoui; Maryam Mobini; Jalal Khoshnoodi; Tannaz Bahadori; Forough Golsaz-Shirazi; Hedieh Moradi Tabriz; Zahra Madjd; Mahmood Jeddi-Tehrani; Amir-Hassan Zarnani; Mohammad Mehdi Amiri; Fazel Shokri
Abstract
Background: Ki67 and P53 are important diagnostic and prognostic biomarkers expressed in several cancers. The current standard method for evaluating Ki67 and P53 in cancer tissues is immunohistochemistry (IHC), and having highly sensitive monoclonal antibodies against these biomarkers is necessary for ...
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Background: Ki67 and P53 are important diagnostic and prognostic biomarkers expressed in several cancers. The current standard method for evaluating Ki67 and P53 in cancer tissues is immunohistochemistry (IHC), and having highly sensitive monoclonal antibodies against these biomarkers is necessary for an accurate diagnosis in the IHC test.Objective: To generate and characterize novel monoclonal antibodies (mAbs) against human Ki67 and P53 antigens for IHC purposes.Methods: Ki67 and P53-specific mAbs were produced by the hybridoma method and screened by enzyme-linked immunosorbent assay (ELISA) and IHC techniques. Selected mAbs were characterized using Western blot and flow cytometry, and their affinities and isotypes were determined by ELISA. Moreover, using the IHC technique in 200 breast cancer tissue samples, we assessed the specificity, sensitivity, and accuracy of the produced mAbs.Results: Two anti-Ki67 (2C2 and 2H1) and three anti-P53 mAbs (2A6, 2G4, and 1G10) showed strong reactivity to their target antigens in IHC. The selected mAbs were also able to recognize their targets by flow cytometry as well as Western blotting using human tumor cell lines expressing these antigens. The specificity, sensitivity, and accuracy calculated for clone 2H1 were 94.2%, 99.0%, and 96.6%, and for clone 2A6 were 97.3%, 98.1%, and 97.5%, respectively. Using these two monoclonal antibodies, we found a significant correlation between Ki67 and P53 overexpression and lymph node metastasis in patients with breast cancer.Conclusion: The present study showed that the novel anti-Ki67 and anti-P53 mAbs could recognize their respective antigens with high specificity and sensitivity and therefore can be used in prognostic studies.
