Sedighian Hamid, Abdi Arash, Goleij Zoleikha, Fasihi-Ramandi Mahdi, Fooladi Abbas Ali Imani
Applied Microbiology Research Center, Biomedicine Technologies Institute, Baqiyatallah University of Medical Sciences, Tehran, Iran.
Department of Physiology, School of Medicine, Shahed University, Tehran, Iran.
Biochem Biophys Rep. 2025 Aug 25;43:102218. doi: 10.1016/j.bbrep.2025.102218. eCollection 2025 Sep.
The delivery of therapeutic agents in a targeted manner shows great potential for improving the degree of efficacy in cancer therapies by minimizing harm for normal tissues and reducing treatment-related complications. Among patients with gastric cancer, the HER2 receptor is expressed in approximately 10-30 % of cases. The scFv(Herceptin)-PE-STXA was cloned into the pET28a vector, followed by protein expression and purification. HEK-293 cells served as the control (HER2-negative), while NCI-N87 cells were utilized as the gastric cancer cells (HER2-positive). The cells were subjected to exposure different concentrations (35 and 50 μg/mL) of immunotoxin, after which growth was assessed using the MTT test. The capability of scFv(Herceptin)-PE-STXA to induce apoptotic pathway in NCI-N87 and HEK-293 cells was estimated through flow cytometry. Furthermore, the affinity for binding the IT, lactate dehydrogenase release, and the measurement of apoptosis pathway enzymes (caspase-9 and caspase-3 activities) were also conducted. The outcomes demonstrated that cytotoxic effects occurred in NCI-N87 cells with a dose-proportional manner approach, whereas no such impact was seen in HEK-293(P < 0.001). Flow cytometry analysis of NCI-N87 cells treated with scFv(Herceptin)-PE-STXA revealed a significant increase in apoptotic rate at dose of 35 and 50 μg/mL (55.6 % and 74.2 %). Additional analysis showed that exposing HER2-expressing NCI-N87 cells to the scFv(Herceptin)-PE-STXA caused a notable to enhance in apoptotic caspases activity (3 and 9) (P < 0.001). Results of our study indicated that the scFv(Herceptin)-PE-STXA induces apoptosis in the NCI-N87 cell line derived from gastric cancer. This study emphasizes the potential of scFv(Herceptin)-PE-STXA to specifically target HER2-expressing cancer cells.
以靶向方式递送治疗剂在通过最小化对正常组织的损害并减少治疗相关并发症来提高癌症治疗的疗效程度方面显示出巨大潜力。在胃癌患者中，HER2受体在约10 - 30%的病例中表达。将单链抗体片段（抗赫赛汀）-PE-STXA克隆到pET28a载体中，随后进行蛋白质表达和纯化。HEK-293细胞用作对照（HER2阴性），而NCI-N87细胞用作胃癌细胞（HER2阳性）。使细胞暴露于不同浓度（35和50μg/mL）的免疫毒素，之后使用MTT试验评估细胞生长。通过流式细胞术估计单链抗体片段（抗赫赛汀）-PE-STXA在NCI-N87和HEK-293细胞中诱导凋亡途径的能力。此外，还进行了与免疫毒素结合的亲和力、乳酸脱氢酶释放以及凋亡途径酶（半胱天冬酶-9和半胱天冬酶-3活性）的测量。结果表明，细胞毒性作用在NCI-N87细胞中以剂量比例方式出现，而在HEK-293细胞中未观察到这种影响（P < 0.001）。用单链抗体片段（抗赫赛汀）-PE-STXA处理的NCI-N87细胞的流式细胞术分析显示，在35和50μg/mL剂量下凋亡率显著增加（55.6%和74.2%）。进一步分析表明，将表达HER2的NCI-N87细胞暴露于单链抗体片段（抗赫赛汀）-PE-STXA会导致凋亡半胱天冬酶活性（3和9）显著增强（P < 0.001）。我们的研究结果表明，单链抗体片段（抗赫赛汀）-PE-STXA在源自胃癌的NCI-N87细胞系中诱导凋亡。本研究强调了单链抗体片段（抗赫赛汀）-PE-STXA特异性靶向表达HER2的癌细胞的潜力。
