Investigation of effects of silver nanoparticles synthesized biologically and chemically on HIF-1 signal pathway in PANC-1 human pancreatic cancer cell line

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal solid tumors and is the fourth leading cause of cancer-related deaths. Since pancreatic cancer has no early symptoms and can be diagnosed at an advanced age in both sexes, the survival rate decreases over the years. There have been remarkable advances in the treatment of pancreatic cancer over the past century, including improvements in surgical techniques and advances in adjuvant and neoadjuvant therapies. However, the incidence of pancreatic cancer is increasing globally and remains a fatal disease. Therefore, there is a need to develop new therapeutic agents to treat advanced pancreatic cancer solid tumors. It is envisaged that green synthesis-mediated silver nanoparticles (AgNPs) may soon be used as novel chemotherapeutic supplements or drugs to treat pancreatic cancer in humans. Nanoparticles play a critical role in the development of effective techniques for various diseases, as they are biocompatible and have a large surface area. In particular, plant-based nanoparticles have the potential to be widely used in medicine, including therapeutic drugs, as they exhibit antibacterial activity, anticancer and antiviral activity. The biological synthesis of nanoparticles shows that it is a much more reliable approach than the use of chemical drugs due to the ubiquitous availability of plant extracts and physiologically active biomolecules. Silver nanoparticles, which have lower cost compared to other nanoparticles, have been used as modern chemotherapeutic drugs to treat pancreatic and various cancers in recent years. The studies to be carried out in this framework will greatly contribute to the treatment of pancreatic cancer. In this thesis study, unlike conventional methods, biological synthesis of silver nanoparticles was carried out using microwave-mediated method extract of Crocus sativus (Saffron) stigma part. Biologically synthesised AgNPs and commercially purchased chemical AgNPs were characterised by UV-Vis, TEM, FESEM-EDX, FTIR, XRD, DLS and Zeta potential methods. PANC-1 human pancreatic cancer and HEK293 cells were treated with increasing concentrations of biologically synthesised AgNP, commercially available chemical AgNP and Crocus sativus (saffron) stigma extract for 24 hours. The cytotoxic effects on Panc-1 human pancreatic cancer and HEK293 normal cell lines were determined by -4,5-dimethyl-thiazolyl-2,5-diphenyltetrazolium bromide (MTT) test. After the IC50 dose was determined by MTT test, changes in mRNA levels of HIF1 transcription factor and VEGFA, FLT-1, GLUT1, Bcl-2 and pro-apoptotic Bax genes, which are target genes related to invasion, metastasis, angiogenesis and glucose metabolism, were analysed by qRT-PCR. As a result of MTT assay, a dose-dependent decrease in cell proliferation, a significant decrease in HIF-1 and its target genes VEGFA, FLT-1, GLUT1, Bcl-2 gene expressions and a significant increase in Bax gene were detected by qRT-PCR. Since the synthesis carried out in this study is a cheap, one-step and environmentally friendly method, it was determined that plant-mediated synthesis approaches are a rather reliable and economical way to synthesise AgNPs. The results of the study showed that biologically synthesised AgNPs are safer than chemical AgNPs because they have less toxic effects on normal HEK293 cells. Biologically synthesised AgNPs may be a promising anticancer drug that overcomes the shortcomings of current cancer therapy, such as resistance to radiation and chemotherapy treatments in pancreatic cancer, as it exhibits effective results on the HIF1 signalling pathway, which is activated in the event of hypoxia that increases the aggressiveness and severity of tumour progression. Keywords: Anticancer activity, Crocus sativus (Saffron), Silver nanoparticle (AgNP), Hypoxia, Microwave-mediated biological synthesis, Pancreatic ductal adenocarcinoma