Phd in TRANSATIONAL MEDICINE AND ONCOLOGY

Thesis title: Synthesis and characterization of nuclear nano-theragnostic formulation for metastatic ovarian cancer treatment

Background Ovarian cancer (OC) is often diagnosed at advanced stages, which significantly limits therapeutic options and contributes to poor prognosis. Luteinizing hormone-releasing hormone (LHRH) receptors are overexpressed in many ovarian tumor cells, making them attractive targets for receptor-mediated drug delivery. Nanoparticles (NPs) offer advantages for targeted delivery of therapeutic and imaging agents. In nuclear medicine, theragnostic nanoparticle systems represent promising platforms for combining diagnosis and therapy. In the present study, poly(lactic-co-glycolic acid) (PLGA) nanoparticles were synthesized and functionalized with the LHRH peptide to develop a targeted nanoparticle system for ovarian cancer applications. Methods PLGA nanoparticles were synthesized using a microfluidic technique, allowing precise control over particle size and formulation reproducibility. Radiolabelling 99mTc was performed to evaluate the feasibility of developing a radiolabelled nanoparticle system. Non-radioactive formulations were prepared under similar conditions and subsequently functionalized with [D-Lys⁶]-LHRH through carbodiimide-mediated conjugation or using PLGA-PEG-NHS polymers. The physicochemical properties of the nanoparticles were characterized by DLS. Conjugation efficiency of LHRH was determined by HPLC. Cellular interaction studies, including binding assays, cellular uptake assays, and cell viability assays, were performed using ovarian cancer cell lines. Results PLGA nanoparticles exhibited particle sizes below 200 nm with relatively low PDI values, indicating a uniform nanoparticle distribution. LHRH conjugation efficiency reached approximately 85–89% using the PLGA-PEG-NHS system. Radiolabelled nanoparticles showed radiochemical purity greater than 99%, confirming successful incorporation of 99mTc. In vitro cell studies demonstrated enhanced cellular interactions of LHRH-functionalized nanoparticles compared with non-functionalized nanoparticles. In particular, ES-2 ovarian cancer cells exhibited significantly higher uptake of LHRH-conjugated nanoparticles, supporting receptor-mediated targeting. Binding studies using A2780 and SKOV-3 ovarian cancer cell lines also showed increased fluorescence intensity for LHRH-functionalized nanoparticles compared with non-targeted formulations, while minimal binding was observed in control cells. Cell viability assays indicated that the nanoparticle formulations were non toxic under the tested conditions. Conclusion The microfluidic synthesis approach enabled reproducible production of PLGA nanoparticles with controlled physicochemical properties. LHRH functionalization significantly improved nanoparticle interaction with ovarian cancer cells, demonstrating the potential of this targeting strategy. The successful incorporation of 99mTc further supports the feasibility of developing LHRH-functionalized PLGA nanoparticles as a nuclear nano-diagnostic platform for ovarian cancer imaging. Future experiments with therapeutic isotopes will possibly expand the role on the NPs for cancer therapy. Keywords Ovarian cancer, PLGA nanoparticles, LHRH targeting, Radiolabelling, Nano-theragnostic, Microfluidic synthesis