Advanced Photonics Assisted Theranostic Platforms For Locoregional Cancer Approaches

This thesis reports the development of an integrated theranostic platform that combines nanomedicine and advanced optical technologies to overcome the major limitations of conventional cancer treatments. Rooted in the principles of theranostics—the concurrent integration of diagnostic and therapeutic functionalities—the proposed approach aims to advance personalized medicine by enhancing therapeutic efficacy while minimizing systemic toxicity. In Chapter 1, the underlying theoretical framework is established by exploring theranostics, defined as the integrated application of diagnostic and therapeutic strategies within a unified methodology to enhance therapeutic efficacy and minimize undesirable systemic side effects. Nanomedicine is identified as a critical enabling technology, leveraging nanoscale materials to improve drug bioavailability, reduce systemic toxicity, and enable precise targeting of diseased tissues. The research prioritizes Polymeric Nanoparticles, specifically those formulated from Poly(lactic-co- glycolic acid) (PLGA), due to their established biocompatibility, biodegradability, and versatility for controlled release and dual targeting (passive via the Enhanced Permeability and Retention (EPR) effect and active via surface functionalization). In Chapter 2 is described the major constraint in light-based therapeutic modalities, offering precise and minimally invasive treatment by selectively targeting tumor cells while sparing healthy tissues. Although limited by shallow tissue penetration, this challenge is effectively overcome through optical fibers, which enable localized light delivery deep within the body via integration into medical tools such as needles and catheters. Recent advances highlight the crucial role of optical fibers in enhancing the precision and efficacy of photothermal and photodynamic therapies, as well as in emerging light- triggered drug release and optoporation techniques. Furthermore, fiber-based theranostic platforms that unite diagnostic and therapeutic functions exemplify the growing potential of these technologies. Overall, optical fiber integration represents a transformative step toward personalized, precision- guided cancer treatment. Due to these considerations, in Chapter 3 is treated the key achievement of this work, consisting in the introduction of a novel "Lab-on-Fiber" (LOF) platform for loco-regional, light-triggered drug delivery. In particular, a cost-effective core-offset optical fiber (coOF) was developed to enable light- triggered therapy. This custom-engineered fiber is capable of hosting nanocarriers on its surface, both on tip and lateral surface, where lateral light scattering promotes efficient and controlled drug release. Nanocarriers were immobilized on the fiber surface using a UV-sensitive photocleavable linker, allowing precise spatiotemporal control and on-demand cargo release. The fiber was functionalized with multifunctional polymeric nanoparticles composed of biocompatible, FDA-approved poly(lactic-co-glycolic acid) (PLGA), serving as efficient drug delivery systems (DDSs) to treat HER2+ breast cancer cells. A major innovation of this work is the development of a dual- functionalized PLGA nanoparticle system for Trastuzumab (TZ), a monoclonal antibody employed in the treatment of HER2-positive breast cancer. In this design, TZ was simultaneously encapsulated within the nanoparticle core and covalently conjugated to its surface, ensuring antibody stability and enabling active targeting of HER2-overexpressing cancer cells. This dual- functional configuration resulted in markedly enhanced antiproliferative activity, HER2 receptor downregulation, and apoptosis induction compared to free TZ. The Lab-on-Fiber system incorporating these nanoparticles was validated in both batch and microfluidic configurations, confirming precise, light-triggered nanoparticle release and demonstrating its potential as a minimally invasive platform for localized cancer therapy. The versatility of PLGA-based nanocarriers was demonstrated in Chapter 4 by the capability of this polymer to encapsulate nutraceutical molecules for cancer treatments and the integration of labeled therapeutic elements for optical imaging capabilities and real-time monitoring of delivery. For Hepatocellular Carcinoma (HCC) therapy, the thesis investigated the encapsulation of Capsaicin (Cap), an antitumor alkaloid, into PLGA NPs to address its limitations in bioavailability and stability. Meticulous optimization of the single-emulsion method resulted in exceptionally small NPs and a remarkable Encapsulation Efficiency (96%), representing a 21% improvement over reported systems. The PLGA-Cap preparation significantly enhanced apoptosis, inducing 40% higher Caspase-3 activation and 58% greater Reactive Oxygen Species (ROS) generation at high concentrations compared to free Cap, suggesting improved therapeutic effectiveness by protecting the drug from rapid degradation. Beyond its therapeutic scope, the thesis explores the potential of nanocarriers for fluorescence imaging in the context of gene therapy for bladder cancer. The incorporation of fluorescence-based imaging enabled real-time tracking of intracellular processes. To investigate the intracellular fate of nanocarriers, Rhodamine B–labeled PLGA nanoparticles were loaded with a peptide nucleic acid (PNA) specifically designed to target the ultraconserved region uc8+, overexpressed in bladder cancer cells. Dual-labeling with spectrally distinct fluorophores— Rhodamine B for the nanoparticle matrix and ATTO 647N for the PNA cargo—enabled simultaneous, high-resolution visualization using confocal fluorescence microscopy. Complementary counterstaining of cellular compartments facilitated detailed mapping of nanoparticle biodistribution, intracellular trafficking, and molecular interactions, providing real-time insights essential for optimizing nanocarrier-based drug delivery. This integration effectively transforms drug delivery into a multifunctional diagnostic and therapeutic platform for personalized cancer management. The final section of the thesis focuses on the diagnostic field, leveraging Fluorescence-based mechanisms for the ultra-sensitive detection of tumor biomarkers, a critical step toward early diagnosis. In Chapter 5, the technique of Metal-Enhanced Fluorescence (MEF) is utilized, exploiting the Localized Surface Plasmon Resonance (LSPR) of metallic nanostructures—specifically Gold Nanoparticles (AuNPs)—to amplify the fluorescence signal of adjacent fluorophores, optimal within the 5–30nm range. This approach drastically enhances the signal-to-noise ratio required for detecting biomarkers present at very low concentrations. A dedicated MEF-based biosensor was developed for the ultrasensitive detection of the oncogenic salivary biomarker microRNA-223-3p, a marker for Oral Squamous Cell Carcinoma (OSCC) discussed in Chapter 6. The system employs Molecular Beacons (MBs) as the biorecognition element. The MB probe, specific for miR-223-3p, is covalently immobilized on the AuNP surface. In the absence of the target, the MB's hairpin structure keeps the fluorophore (ATTO 647N) close to the gold, inducing fluorescence quenching. Upon hybridization with the target miRNA, the MB undergoes a conformational change that opens the hairpin, physically distancing the fluorophore to the optimal MEF range (11 nm), resulting in a strongly amplified fluorescence signal ("turn-on" detection). Finite-element simulations and experimental studies revealed a dual fluorescence quenching/enhancement mechanism, resulting in significant signal amplification upon target recognition—highlighting its potential for early-stage, non-invasive cancer diagnosis. Collectively, this thesis establishes a comprehensive and versatile theranostic framework integrating nanotechnology-driven drug delivery, optical modulation, and plasmonic biosensing. The proposed Lab-on-Fiber platform represents a transformative step toward personalized, real-time monitored cancer therapy, while the MEF-based biosensor lays the foundation for next-generation molecular diagnostics with strong translational potential in clinical oncology.