Synthesis, characterization, and investigation of the anticancer activity of different mof-based drug delivery systems

In this thesis, two advanced multifunctional metal-organic framework (MOF)-based drug delivery systems were developed for targeted and controlled release of Capecitabine (CAP), a well-known chemotherapeutic agent widely used in the treatment of breast cancer. Metal–organic frameworks (MOFs) are a class of crystalline hybrid materials formed by the coordination of metal ions or clusters with organic ligands. Among them, MIL-100(Fe) and UiO-66(Zr) have emerged as two of the most studied and versatile frameworks, especially in biomedical applications due to their structural tunability, biocompatibility, and high porosity. MIL-100(Fe) is constructed from trivalent iron (Fe³⁺) ions coordinated with trimesic acid (1,3,5-benzenetricarboxylic acid, BTC), forming a mesoporous structure with exceptionally high surface area (>1500 m²/g) and two types of large cages interconnected by microporous windows. The synthesis of MIL-100(Fe) can be carried out via conventional hydrothermal or solvothermal routes using aqueous or mildly acidic media, although microwave-assisted synthesis is increasingly adopted due to reduced reaction times and enhanced crystallinity. Moreover, green synthesis approaches using water or ethanol as solvents have been developed to improve biocompatibility and reduce environmental impact. UiO-66(Zr), on the other hand, is a microporous MOF based on zirconium (IV) clusters (Zr₆O₄(OH)₄) and terephthalic acid (1,4-benzenedicarboxylic acid, BDC). This MOF belongs to the UiO (University of Oslo) series and is distinguished by its 12-connected nodes, which result in an extraordinarily stable framework that can withstand extreme pH, high temperature, and biological conditions. Microwave or ultrasound-assisted syntheses have also been explored to obtain nano-sized particles suitable for biomedical applications. More recently, aqueous-phase syntheses have been developed to increase environmental compatibility and clinical translatability. The first nanocomposite system comprises MIL-100(Fe) as the MOF carrier, valonia tannin (VT) as a natural polyphenolic linker, and glucose oxidase (GOx) as a supportive enzyme to induce a starvation therapy effect in the tumor microenvironment. The second system uses UiO-66(Zr) modified with catechin (CAT), another natural polyphenol, and further functionalized with D-mannose (D-MAN) to enhance targeting capabilities via receptor-mediated uptake. The rationale behind using MOFs lies in their intrinsic properties, including high surface area, tunable porosity, and ease of functionalization, enabling efficient drug loading, protection, and pH-responsive release. In both systems, the post-synthetic modification approach allowed functional components (VT, CAT, GOx, and D-MAN) to be stably anchored onto the MOF surface or within the porous structure without impairing the structural integrity of the frameworks. Characterization studies using Fourier Transform Infrared Spectroscopy (FT-IR) confirmed successful coordination of polyphenols with MOF surfaces and encapsulation of CAP. X-ray diffraction (XRD) patterns supported the crystallinity and phase purity of the synthesized MOFs. Thermogravimetric analysis (TGA) showed multi-step mass losses corresponding to decomposition of organic moieties. Scanning electron microscopy (SEM) revealed homogenous nanoparticle morphology, while UV-VIS spectrophotometry and zeta potential analyses provided insights into encapsulation behavior and colloidal stability. Drug release experiments conducted in buffer solutions at pH 7.4 and pH 5.5 revealed a significantly higher release rate in acidic medium, mimicking tumor microenvironments, thus verifying pH-responsive behavior. This confirms the potential of both MOF systems to release CAP preferentially at tumor sites while minimizing premature release in healthy tissues. In vitro cytotoxicity studies on MDA-MB-231 breast cancer cells showed significant suppression of cancer cell viability for both systems. The GOx-loaded variant not only facilitated ROS generation and intracellular acidification but also limited glucose availability in cancer cells, thus simulating a starvation therapy mechanism. On the other hand, the D-MAN functionalized system exhibited enhanced internalization and cytotoxicity due to mannosylation-mediated receptor targeting, confirming the efficacy of selective delivery. Furthermore, structural-functional correlations between MOF components and their biological outcomes were analyzed in-depth. The integration of VT and CAT not only stabilized the MOF structures in physiological environments but also contributed to enhanced bioactivity. VT, rich in galloyl groups, showed strong metal coordination and radical scavenging capabilities, whereas CAT, a flavonoid antioxidant, reduced oxidative stress in cancer cells while also forming stable hydrogen bonds with the carrier matrix. The GOx-catalyzed glucose oxidation pathway significantly enhanced intracellular hydrogen peroxide levels, leading to oxidative stress and increased apoptosis in tumor cells. Simultaneously, gluconic acid produced by GOx acidified the microenvironment, promoting pH-triggered CAP release and enhancing the overall therapeutic efficiency. These synergistic effects underscore the advantage of enzymatic modulation in MOF-based drug delivery. In the UiO-66@CAT@CAP@D-MAN system, mannose-functionalization played a critical role in selective cancer targeting. Mannose receptors (CD206) are overexpressed in many breast cancer subtypes and macrophage populations within tumor tissues. The engineered MOF surfaces demonstrated selective uptake by MDA-MB-231 cells, indicating potential for active targeting via lectin-mediated endocytosis. Flow cytometry and fluorescence microscopy images supported these results by showing increased internalization of D-MAN-modified particles. In conclusion, the presented MOF-based drug delivery systems, MIL-100@VT@CAP@GOx and UiO-66@CAT@CAP@D-MAN, represent a highly modular and adaptable therapeutic approach. They successfully integrate principles of materials science, cancer biology, and pharmaceutical nanotechnology. Future directions include in vivo testing and optimization of pharmacokinetics, scaling up for clinical-grade synthesis, and exploring combinatory therapies using these carriers as co-delivery platforms for multiple anticancer agents.Taken together, this study provides compelling evidence that integrating polyphenols, metabolic enzymes, and targeting sugars into MOF structures leads to a significant enhancement in therapeutic potential. These findings establish a novel paradigm in MOF-based drug delivery, positioning such systems as potential frontrunners in the next generation of cancer therapeutics. The approach detailed in this thesis opens new avenues for personalized and precision medicine by offering controllable, biocompatible, and highly effective nanoscale delivery vehicles tailored for complex pathologies like triple-negative breast cancer. Beyond the primary findings, this thesis also contributes to the broader understanding of how metal-organic frameworks can be engineered as multifunctional drug delivery systems by integrating diverse biochemical strategies. The combination of MOF structural design, polyphenol-mediated biofunctionalization, enzyme-responsive drug release, and receptor-targeted surface modification exemplifies a next-generation drug delivery paradigm. Additionally, the integration of VT and CAT demonstrates a significant advancement over traditional synthetic ligands. These natural polyphenols not only confer enhanced biocompatibility and reduced immunogenicity but also actively participate in the therapeutic mechanism via antioxidant or pro-apoptotic interactions. Their multifunctionality makes them ideal candidates in the era of green nanomedicine, which prioritizes non-toxic, sustainable components for therapeutic nanostructure design. The thesis further demonstrates that MOF-based composites can maintain stability under physiological conditions while selectively degrading under pathological stimuli. This attribute is critical for in vivo applications where premature drug leakage can compromise therapeutic outcomes and increase systemic toxicity. Data obtained from zeta potential measurements and SEM imaging under different pH conditions confirmed that both systems exhibit structural resilience in neutral pH and initiate disintegration in acidic microenvironments. Another notable feature of this study is the successful dual-functionalization strategy. By integrating both a therapeutic enzyme (GOx) and a targeting ligand (D-MAN) into the carrier design, a multi-level mechanism of action was established: enzyme-initiated microenvironment modulation enhanced drug activity, while ligand-receptor interactions facilitated selective uptake. Such dual-targeting or dual-stimuli-responsive platforms are at the frontier of nanomedicine and hold potential for combination therapy and personalized treatment regimens. Moreover, this work opens new opportunities for co-delivery systems. The porosity and tunable structure of MOFs such as MIL-100 and UiO-66 could allow co-encapsulation of synergistic drug combinations or genetic materials (e.g., siRNA or CRISPR-Cas components), which can be released in a temporally and spatially controlled manner. Preliminary studies on CAP encapsulation efficiency and stability provide a foundational protocol that could be adapted for broader drug classes. In terms of translational potential, the modularity of the synthetic approach—using microwave-assisted solvothermal synthesis followed by straightforward post-synthetic modification—makes scale-up feasible for preclinical studies. The components employed are either FDA-approved or biocompatible, which supports the clinical relevance of these systems. Additional studies should now focus on pharmacokinetics, biodistribution, and immunogenicity using in vivo animal models. xxvi The work presented here contributes not only to drug delivery research but also to the interdisciplinary integration of materials science, enzymology, tumor biology, and pharmaceutical technology. By bridging these domains, the thesis sets the stage for a more intelligent, responsive, and bio-integrated drug delivery future. In conclusion, the MIL-100@VT@CAP@GOx and UiO-66@CAT@CAP@D-MAN platforms are more than passive carriers; they represent dynamic therapeutic systems tailored for selective cancer treatment. Their responsiveness to microenvironmental stimuli, high loading efficiency, low cytotoxicity to normal cells, and targeted uptake make them strong candidates for future drug delivery innovations. This study's approach can serve as a blueprint for designing next-generation MOF-based systems that are customizable, scalable, and clinically applicable.