Graphene Oxide as a Nanocarrier for Biochemical Molecules: Current Understanding and Trends

The development of an advanced and efficient drug delivery system with significant improvement in its efficacy and enhanced therapeutic value is one of the critical challenges in modern medicinal biology. The integration of nanomaterial science with molecular and cellular biology has helped in the advancement and development of novel drug delivery nanocarrier systems with precision and decreased side effects. The design and synthesis of nanocarriers using graphene oxide (GO) have been rapidly growing over the past few years. Due to its remarkable physicochemical properties, GO has been extensively used in efforts to construct nanocarriers with high specificity, selectivity, and biocompatibility, and low cytotoxicity. The focus of this review is to summarize and address recent uses of GO-based nanocarriers and the improvements as efficient drug delivery systems. We briefly describe the concepts and challenges associated with nanocarrier systems followed by providing critical examples of GO-based delivery of drug molecules and genes. Finally, the review delivers brief conclusions on the current understanding and prospects of nanocarrier delivery systems.O

Mechanogrowth factor (IGF-1Ec) and flourescent nanoparticles in colorectal cancer

The IGF-1 axis was an area of significant interest in cancer therapy following promising preclinical studies but led to disappointing clinical trials. Further scrutinization of this pathway is, therefore, warranted. The IGF-1 axis has been demonstrated to inhibit autophagy via the Akt/PI3K pathway and induce autophagy via the ERK pathway. Autophagy has been associated with chemotherapy resistance in tumour cells. My work in this thesis involved investigating the expression of an iso-form of IGF-1 referred to as IGF1-Ec or Mechanogrowth Factor (MGF) in colorectal cancer tissues and polyps with immunohistochemistry. Further work was done with fluorescent nanoparticles which have exciting potential to improve the diagnostic yield of investigations including colonosco-py, improve immunohistochemistry assessment of tissue biopsies and help in surgery with in-traoperative delineation of tumours. In addition, I investigated the relationship between autopha-gy and apoptosis with a view towards developing a model for further work in investigating the ef-fect of MGF in autophagy. Semi-quantitative immunohistochemistry for MGF on colonic tissues including normal, polyp and cancer tissues demonstrated a significantly higher expression of MGF in colonic polyps (with higher expression with worsening dysplasia, p=0.001) and cancer compared to normal colon tissues (p<0.001). Semiconductor CdTe quantum dots and gold nanoparticles were synthesised and conju-gated to the MGF peptide and antibody. Gold nanoparticles were successfully characterised with immunodots and applied to the HT29 and SW620 colorectal cancer cell lines and to tissues includ-ing normal, colon cancer and polyp tissues reflecting the results from conventional immunohisto-chemistry. Autophagy inducers were administered to the cell lines HT29 and SW620 and inhibited with the use of Bafilomycin and 3-MA. Immunohistochemistry for LC3B was used to confirm the induction of autophagy, and cell viability studies were used to demonstrate significantly increased cell viability with autophagy induction and significant reduction of cell viability with inhibition of autophagy (p<0.01 at 24 hours). This model can be subjected to the application of MGF and assess its effects on cell viability. MGF is overexpressed in colonic polyps and cancer with low levels of expression in normal colon tissues offering an opportunity for the use of fluorescent gold nanoparticles to augment polyp and cancer detection in colonoscopy and intraoperative tumour delineation

Evaluation and Adaptation of Live-Cell Interferometry for Applications in Basic, Translational, and Clinical Research

Cell mass is an important indicator of cell health and status. A diverse set of techniques have been developed to precisely measure the masses of single cells, with varying degrees of technical complexity and throughput. Here, the development of a non-invasive, label-free optical technique, termed Live-Cell Interferometry (LCI), is described. Several applications are presented, including an evaluation of LCI’s utility for assessing drug response heterogeneity in patient-derived melanoma lines and the measurement of CD3+ T cell kinetics during hematopoietic stem cell transplantation. The characterization of mast cells during degranulation, the measurement of viral reactivation kinetics in Kaposi’s Sarcoma, and drug response studies in patient-derived xenograft models of triple-negative breast cancer are also discussed. Taken together, data from these studies highlight LCI’s versatility as a tool for clinical, translational, and basic research applications

Aptamer-modified nanomaterials: Principles and applications

Aptamers are promising alternative binders that can substitute antibodies in various applications. Due to the advantages of aptamers, namely their high affinity, specificity and stability, along with the benefits originating from the chemical synthesis of aptamers, they have attracted attention in various applications including their use on nanostructured material. This necessitates the immobilization of aptamers on a solid support. Since aptamer immobilization may interfere with its binding properties, the immobilization of aptamers has to be investigated and optimized. Within this review, we give general insights into the principles and factors controlling the binding affinity of immobilized aptamers. Specific features of aptamer immobilization on nanostructured surfaces and nanoparticles are highlighted and a brief overview of applications of aptamer-modified nanostructured materials is given

Nanotechnology and supramolecular chemistry in controlled release and molecular recognition proceses for biomedical applications"

Tesis por compendioLa presente tesis doctoral, titulada "Nanotecnología y química supramolecular en procesos de liberación controlada y reconocimiento molecular para aplicaciones biomédicas", se centra en dos temas importantes: el reconocimiento molecular y los procesos de liberación controlada. Esta tesis doctoral está estructurada en cuatro capítulos. El primer capítulo introduce el concepto de materiales híbridos orgánicos-inorgánicos funcionalizados con puertas moleculares y sus aplicaciones biomédicas como nanomateriales para dirigir y controlar la liberación controlada de fármacos. Además se introduce una breve descripción sobre sensors colorimétricos basados en la base de la quimica supramolecular, particularmente en los procesos de reconocimiento molecular. En particular, el capítulo 2 describe la preparacion de cinco nanodispositivos que responden a enzimas. Estos materiales híbridos se componen de dos unidades principales: un soporte mesoporoso basado en sílice inorgánica, capaz de encapsular moléculas orgánicas y un compuesto orgánico anclado en la superficie externa del soporte mesoporoso inorgánico que actúa como puerta molecular. Todos los sistemas propuestos utilizan puertas moleculares peptídicas que responden a temperatura o enzimas como estímulo. La segunda parte de esta tesis doctoral se centra en el diseño y desarrollo de un nuevo compuesto químico capaz de detectar monóxido de carbono in vivo. En resumen, para todos los resultados antes mencionados podemos decir que esta tesis doctoral constituye una contribución científica original al desarrollo de la química supramolecular. Sus resultados derivados de los estudios presentados dejan rutas abiertas para continuar el estudio y el desarrollo de nuevos materiales híbridos y sensors químicos más eficientes para aplicaciones biomédicas y terapeuticas.This PhD thesis entitled "Nanotechnology and supramolecular chemistry in controlled release and molecular recognition processes for biomedical applications", is focused on two important subjects: molecular recognition and controlled delivery processes. This PhD thesis is structured in four chapters. The first chapter introduces the concept of organic-inorganic hybrid materials containing switchable "gate-like" ensembles and their biomedical applications as nanomaterials for targeting and control drug delivery. Furthermore, is introduced a short review about chromo-fluorogenic chemosensors based on basic principles of supramolecular chemistry, particulary in molecular recognition processes. In particular, in chapter 2 is focus on the development of enzymatic-driven nanodevices. These hybrid materials are composed of two main units: an inorganic silica based mesoporous scaffold, able to store organic molecules and an organic compound anchored on the external surface of the inorganic mesoporous support than acts as molecular gate. All the systems proposed use peptidic gates that respond to temperature or enzimatic stimulis. The second part of this PhD thesis is focused on the design and development of a new chemical compound capable of detecting carbon monoxide in vivo. In summary, for all the results above mentioned we can say that this PhD thesis constitutes an original scientific contribution to the development of supramolecular chemistry. Its results derived from the studies presented leaves open routes to continue the study and development of new hybrid materials and more efficient chemical sensors with biomedical and therapeutic applications.La present tesi doctoral, titulada "Nanotecnologia i química supramolecular en processos d'alliberament controlat i reconeixement molecular per a aplicacions biomèdiques", es centra en dos temes importants de la química: el reconeixement molecular i els processos d'alliberament controlat. Aquesta tesi doctoral està estructurada en quatre capítols. El primer capítol introdueix el concepte de materials híbrids orgànics-inorgànics funcionalitzats amb portes moleculars i les seves aplicacions biomèdiques com nanomaterials per dirigir i controlar l'alliberament controlat de fàrmacs. A més s'introdueix una breu descripció sobre sensors colorimètrics fonamentats en la base de la química supramolecular, particularment en els processos de reconeixement molecular. En particular, el capítol 2 descriu la preparació de cinc nanodispositius que responen a enzims. Aquests materials híbrids es componen de dues unitats principals: un suport mesoporos basat en sílice inorgànica, capaç d'encapsular molècules orgàniques i un compost orgànic ancorat a la superfície externa del suport mesoporós inorgànic que actua com a porta molecular. La segona part d'aquesta tesi doctoral es centra en el disseny i desenvolupaent d'un nou compost químic capaç de detectar monòxid de carboni in vivo. En resum, per a tots els resultats abans mencionats podem dir que esta tesi doctoral constituïx una contribució científica original al desenvolupament de la química supramolecular. Els seus resultats derivats dels estudis presentats deixen rutes obertes per a continuar l'estudi i el desenvolupament de nous materials hibrids i sensors químics més eficients per a aplicacions biomèdiques i terapeutiques.De La Torre Paredes, C. (2017). Nanotechnology and supramolecular chemistry in controlled release and molecular recognition proceses for biomedical applications" [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/94043TESISCompendi

Photoactivatable Organic and Inorganic Nanoparticles in Cancer Therapeutics and Biosensing

In photodynamic therapy a photosensitizer drug is administered and is irradiated with light. Upon absorption of light the photosensitizer goes into its triplet state and transfers energy or an electron to oxygen to form reactive oxygen species (ROS). These ROS react with biomolecules in cells leading to cell damage and cell death. PDT has interested many researchers because of its non-invasiveness as compared to surgery, it leaves little to no scars, it is time and cost effective, it has potential for targeted treatment, and can be repeated as needed. Different photosensitizers such as porphyrines, chlorophylls, and dyes have been used in PDT to treat various cancers, skin diseases, aging and sun-damaged skin. These second generation sensitizers have yielded reduced skin sensitivity and improved extinction coefficients (up to ~ 105 L mol-1 cm-1). While PDT based on small molecule photosensitizers has shown great promise, several problems remain unsolved. The main issues with current sensitizers are (i) hydrophobicity leading to aggregation in aqueous media resulting in reduced efficacy and potential toxicity, (ii) dark toxicity of photosensitizers, (iii) non-selectivity towards malignant tissue resulting in prolonged cutaneous photosensitivity and damage to healthy tissue, (iv) limited light absorption efficiency, and (v) a lack of understanding of where the photosensitizer ends up in the tissue. In this dissertation research program, these issues were addressed by the development of conducting polymer nanoparticles as a next generation of photosensitizers. This choice was motivated by the fact that conducting polymers have large extinction coefficients ( \u3e 107 L mol-1 cm-1), are able to undergo intersystem crossing to the triplet state, and have triplet energies that are close to that of oxygen. It was therefore hypothesized that such polymers could be effective at generating ROS due to the large excitation rate that can be generated. Conducting polymer nanoparticles (CPNPs) composed of the conducting polymer poly[2-methoxy-5-(2-ethylhexyl-oxy)-p-phenylenevinylene] (MEH-PPV) were fabricated and studied in-vitro for their potential in PDT application. Although not fully selective, the nanoparticles exhibited a strong bias to the cancer cells. The formation of ROS was proven in-vitro by staining of the cells with CellROX Green Reagent, after which PDT results were quantified by MTT assays. Cell mortality was observed to scale with nanoparticle dosage and light dosage. Based on these promising results the MEH-PPV nanoparticles were developed further to allow for surface functionalization, with the aim of targeting these NPs to cancer cell lines. For this work targeting of cancers that overexpress folate receptors (FR) were considered. The functionalized nanoparticles (FNPs) were studied in OVCAR3 (ovarian cancer cell line) as FR+, MIA PaCa2 (pancreatic cell line) as FR-, and A549 (lung cancer cell line) having marginal FR expression. Complete selectivity of the FNPs towards the FR+ cell line was found. Quantification of PDT results by MTS assays and flow cytometry show that PDT treatment was fully selective to the FR+ cell line (OVCAR3). No cell mortality was observed for the other cell lines studied here within experimental error. Finally, the issue of confirming and quantifying small molecule drug delivery to diseased tissue was tackled by developing quantum dot (Qdot) biosensors with the aim of achieving fluorescence reporting of intracellular small molecule/drug delivery. For fluorescence reporting prior expertise in control of the fluorescence state of Qdots was employed, where redox active ligands can place the Qdot in a quenched OFF state. Ligand attachment was accomplished by disulfide linker chemistry. This chemistry is reversible in the presence of sulfur reducing biomolecules, resulting in Qdots in a brightly fluorescent ON state. Glutathione (GSH) is such a biomolecule that is present in the intracellular environment. Experimental in-vitro data shows that this design was successfully implemented

Engineering Graphene Oxide-based Nanostructures for DNA sensors

Various nanostructures have been explored in DNA biosensors to convert the hybridization of DNA sequences to easily measurable processes, including optical, mechanical, magnetic, or electrochemical process. In this thesis, graphene oxide, a two-dimensional nanostructure, is applied in quenching the fluorescence of a core-shell nanoparticles modified with targeted DNA sequences. The core-shell nanoparticles, iron oxide (Fe3O4) core, and fluorescent silica (SiO2) shell, were produced through a wet chemical process which can directly link to a targeted DNA sequence (DNA-t), and the graphene oxide nanosheets were produced by the oxidation of graphite. In the meantime, a complementary- DNA single strand (DNA-c) is designed to interact with graphene oxide. Two different mechanisms have been investigated in the sensing system; (1) Ionic interaction between the DNA sequences and nanostructures through cationic bridging; and (2) covalent binding between the DNA sequences and nanostructures. In the cationic bridge system, the fluorescence intensity changes with the concentration of DNA-t in the range of 0 to 30 µM with the limitation detection at 0.25 µM without graphene oxide; the other system can detect DNA-t in the range of 0 to 4 µM with limitation detection at 0.41 µM. In addition, the effect of concentration of graphene oxide on the fluorescence intensity of core-shell nanoparticles has been investigated. We hope that the the validation strategy by engineering the two dimensional nanostructured system can be further applied towards more efficient Cancer diagnosis