Browsing by Author "Costa, Elisabete"
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- 3D tumor spheroids as in vitro models to mimic in vivo human solid tumors resistance to therapeutic drugsPublication . Nunes, Ana S.; Barros, Andreia; Costa, Elisabete; Moreira, André; Correia, I.J.Three-dimensional cell culture models, such as spheroids, can be used in the process of the development of new anticancer agents because they are able to closely mimic the main features of human solid tumors, namely their structural organization, cellular layered assembling, hypoxia, and nutrient gradients. These properties imprint to the spheroids an anticancer therapeutics resistance profile, which is similar to that displayed by human solid tumors. In this review, an overview of the drug resistance mechanisms observed in 3D tumor spheroids is provided. Furthermore, comparisons between the therapeutics resistance profile exhibited by spheroids, and 2D cell cultures are presented. Finally, examples of the therapeutic approaches that have been developed to surpass the drug resistance mechanisms exhibited by spheroids are described.
- Breast cancer targeted photothermal therapy mediated by hyaluronic acid functionalized reduced graphene oxidePublication . Sousa, Ana Rita Lima; Diogo, Duarte de Melo; Alves, Cátia Gomes; Costa, Elisabete; Louro, Ricardo; Mendonça, António G.; Correia, I.J.The use of graphene-based nanomaterials in cancer photothermal therapy (PTT) is an emerging alternative to the currently available cancer treatments. In this regard, reduced graphene oxide (rGO) has been widely explored for cancer PTT due to its excellent photothermal capacity. However, rGO has some limitations, such as low colloidal stability and water insolubility, as well as absence of targeting capacity towards cancer cells. Herein, rGO produced by an environmentally- friendly method was functionalized with an amphiphilic polymer based on hyaluronic acid (HA-rGO) through hydrophobic-hydrophobic interactions for application in targeted breast cancer PTT. The functionalization improved rGO colloidal stability and cytocompatibility towards normal and breast cancer cells, as well as conferred targeting capacity towards CD44 overexpressing breast cancer cells. In addition, the photothermal effect mediated by HA-rGO upon laser irradiation reduced breast cancer cells’ viability. Overall, HA-rGO demonstrated a great potential for being used on-demand and selective treatment of breast cancer cells.
- Environmentally-friendly reduced graphene oxide functionalized with hyaluronic acid for targeted cancer photothermal therapyPublication . Sousa, Ana Rita Lima; Diogo, Duarte de Melo; Alves, Cátia; Costa, Elisabete; Ferreira, Paula; Louro, Ricardo; Mendonça, António; Correia, I.J.Reduced Graphene Oxide (rGO) is one of the most promising nanomaterials for cancer photothermal therapy (PTT) due to its high near infrared (NIR) absorption. However, the rGO producing methods uses dangerous reducing agents, resulting in poor biocompatibility. Additionally, rGO also displays poor colloidal stability and is unable to target cancer cells. These limitations can be improved by using environmentally-friendly reduction methods and by functionalizing this nanomaterial with amphiphilic polymers. In this work, the production of reduced Graphene Oxide (rGO) rGO was performed by using an environmentally-friendly method (reduction with L-ascorbic Acid (LAA)). Then, the obtained rGO was functionalized with an amphiphilic polymer based on hyaluronic acid (HA-g-PMAO) for application in CD44-targeted breast cancer PTT
- Establishment of 2D Cell Cultures Derived From 3D MCF‐7 Spheroids Displaying a Doxorubicin Resistant ProfilePublication . Nunes, Ana S.; Costa, Elisabete; Barros, Andreia; Diogo, Duarte de Melo; Correia, I.J.In vitro 3D cancer spheroids generally exhibit a drug resistance profile similar to that found in solid tumors. Due to this property, these models are an appealing for anticancer compounds screening. Nevertheless, the techniques and methods aimed for drug discovery are mostly standardized for cells cultured in 2D. The development of 2D cell culture models displaying a drug resistant profile is required to mimic the in vivo tumors, while the equipment, techniques, and methodologies established for conventional 2D cell cultures can continue to be employed in compound screening. In this work, the response of 3D-derived MCF-7 cells subsequently cultured in 2D in medium supplemented with glutathione (GSH) (antioxidant agent found in high levels in breast cancer tissues and a promoter of cancer cells resistance) to Doxorubicin (DOX) is evaluated. These cells demonstrated a resistance toward DOX closer to that displayed by 3D spheroids, which is higher than that exhibited by standard 2D cell cultures. In fact, the 50% inhibitory concentration (IC50 ) of DOX in 3D-derived MCF-7 cell cultures supplemented with GSH is about eight-times higher than that obtained for conventional 2D cell cultures (cultured without GSH), and is only about two-times lower than that attained for 3D MCF-7 spheroids (cultured without GSH). Further investigation revealed that this improved resistance of 3D-derived MCF-7 cells may result from their increased P-glycoprotein (P-gp) activity and reduced production of intracellular reactive oxygen species (ROS).
- Hyaluronic acid and vitamin E polyethylene glycol succinate functionalized gold-core silica shell nanorods for cancer targeted photothermal therapyPublication . Jacinto, Telma A.; Rodrigues, Ana Carolina Félix; Moreira, André F.; Miguel, Sónia P.; Costa, Elisabete; Ferreira, Paula; Correia, I.J.Gold-core mesoporous silica shell (AuMSS) nanorods unique physicochemical properties makes them versatile and promising nanomedicines for cancer photothermal therapy. Nevertheless, these nanomaterials present a reduced half-life in the blood and poor specificity towards the tumor tissue. Herein, d-α-Tocopherol polyethylene glycol 1000 succinate (TPGS) and Hyaluronic Acid (HA) were combined for the first time to improve the AuMSS nanorods biological performance. The obtained results revealed that AuMSS surface functionalization induced the surface charge neutralization, from -28 ± 10 mV to −3 ± 5 mV and −10 ± 4 mV for AuMSS-TPGS-HA (1:1) and (4:1) formulations, without impacting on nanomaterials’ photothermal capacity. Moreover, the AuMSS functionalization improved the nanomaterials hemocompatibility and selectivity towards the cancer cells, particularly in the AuMSS-TPGS-HA (4:1) formulation. Furthermore, both formulations were able to mediate an on-demand photothermal effect, that induced the HeLa cancer cells death, confirming its potential for being applied as targeted multifunctional theragnostic nanomedicines.
- Hyaluronic acid functionalized nanoparticles loaded with IR780 and DOX for cancer chemo-photothermal therapyPublication . Alves, Cátia; Diogo, Duarte Miguel De Melo; Sousa, Ana Rita Lima; Costa, Elisabete; Correia, IlidioIR780 is a near infrared (NIR) dye with a huge potential to be applied in cancer phototherapy and imaging. However, IR780 poor water solubility and acute cytotoxicity limit its direct use in cancer theragnostic. Herein, a novel Hyaluronic acid (HA)-based amphiphilic polymer was used, for the first time, in the preparation of polymeric nanoparticles (HPN) encapsulating IR780 aimed to be applied in breast cancer therapy. Furthermore, HPN co-encapsulating IR780 and Doxorubicin (DOX) were also produced in order to further enhance the therapeutic effectiveness of this nanoformulation. The results revealed that HPN were able to successfully encapsulate IR780 (IR-HPN) and the IR780-DOX combination (IR/DOX-HPN). Furthermore, the encapsulation of IR780 in HPN improved its absorption at 808 nm by about 2.2-fold, thereby enhancing its photothermal potential, as well as its cytocompatibility. The 2D in vitro cell uptake studies demonstrated that the nanostructures displayed a higher internalization by breast cancer cells than by normal cells. In addition, the assays performed in 3D in vitro models of breast cancer revealed that HPN can penetrate into spheroids. Furthermore, the 3D in vitro studies also demonstrated that the combined application of IR-HPN and NIR light was unable to induce cytotoxicity on spheroids. In contrast, IR/DOX-HPN produced a decrease on spheroids cells' viability, and their combination with NIR light induced an even stronger therapeutic effect, thus revealing the potential of these nanoparticles for cancer chemo-phototherapy.
- Influence of ClearT and ClearT2 Agitation Conditions in the Fluorescence Imaging of 3D SpheroidsPublication . Silva, Daniel N.; Costa, Elisabete; Rodrigues, Ana Carolina Félix; Diogo, Duarte de Melo; Correia, I.J.; Moreira, André F.3D tumor spheroids have arisen in the last years as potent tools for the in vitro screening of novel anticancer therapeutics. Nevertheless, to increase the reproducibility and predictability of the data originated from the spheroids it is still necessary to develop or optimize the techniques used for spheroids' physical and biomolecular characterization. Fluorescence microscopy, such as confocal laser scanning microscopy (CLSM), is a tool commonly used by researchers to characterize spheroids structure and the antitumoral effect of novel therapeutics. However, its application in spheroids' analysis is hindered by the limited light penetration in thick samples. For this purpose, optical clearing solutions have been explored to increase the spheroids' transparency by reducing the light scattering. In this study, the influence of agitation conditions (i.e., static, horizontal agitation, and rotatory agitation) on the ClearT and ClearT2 methods' clearing efficacy and tumor spheroids' imaging by CLSM was characterized. The obtained results demonstrate that the ClearT method results in the improved imaging of the spheroids interior, whereas the ClearT2 resulted in an increased propidium iodide mean fluorescence intensity as well as a higher signal depth in the Z-axis. Additionally, for both methods, the best clearing results were obtained for the spheroids treated under the rotatory agitation. In general, this work provides new insights on the ClearT and ClearT2 clearing methodologies and their utilization for improving the reproducibility of the data obtained through the CLSM, such as the analysis of the cell death in response to therapeutics administration.
- Microneedle-based delivery devices for cancer therapy: a reviewPublication . Moreira, André; Rodrigues, Ana Carolina Félix; Jacinto, Telma A.; Miguel, Sónia; Costa, Elisabete; Correia, I.J.Macroscale delivery systems that can be locally implanted on the tumor tissue as well as avoid all the complications associated to the systemic delivery of therapeutics have captured researchers' attention, in recent years. Particularly, the microneedle-based devices can be used to efficiently deliver both small and macro-molecules, like chemotherapeutics, proteins, and genetic material, along with nanoparticle-based anticancer therapies. Such capacity prompted the application of microneedle devices for the development of new anticancer vaccines that can permeate the tumor tissue and simultaneously improve the effectiveness of therapeutic agents. Based on the promising results demonstrated by the microneedle systems in the local administration of anticancer therapeutics, this review summarizes the different microneedle formulations developed up to now aimed for application on cancer therapy (mphasizing those produced with polymers). Additionally, the microneedles' general properties, type of therapeutic approach and its main advantages are also highlighted.
- Poly (vinyl alcohol)/chitosan layer-by-layer microneedles for cancer chemo-photothermal therapyPublication . Moreira, André F.; Rodrigues, Ana Carolina Félix; Jacinto, Telma A.; Miguel, Sónia P.; Costa, Elisabete; Correia, I.J.The combination of photothermal and chemo- therapies displays a high potential to increase the efficacy of the cancer treatments or even promote their eradication. In this study, the micromoulding and electrospraying techniques were combined to produce polyvinylpyrrolidone microneedles coated with chitosan and poly (vinyl alcohol) for mediating the delivery of doxorubicin and AuMSS nanorods (Dox@MicroN) to cancer cells. The microneedles' physicochemical characterization demonstrated that the electrospraying technique can be used to produce a layer-by-layer coating consisting of layers of doxorubicin-loaded chitosan and AuMSS enriched poly (vinyl alcohol). Further, the Dox@MicroN patches presented a good photothermal capacity leading to a temperature increase of 12 °C under near-infrared irradiation (808 nm, 1.7 W/cm-2 for 5 min), which in conjugation with the chitosan' pH sensitivity could be used to control the doxorubicin release. Moreover, the microneedles were able to penetrate the tumor-mimicking agarose gel and promote a layer dependent drug release. Additionally, the Dox@MicroN patches' capacity to simultaneously mediate the chemo- and photothermal-therapies rendered a superior cytotoxic effect against the cervical cancer cells. Overall, the Dox@MicroN patches demonstrated to be a simple macroscale delivery device that can be used to mediate the local administration of new drug-photothermal combinations, avoiding all the issues related to the systemic administration of anti-cancer therapeutics.