Nanosized drug delivery systems (DDS) have been studied as a novel strategy against cancer due to their potential to simultaneously decrease drug inactivation and systemic toxicity and increase passive and/or active drug accumulation within the tumor(s). Triterpenes are plant-derived compounds with interesting therapeutic properties. Betulinic acid (BeA) is a pentacyclic triterpene that has great cytotoxic activity against different cancer types. Herein, we developed a nanosized protein-based DDS of bovine serum albumin (BSA) as the drug carrier combining two compounds, doxorubicin (Dox) and the triterpene BeA, using an oil-water-like micro-emulsion method. We used spectrophotometric assays to determine protein and drug concentrations in the DDS. The biophysical properties of these DDS were characterized using dynamic light scattering (DLS) and circular dichroism (CD) spectroscopy, confirming nanoparticle (NP) formation and drug loading into the protein structure, respectively. The encapsulation efficiency was 77% for Dox and 18% for BeA. More than 50% of both drugs were released within 24 h at pH 6.8, while less drug was released at pH 7.4 in this period. Co-incubation viability assays of Dox and BeA alone for 24 h demonstrated synergistic cytotoxic activity in the low μM range against non-small-cell lung carcinoma (NSCLC) A549 cells. Viability assays of the BSA-(Dox+BeA) DDS demonstrated a higher synergistic cytotoxic activity than the two drugs with no carrier. Moreover, confocal microscopy analysis confirmed the cellular internalization of the DDS and the accumulation of the Dox in the nucleus. We determined the mechanism of action of the BSA-(Dox+BeA) DDS, confirming S-phase cell cycle arrest, DNA damage, caspase cascade activation, and downregulation of epidermal growth factor receptor (EGFR) expression. This DDS has the potential to synergistically maximize the therapeutic effect of Dox and diminish chemoresistance induced by EGFR expression using a natural triterpene against NSCLC.
Cancer is the second largest cause of death worldwide with the number of new cancer cases predicted to grow significantly in the next decades. Cytotoxic drugs are the main therapy against cancer; however, they are frequently associated with severe side‐effects related to systemic toxicity and lack of tumor specificity. As consequence, the use of proteins as drug carriers has had great impact in the development of new approaches as drug delivery system (DDS) nanoparticles (NPs). For example, Abraxane®, the first FDA‐approved DDS NPs, is composed of serum albumin protein and paclitaxel drug, and has demonstrated higher tumor accumulation, patient toleration, and response rate than the free paclitaxel. Due to this, the aim of this research project is the development of a synergistic DDS NPs using serum albumin (BSA), as the drug's carrier, coupling two cytotoxic drugs: doxorubicin (DOX), and the natural triterpene betulinic acid (BeA). This system BSA[(Dox)(BeA)] was done by using water in oil (W/O)‐like emulsion followed by heat and ultrasonication. To characterize the BSA and Dox concentration in the DDS, colorimetric assays were performed. DDS's size was determined ~ 100 nm using dynamic light scattering. All the developed DDS demonstrated a strong IC50 in the μM range after 24h incubated with lung (A549) and resistant‐ovarian (A2780‐CP20) cancer cells. A complete discussion of the results will be presented. These DDS have potential to minimize drug systemic toxicity and increase drug bioavailability.
Lung cancer is the second most common cancer and the leading cause of cancer mortality in the U.S. For decades, many chemotherapeutic agents have been used in clinical settings, however, issues regarding low therapeutic index and acquired resistance to drugs (e.g. Cisplatin and Doxorubicin) have arisen. In this way, the use of macromolecules and lipids as the drug carriers, as targeting ligands or as therapeutic agents has had great impact in the development of drug delivery system (DDS) nanomedicines to improve these drawbacks. Herein we developed DDS using biocompatible macromolecules, i.e. serum albumin (BSA), lysozyme (Lyz) and methyl‐β‐cyclodextrin (mβCD) as drug carriers and linoleic acid (LinOA) as the coating. The cytotoxic agents, doxorubicin (DOX), berberine (Ber) and curcumin (Curc) were conjugated to each carrier using oil in water in oil (O/W/O)‐like emulsion system followed by heat and ultrasonication. DDS have three drugs types incorporated in the carrier structure by non‐covalent interactions. To characterize the incorporation of the drugs, fluorimetric measurements were done. The sizes of the DDS were determined ~ 100‐1000 nm using dynamic light scattering. All the developed DDS demonstrated cytotoxic patterns in the μM range after 24h incubated with lung cancer cells (A549). These DDS have potential to minimize drug systemic toxicity and increase drug bioavailability. Future experiments will be performed to determine the impact to reduce multidrug resistance.
In the US, lung cancer is the second most common cancer and the leading cause of cancer mortality in both men and women. Many cytotoxic agents for cancer chemotherapy are being developed under in vitro and in vivo studies, however, the main problem is the low therapeutic index. Another important aspect is the acquired resistance to drugs (e.g. Cisplatin) by cancer cells. The aim of this research project is the development of drug delivery systems (DDS) using bovine serum albumin (BSA) as the drug's carrier and cisplatin (CisPt) and curcumin (Curc) as the drugs. Heat and ultrasonication were used to promote partial unfolding of the protein in a nanoparticle rearrangement and intercalation of the drug moieties into the carrier protein. The DDS developed include BSA‐CisPt and BSA‐Curc. To characterize the ratio of the conjugation of these systems colorimetric assays were performed. Braford assay was used for BSA, for Curc, its intrinsic absorbance and for CisPt, o‐phenylenediamine/heat assay. The preliminary results of the DDS report a ratio of BSA‐Cisplatin 1:25 and BSA‐Curcumin 1:30. Both DDS demonstrated cytotoxicity on lung cancer cells (A549) of an IC50 in the μM range after 24h. In future experiments the DDS’ size will be determined utilizing a dynamic light scattering technique. In addition, other molecules such as folic acid and polyethylene glycol will be added to improve specificity and stability respectively. These DDS have potential to minimize drug systemic toxicity, increased bioavailability and reduce multidrug resistance.Support or Funding InformationSan Juan Bautista Institutional funds and applying to travel awards.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
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