Simón Guerrero

Delivery of gold nanoparticles to the brain by conjugation with a peptide that recognizes the transferrin receptor

The treatment of Alzheimers disease and many other brain-related disorders is limited because of the presence of the blood-brain barrier, which highly regulate the crossing of drugs. Metal nanoparticles have unique features that could contribute to the development of new therapies for these diseases. Nanoparticles have the capacity to carry several molecules of a drug; furthermore, their unique physico-chemical properties allow, for example, photothermal therapy to produce molecular surgery to destroy tumor cells and toxic structures. Recently, we demonstrated that gold nanoparticles conjugated to the peptide CLPFFD are useful to destroy the toxic aggregates of β-amyloid, similar to the ones found in the brains of patients with Alzheimers disease. However, nanoparticles, like many other compounds, have null or very low capacity to cross the blood-brain barrier. In order to devise a strategy to improve drug delivery to the brain, here we introduced the peptide sequence THRPPMWSPVWP into the gold nanoparticle-CLPFFD conjugate. This peptide sequence interacts with the transferrin receptor present in the microvascular endothelial cells of the blood-brain barrier, thus causing an increase in the permeability of the conjugate in brain, as shown by experiments in vitro and in vivo. Our results are highly relevant for the therapeutic applications of gold nanoparticles for molecular surgery in the treatment of neurodegenerative diseases such as Alzheimers disease.

Improving the brain delivery of gold nanoparticles by conjugation with an amphipathic peptide

BACKGROUND & AIMS Gold nanoparticles (GNPs) have promising applications for drug delivery as well as for the diagnosis and treatment of several pathologies, such as those related to the CNS. However, GNPs are retained in a number of organs, such as the liver and spleen. Owing to their negative charge and/or processes of opsonization, GNPs are retained by the reticuloendothelial system, thereby decreasing their delivery to the brain. It is therefore crucial to modify the nanoparticle surface in order to increase its lipophilicity and reduce its negative charge, thus achieving enhanced delivery to the brain. RESULTS In this article, we have shown that conjugation of 12 nm GNPs with the amphipathic peptide CLPFFD increases the in vivo penetration of these particles to the rat brain. The C(GNP)-LPFFD conjugates showed a smaller negative charge and a greater hydrophobic character than citrate-capped GNPs of the same size. We administered intraperitoneal injections of citrate GNPs and C(GNP)-LPFFD in rats, and determined the gold content in the tissues by neutron activation. Compared with citrate GNPs, the C(GNP)-LPFFD conjugate improved the delivery to the brain, increasing the concentration of gold by fourfold, while simultaneously reducing its retention by the spleen 1 and 2 h after injection. At 24 h, the conjugate was partially cleared from the brain, and mainly accumulated in the liver. The C(GNP)-LPFFD did not alter the integrity of the blood-brain barrier, and had no effect on cell viability.

Synthesis and in vivo evaluation of the biodistribution of a 18F-labeled conjugate gold-nanoparticle-peptide with potential biomedical application.

Gold nanoparticles (AuNPs) have been extensively used in biological applications because of their biocompatibility, size, and ease of characterization, as well as an extensive knowledge of their surface chemistry. These features make AuNPs readily exploitable for biomedical applications, including drug delivery and novel diagnostic and therapeutic approaches. In a previous work, we studied ex vivo distribution of the conjugate C(AuNP)-LPFFD for its potential uses in the treatment of Alzheimers disease. In this study, we covalently labeled the conjugate with [(18)F]-fluorobenzoate to study the in vivo distribution of the AuNP by positron emission tomography (PET). After intravenous administration in rat, the highest concentration of the radiolabeled conjugate was found in the bladder and urine with a lower proportion in the intestine, demonstrating progressive accumulation compatible with biliary excretion of the conjugate. The conjugate also accumulated in the liver and spleen. PET imaging allowed us to study the in vivo biodistribution of the AuNPs in a noninvasive and sensitive way using a reduced number of animals. Our results show that AuNPs can be covalently and radioactively labeled for PET biodistribution studies.

Gold Nanoparticles and Microwave Irradiation Inhibit Beta-Amyloid Amyloidogenesis

Peptide-Gold nanoparticles selectively attached to β-amyloid protein (Aβ) amyloidogenic aggregates were irradiated with microwave. This treatment produces dramatic effects on the Aβ aggregates, inhibiting both the amyloidogenesis and the restoration of the amyloidogenic potential. This novel approach offers a new strategy to inhibit, locally and remotely, the amyloidogenic process, which could have application in Alzheimer’s disease therapy. We have studied the irradiation effect on the amyloidogenic process in the presence of conjugates peptide-nanoparticle by transmission electronic microscopy observations and by Thioflavine T assays to quantify the amount of fibrils in suspension. The amyloidogenic aggregates rather than the amyloid fibrils seem to be better targets for the treatment of the disease. Our results could contribute to the development of a new therapeutic strategy to inhibit the amyloidogenic process in Alzheimer’s disease.

Peptides and proteins used to enhance gold nanoparticle delivery to the brain: preclinical approaches.

An exciting and emerging field in nanomedicine involves the use of gold nanoparticles (AuNPs) in the preclinical development of new strategies for the treatment and diagnosis of brain-related diseases such as neurodegeneration and cerebral tumors. The treatment of many brain-related disorders with AuNPs, which possess useful physical properties, is limited by the blood–brain barrier (BBB). The BBB highly regulates the substances that can permeate into the brain. Peptides and proteins may represent promising tools to improve the delivery of AuNPs to the central nervous system (CNS). In this review, we summarize the potential applications of AuNPs to CNS disorders, discuss different strategies based on the use of peptides or proteins to improve the delivery of AuNPs to the brain, and examine the intranasal administration route, which bypasses the BBB. We also analyze the potential neurotoxicity of AuNPs and the perspectives and new challenges concerning the use of peptides and proteins to enhance the delivery of AuNPs to the brain. The majority of the work described in this review is in a preclinical stage of experimentation, or in select cases, in clinical trials in humans. We note that the use of AuNPs still requires substantial study before being translated into human applications. However, for further clinical research, the issues related to the potential use of AuNPs must be analyzed.

Stable Conjugates of Peptides with Gold Nanorods for Biomedical Applications with Reduced Effects on Cell Viability

Gold nanorods used in therapy and diagnosis must be nontoxic and stable in biological media and should be specific for the target. The complete combination of these three factors has hindered the use of gold nanorods as carriers in biological and biomedical applications. In this study, we produced a conjugate of gold nanorods with the peptide CLPFFD that recognizes toxic β-amyloid aggregates present in Alzheimers disease, demonstrates colloidal stability, maintains plasmonic properties, and shows no effects on cell viability in the SH-SY5Y cell line. Furthermore, the irradiation of β-amyloid in the presence of the conjugate with near-infrared region irradiation energy reduces the amyloidogenic process reducing also its cytotoxicity. The nanorods were synthesized following the seed-mediated method in cetyltrimethylammonium bromide (CTAB) and were conjugated with the N-terminal cysteine peptide, CLPFFD. The conjugate was exhaustively characterized using different techniques (Absorption spectroscopy, X-ray photoelectron spectroscopy, electron energy loss spectroscopy, and zeta potential). The effects on cell viability and cell penetration by transmission electron microscopy of the conjugate were evaluated. The chemisorption of the peptide on the surface of gold nanorods increases their stability and reduces their effects on cell viability.

Flow Chemistry to Control the Synthesis of Nano and Microparticles for Biomedical Applications

In this article we review the flow chemistry methodologies for the controlled synthesis of different kind of nano and microparticles for biomedical applications. Injection mechanism has emerged as new alternative for the synthesis of nanoparticles due to this strategy allows achieving superior levels of control of self-assemblies, leading to higher-ordered structures and rapid chemical reactions. Self-assembly events are strongly dependent on factors such as the local concentration of reagents, the mixing rates, and the shear forces, which can be finely tuned, as an example, in a microfluidic device. Injection methods have also proved to be optimal to elaborate microsystems comprising polymer solutions. Concretely, extrusion based methods can provide controlled fluid transport, rapid chemical reactions, and cost-saving advantages over conventional reactors. We provide an update of synthesis of nano and microparticles such as core/shell, Janus, nanocrystals, liposomes, and biopolymeric microgels through flow chemistry, its potential bioapplications and future challenges in this field are discussed.

Encapsulation of Gold Nanostructures and Oil-in-Water Nanocarriers in Microgels with Biomedical Potential

Here we report the incorporation of gold nanostructures (nanospheres or nanorods, functionalized with carboxylate-end PEG) and curcumin oil-in-water (O/W) nanoemulsions (CurNem) into alginate microgels using the dripping technique. While gold nanostructures are promising nanomaterials for photothermal therapy applications, CurNem possess important pharmacological activities as reported here. In this sense, we evaluated the effect of CurNem on cell viability of both cancerous and non-cancerous cell lines (AGS and HEK293T, respectively), demonstrating preferential toxicity in cancer cells and safety for the non-cancerous cells. After incorporating gold nanostructures and CurNem together into the microgels, microstructures with diameters of 220 and 540 µm were obtained. When stimulating microgels with a laser, the plasmon effect promoted a significant rise in the temperature of the medium; the temperature increase was higher for those containing gold nanorods (11–12 °C) than nanospheres (1–2 °C). Interestingly, the incorporation of both nanosystems in the microgels maintains the photothermal properties of the gold nanostructures unmodified and retains with high efficiency the curcumin nanocarriers. We conclude that these results will be of interest to design hydrogel formulations with therapeutic applications.

Functionalization of stable fluorescent nanodiamonds towards reliable detection of biomarkers for Alzheimer’s disease

AbstractBackgroundStable and non-toxic fluorescent markers are gaining attention in molecular diagnostics as powerful tools for enabling long and reliable biological studies. Such markers should not only have a long half-life under several assay conditions showing no photo bleaching or blinking but also, they must allow for their conjugation or functionalization as a crucial step for numerous applications such as cellular tracking, biomarker detection and drug delivery.ResultsWe report the functionalization of stable fluorescent markers based on nanodiamonds (NDs) with a bifunctional peptide. This peptide is made of a cell penetrating peptide and a six amino acids long β-sheet breaker peptide that is able to recognize amyloid β (Aβ) aggregates, a biomarker for the Alzheimer disease. Our results indicate that functionalized NDs (fNDs) are not cytotoxic and can be internalized by the cells. The fNDs allow ultrasensitive detection (at picomolar concentrations of NDs) of in vitro amyloid fibrils and amyloid aggregates in AD mice brains.ConclusionsThe fluorescence of functionalized NDs is more stable than that of fluorescent markers commonly used to stain Aβ aggregates such as Thioflavin T. These results pave the way for performing ultrasensitive and reliable detection of Aβ aggregates involved in the pathogenesis of the Alzheimer disease.

Biomimetic quantum dot-labeled B16F10 murine melanoma cells as a tool to monitor early steps of lung metastasis by in vivo imaging

Background Numerous studies have proposed the use of fluorescent semiconductor nanoparticles or quantum dots (QDs) as novel tools to label cells and tumors. However, QD applications are limited by their toxicity in biological systems and little is known about whether QDs affect the capacity of cancer cells to metastasize. Previously, we described the “biomimetic” synthesis of CdTe-QDs (QDs-glutathione [GSH]) with increased biocompatibility and the potential utility in labeling cells. Purpose In order to determine the feasibility of using QDs-GSH as a tool for tracking tumor cells during early metastasis, we characterized here for the first time, the in vitro and in vivo effects of the incorporation of green or red biomimetic QDs-GSH into B16F10 cells, a syngeneic mouse melanoma line for metastasis assays in C57BL/6 mice. Methods B16F10 cells were labeled with green or red biomimetic QDs-GSH in the presence or absence of n-acetylcysteine. Then, migration, invasion and proliferation of labeled B16F10 were evaluated in vitro. Finally, the B16F10 cells labeled with red QDs-GSH were used to monitor in vivo lung metastasis at early time points (5 minutes to 24 hours) or after 21 days in C57BL/6 mice. Results We developed a methodology that allows obtaining QDs-GSH-labeled B16F10 cells (nearly 100% viable labeled cells), which remained viable for at least 5 days and migrated similarly to control cells. However, proliferation, invasion, and the capacity to form metastatic nodules in the lungs were severely attenuated. Fluorescence imaging revealed that distribution/accumulation of QDs-GSH-labeled B16F10 cells could be tracked following injection into C57BL/6 mice (syngeneic preclinical metastasis model) and that these cells preferentially accumulated in the perialveolar area in lungs as early as 5 minutes post-injection. Conclusion The methodology described here represents a useful alternative for monitoring initial events during tumor cell metastasis.