Yaswanth Kuthati

Killing cancer cells by delivering a nanoreactor for inhibition of catalase and catalytically enhancing intracellular levels of ROS

Intracellular hydrogen peroxide levels have the potential to be exploited in cancer therapy. We have synthesized uniform and size-controlled copper-impregnated mesoporous silica nanoparticles (Cu–MSN) containing a catalase inhibitor, 3-amino-1,2,4-triazole (AT) for ROS-mediated apoptosis in cancer cells. Copper species in the framework and aminotriazole loading were confirmed by electron spin resonance (ESR) and nuclear magnetic resonance (NMR) spectroscopies, respectively. The metal–ligand binding between the copper and AT are sensitive to the endosomal environment (pH = 5.5) for the release of AT to inhibit cytosolic catalase activity. The subsequent enhanced level of intracellular hydrogen peroxide, after catalase inhibition, is transformed into toxic, reactive oxygen species (ROS) by the catalysis of Cu(II) on MSN. The intracellularly delivered Cu–MSN–AT exhibited significant activity against colon carcinoma (HT-29 cell line) substantiated by increased levels of ROS, where AT drove up the hydrogen peroxide concentration and also the level of free radicals through a Fenton-like reaction. The trigger of cell apoptosis was induced from ROS attack to lipid membranes in order for further radical propagation to cause lipid peroxidation and eventually, a decrease in the membrane fluidity and mitochondrial membrane potential. As a result, an increase in the membrane permeability caused the release of cytochrome c into the cytoplasm and further activation of apoptotic cascades to trigger DNA fragmentation. The design of the Cu–MSN–AT system takes advantage of a synergistic effect to inhibit antioxidant defenses and catalyze the activation of lethal ROS by the framework of copper ions to kill cancer cells which represents a novel chemotherapeutic strategy for ameliorating the toxic side effects from non-specific ROS generation in traditional chemotherapeutic agents.

Layered double hydroxide nanoparticles to enhance organ-specific targeting and the anti-proliferative effect of cisplatin

To evaluate the role of charge in the nanoparticle distribution we modified the external surface of layered double hydroxide nanoparticles with various organic groups bearing different charges and further a near-infrared (NIR) fluorescent dye (Cy5.5) is conjugated in the layered structure to assess the biodistribution. The functionalized nanocomposites performed as highly efficient contrast agents since Cy5.5 molecule stabilization inside the layered structure can safeguard them from metabolization in the physiological environments. The cell viability, lactate dehydrogenase and hemolytic assays showed no cytotoxicity with an exceptionally low release of both lactate dehydrogenase and hemoglobin from the treated cells. The in vivo biodistribution results disclosed a high accumulation of positive amino-layered double hydroxides (LDHs) in the lungs. In contrast, there is a rapid clearance of negatively charged carboxylate-LDHs from blood flow by liver uptake. Interestingly neutral LDH-PEG5000 showed enhanced blood circulation time, without high fluorescent accumulation in the major organs. In vitro cellular uptake studies from flow cytometry are relevant to the interactions between the nanoparticle surfaces and various cell types and the data are relevant to effects observed for in vivo biodistribution. To further demonstrate that surface functionalization on LDH nanoparticles can promote targeted drug release, we further immobilized hydroxo-substituted cisplatin (CP) on carboxylate-modified LDHs by coordination bonding. Due to the ideal cleaving property of the carboxylate group the coordinated CP can be efficiently released by the increase of acidic proton and Cl- concentration in the endosomal environment. Functionalized LDHs can be successfully employed as targeted drug delivery systems. When the LDH-CP complex accumulate primarily in the targeted organ, the high positive charge on the framework of LDHs cause susceptibility to rapid endocytosis, which facilitates sustained drug release with minimal systemic toxicity providing the apt treatment in the targeted organ.

Hierarchical coated metal hydroxide nanoconstructs as potential controlled release carriers of photosensitizer for skin melanoma

Inorganic nanostructured ensembles containing an anionic clay matrix with layered double hydroxide (LDH) were designed in nanooncology for photosensitizer delivery. A core–shell strategy was combined with host–guest chemistry by the intercalation of indole-3-acetic acid (IAA). This single formulation, with good drug loading percentage and a compact liposome coating (LDH–IAA–Lipo) over and around the positively charged layered surface establishes a controlled release property for the treatment of skin melanoma. All the data regarding synthesis, physical characterization, chemical stability by thermogravimetric analysis and coat stability by leaching test in solvent mixture containing triton X-100 and biological buffer was obtained. IAA was estimated using high-performance liquid chromatography (HPLC) under optimized conditions with an admirable outcome. An improvement in cytotoxic properties under visible light exposure has been confirmed by MTT, ROS levels, DNA fragmentation using comet assay and apoptosis by analysis of mitochondrial membrane potential (MMP) using the MitoProbe JC-1 assay. In contrast, this formulation depicted cytocompatibility in a normal fibroblast (3T3) cell line. Photodynamic therapy (PDT) can be suggestible for long term therapy since the combinatorial efficiency of drug molecules in addition to light irradiation was dramatically evidenced to treat melanoma effectively.

pH-Triggered Controllable Release of Silver–Indole-3 Acetic Acid Complexes from Mesoporous Silica Nanoparticles (IBN-4) for Effectively Killing Malignant Bacteria

An efficient approach for the antimicrobial agent delivery specifically at acidic pH has been proposed. At the outset, functionalized mesoporous nanoparticles (NPs) were examined to verify the success of synthesis while considering the structural properties by various characterizations. The NPs were immobilized with silver-indole-3 acetic acid hydrazide (IAAH-Ag) complexes via a pH-sensitive hydrazone bond, which functioned as a model drug. When the transitional metal complexes with IBN-4-IAAH-Ag were exposed to acidic pH (near pH 5.0), the silver ions were preferentially released (70%) in a controlled manner up to 12 h by pH-sensitive denial of hydrazone bonds. In contrary, a low drug release (about 25%) was seen in physiological buffer (pH 7.4) demonstrating the pH sensitive release of this drug. Furthermore, the antibacterial efficacy of this unique structured sample was tested against the planktonic cells and biofilms of Gram-positive and Gram-negative bacteria with field emission scanning electron microscope in turn measuring the growth curves, formation of lethal reactive oxygen species, protein leakage, and DNA damage. The synthesized pH-sensitive IAAH-Ag complex was found to have high antimicrobial efficacy against multidrug resistant clinical isolates both in planktonic and biofilm states. Going forward, the synthesized nanoconjugates proved a good in vivo efficacy in treating the bacterial infection of mice. These new metal complex-conjugated NPs through a pH-sensitive hydrazone bond opened up a new avenue for the design and synthesis of the next generation antibacterial agents, which would act as an alternative to antibiotics.

Utilization of Enzyme-Immobilized Mesoporous Silica Nanocontainers (IBN-4) in Prodrug-Activated Cancer Theranostics

To develop a carrier for use in enzyme prodrug therapy, Horseradish peroxidase (HRP) was immobilized onto mesoporous silica nanoparticles (IBN-4: Institute of Bioengineering and Nanotechnology), where the nanoparticle surfaces were functionalized with 3-aminopropyltrimethoxysilane and further conjugated with glutaraldehyde. Consequently, the enzymes could be stabilized in nanochannels through the formation of covalent imine bonds. This strategy was used to protect HRP from immune exclusion, degradation and denaturation under biological conditions. Furthermore, immobilization of HRP in the nanochannels of IBN-4 nanomaterials exhibited good functional stability upon repetitive use and long-term storage (60 days) at 4 °C. The generation of functionalized and HRP-immobilized nanomaterials was further verified using various characterization techniques. The possibility of using HRP-encapsulated IBN-4 materials in prodrug cancer therapy was also demonstrated by evaluating their ability to convert a prodrug (indole-3-acetic acid (IAA)) into cytotoxic radicals, which triggered tumor cell apoptosis in human colon carcinoma (HT-29 cell line) cells. A lactate dehydrogenase (LDH) assay revealed that cells could be exposed to the IBN-4 nanocomposites without damaging their membranes, confirming apoptotic cell death. In summary, we demonstrated the potential of utilizing large porous mesoporous silica nanomaterials (IBN-4) as enzyme carriers for prodrug therapy.

Multi-laminated metal hydroxide nanocontainers for oral-specific delivery for bioavailability improvement and treatment of inflammatory paw edema in mice

Multiple layers of pH-sensitive enteric copolymers were coated over layered double hydroxide (LDH) nanoparticles for controllable drug release and improved solubility of hydrophobic drugs. The nano-sized LDH carriers significantly improved the accessibility of sulfasalazine molecules that have positively charged frameworks. In addition, the successful encapsulation of negatively charged enteric copolymers was achieved via electrostatic attractions. The multi-layered enteric polymer coating could potentially protect nanoparticle dissolution at gastric pH and accelerate the dissolution velocity, which would improve the drug bioavailability in the colon. Next, biological studies of this formulation indicated a highly protective effect from the scavenging of superoxide free radicals and diethyl maleate (DEM) induced lipid peroxidation, which are major cell signalling pathways for inflammation. The histological view of the liver and kidney sections revealed that the nanoformulation is safe and highly biocompatible. The animal studies conducted via paw inflammation induced by complete Freunds adjuvant (CFA) revealed that enteric-coated LDH-sulfasalazine nanoparticles provided a sustained release that maintained the sulfasalazine concentrations in a therapeutic window. Therefore, this nanoformulation exhibited preferential efficacy in reducing the CFA-induced inflammation especially at day 4.

Overcoming multidrug resistance through co-delivery of ROS-generating nano-machinery in cancer therapeutics

The use of nanotechnology to overcome multidrug resistance (MDR) in cancer cells has been predominant. Herein, we report the conjugation of copper(ii)-doxorubicin complexes on the surfaces of layered double hydroxide nanoparticles (LDHs) along with ascorbic acid intercalation in the gallery space to demonstrate synergistic effects to conquer MDR. The pH-sensitive release of doxorubicin (Dox) and the sustained release of ascorbic acid (AA) generate high amounts of hydrogen peroxide intracellularly that concomitantly results in conversion to cytotoxic free radicals through a copper(ii)-catalyzed Fenton-like reaction. Therefore, the combination of the chemotherapeutic agent (Dox) and free radical attack can devastate the MDR for effective cancer treatment through the co-delivery system.

Synthesis and Characterization of Chitosan-Coated Near-Infrared (NIR) Layered Double Hydroxide-Indocyanine Green Nanocomposites for Potential Applications in Photodynamic Therapy.

We designed a study for photodynamic therapy (PDT) using chitosan coated Mg–Al layered double hydroxide (LDH) nanoparticles as the delivery system. A Food and Drug Administration (FDA) approved near-infrared (NIR) fluorescent dye, indocyanine green (ICG) with photoactive properties was intercalated into amine modified LDH interlayers by ion-exchange. The efficient positively charged polymer (chitosan (CS)) coating was achieved by the cross linkage using surface amine groups modified on the LDH nanoparticle surface with glutaraldehyde as a spacer. The unique hybridization of organic-inorganic nanocomposites rendered more effective and successful photodynamic therapy due to the photosensitizer stabilization in the interlayer of LDH, which prevents the leaching and metabolization of the photosensitizer in the physiological conditions. The results indicated that the polymer coating and the number of polymer coats have a significant impact on the photo-toxicity of the nano-composites. The double layer chitosan coated LDH–NH2–ICG nanoparticles exhibited enhanced photo therapeutic effect compared with uncoated LDH–NH2–ICG and single layer chitosan-coated LDH–NH2–ICG due to the enhanced protection to photosensitizers against photo and thermal degradations. This new class of organic-inorganic hybrid nanocomposites can potentially serve as a platform for future non-invasive cancer diagnosis and therapy.

Phototherapeutic spectrum expansion through synergistic effect of mesoporous silica trio-nanohybrids against antibiotic-resistant gram-negative bacterium

The extensive impact of antibiotic resistance has led to the exploration of new anti-bacterial modalities. We designed copper impregnated mesoporous silica nanoparticles (Cu-MSN) with immobilizing silver nanoparticles (SNPs) to apply photodynamic inactivation (PDI) of antibiotic-resistant E. coli. SNPs were decorated over the Cu-MSN surfaces by coordination of silver ions on diamine-functionalized Cu-MSN and further reduced to silver nanoparticles with formalin. We demonstrate that silver is capable of sensitizing the gram-negative bacteria E. coli to a gram-positive specific phototherapeutic agent in vitro; thereby expanding curcumins phototherapeutic spectrum. The mesoporous structure of Cu-MSN remains intact after the exterior decoration with silver nanoparticles and subsequent curcumin loading through an enhanced effect from copper metal-curcumin affinity interaction. The synthesis, as well as successful assembly of the functional nanomaterials, was confirmed by various physical characterization techniques. Curcumin is capable of producing high amounts of reactive oxygen species (ROS) under light irradiation, which can further improve the silver ion release kinetics for antibacterial activity. In addition, the positive charged modified surfaces of Cu-MSN facilitate antimicrobial response through electrostatic attractions towards negatively charged bacterial cell membranes. The antibacterial action of the synthesized nanocomposites can be activated through a synergistic mechanism of energy transfer of the absorbed light from SNP to curcumin.

Encapsulation of palladium porphyrin photosensitizer in layered metal oxide nanoparticles for photodynamic therapy against skin melanoma

Abstract We designed a biodegradable nanocarrier of layered double hydroxide (LDH) for photodynamic therapy (PDT) based on the intercalation of a palladium porphyrin photosensitizer (PdTCPP) in the gallery of LDH for melanoma theragnosis. Physical and chemical characterizations have demonstrated the photosensitizer was stable in the layered structures. In addition, the synthesized nanocomposites rendered extremely efficacious therapy in the B16F10 melanoma cell line by improving the solubility of the hydrophobic PdTCPP photosensitizer. The detection of singlet oxygen generation under irradiation at the excitation wavelength of a 532 nm laser was indeed impressive. Furthermore, the in vivo results using a tumour xenograft model in mice indicated the apparent absence of body weight loss and relative organ weight variation to the liver and kidney demonstrated that the nanocomposites were biosafe with a significant reduction in tumour volume for the anti-cancer efficacy of PDT. This drug delivery system using the nanoparticle–photosensitizer hybrid has great potential in melanoma theragnosis.