A liposomal radionanoformulation for targeted drug delivery and real time monitoring by radionuclide imaging for HER2 overexpressing cancers.

Liposomal formulations carrying chemotherapeutic drugs have demonstrated great potential as effective drug delivery systems. Smart nanoformulations decorated with targeting agents and probes are desired for site specific delivery of drugs and real time monitoring. In this study, we aimed to develop liposomal formulation loaded with doxorubicin and tagged with trastuzumab antibody (Ab) for targeting human epidermal growth factor receptor 2 (HER2) positive tumors. Liposomes were prepared by ethanol injection method using modified lipids to conjugate trastuzumab and radiolabel with Tc-99m radioisotope using DTPA for imaging by single photon emission computed tomography (SPECT). Doxorubicin was loaded using the active pH gradient method. The conjugation of Ab to liposomes was validated by SDS-PAGE and MALDI-MS. 99m Tc labeled liposomes encapsulating doxorubicin conjugated with antibody (99m Tc-Lip-Ab-Dox) and 99m Tc labeled liposomes encapsulating doxorubicin (99m Tc-Lip-Dox) were found to be stable in blood plasma and saline using chromatography method. The specificity of 99m Tc-Lip-Ab-Dox against HER2 receptor was evident from cell uptake and inhibition studies. Results also corroborated with confocal microscopy studies. In vivo studies in tumor bearing severe combined immunodeficient mice by SPECT imaging and biodistribution studies revealed higher uptake of 99m Tc-Lip-Ab-Dox in tumor and less accumulation in the liver compared to 99m Tc-Lip-Dox. In conclusion, liposomal nanoformulation for immunotargeting and monitoring of drug delivery was successfully formulated and evaluated. Encouraging results in preclinical studies were obtained with the radioformulation. Such smart radioformulations will not only serve the purpose of site-specific controlled release of drugs at the target site but also aid in optimizing the drug doses and schedule of cancer treatment by monitoring pharmacokinetics.

Effective Topical Delivery of H-AgNPs for Eradication of Klebsiella pneumoniae-Induced Burn Wound Infection.

The aim of the present study was to explore the therapeutic efficacy of microemulsion-based delivery of histidine-capped silver nanoparticles in eradicating Klebsiella pneumoniae-induced burn wound infection. The developed microemulsion was characterized on the basis of differential light scattering, phase separation, refractive index, and specific conductance. Emulgel was prepared and characterized on the basis of thixotropy, texture, differential scanning calorimetry, and release kinetics. Emulgel was further evaluated in skin irritation and in vivo studies, namely full-thickness K. pneumoniae-induced burn wound infection treatment via topical route. Efficacy of treatment was evaluated in terms of bacterial load, histopathology, wound contraction, and other infection markers. The developed emulgel provided significant in vivo antibacterial activity of histidine-capped silver nanoparticle preparations via topical route and resulted in reduction in bacterial load, wound contraction, and enhanced skin healing as well as decrement of inflammatory markers such as malondialdehyde, myeloperoxidase, and reactive nitrogen intermediate compared to untreated animals. The present study encourages the further employment of histidine-capped silver nanoparticles along with microemulsion-based drug delivery system in combating antibiotic-resistant topical infections.

Controlling the transverse proton relaxivity of magnetic graphene oxide.

The engineering of materials with controlled magnetic properties by means other than a magnetic field is of great interest in nanotechnology. In this study, we report engineered magnetic graphene oxide (MGO) in the nanocomposite form of iron oxide nanoparticles (IO)-graphene oxide (GO) with tunable core magnetism and magnetic resonance transverse relaxivity (r2). These tunable properties are obtained by varying the IO content on GO. The MGO series exhibits r2 values analogous to those observed in conventional single core and cluster forms of IO in different size regimes-motional averaging regime (MAR), static dephasing regime (SDR), and echo-limiting regime (ELR) or slow motion regime (SMR). The maximum r2 of 162 ± 5.703 mM-1s-1 is attained for MGO with 28 weight percent (wt%) content of IO on GO and hydrodynamic diameter of 414 nm, which is associated with the SDR. These findings demonstrate the clear potential of magnetic graphene oxide for magnetic resonance imaging (MRI) applications.

Inducing cell death in vitro in cancer cells by targeted delivery of cytochrome c via a transferrin conjugate.

One of the major drawbacks of many of the currently used cancer drugs are off-target effects. Targeted delivery is one method to minimize such unwanted and detrimental events. To actively target lung cancer cells, we have developed a conjugate of the apoptosis inducing protein cytochrome c with transferrin because the transferrin receptor is overexpressed by many rapidly dividing cancer cells. Cytochrome c and transferrin were cross-linked with a redox sensitive disulfide bond for the intra-cellular release of the protein upon endocytosis by the transferrin receptor. Confocal results demonstrated the cellular uptake of the cytochrome c-transferrin conjugate by transferrin receptor overexpressing A549 lung cancer cells. Localization studies further validated that this conjugate escaped the endosome. Additionally, an in vitro assay showed that the conjugate could induce apoptosis by activating caspase-3. The neo-conjugate not only maintained an IC50 value similar to the well known drug cisplatin (50 µM) in A549 cancer cells but also was nontoxic to the normal lung (MRC5) cells. Our neo-conjugate holds promise for future development to target cancers with enhanced transferrin receptor expression.

Purification and characterization of a cytochrome c with novel caspase-3 activation activity from the pathogenic fungus Rhizopus arrhizus.

BACKGROUND: Members of Rhizopus species are the most common cause of mucormycosis, a rare but often fatal fungal infection. Host induced pathogen apoptosis and pathogen induced host cell apoptosis are often involved in fungal infections. In many organisms, the release of mitochondrial cytochrome c can trigger apoptosis by activating caspase proteases, but the role of fungal cytochrome c in apoptosis remains unknown. RESULTS: DNA sequence encoding Rhizopus arrhizus cytochrome c was cloned and expressed in E. coli. Both native and recombinant cytochrome c were purified using ion exchange followed by gel filtration chromatography. The identities of purified proteins were confirmed by MALDI-MS and UV-Visible spectroscopy. For the first time, we demonstrated that Rhizopus arrhizus cytochrome c could activate human capspase-3 in HeLa cell extracts. We also found that Rhizopus arrhizus cytochrome c has redox potential, peroxidase activity, and spectral properties similar to human and horse cytochrome c proteins. CONCLUSIONS: Rhizopus arrhizus cytochrome c can activate human caspase-3 in HeLa cell extracts and it possesses similar physical and spectral properties as human and horse cytochrome c. This protein was found to have a previously unknown potential to activate human caspase-3, an important step in the apoptosis cascade.