Table of Content
Precision Nanomedicine, 2018 July; Vol. 1, Issue 2
About the cover: Fluorescence lifetime imaging microscopy (FLIM) and single-molecule super-resolution microscopy (SRM) illustrate the intracellular fate of Sn-2 phosphatidylcholine prodrugs.
Featured Research Article
Cellular Trafficking of Sn-2 Phosphatidylcholine Prodrugs Studied with Fluorescence Lifetime Imaging and Super-resolution Microscopy
Dolonchampa Maji PhD, Jin Lu PhD, Pinaki Sarder Phd, Anne H. Schmieder MS, Grace Cui MS, Xiaoxia Yang BSe, Dipanjan Pan PhDf, Matthew D. Lew PhDc, Samuel Achilefu PhDa,b and Gregory M. Lanza MD PhD*
aOptical Radiology Lab, Department of Radiology, Washington University School of Medicine, St. Louis, MO 63110, USA
bDepartment of Biomedical Engineering, Washington University in St. Louis, MO 63130, USA
cDepartment of Electrical and Systems Engineering, Washington University in St. Louis, St. Louis, MO 63130, USA
dDepartment of Pathology and Anatomical Sciences, Jacobs School of Medicine & Biomedical Sciences, University of Buffalo, Buffalo, NY 14203
eDivision of Cardiology, Department of Medicine, Washington University School of Medicine, St. Louis, MO, USA
fDepartment of Bioengineering, University of Illinois at Urbana-Champaign, Champaign, IL, USA
Submitted: July 18, 2018 Accepted: July 30, 2018
While the in vivo efficacy of Sn-2 phosphatidylcholine prodrugs incorporated into targeted, non-pegylated lipid-encapsulated nanoparticles was demonstrated in prior preclinical studies, the microscopic details of cell prodrug internalization and trafficking events are unknown. Classic fluorescence microscopy, fluorescence lifetime imaging microscopy, and single-molecule super-resolution microscopy were used to investigate the cellular handling of doxorubicin-prodrug and AlexaFluor-488-prodrug. Sn-2 phosphatidylcholine prodrugs delivered by hemifusion of nanoparticle and cell phospholipid membranes functioned as phosphatidylcholine mimics, circumventing the challenges of endosome sequestration and release. Phosphatidylcholine prodrugs in the outer cell membrane leaflet translocated to the inner membrane leaflet by ATP-dependent and ATP-independent mechanisms and distributed broadly within the cytosolic membranes over the next 12 h. A portion of the phosphatidylcholine prodrug populated vesicle membranes trafficked to the perinuclear Golgi/ER region, where the drug was enzymatically liberated and activated. Native doxorubicin entered the cells, passed rapidly to the nucleus, and bound to dsDNA, whereas DOX was first enzymatically liberated from DOX-prodrug within the cytosol, particularly in the perinuclear region, before binding nuclear dsDNA. Much of DOX-prodrug was initially retained within intracellular membranes. In vitro anti-proliferation effectiveness of the two drug delivery approaches was equivalent at 48 h, suggesting that residual intracellular DOX-prodrug may constitute a slow-release drug reservoir that enhances effectiveness. We have demonstrated that Sn-2 phosphatidylcholine prodrugs function as phosphatidylcholine mimics following reported pathways of phosphatidylcholine distribution and metabolism. Drug complexed to the Sn-2 fatty acid is enzymatically liberated and reactivated over many hours, which may enhance efficacy over time.
From the Clinical Editor: Sn-2 phosphatidylcholine prodrugs have been shown to be effective systemically for the delivery of a variety of drugs. Nonetheless, little is known about the pharmacokinetics in terms of cell uptake release and activation. Here, the authors have carried out experiments utilizing fluorescence microscopy, fluorescence lifetime imaging, and single-molecule super-resolution imaging techniques and show that the series of events that occur after the prodrugs had contacted with cellular membranes. The data has given valuable new understanding of prodrug pharmacokinetics.
Keng Wooi Ng1*, S. Moein Moghimi1,2*
1School of Pharmacy, Faculty of Medical Sciences, Newcastle University, King George VI Building, Newcastle upon Tyne NE1 7RU, United Kingdom
2Institute of Cellular Medicine, Newcastle University, Framlington Place, Newcastle upon Tyne NE2 4HH, United Kingdom
Submitted: July 9, 2018 Accepted: July 26, 2018
Wearable skin biosensors have important applications in health monitoring, medical treatment and theranostics. There has been a rapid growth in the development of novel biosensing and bioanalytical techniques in recent years, much of it underpinned by recent advancements in nanotechnology. As the two related disciplines continue to co-evolve, we take a timely look at some notable developments in skin biosensing/bioanalysis, scan the horizon for emerging nanotechnologies, and discuss how they may influence the future of biosensing/bioanalysis in the skin.
Rongxin Li1†, Deepanjali Dattatray Gurav2†, Jingjing Wan1*, and Kun Qian2*
1School of Chemistry and Molecular Engineering, East China Normal University, Shanghai, 200062, P. R. China
2School of Biomedical Engineering, Med-X Research Institute, Shanghai Jiao Tong University, Shanghai, 200030, P. R. China
Submitted: July 24, 2018; Accepted: July 31, 2018
Precision diagnostics relies on omics analysis by mass spectrometry to overcome the limitation in accuracy by an individual biomarker, due to the complex nature of diseases. Recent development in nanotechnology markedly enhanced sample treatment and detection efficiency of this method. Herein, we foresee a coming era of precision diagnostics based on nano-assisted mass spectrometry. Some important progress in the field includes detection of (1) nucleic acids for genetic analysis; (2) proteins/peptides for proteomic analysis; and (3) small molecules for metabolic analysis. We anticipate that this review will be a reminder for both young and experienced researchers about the future of diagnostics and call for attention worldwide.
Diana Rafael1,2*, Fernanda Andrade2,3*, Sara Montero2, Petra Gener2,3, Joaquin Seras-Franzoso2, Francesc Martínez2, Patricia González2,3, Helena Florindo1, Diego Arango4, Joan Sayós5, Ibane Abasolo2,3,6, *Mafalda Videira1, and *Simó Schwartz Jr.2,3,
1Research Institute for Medicines and Pharmaceutical Sciences, Faculdade de Farmácia, Universidade de Lisboa (iMed.ULisboa), Lisbon, Portugal
2Drug Delivery and Targeting Group, Molecular Biology and Biochemistry Research Centre for Nanomedicine (CIBBIM-Nanomedicine), Vall d’Hebron Institut de Recerca, Universitat Autònoma de Barcelona, Barcelona, Spain
3Networking Research Centre for Bioengineering, Biomaterials, and Nanomedicine (CIBER-BBN), Instituto de Salud Carlos III, Zaragoza, Spain
4Biomedical Research in Digestive Tract Tumors, CIBBIM-Nanomedicine, Vall d’Hebron Institut de Recerca, Universitat Autònoma de Barcelona, Barcelona, Spain
5Immune Regulation and Immunotherapy, CIBBIM-Nanomedicine, Vall d’Hebron Institut de Recerca, Universitat Autònoma de Barcelona, Barcelona, Spain
6Functional Validation & Preclinical Research (FVPR), CIBBIM-Nanomedicine, Vall d’Hebron Institut de Recerca, Universitat Autònoma de Barcelona, Barcelona, Spain
Submitted: June 22, 2018 Accepted: June 29, 2018
The search for an ideal gene delivery system is a long and laborious process in which several factors from the first idea to final formulation, including main challenges, peaks and troughs, should be deeply taken into consideration to ensure adequate biological safety and in vivo efficacy endpoints. Arachidonate 5-lipoxygenase (ALOX5), a crucial player related with cancer development and in particular with cancer stem cells malignancy. In this work we describe the process behind the development of a small interfering RNA (siRNA) delivery system to inhibit ALOX5 in cancer stem cells (CSC), as a model target gene. We started by screening chitosan polyplexes, among different types of chitosan in different complexation conditions. Due to the low silencing efficacy obtained, chitosan polyplexes were combined with Pluronic®-based polymeric micelles with recognized advantages regarding gene transfection. We tested different types of polymeric particles to improve the biological efficacy of chitosan polyplexes. Nevertheless, limited transfection efficiency was still detected. The well-established polyethylenimine (PEI) cationic polymer was used in substitution of chitosan, in combination with polymeric micelles, originating PEI-siRNA-Pluronic® systems. The presence of Pluronic® F127 in the formulation showed to be of utmost importance, because not only the silencing activity of the polyplexes was improved, but also PEI-associated toxicity was clearly reduced. This allowed to increase the amount of PEI inside the system and its overall efficacy. Indeed, different types of PEI, N/P ratios and preparation methods were tested until an optimal formulation composed by PEI 10k branched-based polyplexes at an N/P ratio of 50 combined with micelles of Pluronic® F127 was selected. This combined micelle presented adequate technological properties, safety profile and biological efficacy, resulting in high ALOX5 gene silencing and strong reduction of invasion and transformation capabilities of a stem cell subpopulation isolated from MDA-MB-231 triple negative breast cancer cells.
From the Clinical Editor: The use of gene-silencing technique for cancer therapy has been met with problems like cellular uptake, degradation, and clearance. The authors here test two different types of polymeric particle formulations as vectors in an attempt to improve the biological efficacy and describe a general algorithm for testing. The data obtained provide more knowledge of how to further progress the field for the benefits of selected cancer patients.
Retinal Multipotent Stem-Cell Derived “MiEye” Spheroid 3D Culture Model for Preclinical Screening of Non-viral Gene Delivery Systems
Ding-Wen Chen and Marianna Foldvari*
School of Pharmacy, University of Waterloo, 10A Victoria St. S., Kitchener, ON N2G 1C5, Canada.
Submitted: July 1, 2018 Accepted: July 11, 2018