Discovery of peptides for ligand-mediated delivery of mRNA lipid nanoparticles to cystic fibrosis lung epithelia.

For cystic fibrosis patients, a lung-targeted gene therapy would significantly alleviate pulmonary complications associated with morbidity and mortality. However, mucus in the airways and cell entry pose huge delivery barriers for local gene therapy. Here, we used phage display technology to select for and identify mucus- and cell-penetrating peptides against primary human bronchial epithelial cells from cystic fibrosis patients cultured at the air-liquid interface.

Optimization of ionizable lipids for aerosolizable mRNA lipid nanoparticles.

Although mRNA lipid nanoparticles (LNPs) are highly effective as vaccines, their efficacy for pulmonary delivery has not yet fully been established. A major barrier to this therapeutic goal is their instability during aerosolization for local delivery. This imparts a shear force that degrades the mRNA cargo and therefore reduces cell transfection.

Using bugs as drugs: Administration of bacteria-related microbes to fight cancer.

Bioengineering of bacteria-related microbes has demonstrated a great potential in targeted cancer therapy. Presently, the major administration routes of bacteria-related microbes for cancer treatment include intravenous injection, intratumoral injection, intraperitoneal injection, and oral delivery. Routes of bacteria administration are critical since different delivery approaches might exert anticancer effects through diverse mechanisms.

Manufacturing Stable Bacteriophage Powders by Including Buffer System in Formulations and Using Thin Film Freeze-drying Technology.

Purpose: Bacteriophage (phage) therapy has re-gained attention lately given the ever-increasing prevalence of multi-drug resistance 'super-bugs'. To develop therapeutic phage into clinically usable drug products, the strategy of solidifying phage formulations has been implemented to diversify the dosage forms and to overcome the storage condition limitations for liquid phage formulations.

Method: In our work, we hypothesize and tested that an advanced technology, thin film freeze-drying (TFFD), can be used to produce phage containing dry powders without significantly losing phage viability.

Aerosolizable siRNA-encapsulated solid lipid nanoparticles prepared by thin-film freeze-drying for potential pulmonary delivery.

Lipid nanoparticles are increasingly used for drug and gene delivery, including the delivery of small interfering RNA (siRNA). Pulmonary delivery of drug molecules carried by lipid nanoparticles directly into the lung may improve the treatment of certain lung diseases. The present study was designed to test the feasibility of engineering aerosolizable dry powder of lipid nanoparticles by thin-film freeze-drying (TFFD).

Aerosolizable Lipid Nanoparticles for Pulmonary Delivery of mRNA through Design of Experiments.

Messenger RNA is a class of promising nucleic acid therapeutics to treat a variety of diseases, including genetic diseases. The development of a stable and efficacious mRNA pulmonary delivery system would enable high therapeutic concentrations locally in the lungs to improve efficacy and limit potential toxicities. In this study, we employed a Design of Experiments (DOE) strategy to screen a library of lipid nanoparticle compositions to identify formulations possessing high potency both before and after aerosolization.

Controlled loading of albumin-drug conjugates ex vivo for enhanced drug delivery and antitumor efficacy.

To harness the intrinsic transport properties of albumin yet improve the therapeutic index of current in situ albumin-binding prodrugs, we developed albumin-drug conjugates with a controlled loading that achieved better antitumor efficacy. Here, model drug monomethyl auristatin E (MMAE) was conjugated ex vivo to Cys34 of albumin via a cathepsin B-sensitive dipeptide linker to ensure that all drug would be bound specifically to albumin. The resulting albumin-drug conjugate with a drug to albumin ratio (DAR) of 1 (ALDC1) retained the native secondary structure of albumin compared to conjugate with a higher DAR of 3 (ALDC3).

Just how prevalent are peptide therapeutic products? A critical review.

How prevalent are peptide therapeutic products? How innovative are the formulations used to deliver peptides? This review provides a critical analysis of therapeutic peptide products and the formulations approved by the United States Food and Drug administration (FDA), the European Medicines Agency (EMA), and the Japanese Pharmaceuticals and Medical Devices Agency (PMDA). This review also provides an in-depth analysis of dosage forms and administration routes for delivering peptide therapeutics, including injectables, oral dosage forms, and other routes of administration. We discuss the function of excipients in parenteral formulations in detail, since most peptide therapeutics are parenterally administered.

Electrostatic driven transport enhances penetration of positively charged peptide surfaces through tumor extracellular matrix.

Drug carriers achieve poor and heterogeneous distribution within solid tumors due to limited transport through the tumor extracellular matrix (ECM). The tumor ECM forms a net negatively charged network that interacts with and hinders the transport of molecules in part due to electrostatic interactions. Traditionally, the surfaces of drug delivery systems are passivated to minimize these interactions, but the mechanism of how charge interactions impact transport and penetration within the tumor microenvironment (TME) is not well understood.

The Stabilizing Excipients in Dry State Therapeutic Phage Formulations.

Phage therapy has gained prominence due to the increasing pathogenicity of "super bugs" and the rise of their multidrug resistance to conventional antibiotics. Dry state formulation of therapeutic phage is attractive to improve their "druggability" by increasing their shelf life, improving their ease of handling, and ultimately retaining their long-term potency. The use and selection of excipients are critical to stabilize phage in solid formulations and protect their viability from stresses encountered during the solidification process and long-term storage prior to use.

Peptides as surface coatings of nanoparticles that penetrate human cystic fibrosis sputum and uniformly distribute in vivo following pulmonary delivery.

Therapeutic delivery of drug and gene delivery systems have to traverse multiple biological barriers to achieve efficacy. Mucosal administration, such as pulmonary delivery in cystic fibrosis (CF) disease, remains a significant challenge due to concentrated viscoelastic mucus, which prevents drugs and particles from penetrating the mucus barrier. To address this problem, we used combinatorial peptide-presenting phage libraries and next-generation sequencing (NGS) to identify hydrophilic, net-neutral charged peptide coatings that enable penetration through human CF mucus ex vivo with ~600-fold better penetration than control, improve uptake into lung epithelial cells compared to uncoated or PEGylated-nanoparticles, and exhibit enhanced uniform distribution and retention in the mouse lung airways.

Identification of peptide coatings that enhance diffusive transport of nanoparticles through the tumor microenvironment.

In solid tumors, increasing drug penetration promotes their regression and improves the therapeutic index of compounds. However, the heterogeneous extracellular matrix (ECM) acts as a steric and interaction barrier that hinders effective transport of therapeutics, including nanomedicines. Specifically, the interactions between the ECM and surface physicochemical properties of nanomedicines (e.

Intracellular nanoparticle delivery by oncogenic KRAS-mediated macropinocytosis.

Background: The family of oncogenes () are the most frequent mutations in cancers and regulate key signaling pathways that drive tumor progression. As a result, drug delivery targeting -driven tumors has been a long-standing challenge in cancer therapy. Mutant activates cancer cells to actively take up nutrients, including glucose, lipids, and albumin, via macropinocytosis to fulfill their energetic requirements to survive and proliferate.

Quantitative PCR of T7 Bacteriophage from Biopanning.

This protocol describes the use of quantitative PCR (qPCR) to enumerate T7 phages from phage selection experiments (i.e., "biopanning").

Mucus-penetrating phage-displayed peptides for improved transport across a mucus-like model.

The objective of this work is to use phage display libraries as a screening tool to identify peptides that facilitate transport across the mucus barrier. Mucus is a complex selective barrier to particles and molecules, limiting penetration to the epithelial surface of mucosal tissues. In mucus-associated diseases such as cystic fibrosis (CF), mucus has increased viscoelasticity and a higher concentration of covalent and non-covalent physical entanglements compared to healthy tissues, which greatly hinders permeability and transport of drugs and particles across the mucosae for therapeutic delivery.

Peptides as drug delivery vehicles across biological barriers.

Peptides are small biological molecules that are attractive in drug delivery and materials engineering for applications including therapeutics, molecular building blocks and cell-targeting ligands. Peptides are small but can possess complexity and functionality as larger proteins. Due to their intrinsic properties, peptides are able to overcome the physiological and transport barriers presented by diseases.

Manufacturing and ambient stability of shelf freeze dried bacteriophage powder formulations.

The severity of multidrug resistance to antibiotics has urged development of alternative treatment approaches, including bacteriophage therapy. Given the complexity of the bacteriophage structure, formulation and stability are primary concerns. Our present work optimized process and formulations of phage powder manufacturing and investigated the stability of lyophilized bacteriophage powders under ambient storage.

Quantification of M13 and T7 bacteriophages by TaqMan and SYBR green qPCR.

TaqMan and SYBR Green quantitative PCR (qPCR) methods were developed as DNA-based approaches to reproducibly enumerate M13 and T7 phages from phage display selection experiments individually and simultaneously. The genome copies of M13 and T7 phages were quantified by TaqMan or SYBR Green qPCR referenced against M13 and T7 DNA standard curves of known concentrations. TaqMan qPCR was capable of quantifying M13 and T7 phage DNA simultaneously with a detection range of 2.

Physicochemical properties of mucus and their impact on transmucosal drug delivery.

Mucus is a selective barrier to particles and molecules, preventing penetration to the epithelial surface of mucosal tissues. Significant advances in transmucosal drug delivery have recently been made and have emphasized that an understanding of the basic structure, viscoelastic properties, and interactions of mucus is of great value in the design of efficient drug delivery systems. Mucins, the primary non-aqueous component of mucus, are polymers carrying a complex and heterogeneous structure with domains that undergo a variety of molecular interactions, such as hydrophilic/hydrophobic, hydrogen bonds and electrostatic interactions.

Carbon nanotubes as in vivo bacterial probes.

With the rise in antibiotic-resistant infections, non-invasive sensing of infectious diseases is increasingly important. Optical imaging, although safer and simpler, is less developed than other modalities such as radioimaging, due to low availability of target-specific molecular probes. Here we report carbon nanotubes (SWNTs) as bacterial probes for fluorescence imaging of pathogenic infections.

Deep, noninvasive imaging and surgical guidance of submillimeter tumors using targeted M13-stabilized single-walled carbon nanotubes.

Highly sensitive detection of small, deep tumors for early diagnosis and surgical interventions remains a challenge for conventional imaging modalities. Second-window near-infrared light (NIR2, 950-1,400 nm) is promising for in vivo fluorescence imaging due to deep tissue penetration and low tissue autofluorescence. With their intrinsic fluorescence in the NIR2 regime and lack of photobleaching, single-walled carbon nanotubes (SWNTs) are potentially attractive contrast agents to detect tumors.

Refactored M13 bacteriophage as a platform for tumor cell imaging and drug delivery.

M13 bacteriophage is a well-characterized platform for peptide display. The utility of the M13 display platform is derived from the ability to encode phage protein fusions with display peptides at the genomic level. However, the genome of the phage is complicated by overlaps of key genetic elements.

M13 virus based detection of bacterial infections in living hosts.

We report a first method for using M13 bacteriophage as a multifunctional scaffold for optically imaging bacterial infections in vivo. We demonstrate that M13 virus conjugated with hundreds of dye molecules (M13-Dye) can target and distinguish pathogenic infections of F-pili expressing and F-negative strains of E. coli.

M13-templated magnetic nanoparticles for targeted in vivo imaging of prostate cancer.

Molecular imaging allows clinicians to visualize the progression of tumours and obtain relevant information for patient diagnosis and treatment. Owing to their intrinsic optical, electrical and magnetic properties, nanoparticles are promising contrast agents for imaging dynamic molecular and cellular processes such as protein-protein interactions, enzyme activity or gene expression. Until now, nanoparticles have been engineered with targeting ligands such as antibodies and peptides to improve tumour specificity and uptake.