Prevention of Colitis-Associated Cancer via Oral Administration of M13-Loaded Lipid Nanoparticles.

Inflammatory bowel disease (IBD), which includes ulcerative colitis (UC) and Crohn's disease, is known to increase the risk of colitis-associated cancer (CAC). CAC has been found to be unresponsive to standard chemotherapy regimens, and the current treatments do not utilize effective small-molecule drugs and colon-targeted delivery systems. Previous studies indicated that the M13-nano-liposome (NL) formulation can effectively target the colon and reshape the gut microbiota in ex vivo cultures, generating altered microbial metabolites that can efficiently prevent chronic UC. In this study, we tested the cancer cell uptake ability of the NL formulation and investigated the potential of the M13-NL formulation to prevent CAC in the azoxymethane (AOM)-exposed IL10-/- mouse model. Our findings demonstrate that oral administration of M13-NL prevents tumor development in AOM-exposed IL10-/- mice, suggesting that M13-NL is a promising oral drug formulation for preventing CAC.

Oral administration of M13-loaded nanoliposomes is safe and effective to treat colitis-associated cancer in mice.

OBJECTIVE: Colitis-associated cancer (CAC) treatment lacks effective small-molecule drugs and efficient targeted delivery systems. Here, we loaded M13 (an anti-cancer drug candidate) to colon-targeting ginger-derived nanoliposomes (NL) and investigated if orally administered M13-NL could enhance the anticancer effects of M13 in CAC mouse models. METHODS: The biopharmaceutical properties of M13 were assessed by physicochemical characterizations. The in vitro immunotoxicity of M13 was assessed against PBMCs using FACS and the mutagenic potential of M13 was evaluated by the Ames assay. The in vitro efficacy of M13 was tested in 2D- and 3D-cultured cancerous intestinal cells. AOM/DSS-induced CAC mice were used to evaluate the therapeutic effects of free M13 or M13-NL on CAC in vivo. RESULTS: M13 has beneficial physiochemical properties, including high stability, and no apparent immunotoxicity or mutagenic potential in vitro. M13 is effective against the growth of 2D- and 3D-cultured cancerous intestinal cells in vitro. The in vivo safety and efficacy of M13 were significantly improved by using NL for drug delivery (p < 0.001). Oral administration of M13-NL exhibited excellent therapeutic effects in AOM/DSS-induced CAC mice. CONCLUSION: M13-NL is a promising oral drug formulation for CAC treatment.

Preparation and Characterization of IL-22 mRNA-loaded Lipid Nanoparticles.

Interleukin-22 (IL-22) has been demonstrated as a critical regulator of epithelial homeostasis and repair; it showed an anti-inflammatory effect against ulcerative colitis. Local microinjection of IL-22 cDNA vector has been shown to be effective in treating ulcerative colitis in mouse models. However, microinjection comes with multiple technical challenges for routine colon-targeted drug delivery. In contrast, oral administration can get around these challenges and provide comparable efficacy. We showed in previous studies that oral administration of new lipid nanoparticles (nLNP)-encapsulated IL-22 mRNA targets the colon region and efficiently ameliorates colitis. This protocol describes the details of preparing and characterizing the nLNP-encapsulated IL-22 mRNA using three major lipids that mimic the natural ginger-derived nanoparticles. It provides an nLNP platform that can be used to orally deliver other types of nucleic acids to the colon.

Oral delivery of IL-22 mRNA-loaded lipid nanoparticles targeting the injured intestinal mucosa: A novel therapeutic solution to treat ulcerative colitis.

Oral mRNA delivery is a promising yet understudied approach for treating inflammatory bowel disease (IBD). Inspired by the colon-targeting ability of ginger-derived nanoparticles (GDNPs), we reversely engineered lipid nanoparticles that comprise the three major lipids identified in GDNPs. When mixed at the ratio found in GDNPs, the selected lipids (phosphatidic acid, monogalactosyldiacylglycerol, and digalactosyldiacylglycerol; 5:2:3) self-assembled into new lipid nanoparticles (nLNPs) in phosphate-buffered saline. We encapsulated IL-22-mRNA within the nLNPs, as enhanced IL-22 expression in the colon is known to have potent anti-inflammatory efficacy against ulcerative colitis (UC). The IL-22 mRNA-loaded nLNPs (IL-22/nLNPs) were found to be about 200 nm in diameter and have a zeta potential of -18 mV. Oral delivery of IL-22/nLNPs elevated the protein expression level of IL-22 in the colonic mucosa of mice. In a mouse model of acute colitis, mice fed with IL-22/nLNPs experienced an accelerated healing process, as indicated by the recovery of more body weight and colon length as well as reduction of the histological index, colonic MPO activity, fecal lipocalin concentration, and mRNA expression levels of pro-inflammatory cytokines (TNF-α, IL-6, and IL-1ß). Our results suggest that our reversely engineered nLNPs is an excellent mRNA delivery platform for treating ulcerative colitis.

The Current Status of Molecular Biomarkers for Inflammatory Bowel Disease.

Diagnosis and prognosis of inflammatory bowel disease (IBD)-a chronic inflammation that affects the gastrointestinal tract of patients-are challenging, as most clinical symptoms are not specific to IBD, and are often seen in other inflammatory diseases, such as intestinal infections, drug-induced colitis, and monogenic diseases. To date, there is no gold-standard test for monitoring IBD. Endoscopy and imaging are essential diagnostic tools that provide information about the disease's state, location, and severity. However, the invasive nature and high cost of endoscopy make it unsuitable for frequent monitoring of disease activity in IBD patients, and even when it is possible to replace endoscopy with imaging, high cost remains a concern. Laboratory testing of blood or feces has the advantage of being non-invasive, rapid, cost-effective, and standardizable. Although the specificity and accuracy of laboratory testing alone need to be improved, it is increasingly used to monitor disease activity or to diagnose suspected IBD cases in combination with endoscopy and/or imaging. The literature survey indicates a dearth of summarization of biomarkers for IBD testing. This review introduces currently available non-invasive biomarkers of clinical importance in laboratory testing for IBD, and discusses the trends and challenges in the IBD biomarker studies.

Prevention of Ulcerative Colitis by Autologous Metabolite Transfer from Colitogenic Microbiota Treated with Lipid Nanoparticles Encapsulating an Anti-Inflammatory Drug Candidate.

Modulating the gut microbiota composition is a potent approach to treat various chronic diseases, including obesity, metabolic syndrome, and ulcerative colitis (UC). However, the current methods, such as fecal microbiota transplantation, carry a risk of serious infections due to the transmission of multi-drug-resistant organisms. Here, we developed an organism-free strategy in which the gut microbiota is modulated ex vivo and microbiota-secreted metabolites are transferred back to the host. Using feces collected from the interleukin-10 (IL-10) knockout mouse model of chronic UC, we found that a drug candidate (M13)-loaded natural-lipid nanoparticle (M13/nLNP) modified the composition of the ex vivo-cultured inflamed gut microbiota and its secreted metabolites. Principal coordinate analysis (PCoA) showed that M13/nLNP shifted the inflamed microbiota composition toward the non-inflamed direction. This compositional modification induced significant changes in the chemical profiles of secreted metabolites, which proved to be anti-inflammatory against in vitro-cultured NF-κß reporter cells. Further, when these metabolites were orally administered to mice, they established strong protection against the formation of chronic inflammation. Our study demonstrates that ex vivo modulation of microbiota using M13/nLNP effectively reshaped the microbial secreted metabolites and that oral transfer of these metabolites might be an effective and safe therapeutic approach for preventing chronic UC.

Preparation, Characterization, and Cell Uptake of PLGA/PLA-PEG-FA Nanoparticles.

Oral administration of colon-targeting nanoformulations holds many advantages over the systemic delivery of free drugs, or traditional nontargeting formulations in the treatment of ulcerative colitis (UC). Currently, the most conventional method for constructing colon-targeting drug delivery systems (DDS) is by integrating the biocompatible materials poly(lactic-co-glycolic acid) (PLGA) and polylactic acid (PLA) into a copolymer. This PLGA/PLA-polyethylene glycol-folic acid (PEG-FA) copolymeric nanoformulation effectively delivers the drugs for uptake by various human colon cancer cells (e.g., HT-29 and HCT-116) and mouse colon cancer cells (CT-26). There is, however, a distinct lack of comprehensive protocols for the construction of such copolymer. This protocol details an easy-to-follow single-step method for the construction of a colon-targeting PLGA/PLA-PEG-FA nanoformulation, which encapsulates a fluorescent dye and demonstrates the visualization of its cell uptake in vitro.

Orally Administered Natural Lipid Nanoparticle-Loaded 6-Shogaol Shapes the Anti-Inflammatory Microbiota and Metabolome.

The past decade has seen increasing interest in microbiota-targeting therapeutic strategies that aim to modulate the gut microbiota's composition and/or function to treat chronic diseases, such as inflammatory bowel disease (IBD), metabolic symptoms, and obesity. While targeting the gut microbiota is an innovative means for treating IBD, it typically requires an extended treatment time, hampering its potential application. Herein, using an established natural-lipid nanoparticle (nLNP) platform, we demonstrate that nLNPs encapsulated with the drug candidate 6-shogaol (6S/nLNP) distinctly altered microbiota composition within one day of treatment, significantly accelerating a process that usually requires five days using free 6-shogaol (6S). In addition, the change in the composition of the microbiota induced by five-day treatment with 6S/nLNP was maintained for at least 15 days (from day five to day 20). The consequent alteration in the fecal metabolic profile stemming from this compositional change manifested as functional changes that enhanced the in vitro anti-inflammatory and wound-healing efficacy of macrophage cells (Raw 264.7) and epithelial cells (Caco-2 BBE1), respectively. Further, this metabolic compositional change, as reflected in an altered metabolic profile, promoted a robust anti-inflammatory effect in a DSS-induced mouse model of acute colitis. Our study demonstrates that, by near-instantly modulating microbiota composition and function, an nLNP-based drug-delivery platform might be a powerful tool for treating ulcerative colitis.

Characterization of the polarity of subcritical water.

The polarity of subcritical water was studied solvatochromically with betaine dye (33) across a temperature range of 30°C-180°C and a pressure range of 13.8bar (200psi) to 124bar (1800psi). It was observed that temperature has a greater effect than pressure on the polarity of subcritical water. In addition, subcritical water was compared with traditional hydro-organic mobile phases and the polarity of subcritical water was found to be comparable to a range of 30%-50% methanol/water and a range of 20%-30% acetonitrile/water mobile phases. It was concluded that subcritical water is more suited to separations involving slightly polar and polar compounds than more nonpolar analytes.