Engineered bacteria to accelerate wound healing: an adaptive, randomised, double-blind, placebo-controlled, first-in-human phase 1 trial.

Background: Impaired wound healing is a growing medical problem and very few approved drugs with documented clinical efficacy are available. CXCL12-expressing lactic acid bacteria, Limosilactobacillus reuteri (ILP100-Topical), has been demonstrated to accelerate wound healing in controlled preclinical models. In this first-in-human study, the primary objective was to determine safety and tolerability of the drug candidate ILP100-Topical, while secondary objectives included assessments of clinical and biologic effects on wound healing by traditionally accepted methods and explorative and traceable assessments. Methods: SITU-SAFE is an adaptive, randomised, double-blind, placebo-controlled, first-in-human phase 1 trial (EudraCT 2019-000680-24) consisting of a single (SAD) and a multiple ascending dose (MAD) part of three dose cohorts each. The study was performed at the Phase 1 Unit, Uppsala University Hospital, Uppsala, Sweden. Data in this article were collected between Sep 20th, 2019 and Oct 20th 2021. In total 240 wounds were induced on the upper arms in 36 healthy volunteers. SAD: 12 participants, 4 wounds (2/arm), MAD: 24 participants, 8 wounds (4/arm). Wounds in each participant were randomised to treatment with placebo/saline or ILP100-Topical. Findings: In all individuals and doses, ILP100-Topical was safe and well-tolerated with no systemic exposure. A combined cohort analysis showed a significantly larger proportion of healed wounds (p = 0.020) on Day 32 by multi-dosing of ILP100-Topical when compared to saline/placebo (76% (73/96) and 59% (57/96) healed wounds, respectively). In addition, time to first registered healing was shortened by 6 days on average, and by 10 days at highest dose. ILP100-Topical increased the density of CXCL12+ cells in the wounds and local wound blood perfusion. Interpretation: The favourable safety profile and observed effects on wound healing support continued clinical development of ILP100-Topical for the treatment of complicated wounds in patients. Funding: Ilya Pharma AB (Sponsor), H2020 SME Instrument Phase II (#804438), Knut and Alice Wallenberg foundation.

Recent progress on biocompatible nanocarrier-based genistein delivery systems in cancer therapy.

Diets with naturally occuring chemopreventive agents are showing good potentials in serving dual purposes: firstly, for maintaining health, and secondly, for emerging as most puissant cost-effective strategy against chronic diseases like cancer. Genistein, one of the active soy isoflavone, is gaining attention due to its ability to impede carcinogenic processes by regulating wide range of associated molecules and signalling mechanisms. Epidemiologic and preclinical evidences suggest that sufficient consumption of soy-based food having genistein can be correlated to the reduction of cancer risk. However, certain adverse effects like poor oral bioavailability, low aqueous solubility and inefficient pharmacokinetics have pushed it down in the list of phytoconstituents currently undergoing successful clinical trials. In order to maximise the utilisation of therapeutic benefits of this phytoestrogen, suitable drug carrier designs are required. Recently, nanocarriers, mainly composed of polymeric materials, are progressively and innovatively exploited with the aim to improve pharmacokinetics and pharmacodynamics of genistein. Here, we have briefly reviewed (a) the targeted molecular mechanisms of geinstein, (b) nanopolymeric approaches opted so far in designing carriers and (c) the reasons behind their restricted clinical applications. Finally, some mechanism-based approaches are proposed presenting genistein as the future paradigm in cancer therapy.

Exfoliated graphene nanosheets: pH-sensitive drug carrier and anti-cancer activity.

A straightforward and facile method for the exfoliation of graphene sheets using poly(vinylpyrrolidone) nanoparticles of an average size of 42nm was developed and their dual role as pH-sensitive drug carrier and anti-cancer agent was evaluated. The cytotoxic impact of the exfoliated nanosheets (GRP-PVP-NP) was examined on various cells (HCT-116, HeLa, SCC-9, NIH-3T3 and HEK-293cells) by a series of assays. Their cytotoxic nature was attributed to affecting the mitochondrial enzyme activity, proliferation capability, and the formation of tight junctions in cancer cells. The endocytosis was found to be internalization mechanism for the cellular uptake of nanosheets. The generation of reactive oxygen species and elicitation of caspase-3 activity which was undoubtedly associated with triggering of oxidative stress speculated to be the dominant cause of the cytotoxic pattern of nanosheets against cancer cells. Additionally, the results also showed the role of the nanosheets as a pH-sensitive drug carrier through drug loading by supramolecular interaction. The efficient release of doxorubicin was seen at low pH and in an environment with a low oxygen concentration, thus under conditions mimicking the typical tumor microenvironment. Therefore, these findings provide the first evidence for a dual function of exfoliated graphene sheets and also elucidate the cytotoxic mechanism responsible for the cancer cell death.

Cancer therapeutics with epigallocatechin-3-gallate encapsulated in biopolymeric nanoparticles.

With the recent quantum leap in chemoprevention by dietary products, their use as cancer therapeutics is garnering worldwide attention. The concept of effortlessly fighting this deadly disease by gulping cups of green tea or swallowing green tea extract capsules is appreciated universally. Epigallocatechin-3-gallate (EGCG), a major polyphenol in green tea, has generated significant interest in controlling carcinogenesis due to its growth-inhibitory efficacy against a variety of cancers by targeting multiple signaling pathways. However, the success of EGCG in preclinical studies is difficult to translate into clinical trials due to issues of low solubility, bioavailability and an uncertain therapeutic window. The laborious and expensive journey of drugs from the laboratory to commercialization can be improved by utilizing nanoparticles as anti-cancer drug carriers. Exploitation of biopolymeric nanoparticles in recent years has improved EGCG's biodistribution, stability and tumor selectivity, revealing its superior chemopreventive effects. This review briefly summarizes recent developments regarding the targets and side effects of EGCG, complications associated with its low bioavailability and critically analyses the application of biopolymeric nanoparticles encapsulating EGCG as a next generation delivery systems.

Delineation of the role of glycosylation in the cytotoxic properties of quercetin using novel assays in living vertebrates.

Quercetin is a plant-derived flavonoid and its cytotoxic properties have been widely reported. However, in nature, quercetin predominantly occurs as various glycosides. Thus far the cytotoxic activity of these glycosides has not been investigated to the same extent as quercetin, especially in animal models. In this study, the cytotoxic properties of quercetin (1), hyperoside (quercetin 3-O-galactoside, 2), isoquercitrin (quercetin 3-O-glucoside, 3), quercitrin (quercetin 3-O-rhamnoside, 4), and spiraeoside (quercetin 4'-O-glucoside, 5) were directly compared in vitro using assays of cancer cell viability. To further characterize the influence of glycosylation in vivo, a novel zebrafish-based assay was developed that allows the rapid and experimentally convenient visualization of glycoside cleavage in the digestive tract. This assay was correlated with a novel human tumor xenograft assay in the same animal model. The results showed that 3 is as effective as 1 at inhibiting cancer cell proliferation in vivo. Moreover, it was observed that 3 can be effectively deglycosylated in the digestive tract. Collectively, these results indicate that 3 is a very promising drug candidate for cancer therapy, because glycosylation confers advantageous pharmacological changes compared with the aglycone, 1. Importantly, the development of a novel and convenient fluorescence-based assay for monitoring deglycosylation in living vertebrates provides a valuable platform for determining the metabolic fate of naturally occurring glycosides.

Protein-coated pH-responsive gold nanoparticles: Microwave-assisted synthesis and surface charge-dependent anticancer activity.

The biocompatibility and ease of functionalization of gold nanoparticles underlie significant potential in biotechnology and biomedicine. Eight different proteins were examined in the preparation of gold nanoparticles (AuNPs) in aqueous medium under microwave irradiation. Six of the proteins resulted in the formation of AuNPs. The intrinsic pH of the proteins played an important role in AuNPs with strong surface plasmon bands. The hydrodynamic size of the nanoparticles was larger than the values observed by TEM and ImageJ. The formation of a protein layer on the AuNPs accounts for this difference. The AuNPs exhibited sensitivity towards varying pH conditions, which was confirmed by determining the difference in the isoelectric points studied by using pH-dependent zeta potential titration. Cytotoxicity studies revealed anticancerous effects of the AuNPs at a certain micromolar concentration by constraining the growth of cancer cells with different efficacies due to the use of different proteins as capping agents. The positively charged AuNPs are internalized by the cells to a greater level than the negatively charged AuNPs. These AuNPs synthesized with protein coating holds promise as anticancer agents and would help in providing a new paradigm in area of nanoparticles.

Suppression of Toll-like receptor 4 activation by caffeic acid phenethyl ester is mediated by interference of LPS binding to MD2.

BACKGROUND AND PURPOSE: Toll-like receptors (TLRs) play a crucial role in recognizing invading pathogens and endogenous danger signal to induce immune and inflammatory responses. Since dysregulation of TLRs enhances the risk of immune disorders and chronic inflammatory diseases, modulation of TLR activity by phytochemicals could be useful therapeutically. We investigated the effect of caffeic acid phenethyl ester (CAPE) on TLR-mediated inflammation and the underlying regulatory mechanism. EXPERIMENTAL APPROACH: Inhibitory effects of CAPE on TLR4 activation were assessed with in vivo murine skin inflammation model and in vitro production of inflammatory mediators in macrophages. In vitro binding assay, cell-based immunoprecipitation study and liquid chromatography-tandem mass spectrometry analysis were performed to determine lipopolysaccharide (LPS) binding to MD2 and to identify the direct binding site of CAPE in MD2. KEY RESULTS: Topical application of CAPE attenuated dermal inflammation and oedema induced by intradermal injection of LPS (a TLR4 agonist). CAPE suppressed production of inflammatory mediators and activation of NFκB and interferon-regulatory factor 3 (IRF3) in macrophages stimulated with LPS. CAPE interrupted LPS binding to MD2 through formation of adduct specifically with Cys133 located in hydrophobic pocket of MD2. The inhibitory effect on LPS-induced IRF3 activation by CAPE was not observed when 293T cells were reconstituted with MD2 (C133S) mutant. CONCLUSIONS AND IMPLICATIONS: Our results show a novel mechanism for anti-inflammatory activity of CAPE to prevent TLR4 activation by interfering with interaction between ligand (LPS) and receptor complex (TLR4/MD2). These further provide beneficial information for the development of therapeutic strategies to prevent chronic inflammatory diseases.