Synthesis and evaluation of antisense oligonucleotides prodrug with G-quadruplex assembly and lysosome escape capabilities for oncotherapy.

The applications of antisense oligonucleotides (ASOs) in rare or common diseases treatment have garnered great attention in recent years. Nevertheless, challenges associated with stability and bioavailability still persist, hampering the efficiency of ASOs. This work presents an ASO prodrug with parallel G-quadruplex assembly and lysosome escape capabilities for oncotherapy. Our findings revealed that the end-assembled quadruplex structure effectively shielded the ASO from enzymatic degradation. Meanwhile, the conjugation of maleimide within the quadruplex enhanced cellular uptake, potentially offering an alternative cell entry mechanism that circumvents lysosome involvement. Notably, an optimized molecule, Mal2-G4-ASO, exhibited remarkable therapeutic effects both in vitro and in vivo. This work presents a promising avenue for enhancing the activity of nucleic acid drugs in oncotherapy and potentially other disease contexts.

Anisamide-conjugated hairpin antisense oligonucleotides prodrug co-delivering doxorubicin exhibited enhanced anticancer efficacy.

Antisense oligonucleotides (ASONs)-based therapeutics offers tremendous promise for the treatment of diverse diseases. However, there is still a need to develop ASONs with enhanced stability against enzymes, improved drug delivery, and enhanced biological potency. In this study, we propose a novel anisamide (AA)-conjugated hairpin oligonucleotide prodrug loading with chemotherapeutic agent (doxorubicin, DOX) (AA-loop-ASON/DOX) for oncotherapy. Results indicated that the introduction of a hairpin conformation and AA ligand in prodrug significantly improved the stability against enzymatic hydrolysis, as well as the cellar uptake of ASONs and DOX. The incorporation of disulfide bonds could trigger mechanical opening, resulting in the release of ASON and DOX in response to the intracellular glutathione (GSH) in tumors. Moreover, the composite of DOX-loading ASONs prodrug exhibited a robust and selective inhibition of tumor cell proliferation. This paper introduces a novel design concept for nucleic acid-based therapeutics, aiming to enhance the delivery of drug and improve biological effectiveness.

Synthesis, Biophysical Properties, and Antitumor Activity of Antisense Oligonucleotides Conjugated with Anisamide.

Antisense oligonucleotides (ASONs) have proven potential for the treatment of various diseases. However, their limited bioavailability restricts their clinical application. New structures with improved enzyme resistance stability and efficient drug delivery are needed. In this work, we propose a novel category of ASONs bearing anisamide conjugation at phosphorothioate sites for oncotherapy. ASONs can be conjugated with the ligand anisamide very efficiently and flexibly in a solution. The conjugation sites and the ligand amount both influence anti-enzymatic stability and cellular uptake, resulting in changes in antitumor activity that are detectable by cytotoxicity assay. The conjugate with double anisamide (T6) was identified as the optimal conjugate, and its antitumor activity and the underlying mechanism were examined further in vitro and in vivo. This paper presents a new strategy for the design of nucleic acid-based therapeutics with improved drug delivery and biophysical and biological efficacy.

Functional diversity of rhizosphere soil microbial communities in response to different tillage and crop residue retention in a double-cropping rice field.

Microbial community functional diversity is a sensitive indicator of soil quality, soil management such as tillage and crop residue which can affect the microbial community functional diversity of paddy field. However, there is still limited information about the influence of different tillage and crop residue management on rhizosphere soil microbial community functional diversity in a double-cropping rice (Oryza sativa L.) field. Therefore, four tillage treatments were set up in paddy field, tillage treatments were included: conventional tillage with residue incorporation (CT), rotary tillage with residue incorporation (RT), no-tillage with residue retention (NT), and rotary tillage with residue removed as control (RTO). And the effects of CT, RT, NT, and RTO treatments on the average well color development (AWCD), genetic diversity indices and carbon source utilization of rhizosphere soil were studied in the present paper. The results showed that the values of AWCD with CT, RT and NT treatments were higher than that of RTO treatment. It was implied that application of crop residue management resulted in the variation of the carbon utilization efficiency of rhizosphere soil microbial communities. At maturity stages of early and late rice, the Richness indices, Shannon indices and McIntosh indices with CT treatment were significantly higher than that of RTO treatment, and with the order as CT>RT>NT>RTO. Principal component analysis (PCA) results indicated that there were significant differences in carbon substrate utilization patterns among different tillage treatments. Carbohydrates and amino acids were the main carbon resources utilized by rhizosphere soil microbes. Therefore, the combined application of tillage with crop residue management could significantly increase the rhizosphere soil microbial community functional diversity in the double-cropping paddy field of southern China.

Utilization of carbon sources in the rice rhizosphere and nonrhizosphere soils with different long-term fertilization management.

Carbon (C) plays an important role in the interaction between plant and rhizosphere microbial communities, but there is still limited information about how C source utilization soil microbial structure responds to soil fertility changes under the double-cropping rice (Oryza sativa L.) system in Southern China paddy fields. Therefore, the effects of long-term (33 years) fertilizer regimes on the characteristics of C utilization in both rhizosphere and nonrhizosphere soils under double-cropping rice fields in Southern China were investigated by using the metagenome sequencing technology. The experiment began in 1986, and included five fertilizer treatments: without fertilizer input (CK), chemical fertilizer alone (MF), rice straw residue and chemical fertilizer (RF), 30% organic matter, and 70% chemical fertilizer (LOM), and 60% organic matter and 40% chemical fertilizer (HOM). The results showed that the relative abundance of Gemmatimonadetes and Planctomycetia in both the rhizosphere and nonrhizosphere soils was increased by application of rice straw residue and organic manure, whereas the relative abundance of Gammaproteobacteria and Nitrospira was promoted by application of inorganic fertilizers. The largest group of clusters of orthologous groups of proteins categories was "amino acid transport and metabolism" with 16.46% unigenes, followed by "general function prediction only" (12.23%). Regarding the gene ontology categories, biological process were the largest category (174 949, 46.40%), followed by cellular component (126 766, 33.62%), and molecular function (110 353, 29.26%). The principal coordinate analysis indicated that different parts of the root zone were the most important factors affecting the variation of C source utilization bacteria community, and the different fertilizer treatments were the second important factor affecting the variation of C source utilization bacteria community. As a result, the application of fertilization practices had significant effects on the abundance and community composition of C source utilization microbes in paddy soils. The results showed that the combined application of rice straw residue or organic manure with chemical fertilizer practices significantly increases the C source utilization of soil microorganisms in double-cropping rice fields.

Impact of long-term fertilization practices on the soil aggregation and humic substances under double-cropped rice fields.

Soil organic matter (SOM) content and soil aggregation are essential components of soil structure, which plays an important role in soil quality and fertility. Also, the SOM content, aggregation, and humus substances in paddy field were affected by application of fertilization practices. However, there is still limited information about the effects of long-term different fertilization practices on soil aggregation and carbon content in the humic acid (C-HAF), fulvic acid (C-FAF), and humin (C-HUM) fractions under double-cropping rice (Oryza sativa L.) system in Southern China paddy fields. Therefore, the effects of long-term fertilizer application on soil aggregation and C-HUM, C-HAF, and C-FAF contents in 0-5-, 5-10-, and 10-20-cm soil depth under double-cropped rice fields in Southern China were investigated. The experiment located at NingXiang County in Hunan Province, China begins in 1986 and the experiment includes five treatments: without fertilizer input (CK), mineral fertilizer alone (MF), rice straw residues and mineral fertilizer (RF), 30% organic matter and 70% mineral fertilizer (LOM), and 60% organic matter and 40% mineral fertilizer (HOM). The results showed that the soil total organic carbon content in paddy soils with RF, LOM, and HOM treatments was significant higher (P < 0.05) than that of the CK treatment at early and late rice maturity stages. The different sizes of soil aggregates with different fertilization treatments were decreased as HOM > LOM > RF > MF > CK. The HOM treatment had the highest percentage of soil aggregates in each size class and the CK treatment had the lowest percentage of soil aggregates in each size class in 0-5-, 5-10-, and 10-20-cm soil depth at early and late rice maturity stages. The soil C-HAF, C-FAF, and C-HUM contents were increased by long-term combined application of manure with mineral fertilizer practices. Meanwhile, the results indicated that the soil C-HAF, C-FAF, and C-HUM contents with RF, LOM, and HOM treatments were significantly higher (P < 0.05) than that of the CK treatment at early and late rice maturity stages. As a result, the soil total organic carbon content, each size class of soil aggregates, and soil C-HAF, C-FAF, and C-HUM contents were increased by long-term combined application of manure with mineral fertilizer in double-cropped rice fields.

A composite hydrogel of chitosan/heparin/poly (γ-glutamic acid) loaded with superoxide dismutase for wound healing.

In chronic wounds, severe oxidative stress hampers the healing of wounds. This study constructed a novel antioxidant-loaded hydrogel for healing diabetic wounds. First, composite hydrogels consisting of chitosan, heparin and poly (γ-glutamic acid) with different ratios were prepared through electrostatic interaction. These hydrogels exhibited good three-dimensional network structures, and the porosity of the hydrogels decreased as the crosslinking density increased. The hydrogels possessed good swelling capacity, and showed typical viscoelastic behavior and good mechanical property in rheological test. Fibroblast proliferation assay demonstrated that the hydrogels were cytocompatible. Then, superoxide dismutase was loaded onto the hydrogel to build a wound dressing with antioxidant property. In diabetic rat models, the dressing proved to accelerate wound healing by promoting wound closure and collagen deposition. In summary, the hydrogel prepared in this study exhibited good physical properties and could promote the repair of chronic trauma in diabetes effectively and therefore is a promising candidate dressing in wound healing.

Synergistic function of Smad4 and PTEN in suppressing forestomach squamous cell carcinoma in the mouse.

The genetic bases underlying esophageal tumorigenesis are poorly understood. Our previous studies have shown that coordinated deletion of the Smad4 and PTEN genes results in accelerated hair loss and skin tumor formation in mice. Herein, we exemplify that the concomitant inactivation of Smad4 and PTEN accelerates spontaneous forestomach carcinogenesis at complete penetrance during the first 2 months of age. All of the forestomach tumors were invasive squamous cell carcinomas (SCCs), which recapitulated the natural history and pathologic features of human esophageal SCCs. A small population of the SCC lesions was accompanied by adenocarcinomas at the adjacent submucosa region in the double mutant mice. The rapid progression of forestomach tumor formation in the Smad4 and PTEN double knockout mice corresponded to a dramatic increase in esophageal and forestomach epithelial proliferation. The decreased expression of p27, p21, and p16 together with the overexpression of cyclin D1 contributed cooperatively to the accelerated forestomach tumorigenesis in the double mutant mice. Our results point strongly to the crucial relevance of synergy between Smad4 and PTEN to suppress forestomach tumorigenesis through the cooperative induction of cell cycle inhibitors.