An acid-activatable fluorouracil prodrug for colorectal cancer synergistic therapy.

5-Fluorouracil has demonstrated certain efficiency in patients with colorectal cancer. However, significant side effects of use by injection are common. To address this issue defects, a reengineered 5'-deoxy-5-fluorocytidine (DFCR) based drug delivery system (POACa) is developed as a prominent tumor-selective nano-activator. Investigations demonstrate that the constructed nano-activator exhibits good biocompatibility and high therapeutic efficiency in mice with subcutaneous and orthotopic SW-480 colorectal tumors, as its activity is strictly dependent on the tumor-associated acid environment and thymidine phosphorylase. These strategies diminish the off-target toxicity and improve the specificity and sensitivity of human colorectal cancer cells to 5-Fu, obtaining potent efficiency by the combination of H2O2 mediated oxidative stress, calcium overload and 5-Fu-induced chemotherapy (the combination index is 0.11). Overall, the engineered nano-activator exhibits a high therapeutic index in vitro and in vivo. STATEMENT OF SIGNIFICANCE: In this study, we designed and prepared a pH-responsive polymer to synchronously deliver DFCR (5'-deoxy-5-fluorocytidine, a prodrug of 5-Fu), Ca2+ and H2O2. The constructed nano-activator was denoted as POACa. (1) To address the problem of premature leakage of cargo by physical embedding, our research modified the inactive prodrug DFCR through chemical bonding. (2) The activation of the prepared nano-activator was strictly dependent on the tumor-associated acid environment and thymidine phosphorylase, providing the drug delivery system with inherent safety. (3) A distinctly low combination index value (0.11) of CaO2 and DFCR indicated that POACa has a prominent tumor suppression effect by tumor calcium overload sensitized chemotherapy and H2O2 mediated cytotoxicity.

Integrated anti-vascular and immune-chemotherapy for colorectal carcinoma using a pH-responsive polymeric delivery system.

Colorectal carcinoma (CRC) has become one of the most prevalent malignant tumors and exploring a potential therapeutic strategy with diminished drug-associated adverse effects to combat CRC is urgent. Herein, we designed a pH-responsive polymer to efficiently encapsulate a stimulator of interferon genes (STING) agonist (5,6- dimethylxanthenone-4-acetic acid, termed ASA404) and a common clinically used chemotherapeutic agent (1-hexylcarbamoyl-5-fluorouracil, termed HCFU). Investigations in vitro demonstrated that polymer encapsulation endowed the system with a pH-dependent disassembly behavior (pHt 6.37), which preferentially selected cancerous cells with a favorable dose reduction (dose reduction index (DRI) of HCFU was 4.09). Moreover, the growth of CRC in tumor-bearing mice was effectively suppressed, with tumor suppression rates up to 94.74%, and a combination index (CI) value of less than one (CI = 0.41 for CT26 cell lines), indicating a significant synergistic therapeutic effect. Histological analysis of the tumor micro-vessel density and enzyme-linked immunosorbent assay (ELISA) tests indicated that the system increased TNF-α and IFN-ß levels in serum. Therefore, this research introduces a pH-responsive polymer-based theranostic platform with great potential for immune-chemotherapeutic and anti-vascular combination therapy of CRC.

Engineered Sucrose Metabolism Improves the Smut Disease Suppression Potency of Pseudomonas sp. ST4.

Sporisorium scitamineum and Ustilago maydis are two fungal pathogens causing severe sugarcane and maize diseases, respectively. Sexual mating of compatible sporidia is essential for these pathogens to form infections dikaryotic mycelia and cause smut diseases. We showed recently that in the presence of exogenous glucose, the Pseudomonas sp. strain ST4 could block the fungal mating and display a strong disease suppression potency on S. scitamineum. With the aim of conferring strain ST4 the ability to metabolize sucrose in plants for glucose production, we identified a strong native promoter pSsrA in strain ST4 and additional promoter elements to facilitate translation and peptide translocation for the construction of a fusion gene encoding sucrose metabolism. The cscA gene encoding sucrose hydrolase from Pseudomonas protegens Pf-5 was fused to the promoter pSsrA, a translational coupler bicistronic design and a Tat signal peptide, which was then cloned into mini-Tn7 transposon. This synthetic gene cassette was integrated into the chromosome of strain ST4, and the resultant engineered strain ST4E was able to hydrolyze sucrose with high efficiency and displayed elevated inhibitory activity on the mating and virulence of S. scitamineum and U. maydis. The findings from this study provide a valuable device and useful clues for the engineering of sucrose metabolism in non- or weak-sucrose-utilizing bacterial strains and present an improved biocontrol agent against plant smut pathogens. IMPORTANCE Sporisorium scitamineum and Ustilago maydis are typical dimorphic fungi causing severe sugarcane and maize smut diseases, respectively. Sexual mating of compatible sporidia is essential for these pathogens to form infections dikaryotic mycelia and cause smut diseases. We previously demonstrated that the biocontrol strain Pseudomonas sp. ST4 could block the fungal mating and displays a strong suppression potency on smut diseases, while it was unable to utilize the host-sourced sucrose for glucose production critical for antifungus efficiency. In this study, we constructed a high-expression gene cassette for minitransposon-mediated genome integration and sucrose hydrolysis in the bacterial periplasmic space. The resultant engineered strain ST4E was able to hydrolyze sucrose and inhibit the mating and hyphal growth of S. scitamineum and U. maydis. These findings provide a valuable tool and useful clues for the engineering of sucrose metabolism in non- or weak-sucrose-utilizing bacterial strains and present an improved biocontrol agent against plant smut pathogens.