Core Role of Hydrophobic Core of Polymeric Nanomicelle in Endosomal Escape of siRNA.

Efficient endosomal escape is the most essential but challenging issue for siRNA drug development. Herein, a series of quaternary ammonium-based amphiphilic triblock polymers harnessing an elaborately tailored pH-sensitive hydrophobic core were synthesized and screened. Upon incubating in an endosomal pH environment (pH 6.5-6.8), mPEG45-P(DPA50-co-DMAEMA56)-PT53 (PDDT, the optimized polymer) nanomicelles (PDDT-Ms) and PDDT-Ms/siRNA polyplexes rapidly disassembled, leading to promoted cytosolic release of internalized siRNA and enhanced silencing activity evident from comprehensive analysis of the colocalization and gene silencing using a lysosomotropic agent (chloroquine) and an endosomal trafficking inhibitor (bafilomycin A1). In addition, PDDT-Ms/siPLK1 dramatically repressed tumor growth in both HepG2-xenograft and highly malignant patient-derived xenograft models. PDDT-Ms-armed siPD-L1 efficiently blocked the interaction of PD-L1 and PD-1 and restored immunological surveillance in CT-26-xenograft murine model. PDDT-Ms/siRNA exhibited ideal safety profiles in these assays. This study provides guidelines for rational design and optimization of block polymers for efficient endosomal escape of internalized siRNA and cancer therapy.

Comparison between 18 F-DCFPyL PET and MRI for the detection of transition zone prostate cancer.

BACKGROUND: We aimed to compare the diagnostic performance of 18 F-DCFPyL positron emission tomography (PET) and multiparameter magnetic resonance imaging (mp-MRI) in detecting transition zone (TZ) prostate cancer (PCa). METHODS: This retrospective study included 20 patients who underwent 18 F-DCFPyL PET/MRI and 32 patients who underwent 18 F-DCFPyL PET/CT and MRI from January 2019 to June 2020. All patients had TZ lesions and underwent prostate biopsies. One senior (reader 1) and one junior (reader 2) nuclear medicine physician evaluated each TZ lesion independently, according to the molecular imaging prostate-specific membrane antigen scoring system and the Prostate Imaging Reporting and Data System version 2.1 (PI-RADS v2.1). The histologic diagnosis of prostate biopsy was used as the reference standard. The diagnostic performance of the two methods was compared in terms of inter-reader agreement and area under the receiver operating characteristic (AUC-ROC) curve. RESULTS: Of the 52 patients, 43 had TZ PCa. For inter-reader agreement, the kappa value was 0.883 for 18 F-DCFPyL PET and 0.393 for mp-MRI. For PET, both readers had the same diagnostic sensitivity, specificity, and accuracy of 93.0%, 77.8%, and 90.4%, respectively. For mp-MRI, the diagnostic sensitivity, specificity, and accuracy was 67.4%, 33.3%, and 61.5% for reader 1, and 51.2%, 44.4%, and 51.9% for reader 2, respectively. PET outperformed mp-MRI for both readers with an AUC of 0.872 for PET versus 0.584 for mp-MRI, p = .0209 for reader 1, and an AUC of 0.860 for PET versus 0.505 for mp-MRI, p = .0213 for reader 2. Among the 43 patients with TZ PCa, 18 F-DCFPyL PET detected a distant bone metastasis missed by the CT in one case and two small lymph node metastases missed by the CT and MRI in another case. CONCLUSIONS: These results suggest that 18 F-DCFPyL PET, which was almost independent of the experience of the readers, was more objective in the evaluation of TZ lesions, and had higher diagnostic value than mp-MRI.

Metabolic parameters of pretreatment 2-[18F]fluoro-D-glucose positron emission tomography for prognosis in patients with gallbladder adenocarcinoma: a cohort study.

Background: The incidence of gallbladder adenocarcinoma (GBA) is relatively low, yet it exhibits a high degree of malignancy and a significantly low 5-year survival rate. The aim of this study was to investigate the prognostic value of pretreatment 2-[18F]fluoro-D-glucose positron emission tomography {2-[18F]FDG PET} parameters in predicting outcomes for patients with GBA. Methods: In total, 67 patients with GBA who underwent 2-[18F]FDG PET/computed tomography (CT) before treatment were retrospectively analyzed at Chinese PLA General Hospital from January 2012 to June 2022. All patients were diagnosed by pathology, and their baseline characteristics and clinical data were collected. The metabolic PET parameters of the primary and metastatic lesions were measured, including the maximum and average standardized uptake values (SUVs), metabolic tumor volume (MTV), and total lesion glycolysis (TLG). The prognostic significance of metabolic parameters and other clinical variables was assessed using Cox proportional hazards regression models. Differences in progression-free survival (PFS) and overall survival (OS) in relation to metabolic parameters were examined using the Kaplan-Meier method. Results: During a median follow-up period of 14.2 months, 43 patients (64.2%) experienced tumor recurrence or progression, and 38 patients (56.7%) died of cancer. In the univariate Cox regression analysis, liver parenchymal invasion (P=0.001), lymph node metastasis (P=0.007), distant metastases (P=0.049), tumor differentiation (P=0.028), surgery (P=0.014), carcinoembryonic antigen (CEA) level (P=0.030), carbohydrate antigen 19-9 (CA19-9) level (P=0.003), TLG (P=0.005), MTV (P<0.001), sum of the TLGs of the primary and metastatic lesions (total TLG, tTLG) (P=0.001), and sum of the MTVs of the primary and metastatic lesions (total MTV, tMTV) (P<0.001) were significant predictors of PFS. In multivariate analysis, MTV was an independent predictor of PFS [hazard ratio (HR) =2.785; 95% confidence interval (CI): 1.204-6.441; P=0.017]. In the univariate Cox regression analysis, liver parenchymal invasion (P=0.001), lymph node metastasis (P=0.027), distant metastases (P=0.036), tumor differentiation (P=0.047), surgery (P=0.002), neutrophil-to-lymphocyte ratio (NLR) (P=0.011), CEA level (P=0.036), CA19-9 level (P<0.001), TLG (P=0.007), MTV (P<0.001), tTLG (P=0.003), and tMTV (P<0.001) were significant predictors of OS. In the multivariate analysis, higher CA19-9 levels >37 U/mL and a greater tMTV (HR =2.961; 95% CI: 1.092-8.024; P=0.033) were predictive of OS. Conclusions: Our study results suggest that pretreatment 2-[18F]FDG PET parameters can not only assist in the diagnosis of patients with GBA but may also serve as predictive factors for the prognosis of these patients and should thus be applied in their treatment.

Regulating intermediates to realize the coupled hydrion with biology polysulfide in wastewater treatment.

The purpose of this study is to clarify the mechanism of the coupled hydrion with biology polysulfide in the simultaneous denitrification and desulfurization process. The coupled hydrion with biology polysulfide, uncoupled hydrion with biology polysulfide and no polysulfide experiments were performed in wastewater with two kinds of sulfide loads (100 and 200 mg/L). When the concentration of thiosulfate was suitable, the free H+ concentration (74.2 and 91.0 mg/L) and the proportion of Thiobacillus denitrificans (85.4% and 59.7%) were both higher under the two kinds of sulfide loading conditions (100 and 200 mg/L), and coupled hydrion with biology polysulfide was realized (the production of elemental sulfur is as high as 33 and 101 mg/L). Further analysis shown that the way of coupled hydrion with biology polysulfide were both: 2.0S2-+6.4NO3-+30.1H++21.7e-→1.0S2-+1.0SO42-+3.2N2+15.0H2O. In addition, for the coupled hydrion with biology polysulfide, more nitrates could be utilized to produce elemental sulfur S0, and the lower ratio of H+/S0 and SO42-/S0 were observed (S2- = 100 mg/L: 2.3 and 0.9; S2- = 200 mg/L: 0.9 and 0.03), which could promote the growth of Thiobacillus denitrificans and increase the proportion of Thiobacillus denitrificans. This maybe one of the reasons why coupled hydrion with biology polysulfide could be achieved.

Possibility for double optimization of siRNA intracellular delivery efficiency and antibacterial activity: Structure screening of pH-sensitive triblock amphiphilic polycation micelles.

Optimal combination of hydrophobic-hydrophilic balance, proton buffering and electrostatic interaction is the key issue for designing polycations as efficient gene vectors and antibacterial agents. Herein, we screened a series of pH-sensitive quaternary ammonium-based amphiphilic triblock copolymers, mPEG2k-P(DPAa/DMAb)-PQAc (TDDE-x), which had different pKa values and proton buffering capacities. Significantly, we found that both the highest siRNA intracellular delivery efficiency and the strongest antibacterial capacity occurred on TDDE-3 micelles with the segment structure of mPEG2k-P(DPA50/DMA56)-PQA55. The TDDE-3/siRNA complex achieved 67% silencing efficiency on H9C2 cells (N/P = 5, 50 nM siRNA), higher than the advanced commercial transfection reagents RNAiMAX (58%) and Lipo2000 (30%). Moreover, TDDE-3 micelles showed quite low MICs of 32 µg/mL and 8 µg/mL against E. coli and S. aureus, respectively. Further studies on the structure-function relationship indicated that TDDE-3 micelles could mediate robust endosome escape and siRNA cytosolic release, and strong bacterial cell membrane-destabilizing function. Undoubtedly, this work reveals the possibility for double optimization of siRNA intracellular delivery efficiency and antibacterial activity of amphiphilic polycations by reasonable structure design, which is significant for low-cost development and clinical translation of efficient multifunctional polycations.

pH-Sensitive Polycations for siRNA Delivery: Effect of Asymmetric Structures of Tertiary Amine Groups.

pH-sensitive polyelectrolytes provide enormous opportunity for siRNA delivery. Especially, their tertiary amine structures can not only bind genes but also act as pH-sensitive hydrophobic structure to control genes release. However, the influence of molecular structures on siRNA delivery still remains elusive, especially for the asymmetric alkyl substituents of the tertiary amine groups. Herein, a library of N-methyl-N-alkyl aminoethyl methacrylate monomers (MsAM) with asymmetric alkyl substituents on the tertiary amine group is synthesized and used to prepare a series of tri-block polycationic copolymers poly(aminoethyl methacrylate)-block-poly (N-methyl-N-alkyl aminoethyl methacrylate)-block-poly(ethylene glycol methacrylate) (PAMA-PMsMA-PEG). And the properties of these polycations and their self-assembled micelles are characterized, including molecular structure, proton buffering capacity, pH-sensitivity, size, and zeta potential. With the length increase of one alkyl substituent, the proton buffering capacity of both monomers and polycations is demonstrated to be narrowed down. The siRNA delivery efficiency and cytotoxicity of these micelles are also evaluated on HepG2 cells. In particular, poly(aminoethyl methacrylate)-block-poly(N-methyl-N-ethyl aminoethyl methacrylate)-block-poly(ethylene glycol methacrylate) (PAMA-PMEMA-PEG) elicited the best luciferase knockdown efficiency and low cytotoxicity. Besides, PAMA-PMEMA-PEG/siRRM2 also induced significant anti-tumor activity in vitro. These results indicated PAMA-PMEMA-PEG has potential for further use in the design of gene vehicles with the improved efficiency of siRNA delivery.

Combating drug-resistant bacterial infection using biodegradable nanoparticles assembled from comb-like polycarbonates grafted with amphiphilic polyquaternium.

Bacterial infection is a serious clinical threat. The misuse of antibiotics has already resulted in the emergence of antibiotic-resistant strains of pathogenic bacteria. Efficient membrane-destructive antibacterial agents are considered as an alternative, promising solution against bacterial infection. Herein, we prepared a new type of comb-like cationic, polyethylene glycol (PEG) block polycarbonates with polyquaternium arms (G-CgQAs). The amphiphilic G-CgQAs could self-assemble into about 60 nm sized nanoparticles (NPs) with positive charges (20~30 mV). G-CgQA-3 NPs with an appropriate hydrophobic-hydrophilic balance in the polyquaternium arms showed antibacterial activity against Gram-negative, Gram-positive, and drug-resistant strains at low concentrations (MIC 64-128 µg mL-1) and low hemolysis (HC50 > 2000 µg mL-1). In vivo anti-infection tests indicated G-CgQA-3 NPs could highly inhibit the growth of vancomycin-resistant bacteria by spraying on wounds. Collectively, G-CgQA NPs hold great promise for the prevention of infection, serving as new antibacterial agents. This study also highlights the significance of a hydrophobic block in positive polyquaternium arms to facilitate the antibacterial activity of cationic, quaternized polymers. The design of comb-like amphiphilic cationic polycarbonates provides a new method for manufacturing antibacterial nano-agents.

Harnessing pH-Sensitive Polycation Vehicles for the Efficient siRNA Delivery.

pH-sensitive hydrophobic segments have been certificated to facilitate siRNA delivery efficiency of amphiphilic polycation vehicles. However, optimal design concepts for these vehicles remain unclear. Herein, by studying the library of amphiphilic polycations mPEG-PAMA50-P(DEAx-r-D5Ay) (EAE5x/y), we concluded a multifactor matching concept (pKa values, "proton buffering capacities" (BCs), and critical micelle concentrations (CMCs)) for polycation vehicles to improve siRNA delivery efficiency in vitro and in vivo. We identified that the stronger BCs in a pH 5.5-7.4 subset induced by EAE548/29 (pKa = 6.79) and EAE539/37 (pKa = 6.20) are effective for siRNA delivery in vitro. Further, the stronger BCs occurred in a narrow subset of pH 5.5-6.5 and the lower CMC attributed to higher siRNA delivery capacity of EAE539/37 in vivo than EAE548/29 after intravenous administration and subcutaneous injection. More importantly, 87.2% gene knockdown efficacy was achieved by EAE539/37 via subcutaneous injection, which might be useful for an mRNA vaccine adjuvant. Furthermore, EAE539/37 also successfully delivered siRRM2 to tumor via intravenous administration and received highly efficient antitumor activity. Taken together, the suitable pKa values, strong BCs occurred in pH 5.5-6.5, and low CMCs were probably the potential solution for designing efficient polycationic vehicles for siRNA delivery.

Oriented oxidation of all alkanes in soils.

In order to investigate the mechanism of the oriented oxidation of all alkanes by regulating organic functional groups, Fenton oxidation was performed in two soils (S1 and S2: total petroleum hydrocarbons (TPH) are 26,281 mg/kg and 12,668 mg/kg). The higher the proportion of hydroxyl radicals (OH) transferred (41 %-58 %), the more the number of oriented oxidation of alkanes, which realized the oriented oxidation of all alkanes. Meanwhile, high oriented oxidation of long alkanes and short alkanes (58 %: 3405 mg/kg and 1729 mg/kg) was observed. Protein Ⅰ in soil organic matter (SOM) was reduced by regulating CH and carboxyl group OH, which indicated that protein Ⅰ was inactive. Protein Ⅰ oxidation after regulation was decreased significantly. Protein Ⅰ was the main active organic matter to capture OH. When the relative reactivity coefficient KTPH/SOM (the ratio of TPH oxidation to SOM oxidation) and KTPH/protein I (the ratio of TPH oxidation to protein Ⅰ oxidation) were higher than 1, low oxidation of SOM and protein Ⅰ was obtained. It indicated that for the oriented oxidation of all alkanes, the high coefficient of relative reactivity for petroleum was the key for the transfer of OH from oxidizing SOM to oxidizing alkanes.

Comb-Like Amphiphilic Polycarbonates with Different Lengths of Cationic Branches for Enhanced siRNA Delivery.

Owing to the biodegradability and good biocompatibility polycarbonates show the versatile class of applications in biomedical fields. While their poor functional ability seriously limited the development of functional polycarbonates. Herein, a new Br-containing cyclic carbonate (MTC-Br) and a polycarbonate atom transfer radical polymerization (ATRP) macro-initiator (PEG-PMTC-Br) is synthesized. Then, by initiating the side-chain ATRP of 2-(dimethyl amino)ethyl methacrylate (DMAEMA) on PEG-PMTC-Br, a series of comb-like amphiphilic cationic polycarbonates, PEG-b-(PMTC-g-PDMAEMA) (GMDMs), with different lengths of cationic branches are successfully prepared. All these poly(ethylene glycol)-b-(poly((5-methyl-2-oxo-1,3-dioxane-5-yl) methyl 2-bromo-2-methylpropanoate/1,3-dioxane-2-one)-g-poly(2-dimethyl aminoethyl methacrylate) (GMDMs) self-assembled nanoparticles (NPs) (≈180 nm, +40 mV) can well bind siRNA to form GMDM/siRNA NPs. The gene silence efficiency of GMDM/siRNA high to 80%, which is even higher than the commercial transfection reagent lipo2000 (76%). But GMDM/siRNA shows lower cell uptake than lipo2000. So, the high gene silence ability of GMDM/siRNA NPs can be attributed to the strong intracellular siRNA trafficking capacity. Therefore, GMDM NPs are potential siRNA vectors and the successful preparation of comb-like polycarbonates also provides a facile way for diverse side-chain functional polycarbonates, expanding the application of polycarbonates.

"Off/on" fluorescence imaging-guided cancer diagnosis and multi-modal therapy.

An efficient theranostic nanoplatform responding to tumour microenvironments with characters of simple and flexible combinations owns great potential in cancer diagnosis and therapy. Herein, a series of triblock copolymers, mPEG-b-PDPA-b-P(nBMA-r-cystamine) (EPB), were synthesized and among them, the structure of EPB-3 was optimized for both fluorescence imaging-guided cancer diagnosis and multi-modal therapy with good biocompatibility. (1) The self-assembled nanoparticles of EPB-3-ICG1 obtained by conjugating one ICG on EPB-3 via S-S bonds effectively performed reduction-sensitive OFF/ON fluorescence signal transition, thus inducing tumour cell-specific amplified fluorescence imaging in vitro and in vivo. (2) By entrapping Au nanorods into the co-assembled NPs of EPB-3 and EPB-3-ICG1, EPB-3-ICG1@Au NPs could synchronously induce strong tumour fluorescence imaging and high local photothermal effect, indicating the potential of imagine-guided photothermal therapy. (3) EPB-3 NPs could efficiently co-load paclitaxel (PTX) and ICG to form stable EPB-3@PTX@ICG NPs, which provided long periods of intracellular pH-sensitive sustainable drug release and highly enhanced apoptosis of 4T1 cells in vitro by the chemo-photothermal effect. Excitingly, a single intravenous injection of EPB-3@PTX@ICG NPs followed by a one-time local near-infrared light (NIR, 808 nm) irradiation treatment for 10 min could lead to significant inhibition of tumour growth, avoiding tumor metastasis and extending the survival of mice. All the above-mentioned results suggest that EPB-3 provides a nanoplatform with the characters of simple structure, convenience of use and flexible combination, holding potential for multi-modal diagnosis and therapy.