Short cell-penetration peptide conjugated bioreducible polymer enhances gene editing of CRISPR system.

CRISPR-based gene therapy offers precise targeting and specific editing of disease-related gene sequences, potentially yielding long-lasting treatment effects. However, efficient delivery remains a significant challenge for its widespread application. In this study, we design a novel short peptide-conjugated bioreducible polymer named TSPscp as a safe and effective delivery vector for the CRISPR system. Our results show that TSPscp markedly boosts transcriptional activation and genome editing activities of multiple CRISPR systems as confirmed by decomposition-seq and Deep-seq, which is resulted from its capability in facilitating delivery of plasmid DNA by promoting cellular uptake and lysosomal escape. Additionally, TSPscp further enhances genome editing of CRISPR by delivery of minicircle DNA, a condensed form of regular plasmid DNA. More importantly, TSPscp significantly improves delivery and genome editing of CRISPR system in vivo. In summary, our study highlights TSPscp as a promising delivery tool for CRISPR applications in vivo.

Nanoscale Metal-Organic Frameworks for Ratiometric Oxygen Sensing in Live Cells.

We report the design of a phosphorescence/fluorescence dual-emissive nanoscale metal-organic framework (NMOF), R-UiO, as an intracellular oxygen (O2) sensor. R-UiO contains a Pt(II)-porphyrin ligand as an O2-sensitive probe and a Rhodamine-B isothiocyanate ligand as an O2-insensitive reference probe. It exhibits good crystallinity, high stability, and excellent ratiometric luminescence response to O2 partial pressure. In vitro experiments confirmed the applicability of R-UiO as an intracellular O2 biosensor. This work is the first report of a NMOF-based intracellular oxygen sensor and should inspire the design of ratiometric NMOF sensors for other important analytes in biological systems.

Nanoscale Coordination Polymers Codeliver Carboplatin and Gemcitabine for Highly Effective Treatment of Platinum-Resistant Ovarian Cancer.

Due to the ability of ovarian cancer (OCa) to acquire drug resistance, it has been difficult to develop efficient and safe chemotherapy for OCa. Here, we examined the therapeutic use of a new self-assembled core-shell nanoscale coordination polymer nanoparticle (NCP-Carbo/GMP) that delivers high loadings of carboplatin (28.0 ± 2.6 wt %) and gemcitabine monophosphate (8.6 ± 1.5 wt %). A strong synergistic effect was observed between carboplatin and gemcitabine against platinum-resistant OCa cells, SKOV-3 and A2780/CDPP, in vitro. The coadministration of carboplatin and gemcitabine in the NCP led to prolonged blood circulation half-life (11.8 ± 4.8 h) and improved tumor uptake of the drugs (10.2 ± 4.4% ID/g at 24 h), resulting in 71% regression and 80% growth inhibition of SKOV-3 and A2780/CDDP tumors, respectively. Our findings demonstrate that NCP particles provide great potential for the codelivery of multiple chemotherapeutics for treating drug-resistant cancer.

Polymeric Micelle-Mediated Delivery of DNA-Targeting Organometallic Complexes for Resistant Ovarian Cancer Treatment.

Three half-sandwich iridium and ruthenium organometallic complexes with high cytotoxicity are synthesized, and their anticancer mechanisms are elucidated. The organometallic complexes can interact with DNA through coordination or intercalation, thereby inducing apoptosis and inhibiting proliferation of resistant cancer cells. The organometallic complexes are then incorporated into polymeric micelles through the polymer-metal coordination between poly(ethylene glycol)-b-poly(glutamic acid) [PEG-b-P(Glu)] and organometallic complexes to further enhance their anticancer effects as a result of the enhanced permeability and retention effect. The micelles with particle sizes of ≈60 nm are more efficiently internalized by cancer cells than the corresponding complexes, and selectively dissociate and release organometallic anticancer agents within late endosomes and lysosomes, thereby enhancing drug delivery to the nuclei of cancer cells and facilitating their interactions with DNA. Thus, the micelles display higher antitumor activity than the organometallic complexes alone with a lack of the systemic toxicity in a mouse xenograft model of cisplatin-resistant human ovarian cancer. These results suggest that the polymeric micelles carrying anticancer organometallic complexes provide a promising platform for the treatment of resistant ovarian cancer and other hard-to-treat solid tumors.

Multi-targeted inhibition of tumor growth and lung metastasis by redox-sensitive shell crosslinked micelles loading disulfiram.

Metastasis, the main cause of cancer related deaths, remains the greatest challenge in cancer treatment. Disulfiram (DSF), which has multi-targeted anti-tumor activity, was encapsulated into redox-sensitive shell crosslinked micelles to achieve intracellular targeted delivery and finally inhibit tumor growth and metastasis. The crosslinked micelles demonstrated good stability in circulation and specifically released DSF under a reductive environment that mimicked the intracellular conditions of tumor cells. As a result, the DSF-loaded redox-sensitive shell crosslinked micelles (DCMs) dramatically inhibited cell proliferation, induced cell apoptosis and suppressed cell invasion, as well as impairing tube formation of HMEC-1 cells. In addition, the DCMs could accumulate in tumor tissue and stay there for a long time, thereby causing significant inhibition of 4T1 tumor growth and marked prevention in lung metastasis of 4T1 tumors. These results suggested that DCMs could be a promising delivery system in inhibiting the growth and metastasis of breast cancer.

Low molecular weight polyethylenimine-graft-Tween 85 for effective gene delivery: synthesis and in vitro characteristics.

The development of safe and efficient gene delivery systems is still a challenge for successful gene therapy. In this work, low molecular weight polyethylenimine (PEI 2K) was modified by Tween 85, which bears three oleate chains. Tween 85 modified PEI 2K (TP) could condense DNA efficiently, and TP/DNA complexes (TPCs) showed high resistance to salt-induced aggregation and enzymatic degradation. In addition, TP did not show the obvious cytotoxicity. The introduction of Tween 85 led to a significant increase in the cellular uptake of complexes with higher transfection efficiency, which was strongly inhibited by the addition of free Tween 85 in MCF-7/ADR cells, but not in MCF-7 cells. These results indicated that TP could be a potentially safe and effective copolymer for gene delivery, and TPCs could be taken up mainly by Tween 85-mediated endocytosis in MCF-7/ADR cells.

Immunostimulatory nanomedicines synergize with checkpoint blockade immunotherapy to eradicate colorectal tumors.

Nanoparticles can potentially stimulate tumour microenvironments to elicit antitumour immunity. Herein, we demonstrate effective immunotherapy of colorectal cancer via systemic delivery of an immunostimulatory chemotherapeutic combination in nanoscale coordination polymer (NCP) core-shell particles. Oxaliplatin and dihydroartemesinin have contrasting physicochemical properties but strong synergy in reactive oxygen species (ROS) generation and anticancer activity. The combined ROS generation is harnessed for immune activation to synergize with an anti-PD-L1 antibody for the treatment of murine colorectal cancer tumours. The favourable biodistribution and tumour uptake of NCPs and the absence of peripheral neuropathy allow for repeated dosing to afford 100% tumour eradication. The involvement of innate and adaptive immune systems elicit strong and long lasting antitumour immunity which prevents tumour formation when cured mice are challenged with cancer cells. The intrinsically biodegradable, well tolerated, and systemically available immunostimulatory NCP promises to enter clinical testing as an immunotherapy against colorectal cancer.

Systemic miRNA delivery by nontoxic nanoscale coordination polymers limits epithelial-to-mesenchymal transition and suppresses liver metastases of colorectal cancer.

Though early detection and treatment of primary tumors has significantly improved in recent years, metastatic disease remains among the most significant challenges in cancer therapy. Cancer cells can disseminate before the primary tumor is detected to form micro or gross metastases, requiring toxic systemic therapies. To prevent and suppress metastases, we have developed a nontoxic, long-circulating nanoscale coordination polymer (NCP) protecting microRNA (miRNA) in circulation and releasing it in tumors. PtIV(en)2 [en = ethylenediamine] containing NCPs (PtEN) can release a nontoxic, kinetically inert PtII(en)2 compound and carbon dioxide which aids the endosomal escape of its miRNA cargo, miR-655-3p. Without the presence of the PtEN core, the miRNA showed cellular uptake but no effect. When transfected into human colorectal HCT116 cells by NCPs, this oligometastatic miRNA limited proliferation and epithelial-to-mesenchymal transition by preventing ß-catenin nuclear translocation and tumor cell invasion. Systemic administrations of PtEN/miR-655-3p sustained effective transfection to reduce liver colonization and tumor burden in a xenogenic hepatic metastatic model of HCT116 without any observable toxicity.

Core-shell nanoscale coordination polymers combine chemotherapy and photodynamic therapy to potentiate checkpoint blockade cancer immunotherapy.

Advanced colorectal cancer is one of the deadliest cancers, with a 5-year survival rate of only 12% for patients with the metastatic disease. Checkpoint inhibitors, such as the antibodies inhibiting the PD-1/PD-L1 axis, are among the most promising immunotherapies for patients with advanced colon cancer, but their durable response rate remains low. We herein report the use of immunogenic nanoparticles to augment the antitumour efficacy of PD-L1 antibody-mediated cancer immunotherapy. Nanoscale coordination polymer (NCP) core-shell nanoparticles carry oxaliplatin in the core and the photosensitizer pyropheophorbide-lipid conjugate (pyrolipid) in the shell (NCP@pyrolipid) for effective chemotherapy and photodynamic therapy (PDT). Synergy between oxaliplatin and pyrolipid-induced PDT kills tumour cells and provokes an immune response, resulting in calreticulin exposure on the cell surface, antitumour vaccination and an abscopal effect. When combined with anti-PD-L1 therapy, NCP@pyrolipid mediates regression of both light-irradiated primary tumours and non-irradiated distant tumours by inducing a strong tumour-specific immune response.

Effective delivery of p65 shRNA by optimized Tween 85-polyethyleneimine conjugate for inhibition of tumor growth and lymphatic metastasis.

To maximize the interference efficacy of pGPU6/Neo-p65 shRNA-expressing pDNA (p65 shRNA) and subsequently more effectively inhibit tumor growth and lymphatic metastasis through blocking the nuclear factor-kappa B (NF-κB) signaling pathway, seven Tween 85-polyethyleneimine (PEI) conjugates (TnPs, n=2, 3, 4, 5, 6, 7 and 8), which differed in the length of the polymethylene [-(CH2)n-] spacer between Tween 85 and PEI, were synthesized and investigated. The results showed that the transfection efficiency and cytotoxicity both increased with the spacer chain length. Then, TnPs with a [-(CH2)6-] spacer (T6P) were chosen to deliver p65 shRNA to a tumor and subsequently inhibit tumor growth and lymphatic metastasis. The T6P/p65 shRNA complex nanoparticles (T6Ns) could significantly down-regulate p65 expression in breast cancer cells, and consequently inhibit cell invasion and disrupt the tube formation. Most importantly, T6Ns accumulated greatly in tumor tissue, and as a result, significantly inhibited the growth and lymphatic metastasis of breast cancer xenograft. All these results indicated that the transfection efficacies of cationic amphiphiles could be significantly modulated by minor structural variations, and that T6P was promising for the effective delivery of p65 shRNA to knock down the expression of the key metastasis-driving genes and inhibit tumor growth and metastasis.

Smart pH-sensitive and temporal-controlled polymeric micelles for effective combination therapy of doxorubicin and disulfiram.

The combination of a chemotherapeutic drug with a multidrug resistance (MDR) modulator has emerged as a promising strategy for treating MDR cancer. To ensure two drugs could be simultaneously delivered to tumor region at the optimum ratio, and the MDR modulator could be released earlier and faster than the chemotherapeutic drug to inactivate P-glycoprotein (P-gp) and subsequently inhibit the pumping out of the chemotherapeutic drug, a smart pH-sensitive polymeric micelles system with high drug loading and precise drug ratio was designed and prepared by conjugating doxorubicin (DOX) to poly(styrene-co-maleic anhydride) (SMA) derivative with adipic dihydrazide (ADH) through a acid-cleavable hydrazone bond, and then encapsulating disulfiram (DSF), a P-gp inhibitor as well as an apoptosis inducer, into the micelles formed by the self-assembly of SMA-ADH-DOX (SAD) conjugate. The pH-sensitive polymeric micelles system enabled a temporal release of two drugs: encapsulated DSF was released fast to inhibit the activity of P-gp and restore cell apoptotic signaling pathways, while conjugated DOX was released in a sustained and pH-dependent manner and highly accumulated in drug resistant cells to exert therapeutic effect, due to the inactivation of P-gp by DSF. The smart co-delivery system was very effective in enhancing the cytotoxicity by increasing the intracellular accumulation of DOX and promoting the apoptotic response, and showed the most effective inhibitory effect on the growth of drug-resistant breast cancer xenografts as compared to other combinations of both drugs. In a word, this smart co-delivery system has significant promise for the clinical therapy of MDR cancer.

The inhibition of metastasis and growth of breast cancer by blocking the NF-κB signaling pathway using bioreducible PEI-based/p65 shRNA complex nanoparticles.

Metastasis is one of the greatest challenges in cancer treatment. In this study, a bioreducible polymer, Tween 85-s-s-polyethyleneimine 2K (TSP), was synthesized and used as a non-viral gene vector for p65 shRNA to block NF-κB signaling pathway, thereby inhibiting the growth and metastasis of breast cancer. The TSP/p65 shRNA complex nanoparticles (TSNs) could significantly down-regulate p65 expression in breast cancer cells due to the rapid degradation of TSP with prompt shRNA release, and consequently not only inhibit cell proliferation and invasion, but also induce cell apoptosis and disrupt the tube formation. Most importantly, TSNs showed high accumulation in tumor and almost completely inhibited the growth and metastasis of the breast cancer xenograft in nude mice induced by MDA-MB-435 cells. All these results indicated the promising of TSP as a non-viral gene vector to knock down p65 expression and inhibit the growth and metastasis of breast cancer.

Amphiphilic graft copolymer based on poly(styrene-co-maleic anhydride) with low molecular weight polyethylenimine for efficient gene delivery.

BACKGROUND AND METHODS: A new amphiphilic comb-shaped copolymer (SP) was synthesized by conjugating poly(styrene-co-maleic anhydride) with low molecular weight polyethyleneimine for gene delivery. Fourier transform infrared spectrum, (1)H nuclear magnetic resonance, and gel permeation chromatography were used to characterize the graft copolymer. RESULTS: The buffering capability of SP was similar to that of polyethyleneimine within the endosomal pH range. The copolymer could condense DNA effectively to form complexes with a positive charge (13-30 mV) and a small particle size (130-200 nm) at N/P ratios between 5 and 20, and protect DNA from degradation by DNase I. In addition, SP showed much lower cytotoxicity than polyethyleneimine 25,000. Importantly, the gene transfection activity and cellular uptake of SP-DNA complexes were all markedly higher than that of complexes of polyethyleneimine 25,000 and DNA in MCF-7 and MCF-7/ADR cell lines. CONCLUSION: This work highlights the promise of SP as a safe and efficient synthetic vector for DNA delivery.