Antimicrobial and Antiviral Nanofibers Halt Co-Infection Spread via Nuclease-Mimicry and Photocatalysis.

The escalating spread of drug-resistant bacteria and viruses is a grave concern for global health. Nucleic acids dominate the drug-resistance and transmission of pathogenic microbes. Here, imidazolium-type poly(ionic liquid)/porphyrin (PIL-P) based electrospun nanofibrous membrane and its cerium (IV) ion complex (PIL-P-Ce) are developed. The obtained PIL-P-Ce membrane exhibits high and stable efficiency in eradicating various microorganisms (bacteria, fungi, and viruses) and decomposing microbial antibiotic resistance genes and viral nucleic acids under light. The nuclease-mimetic and photocatalytic mechanisms of the PIL-P-Ce are elucidated. Co-infection wound models in mice with methicillin-resistant S. aureus and hepatitis B virus demonstrate that PIL-P-Ce integrate the triple effects of cationic polymer, photocatalysis, and nuclease-mimetic activities. As revealed by proteomic analysis, PIL-P-Ce shows minimal phototoxicity to normal tissues. Hence, PIL-P-Ce has potential as a "green" wound dressing to curb the spread of drug-resistant bacteria and viruses in clinical settings.

Functional metal-phenolic cortical cytoskeleton for artificial cells.

Cortex-like cytoskeleton, a thin layer of cross-linked cytoplasmic proteins underlying the cell membrane, plays an essential role in modulating membrane behavior and cell surface properties. However, bottom-up construction of functional cortex-like cytoskeleton in artificial cells remains a challenge. Here, we present metal-phenolic networks as artificial cortical cytoskeletons in liposome-based artificial cells. The metal-phenolic cytoskeleton-reinforced artificial cells exhibit long-term stability, enhanced resistance to a variety of harsh environments, tunable permeability, and well-controlled morphologies. We anticipate that our stable artificial cell models will stride forward to practical applications of liposome-based microsystem.

Photoswitchable Molecular Communication between Programmable DNA-Based Artificial Membraneless Organelles.

Spatiotemporal organization of distinct biological processes in cytomimetic compartments is a crucial step towards engineering functional artificial cells. Mimicking controlled bi-directional molecular communication inside artificial cells remains a considerable challenge. Here we present photoswitchable molecular transport between programmable membraneless organelle-like DNA coacervates in a synthetic microcompartment. We use droplet microfluidics to fabricate membraneless non-fusing DNA coacervates by liquid-liquid phase separation in a water-in-oil droplet, and employ the interior DNA coacervates as artificial organelles to imitate intracellular communication via photo-regulated uni- and bi-directional transfer of biomolecules. Our results highlight a promising new route to assembly of multicompartment artificial cells with functional networks.

Efficient disruption of bcr-abl gene by CRISPR RNA-guided FokI nucleases depresses the oncogenesis of chronic myeloid leukemia cells.

BACKGROUND: The bcr-abl fusion gene encodes BCR-ABL oncoprotein and plays a crucial role in the leukemogenesis of chronic myeloid leukemia (CML). Current therapeutic methods have limited treatment effect on CML patients with drug resistance or disease relapse. Therefore, novel therapeutic strategy for CML is essential to be explored and the CRISPR RNA-guided FokI nucleases (RFNs) meet the merits of variable target sites and specificity of cleavage enabled its suitability for gene editing of CML. The RFNs provide us a new therapeutic direction to obliterate this disease. METHODS: Guide RNA (gRNA) expression plasmids were constructed by molecular cloning technique. The modification rate of RFNs on bcr-abl was detected via NotI restriction enzyme digestion and T7 endonuclease 1 (T7E1) assay. The expression of BCR-ABL and its downstream signaling molecules were determined by western blotting. The effects of RFNs on cell proliferation and apoptosis of CML cell lines and CML stem/progenitor cells were evaluated by CCK-8 assay and flow cytometry. In addition, murine xenograft model was adopted to evaluate the capacity of RFNs in attenuating the tumorigenic ability of bcr-abl. RESULTS: The RFNs efficiently disrupted bcr-abl and prematurely terminated its translation. The destruction of bcr-abl gene suppressed cell proliferation and induced cell apoptosis in CML lines and in CML stem/progenitor cells. Moreover, the RFNs significantly impaired the leukemogenic capacity of CML cells in xenograft model. CONCLUSION: These results illustrate that the RFNs can target to disrupt bcr-abl gene and may provide a new therapeutic option for CML patients affiliated by drug resistance or disease relapse.

Colorimetric determination of BCR/ABL fusion genes using a nanocomposite consisting of Au@Pt nanoparticles covered with a PAMAM dendrimer and acting as a peroxidase mimic.

A colorimetric assay is described for the detection of BCR/ABL fusion genes. Polyamidoamine (PAMAM) dendrimers were placed on peroxidase (POx) mimicking Au@Pt nanoparticles to form a nanocomposite of type Au@Pt-PAMAM. Capture DNA probe is a designed nucleic acid strand that specifically binds target DNA to the surface of the electrode. The capture probe was attached to magnetic beads via biotin and avidin interaction. The hairpin structure of the capture probe can only be opened by the complementary BCR/ABL DNA. This results in a highly specific assay. The POx-mimicking property of the Au@Pt-PAMAM causes the formation of a blue dye by reaction of H2O2 and 3,3,3',3'-tetramethylbenzidine (TMB) which is measured by a microplate reader. Under optimum conditions, the absorbance increases linearly the 1 pM to 100 nM BCR/ABL concentration range, and the detection limit is as low as 190 fM. The method is highly selective and was successfully applied to the determination of fusion genes in spiked real samples. Conceivably, it possesses a large potential in clinical testing of patients suffering from chronic myeloid leukemia. Graphical abstract Au@PtNP, an efficient catalyst, is bound with polyamidoamine (PAMAM) dendrimer to amplify the colorimetric signal. With the introduction of streptavidin-magnetic beads to remove non-specific signals, a novel colorimetric sensor is constructed to detect BCR/ABL fusion genes.

Induction of apoptosis in imatinib sensitive and resistant chronic myeloid leukemia cells by efficient disruption of bcr-abl oncogene with zinc finger nucleases.

BACKGROUND: The bcr-abl fusion gene is the pathological origin of chronic myeloid leukemia (CML) and plays a critical role in the resistance of imatinib. Thus, bcr-abl disruption-based novel therapeutic strategy may warrant exploration. In our study, we were surprised to find that the characteristics of bcr-abl sequences met the design requirements of zinc finger nucleases (ZFNs). METHODS: We constructed the ZFNs targeting bcr-abl with high specificity through simple modular assembly approach. Western blotting was conducted to detect the expression of BCR-ABL and phosphorylation of its downstream STAT5, ERK and CRKL in CML cells. CCK8 assay, colony-forming assay and flow cytometry (FCM) were used to evaluate the effect of the ZFNs on the viablity and apoptosis of CML cells and CML CD34+ cells. Moreover, mice model was used to determine the ability of ZFNs in disrupting the leukemogenesis of bcr-abl in vivo. RESULTS: The ZFNs skillfully mediated 8-base NotI enzyme cutting site addition in bcr-abl gene of imatinib sensitive and resistant CML cells by homology-directed repair (HDR), which led to a stop codon and terminated the translation of BCR-ABL protein. As expected, the disruption of bcr-abl gene induced cell apoptosis and inhibited cell proliferation. Notably, we obtained similar result in CD34+ cells from CML patients. Moreover, the ZFNs significantly reduced the oncogenicity of CML cells in mice. CONCLUSION: These results reveal that the bcr-abl gene disruption based on ZFNs may provide a treatment choice for imatinib resistant or intolerant CML patients.

Inhibition of YAP suppresses CML cell proliferation and enhances efficacy of imatinib in vitro and in vivo.

BACKGROUND: Yes-associated protein (YAP), an essential component of Hippo pathway, was identified as an oncoprotein which participated in the progression of various malignancies. However, its role in chronic myeloid leukemia (CML) remains to be further clarified. METHODS: The expression of YAP in CML cells was determined by western blotting. Next, the effects of YAP knockdown and YAP inhibitor on CML cells were evaluated by MTT assay, flow cytometry (FCM) and Wright's staining. Moreover, K562 induced mice model was employed to further investigate the role of YAP in vivo. RESULTS: YAP was overexpressed in CML cells. Knockdown of YAP by si-RNA or inhibition the function of YAP using verteporfin (VP) not only inhibited the proliferation, induced the apoptosis of CML cells but also reduced the expression of YAP target genes c-myc and survivin. Additionally, VP enhanced the efficacy of imatinib (IM) in vitro and suppressed leukemogenesis in vivo. CONCLUSION: Our results indicate that YAP may play an important role in the proliferation and leukemogenesis of CML cells. Genetic or pharmacological inhibition of YAP provides a novel treatment strategy for CML.