Aromatic Ring-Fused Dipyrrins: Programmed [4 + 2]-Cycloaddition Pathway with Regio-selectivity upon Alkylamino-Substitution.

Sulfolenodipyrrins are employed as building blocks to concisely and efficiently construct aromatic rings (e.g., naphthoquinone, anthraquinone, fullerenes, and phthalimide) from fused dipyrrins by programmed [4 + 2]-cycloaddition reactions. Notably, alkylamino-substitution at the α-position not only enhances the reactivity of sulfolenodipyrrins but also results in the regio-selectivity of the cycloaddition reactions. Theoretical calculations in terms of frontier orbitals of dienes, energy of dienes, steric hindrance, and aromaticity have been conducted to understand the reason in depth. Additionally, the fusion of aromatic groups enables bathochromic absorption with up to ∼130 nm for the monoadducts and to ∼200 nm for the bis-adducts. The phthalimide annulation dipyrrin displays red emission, while the other mono- or bis-adducts do not, owing to the presence of typical acceptors such as quinone analogs or fullerene.

Design, synthesis and biological evaluation of 2-((4-sulfamoylphenyl)amino)-pyrrolo[2,3-d]pyrimidine derivatives as CDK inhibitors.

To explore the potential use of CDK inhibitors in pancreatic ductal adenocarcinoma (PDAC) therapy, a series of novel 2-((4-sulfamoylphenyl)amino)-pyrrolo[2,3-d]pyrimidine derivatives was designed, synthesised, and investigated for inhibition on both CDK kinase activity and cellular proliferation of pancreatic cancer. Most of new sulphonamide-containing derivatives demonstrated strong inhibitory activity on CDK9 and obvious anti-proliferative activity in cell culture. Moreover, two new compounds suppressed cell proliferation of multiple human pancreatic cancer cell lines. The most potent compound 2g inhibited cancer cell proliferation by blocking Rb phosphorylation and induced apoptosis via downregulation of CDK9 downstream proteins Mcl-1 and c-Myc in MIA PaCa-2 cells. CDK9 knockdown experiment suggests its anti-proliferative activity is mainly mediated by CDK9. Additionally, 2g displayed moderate tumour inhibition effect in AsPC-1 derived xenograft mice model. Altogether, this study provided a new start for further optimisation to develop potential CDK inhibitor candidates for PDAC treatment by alone or combination use.

Isolation, Structure Elucidation, and First Total Synthesis of Quinomycins K and L, Two New Octadepsipeptides from the Maowei Sea Mangrove-Derived Streptomyces sp. B475.

Mangrove actinomycetia have been proven to be one of the promising sources for discovering novel bioactive natural products. Quinomycins K (1) and L (2), two rare quinomycin-type octadepsipeptides without intra-peptide disulfide or thioacetal bridges, were investigated from the Maowei Sea mangrove-derived Streptomyces sp. B475. Their chemical structures, including the absolute configurations of their amino acids, were elucidated by a combination of NMR and tandem MS analysis, electronic circular dichroism (ECD) calculation, advanced Marfey's method, and further unequivocally confirmed by the first total synthesis. The two compounds displayed no potent antibacterial activity against 37 bacterial pathogens and had no significant cytotoxic activity against H460 lung cancer cells.

Loading Phlorins with Fullerenes by a [4 + 2]-Cycloaddition Reaction: Regulation of the Regioselectivity by Pyrrole Linkage Modes.

Sulfolenopyrrole-based normal and N-confused phlorins have been constructed to address the seldom touched phlorin functionalization and simultaneously explore the effect of the pyrrole linkage modes (αα, αß) on the [4 + 2] cycloaddition reaction. The common sulfolenophlorin 1 contains two sulfolenopyrroles with the same reactivity upon tautomerization and undergoes stepwise [4 + 2]-cycloaddition with fullerene to furnish monoadduct 1-C60 and bisadduct 1-2C60 with a total yield up to 76%. By contrast, the presence of the confused pyrrole in 2 fixes the π-system owing to the low tendency to tautomerize and enables the two sulfolenopyrroles to exhibit in different fashions (i.e., normal NH-type and imino-type). Notably, under milder conditions (120 °C), the monofullerenoadduct 2-C60 forms rapidly and has been isolated from the [4 + 2] cycloaddition reaction of 2 and fullerene as the predominant fraction, accompanied by a trace amount of bisadduct 2-2C60. Raising the temperature to 140 °C did not improve the yield of 2-2C60. The structural analysis of 2-C60 indicates the attachment of fullerene at the iminopyrrole part. The high regioselectivity in the [4 + 2] cycloaddition of the imino-type sulfolenopyrrole unit has been rationalized thermodynamically by the DFT calculation on the relative energy of the two diene intermediates.

Porphyrindiene-Based Tandem Diels-Alder Reaction for Preparing Low-Symmetry π-Extended Porphyrins with Push-Pull Skeletons.

Tandem Diels-Alder reactions of masked porphyrindienes (i.e., sulfolenoporphyrins) with benzoquinones and stilbenes, followed by aromatization, have been developed to load porphyrin with mixed annulation units (i.e., terphenyl and naphthoquinone), furnishing the low-symmetry π-extended porphyrins (DxAy) with push-pull skeletons. All low-symmetrical chromophores display panchromatic absorption spectra, which look like a spectral combination of symmetrical congeners (D4/A4) in a certain ratio. Among them, tD2A2 with trans-arrangement of push/pull units possesses the largest maximum centered at 766 nm with the onset around 900 nm. The fusion of the electron-deficient naphthoquinone moiety on the porphyrin core results in the approximately quantitative regulation of the Eox1 and HOMOs (i.e., 0.10-0.13 V increase for the Eox1 and 0.14-0.16 eV decrease for the HOMOs per naphthoquinone unit). In brief, this work provides a new way to construct low-symmetry π-extended porphyrins with tunable properties resorting to the ratios and locations of the annulated push-pull units.

Polysaccharides of Brassica rapa L. attenuate tumor growth via shifting macrophages to M1-like phenotype.

Macrophages are the major tumor-infiltrating leukocytes, and tumor-associated macrophages (TAM) play a critical role in cancer-related inflammation since they show alternative polarization to M1 (tumor-inhibited macrophages) or M2 (tumor-promoted macrophages) phenotype. Brassica rapa L. (B. rapa) has been clinically proven to have anti-tumor and immunity-enhancing activity, and the polysaccharides of B. rapa (BRP) have been reported to have an immunoregulatory effect on macrophages. In this study, we focus on macrophage polarization to investigate the mechanism of anti-tumor response of BRP in vivo and in vitro. We found that BRP improved the expression of M1 markers, including iNOS, COX-2, HLA-DR, CD11b and M1-related cytokines. The expression of M2 markers Arg-1, CD206 and CD163 induced by IL-4 were inhibited by BRP treatment, resulting in the inhibition of tumor growth both in vivo and in co-culture experiments in vitro. The activation of STAT signaling pathway were significantly regulated by BRP, which are important signals in TAM polarization. Overall, the results indicated that BRP has anti-tumor effect through mediating macrophage polarization.

Enhancing the Therapeutic Efficacy of Gefitinib in Human Non-Small-Cell Lung Cancer through Drug Combination.

The interaction between tumor cells and the tumor microenvironment (TME) significantly influences tumorigenesis, so TME-targeted therapy has attracted widespread attention. We have previously demonstrated that the combination of dipyridamole, bestatin, and dexamethasone (DBD mix, DBDx) is effective against heterogeneous human pancreatic cancer and hepatocellular carcinoma in mouse xenograft models. To further expand the therapeutic potential of this drug combination, herein, we investigated the antitumor efficacy and the underlying mechanism of DBDx and the combination of DBDx and gefitinib in different mouse xenograft models of human non-small-cell lung cancer (NSCLC). Three human cancer cell lines H460, PG, and A431 were used to determine the apoptosis and growth inhibition induced by DBDx, gefitinib, and their combinations. Changes in epidermal growth factor receptor (EGFR) signaling pathway-related proteins were analyzed following treatment using western blotting. In vitro, DBDx strongly inhibited the proliferation of tumor cells, whereas the combined treatment exhibited a significant synergistic effect. Compared with DBDx, the combination treatment further induced apoptosis and downregulated the expression of molecules associated with EGFR signaling pathway. In vivo, compared with DBDx alone, the combination treatment distinctly inhibited tumor growth in mouse xenograft models of human NSCLC. Overall, our results indicate that the combination of DBDx and gefitinib in the treatment of human NSCLC is very promising, which warrants further translational studies.

Metabolomics Tools Assisting Classic Screening Methods in Discovering New Antibiotics from Mangrove Actinomycetia in Leizhou Peninsula.

Mangrove actinomycetia are considered one of the promising sources for discovering novel biologically active compounds. Traditional bioactivity- and/or taxonomy-based methods are inefficient and usually result in the re-discovery of known metabolites. Thus, improving selection efficiency among strain candidates is of interest especially in the early stage of the antibiotic discovery program. In this study, an integrated strategy of combining phylogenetic data and bioactivity tests with a metabolomics-based dereplication approach was applied to fast track the selection process. A total of 521 actinomycetial strains affiliated to 40 genera in 23 families were isolated from 13 different mangrove soil samples by the culture-dependent method. A total of 179 strains affiliated to 40 different genera with a unique colony morphology were selected to evaluate antibacterial activity against 12 indicator bacteria. Of the 179 tested isolates, 47 showed activities against at least one of the tested pathogens. Analysis of 23 out of 47 active isolates using UPLC-HRMS-PCA revealed six outliers. Further analysis using the OPLS-DA model identified five compounds from two outliers contributing to the bioactivity against drug-sensitive A. baumannii. Molecular networking was used to determine the relationship of significant metabolites in six outliers and to find their potentially new congeners. Finally, two Streptomyces strains (M22, H37) producing potentially new compounds were rapidly prioritized on the basis of their distinct chemistry profiles, dereplication results, and antibacterial activities, as well as taxonomical information. Two new trioxacarcins with keto-reduced trioxacarcinose B, gutingimycin B (16) and trioxacarcin G (20), together with known gutingimycin (12), were isolated from the scale-up fermentation broth of Streptomyces sp. M22. Our study demonstrated that metabolomics tools could greatly assist classic antibiotic discovery methods in strain prioritization to improve efficiency in discovering novel antibiotics from those highly productive and rich diversity ecosystems.

Vegetation successions of coastal wetlands in southern Laizhou Bay, Bohai Sea, northern China, influenced by the changes in relative surface elevation and soil salinity.

Vegetation successions of coastal wetlands were influenced by the changes in relative surface elevation and soil salinity. In this study, the vegetation successions of coastal wetlands in southern Laizhou Bay and the factors influencing the successions were investigated by quadrat survey. The changes of relative surface elevation and soil salinity in coastal wetlands of the study region were caused by climate change, sea-level rise, coastal erosion, sedimentation, neotectonism, storm surge, seawater intrusion, invasion of Spatina alterniflora, and utilization of underground brine. The changes led to the regressive vegetation succession of coastal wetlands without the protection of sea embankment and the progressive vegetation succession of coastal wetlands with the protection of sea embankment. The invasion of S. alterniflora resulted in the regressive vegetation succession of wetlands in the riparian zone. The successions weakened the coastal wetlands' ecological capacities of carbon sequestration, pollutant purification, and resisting marine disasters, decreasing their species diversity. Some measures were proposed to resist the adverse impact of successions, such as introducing passenger water, storing water in flood season, digging 200 hm2 of ponds, and planting Salix matsudana and Tamarix chinensis around the ponds.

The novel quinolizidine derivate IMB-HDC inhibits STAT5a phosphorylation at 694 and 780 and promotes DNA breakage and cell apoptosis via blocking STAT5a nuclear translocation.

Sophoridine is a quinolizidine natural product and the exploration of its derivatives has been carried out, and the potent anticancer compound IMB-HDC was acquired. Although previous studies have revealed that some sophoridine derivatives could induce DNA breakage, the underlying mechanisms of inhibition of DNA damage repair (ATR inactivation) and the apoptosis independent of p53, have not been elucidated. Our research reveals a novel DNA response mechanism different from general DNA-damaging agents, and that sophoridine derivate inhibits the phosphorylation of Tyr694 and Ser780 of STAT5a to induce the lessened shuttle from the cytoplasm to the nucleus, and leads to the decreased nuclear STAT5a and subsequently inhibits the expression of STAT5a target gene RAD51 that contributes to the checkpoint activation, thus inhibiting ATR activation. Meanwhile, IMB-HDC that induced the diminished expression of STAT5a target gene contributes to proliferation and leads to apoptosis. More importantly, we give the first evidence that promoting the effect of Tyr694 phosphorylation on nuclear location and subsequent STAT5a target gene transcription depends on Ser780 increased or unchanged phosphorylation and was not correlated with Ser726 phosphorylation.

The ionophore antibiotic gramicidin A inhibits pancreatic cancer stem cells associated with CD47 down-regulation.

BACKGROUND: Pancreatic cancer stem cells (CSCs), a special population of cells, renew themselves infinitely and resist to various treatment. Gramicidin A (GrA), an ionophore antibiotic derived from microorganism, can form channels across the cell membrane and disrupt cellular ionic homeostasis, leading to cell dysfunction and death. As reported, the ionophore antibiotic salinomycin (Sal) has been proved to kill CSCs effectively. Whether GrA owns the potential as a therapeutic drug for CSCs still remains unknown. This study investigated the effect of GrA on pancreatic CSCs and the mechanism. METHODS: Tumorsphere formation assay was performed to assess pancreatic CSCs self-renewal potential. In vitro hemolysis assay was determined to test the borderline concentration of GrA. CCK-8 assay was used to detect pancreatic cancer cell proliferation capability. Flow cytometry was performed to detect cell apoptosis and mitochondrial membrane potential. Scanning and transmission electron microscopy was used to observe ultrastructural morphological changes on cell membrane surface and mitochondria, respectively. Western blot analysis was used to determine relative protein expression levels. Immunofluorescence staining was performed to observe CD47 re-distribution. RESULTS: GrA at 0.05 µM caused tumorspheres disintegration and decrease in number of pancreatic cancer BxPC-3 and MIA PaCa-2 cells. GrA and Sal both inhibited cancer cell proliferation. The IC50 values of GrA and Sal for BxPC-3 cells were 0.025 µM and 0.363 µM; while for MIA PaCa-2 cells were 0.032 µM and 0.163 µM, respectively. Compared on equal concentrations, the efficacy of GrA was stronger than that of Sal. GrA at 0.1 µM or lower did not cause hemolysis. GrA induced ultrastructural changes, such as the decrease of microvilli-like protrusions on cell surface membrane and the swelling of mitochondria. GrA down-regulated the expression levels of CD133, CD44, and CD47; in addition, CD47 re-distribution was observed on cell surface. Moreover, GrA showed synergism with gemcitabine in suppressing cancer cell proliferation. CONCLUSIONS: The study found that GrA was highly active against pancreatic CSCs. It indicates that GrA exerts inhibitory effects against pancreatic CSCs associated with CD47 down-regulation, implying that GrA might play a positive role in modulating the interaction between macrophages and tumor cells.

Yellow Dioxobilin-Type Tetrapyrroles from Chlorophyll Breakdown in Higher Plants-A New Class of Colored Phyllobilins.

In senescent leaves chlorophyll (Chl) catabolites typically accumulate as colorless tetrapyrroles, classified as formyloxobilin-type (or type-I) or dioxobilin-type (type-II) phyllobilins (PBs). Yellow type-I Chl catabolites (YCCs) also occur in some senescent leaves, in which they are generated by oxidation of colorless type-I PBs. A yellow type-II PB was recently proposed to occur in extracts of fall leaves of grapevine (Vitis vinifera), tentatively identified by its mass and UV/Vis absorption characteristics. Here, the first synthesis of a yellow type-II Chl catabolite (DYCC) from its presumed natural colorless type-II precursor is reported. A homogenate of a Spatiphyllum wallisii leaf was used as "green" means of effective and selective oxidation. The synthetic DYCC was fully characterized and identified with the yellow grapevine leaf pigment. As related yellow type-I PBs do, the DYCC functions as a reversible photoswitch by undergoing selective photo-induced Z/E isomerization of its C15=C16 bond.

Depressive Symptoms in Elderly Chinese Primary Care Patients: Prevalence and Sociodemographic and Clinical Correlates.

OBJECTIVE: To estimate the prevalence of depressive symptoms (depression thereafter) and to identify the sociodemographic and clinical correlates of depression in a sample of elderly patients treated in the primary care setting in Wuhan, China. BACKGROUND: Primary care is an opportune setting for the management of late-life depression in China, but there have been no representative studies on the clinical epidemiology of depression in elderly Chinese primary care patients. METHODS: In total, 752 elderly patients (≥ 65 years) were consecutively recruited from 13 primary care centers in Wuhan, China, and interviewed with a standardized questionnaire. Depression was assessed with the 15-item Geriatric Depression Scale (GDS-15). RESULTS: Of the elderly Chinese primary care patients, 30.6% had depression (GDS-15 ≥ 5). Correlates of depression were an education level of primary school or less (odds ratio [OR]: 1.94, 95% confidence interval [CI]: 1.36-2.77, P < .001), poor financial status (OR: 2.19, 95% CI: 1.16-4.15, P = .016), lack of an exercise habit (OR: 1.40, 95% CI: 1.06-1.74, P = .023), 2 or more chronic medical conditions (OR: 1.90, 95% CI: 1.34-2.69, P < .001), and loneliness (OR: 3.53, 95% CI: 2.46-5.08, P < .001). CONCLUSIONS: Depression is prevalent among elderly Chinese primary care patients, indicating that elderly patients treated in primary care have a high level of need for mental health services in China. There is an urgent need to integrate mental health services into primary health care.

Introduction of an isoxazoline unit to the ß-position of porphyrin via regioselective 1,3-dipolar cycloaddition reaction.

Isoxazoline-linked porphyrins have been synthesized by a regioselective 1,3-dipolar cycloaddition reaction between vinylporphyrin 2 and nitrile oxides. The steric interaction directed the reaction trajectory, in which only the product with a link between the 5-position of the isoxazoline and the ß-position of porphyrin was observed. The isoxazoline-porphyrins 3a,b have been characterized by absorption, emission, 1H NMR and mass spectra. Later, the crystal structure of 3a was obtained and confirmed the basic features of the NMR-derived structure. Furthermore, a pair of enantiomers of 3a presented in the crystal, which formed a dimeric complex through intermolecular coordination between the Zn2+ center and the carbonyl group of the second molecule.

Novel Types of Hypermodified Fluorescent Phyllobilins from Breakdown of Chlorophyll in Senescent Leaves of Grapevine (Vitis vinifera).

The tetrapyrrolic chlorophyll catabolites (or phyllobilins, PBs) were analyzed in yellow fall leaves of the grape Chardonnay, a common Vitis vinifera white wine cultivar. The major fractions in leaf extracts of V. vinifera, tentatively assigned to PBs, were isolated and their structures elucidated. The dominant fraction is a dioxobilin-type non-fluorescent Chl-catabolite of a previously observed type. Two less polar fluorescent PBs were characterized as a novel dioxobilin-type fluorescent Chl-catabolite with a bicyclo-1',6'-glycosyl architecture, and its new fluorescent formyloxobilin-type analogue. The discovery of persistent hypermodified fluorescent PBs with the architecture of bicyclo-[17.3.1]-PBs (bcPBs), suggests the activity of an unknown enzyme that forges the 20-membered macroring at the tetrapyrrolic core of a fluorescent PB. bcPBs may play specific physiological roles in grapevine plants and represent endogenous anti-infective agents, as found similarly for other organic bicyclo-[n.3.1]-1',6'-glycosyl derivatives.

A Macropinocytosis-Intensifying Albumin Domain-Based scFv Antibody and Its Conjugate Directed against K-Ras Mutant Pancreatic Cancer.

Enhanced macropinocytosis has been found in K-Ras mutant pancreatic cancer cells, through which albumin can massively enter into the K-Ras-driven cancer cells, suggesting its role in serving as a macropinocytosis-intensifying drug delivery carrier. In the present study, a novel recombinant protein Fv-LDP-D3 and its reconstituted analogue Fv-LDP-D3-AE were designed and prepared. Fv is the fragment of an anti-EGFR antibody, D3 is the domain III of human serum albumin (HSA), LDP is the apoprotein of the antitumor antibiotic lidamycin (LDM), and AE is an extremely cytotoxic enediyne chromophore derived from LDM. As shown, the recombinant protein Fv-LDP-D3 presented intensive and selective binding capacity to pancreatic cancer cells and inhibited cell proliferation by blocking EGFR signaling. Moreover, Fv-LDP-D3 showed prominent tumor imaging in pancreatic carcinoma xenograft. The reconstituted, enediyne-integrated analogue Fv-LDP-D3-AE displayed highly potent cytotoxicity to pancreatic cancer cells through apoptosis induction and G2/M arrest. Fv-LDP-D3 and Fv-LDP-D3-AE markedly inhibited the tumor growth of the pancreatic carcinoma AsPC-1 xenograft. Study results indicated that the novel recombinant protein displays both EGFR-targeting and macropinocytosis-intensifying attributes, presenting a new format of scFv antibody that integrates with albumin domain III. It might be a feasible strategy to develop targeted drugs for K-Ras mutant pancreatic cancer.

EGFR-targeting, ß-defensin-tailored fusion protein exhibits high therapeutic efficacy against EGFR-expressed human carcinoma via mitochondria-mediated apoptosis.

Defensins play an essential role in innate immunity. In this study, a novel recombinant ß-defensin that targets the epidermal growth factor receptor (EGFR) was designed and prepared. The EGFR-targeting ß-defensin consists of an EGF-derived oligopeptide (Ec), a ß-defensin-1 peptide (hBD1) and a lidamycin-derived apoprotein (LDP), which serves as the "scaffold" for the fusion protein (Ec-LDP-hBD1). Ec-LDP-hBD1 effectively bound to EGFR highly expressed human epidermoid carcinoma A431 cells. The cytotoxicity of Ec-LDP-hBD1 to EGFR highly expressed A431 cells was more potent than that to EGFR low-expressed human lung carcinoma A549 and H460 cells (the IC50 values in A431, A549, and H460 cells were 1.8 ± 0.55, 11.9 ± 0.51, and 5.19 ± 1.21 µmol/L, respectively); in addition, the cytotoxicity of Ec-LDP-hBD1 was much stronger than that of Ec-LDP and hBD1. Moreover, Ec-LDP-hBD1 suppressed cancer cell proliferation and induced mitochondria-mediated apoptosis. Its in vivo anticancer action was evaluated in athymic mice with A431 and H460 xenografts. The mice were administered Ec-LDP-hBD1 (5, 10 mg/kg, i.v.) two times with a weekly interval. Administration of Ec-LDP-hBD1 markedly inhibited the tumor growth without significant body weight changes. The in vivo imaging further revealed that Ec-LDP-hBD1 had a tumor-specific distribution with a clear image of localization. The results demonstrate that the novel recombinant EGFR-targeting ß-defensin Ec-LDP-hBD1 displays both selectivity and enhanced cytotoxicity against relevant cancer cells by inducing mitochondria-mediated apoptosis and exhibits high therapeutic efficacy against the EGFR-expressed carcinoma xenograft. This novel format of ß-defensin, which induces mitochondrial-mediated apoptosis, may play an active role in EGFR-targeting cancer therapy.

Loneliness in Chinese older adults in primary care: prevalence and correlates.

BACKGROUND: Loneliness is a significant public health concern among older adults (OA) given its association with a wide range of adverse health outcomes. Primary care is an opportune setting to manage loneliness. However, the epidemiology of loneliness in Chinese OA treated in primary care remains unclear. The present study investigated the prevalence and correlates of loneliness in OA treated in Chinese primary care. METHODS: A total of 744 OA patients (65+ years) were consecutively recruited from 13 primary care clinics in Wuhan, China, and interviewed with a standardized questionnaire, concerning sociodemographic characteristics, lifestyle, relationships with family and others, physical health, and sensory impairments. Consistent with prior research on the construct, loneliness was measured with a single-item self-report question. Logistic regression was used to identify correlates of loneliness. RESULTS: Of primary care OA patients, 26.2% endorsed loneliness. Factors significantly and independently associated with loneliness included 75+ age group (odds ratio [OR]: 1.61, 95% confidence interval [CI]: 1.07, 2.44, P: 0.023), being illiterate (OR: 2.07, 95%CI: 1.26, 3.42, P: 0.004), unmarried (OR: 2.30, 95%CI: 1.40, 3.78, P: 0.001), living alone (OR: 4.37, 95%CI: 2.27, 8.41, P < 0.001), having fair and poor family (OR: 2.44, 95%CI: 1.48, 4.00, P < 0.001) and non-family relationships (OR: 1.75, 95%CI: 1.10, 2.78, P: 0.019), and ≥2 chronic medical conditions (OR: 2.91, 95%CI: 1.22, 6.95, P: 0.016). CONCLUSIONS: Loneliness is common in Chinese primary care OA. The high prevalence and many negative health consequences of loneliness for OA highlight the importance of routine screening, assessment, and interventions to reduce loneliness in the primary health-care setting.

A CD13-targeting peptide integrated protein inhibits human liver cancer growth by killing cancer stem cells and suppressing angiogenesis.

CD13 is a marker of angiogenic endothelial cells, and recently it is proved to be a biomarker of human liver cancer stem cells (CSCs). Herein, the therapeutic effects of NGR-LDP-AE, a fusion protein composed of CD13-targeting peptide NGR and antitumor antibiotic lidamycin, on human liver cancer and its mechanism were studied. Western blot and immunofluorescence assay demonstrated that CD13 (WM15 epitope) was expressed in both human liver cancer cell lines and vascular endothelial cells, while absent in normal liver cells. MTT assay showed that NGR-LDP-AE displayed potent cytotoxicity to cultured tumor cell lines with IC50 values at low nanomolar level. NGR-LDP-AE inhibited tumorsphere formation of liver cancer cells, and the IC50 values were much lower than that in MTT assay, indicating selectively killing of CSCs. In endothelial tube formation assay, NGR-LDP-AE at low cytotoxic dose significantly inhibited the formation of intact tube networks. Animal experiment demonstrated that NGR-LDP-AE inhibited the growth of human liver cancer xenograft. Immunohistochemical analysis showed that NGR-LDP-AE induced the down-regulation of CD13. In vitro experiment using cultured tumor cells also confirmed this result. NGR-LDP-AE activated both apoptotic and autophagic pathways in cultured tumor cells, while the induced autophagy protected cells from death. Conclusively, NGR-LDP-AE exerts its antitumor activity via killing liver CSCs and inhibiting angiogenesis. With one targeting motif, NGR-LDP-AE acts on both liver CSCs and angiogenic endothelial cells. It is a promising dual targeting fusion protein for liver cancer therapy, especially for advanced or relapsed cancers.

Recombinant EGFR/MMP-2 bi-targeted fusion protein markedly binding to non-small-cell lung carcinoma and exerting potent therapeutic efficacy.

Overexpression of EGFR and MMP-2 plays an essential role in the initiation and progression of non-small-cell lung carcinoma (NSCLC). In this study, a novel format of EGFR/MMP-2 bi-targeted fusion protein Ec-LDP-TIMP2 and its enediyne-integrated analogue Ec-LDP(AE)-TIMP2 have been prepared by genetic engineering and molecular reconstitution. The Ec-LDP(AE)-TIMP2 comprises endogenous inhibitor of matrix metalloproteinase 2 (TIMP2), EGF-derived oligopeptide (Ec), lidamycin apoprotein (LDP), and the extremely potent cytotoxic enediyne (AE). By tissue microarray, Ec-LDP-TIMP2 showed high binding intensity and selectivity to human NSCLC specimens as compared with the matched non-cancerous tissues. By in vivo imaging, Ec-LDP-TIMP2 displayed prominent tumor-specific distribution in human NSCLC H460 xenograft. Particularly, Ec-LDP(AE)-TIMP2 inhibited tumor growth of H460 xenograft in athymic mice more striking. At doses of 0.2 and 0.4mg/kg, Ec-LDP(AE)-TIMP2 suppressed tumor growth by 74% and 89%, respectively. No histopathological changes were found in various organs of treated animals, suggesting that the effective dosage was tolerated. In summary, the ligand-based and enediyne-integrated fusion protein displaying extremely potent cytotoxicity might be highly effective for NSCLC therapy and useful as a carrier for drug delivery.