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Cancer is the leading cause of death worldwide. Drugs play a pivotal role in cancer treatment, but the complex biological processes of cancer cells seriously limit the efficacy of various anticancer drugs. Autophagy, a self-degradative system that maintains cellular homeostasis, universally operates under normal and stress conditions in cancer cells. The roles of autophagy in cancer treatment are still controversial because both stimulation and inhibition of autophagy have been reported to enhance the effects of anticancer drugs. Thus, the important question arises as to whether we should try to strengthen or suppress autophagy during cancer therapy. Currently, autophagy can be divided into four main forms according to its different functions during cancer treatment: cytoprotective (cell survival), cytotoxic (cell death), cytostatic (growth arrest), and nonprotective (no contribution to cell death or survival). In addition, various cell death modes, such as apoptosis, necrosis, ferroptosis, senescence, and mitotic catastrophe, all contribute to the anticancer effects of drugs. The interaction between autophagy and these cell death modes is complex and can lead to anticancer drugs having different or even completely opposite effects on treatment. Therefore, it is important to understand the underlying contexts in which autophagy inhibition or activation will be beneficial or detrimental. That is, appropriate therapeutic strategies should be adopted in light of the different functions of autophagy. This review provides an overview of recent insights into the evolving relationship between autophagy and cancer treatment.

The poor stability of peptides against trypsin largely limits their development as potential antibacterial agents. Here, to obtain a peptide with increased trypsin stability and potent antibacterial activity, TICbf-14 derived from the cationic peptide Cbf-14 was designed by the addition of disulfide-bridged hendecapeptide (CWTKSIPPKPC) loop. Subsequently, the trypsin stability and antimicrobial and antibiofilm activities of this peptide were evaluated. The possible mechanisms underlying its mode of action were also clarified. The results showed that TICbf-14 exhibited elevated trypsin inhibitory activity and effectively mitigated lung histopathological damage in bacteria-infected mice by reducing the bacterial counts, further inhibiting the systemic dissemination of bacteria and host inflammation. Additionally, TICbf-14 significantly repressed bacterial swimming motility and notably inhibited biofilm formation. Considering the mode of action, we observed that TICbf-14 exhibited a potent membrane-disruptive mechanism, which was attributable to its destructive effect on ionic bridges between divalent cations and LPS of the bacterial membrane. Overall, TICbf-14, a bifunctional peptide with both antimicrobial and trypsin inhibitory activity, is highly likely to become an ideal candidate for drug development against bacteria.

Antimicrobial peptides (AMPs), which hold tremendous promise in overcoming the emergence of drug resistance, are limited in wide clinical applications due to their instability, especially against trypsin. Herein, we designed six peptide mutants based on the cathelicidin CATHPb2, followed by screening. Pb2-1, which showed the best activity against drug-resistant bacteria among these mutants, was selected to be combined with the trypsin inhibitory loop ORB-C to obtain two hybrid peptides: PCL-1 and Pb2-1TI. Notably, both of the hybrid peptides exhibited a remarkable enhancement in trypsin resistance compared with Pb2-1. The tests showed that PCL-1 displayed broad-spectrum antimicrobial activity that was superior to that of Pb2-1TI. In addition, PCL-1 had relatively lower cytotoxicity than Pb2-1TI towards the L02 and HaCaT cell lines and negligible hemolysis, as well as tolerance to high concentrations of salt, extreme pH, and temperature variations. In vivo, PCL-1 effectively improved the survival rate of mice that were systemically infected with drug-resistant Escherichia coli through efficient bacterial clearance from the blood and organs. With regard to mode of action, PCL-1 damaged the integrity of the bacterial cell membrane and attached to the membrane surface while bound to bacterial genomic DNA to eventually kill the bacteria. Altogether, the trypsin-resistant peptide PCL-1 is expected to be a candidate for the clinical treatment of bacterial infections.

Infections induced by fungi, especially the drug-resistant fungi, are difficult clinical problems. Conventional antifungal treatment is effective but due to resistance, treatment failure, and treatment-related toxicity, there is a need for new antifungal drugs. In this study, SA-2 (YYRRLLRVLRRRW) was derived from Cystatin-SA, a saliva protein with a molecular weight of 14 kDa. Meanwhile, the structure-activity of SA-2 and its mutants was also studied. We detected the antimicrobial activity and cytotoxicity of SA-2 and found that SA-2 had a low cytotoxicity toward mammalian cells but a good inhibitory effect on Candida albicans (C. albicans) and Cryptococcus neoformans (C. neoformans), with MIC values of 16-64 μg/mL and 8-32 μg/mL, respectively. Interestingly, SA-2 effectively killed fluconazole-resistant C. neoformans and C. albicans within 12 h. This antifungal activity against fluconazole-resistant fungi was comparable to that of amphotericin B. In addition, the C. neoformans-infected mice model was established to evaluate the anti-infective activity of SA-2 in vivo. Results showed that SA-2 significantly reduced the counts of fungi in lung and brain tissues to protect fluconazole-resistant C. neoformans-infected mice from death without changing mice body weights. Moreover, the dramatically increased pro-inflammatory cytokines TNF-α, IL-6 and IL-1β induced by intranasal infection of C. neoformans could be obviously declined due to the treatment of SA-2, which may be attributed to the elimination of C. neoformans in time in the infected tissue. For the mode of actions underlying SA-2 against C. neoformans, we found that the cationic peptide SA-2 could adhere to the negatively charged fungal cell membrane to increase the surface potential of C. neoformans in a dose-dependent manner, and finally disrupted the integrity of fungal cell membrane, reflecting as a 60% positive rate of propidium iodide uptake of C. neoformans cells after SA-2 (4 × MIC) treatment. Our study indicated that SA-2 has the potential to develop as a new therapeutic agent against infection induced by drug-resistant fungi.

Cbf-14 (RLLRKFFRKLKKSV), a designed antimicrobial peptide derived from the cathelicidin family, is effective against drug-resistant bacteria. Structurally related peptide impurities in peptide medicines probably have side effects or even toxicity, thus impurity profiling research during the entire production process is indispensable. In this study, a simple liquid chromatography-high-resolution mass spectrometry (LC-HRMS) method using a quadrupole time-of-flight (Q-TOF) mass spectrometer was developed for separation, identification, and characterization of structurally related peptide impurities in Cbf-14. A total of one process-related impurity and thirty-two degradation products were identified, and seven of them have been synthesized and confirmed. These impurities have not been declared in custom synthetic peptides. The degradation products were divided into five categories: fifteen Cbf-14 hydrolysates, five Cbf-14 isomers, four acetyl-Cbf-14 isomers, two aldimine derivatives, and six oxidized impurities. Combined with the peptide synthesis and the stress-testing studies, the origins and the formation mechanisms of these impurities were elucidated, which provides a unique insight for the follow-up quality study of Cbf-14 and other peptide products.

A polysaccharide (ESPS) purified from Eupolyphaga sinensis Walker by ion exchange chromatography and gel chromatography was investigated, including its structure characterization and antitumor activity. The results showed that ESPS was composed of rhamnose, fucose, arabinose, xylose, glucose, and galactose in a molar ratio of 7.4: 3.1: 13.9: 9.3: 39.7: 26.5, with the mean weight (Mw) of 2.14 × 104 Da; the main chain of ESPS was mainly composed of → 4) - α - D - Glcp - (1 → and → 3) - β - D - Galp - (1 →, and the side chains were connected to the main chain through the O-6 atom of glucose and O-4 and O-6 atom of galactose. In addition, ESPS promoted the lymphocyte proliferation and inhibited liver cancer cells growth through enhancing lymphocyte activity in vitro, mainly NK cells. Moreover, ESPS markedly stimulated immunity in H22-bearing mice by increasing the spleen and thymus indices and effectively inhibited H22 cell growth in vivo. These data indicated that ESPS was a polysaccharide component possessing high anti-hepatocellular carcinoma activity, representing a potential immunotherapy candidate for the treatment of liver cancer.

The structural modification of existing AMPs is an effective strategy to develop antimicrobial agents with high-efficiency, low-cost and low-toxicity antimicrobial agents. Methods: Here, we truncated 14-amino-acids at the N-terminus of MSI-78 to obtain MSI and further modified MSI to obtain four peptide analogs: MSI-1, MSI-2, MSI-3 and MSI-4. These peptide mutants were evaluated regarding their antibacterial activity against various sensitive or resistant bacteria; toxicity against mammalian cells or mice; and stability against violent pH, temperature variations and high NaCl concentrations. Finally, we also elucidated the possible mechanisms underlying its mode of action. Results: The results showed that MSI-1 and MSI-3 displayed activity that was superior to that of MSI-78 with MICs of 4-16 μg/ml and MBCs of 8-64 μg/ml, respectively, especially against drug-resistant bacteria, due to the increase in percent helicity and amphiphilicity. However, MSI-3, with higher hydrophobicity and antibacterial activity, had a relatively higher hemolysis rate and toxicity than MSI-1. MSI-1 exerted rapid bactericidal activity and effectively improved the survival rate and wound closure in penicillin-resistant E. coli-infected mice by eliminating bacterial counts in mouse organs or subeschar, further inhibiting the systemic dissemination of bacteria. Additionally, MSI-1 displayed perfect stability against violent pH, temperature variations and high NaCl concentrations and has the ability to circumvent the development of drug resistance. In terms of the mode of action, we found that at the super-MIC level, MSI-1 exhibited direct antimicrobial activity by disrupting the integrity of the bacterial cell membrane, while at the sub-MIC level, it bound to bacterial DNA to inhibit DNA replication and protein expression and ultimately disrupted bacterial biological function. Conclusions: This novel peptide MSI-1 could be a potential candidate for drug development against infection induced by drug-resistant bacteria.

Graphical abstract, Cluster of differentiation 47 (CD47)/signal regulatory protein alpha (SIRPα) is a negative innate immune checkpoint signaling pathway that restrains immunosurveillance and immune clearance, and thus has aroused wide interest in cancer immunotherapy. Blockade of the CD47/SIRPα signaling pathway shows remarkable antitumor effects in clinical trials. Currently, all inhibitors targeting CD47/SIRPα in clinical trials are biomacromolecules. The poor permeability and undesirable oral bioavailability of biomacromolecules have caused researchers to develop small-molecule CD47/SIRPα pathway inhibitors. This review will summarize the recent advances in CD47/SIRPα interactions, including crystal structures, peptides and small molecule inhibitors. In particular, we have employed computer-aided drug discovery (CADD) approaches to analyze all the published crystal structures and docking results of small molecule inhibitors of CD47/SIRPα, providing insight into the key interaction information to facilitate future development of small molecule CD47/SIRPα inhibitors.

Mitotic catastrophe (MC) is a newly identified type of anticancer mechanism for multidrug resistance (MDR) prevention. However, the long cellular death process resulting from MC is not beneficial for anticancer treatment. BZML is a novel colchicine-binding site inhibitor which can overcome MDR by inducing MC; however, BZML-induced MC cells underwent a long cellular death process. Thus, to improve anticancer therapies based on drug-induced MC, BZML-induced MC was served as a model to further study the underlying molecular mechanisms in the process of MC. Here, BZML could induce p53-dependent senescence in A549/Taxol cells, a MDR cell line. This senescence was a secondary effect of MC in overcoming MDR. During MC, BZML-induced destruction of protein-degradation system contributed not only to an increase of p53 protein but also to the accumulation of survivin in nucleus of A549/Taxol cells. Importantly, the nuclear accumulation of survivin was not the inducer but the result of BZML-induced MC, and it promoted the survival of senescent cells. Moreover, it provided additional vulnerability and critical opportunities for sequentially applied therapies. Further, targeting survivin with YM155 accelerated the death of MC cells by timely eliminating therapy-induced senescent cells and strengthening the efficiency of BZML in overcoming MDR in A549/Taxol cells. Collectively, nuclear accumulation of survivin delayed cellular death during MC by promoting the survival of BZML-induced senescent A549/Taxol cells. Moreover, “one-two punch” approach to cancer treatment based on combination therapy with YM155 for survivin suppression might be a new strategy for potentiating MC to overcome MDR.

Background: Mitotic catastrophe (MC) of cancer cells induced by BZML, a novel colchicine-binding site inhibitor, exerts a significant advantage in overcoming multidrug resistance (MDR) in NSCLC. However, the long cellular death process resulting from MC is not beneficial for anticancer treatment. Here, we study the mechanisms underlying MC occurrence and development to promote the development of anticancer therapies based on drug-induced MC.Methods: Cellular senescence was confirmed by morphological features, SA-β-Gal and C12FDG staining. Cell cycle analysis and Hoechst 33342 staining were used to detect MC. Relevant signal transduction pathways and protein location were detected by qRT-PCR, westren blot and immunofluorescence. The half-life of proteins was evaluated using the protein synthesis inhibitor cycloheximide. Flow cytometry, MTT assay, crystal violet staining, Hoechst 33342 staining and cell division detection were performed to determine the effects of BZML and/or YM155 on cell fate. Results: We found that BZML induced p53-dependent cellular senescence in A549/Taxol cells, but not in A549, H1299 and MDA-MB-231 cells. Interestingly, BZML-induced senescence was a secondary effect of MC. In addition, the destruction of the protein-degradation system induced by BZML contributed not only to an increase in p53 protein but also to the accumulation of survivin in the nucleus of A549/Taxol cells. However, in A549 cells, the overexpression of survivin had no effect on apoptosis resistance against BZML and failed to promote BZML-induced MC. The inhibition of survivin did not prevent MC occurrence. Unexpectedly, targeting survivin with YM155 accelerated the death of the MC cells by eliminating senescent cells and strengthening the efficiency of BZML in overcoming the MDR of A549/Taxol cells.Conclusions: Our data suggest that nuclear accumulation of survivin can delay cellular death during MC by promoting the survival of senescent BZML-treated A549/Taxol cells. Further, depending on the dose sequence, combination therapy with YM155 to inhibit survivin might be a new strategy for potentiating BZML-induced MC to overcome MDR during cancer treatment.

Background The antibiotic resistance and biofilm formation of pathogenic microbes exacerbate the difficulties of anti-infection therapy in the clinic. The structural modification of antimicrobial peptides (AMP) is an effective strategy to develop novel anti-infective agents. Method Seventeen amino acids (AA) in the longer chain of EeCentrocin 1 (from the edible sea-urchin Echinus esculentus) were truncated and underwent further modification. To produce lead peptides with low toxicity and high efficacy, the antimicrobial activity or cytotoxicity of peptides was evaluated against various multidrug-resistant bacteria/fungi or mammalian cells in vivo/ in vitro. In addition, the stability and modes of action of the lead peptide were investigated. Findings EC1-17KV displayed potent activity and an expanded antimicrobial spectrum, especially against drug-resistant gram-negative bacteria and fungi, attributable to its enhanced amphiphilicity and net charge. In addition, it exhibits bactericidal/fungicidal activity and effectively increased the animal survival rate and mitigated the histopathological damage induced by multidrug-resistant P. aeruginosa or C. albicans in infected mice or G. mellonella. Moreover, EC1-17KV had a poor ability to induce resistance in bacteria and fungi and exhibited desirable high-salt/high-temperature tolerance properties. In bacteria, EC1-17KV promoted divalent cation release to damage bacterial membrane integrity. In fungi, it changed C. albicans membrane fluidity to increase membrane permeabilization or reduced hyphal formation to suppress biofilm formation. Interpretation EC1-17KV is a promising lead peptide for the development of antimicrobial agents against antibiotic resistant bacteria and fungi. Funding This work was funded by the National Natural Science Foundation of China (No. 81673483, 81803591); National Science and Technology Major Project Foundation of China (2019ZX09721001-004-005); National Key Research and Development Program of China (2018YFA0902000); "Double First-Class" University project (CPU2018GF/GY16); Natural Science Foundation of Jiangsu Province of China (No. BK20180563); and A Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions.

The development of novel antifungal agents with high efficacy, low drug tolerance and few side effects is urgent. MSI-1 (GIWKFLKKAKKFWK-NH2), a cationic antimicrobial peptide, may be an attractive antifungal agent because of its structural characteristics, perfect stability against pH and high-temperature/salt, low toxicity towards mammalian cells and low potential for emergence of drug tolerance. In this study, the antifungal activity of MSI-1 in vitro and in a murine model of cryptococcal meningoencephalitis was evaluated. Zeta potential assay, flow cytometry, fluorescence microscope, transmission electron microscopy and microscale thermophoresis were performed to clarify the mechanisms underlying MSI-1 against C. neoformans. The results showed that MSI-1 exerted effective anti-cryptococcal activity in vitro, with MICs of 8−16 μg/mL and MFCs of 8−32 μg/mL, and in a C neoformans-infected mouse model, with significantly improved animal survival, decreased production of pro-inflammatory cytokines and alleviated lung injury, because the potent and rapid fungicidal activity of MSI-1 could effectively eliminate fungal counts in mouse organs. We confirmed that the positively charged peptide bound to C. neoformans by electrostatic attraction after interacting with glucuronoxylomannan (the primary component of C. neoformans capsule). Subsequently, MSI-1 increased the membrane fluidity of fungal cells and the cell membrane permeability, causing destabilized membrane integrity and leading to the final death of fungi. Collectively, MSI-1 possessed potent anti-cryptococcal activity via its notable membrane disruption effect and may be a potential candidate for use in antifungal infection induced by C. neoformans, especially azole-resistant cryptococcus.

Development of novel therapeutic strategies and antibacterial agents against antibiotic-resistant Staphylococcus aureus (S. aureus) is urgent. In this study, antibacterial activities and possible mechanisms of peptide MSI-1 against multiple drug-resistant S. aureus were investigated. Results demonstrated that MSI-1 had potent bacteriostatic activity and bactericidal efficiency against S. aureus, including methicillin-resistant S. aureus (MRSA), vancomycin-intermediate S. aureus (VISA) and vancomycin-resistant S. aureus (VRSA), with minimum inhibitory concentrations (MICs) ranging from 4 to 16 μg/mL and bactericidal times from 2-12 h. MSI-1 exhibited a low incidence of developing resistance and additive effects with vancomycin to overcome MRSA and VRSA. Moreover, MSI-1, even at sub-MIC concentrations, inhibited staphyloxanthin (STX) production of S. aureus. This inhibitory effect was unique and effectively sensitized S. aureus to host immune defense. In terms of its modes of action, MSI-1 disrupted the cell membrane of S. aureus by binding to negatively-charged lipoteichoic acid to exert a direct bactericidal effect. Interestingly, MSI-1 interacted with 4,4'-diapophytoene desaturase (CrtN) of S. aureus via ionic bonds, hydrogen bonds, and Pi-Pi or Pi-alkyl interactions, and alanine substitution of the key amino acids contributed to these interactions weakened this STX production inhibition. Thus, in a MRSA-induced skin infection in mice and MRSA/VRSA-induced systemic infection in Galleria mellonella,MSI-1 alleviated staphylococcal scalded skin syndrome to promote mouse skin wound repair and mitigated staphylococcus infection-induced immune melanization to enhance G. mellonella survival. Collectively, MSI-1 has potent antibacterial activity against drug-resistant S. aureus by affecting bacterial viability and exerting its anti-virulence effects. It can be developed as a new antibacterial agent to resist refractory S. aureus infection.

The currently available drugs against influenza A virus primarily target neuraminidase (NA) or the matrix protein 2 (M2) ion channel. The emergence of drug-resistant viruses requires the development of new antiviral chemicals. Our study applied a cell-based approach to evaluate the antiviral activity of a series of newly synthesized benzoic acid derivatives, and 4-(2,2-Bis(hydroxymethyl)-5-oxopyrrolidin-l-yl)-3-(5-cyclohexyl-4H-1,2,4-triazol-3-yl)amino). benzoic acid, termed NC-5, was found to possess antiviral activity. NC-5 inhibited influenza A viruses A/FM/1/47 (H1N1), A/Beijing/32/92 (H3N2) and oseltamivir-resistant mutant A/FM/1/47-H275Y (H1N1-H275Y) in a dose-dependent manner. The 50% effective concentrations (EC50) for H1N1 and H1N1-H275Y were 33.6 &mu, M and 32.8 &mu, M, respectively, which showed that NC-5 had a great advantage over oseltamivir in drug-resistant virus infections. The 50% cytotoxic concentration (CC50) of NC-5 was greater than 640 &mu, M. Orally administered NC-5 protected mice infected with H1N1 and H1N1-H275Y, conferring 80% and 60% survival at 100 mg/kg/d, reducing body weight loss, and alleviating virus-induced lung injury. NC-5 could suppress NP and M1 protein expression levels during the late stages of viral biosynthesis and inhibit NA activity, which may influence virus release. Our study proved that NC-5 has potent anti-influenza activity in vivo and in vitro, meaning that it could be regarded as a promising drug candidate to treat infection with influenza viruses, including oseltamivir-resistant viruses.

Baicalin was identified as a neuraminidase (NA) inhibitor displaying anti-influenza A virus (IAV) activity. However, its poor solubility in saline has limited its use in the clinic. We generated sodium baicalin and showed that it exhibited greatly increased solubility in saline. Its efficacy against oseltamivir-resistant mutant A/FM/1/47-H275Y (H1N1-H275Y) was evaluated in vitro and in vivo. Results showed that 10 μM of sodium baicalin inhibited A/FM/1/47 (H1N1), A/Beijing/32/92 (H3N2) and H1N1-H275Y in MDCK cells in a dose-dependent manner, with inhibitory rates of 83.9, 75.9 and 47.7%, respectively. Intravenous administration of sodium baicalin at 100 mg/kg/d enabled the survival of 20% of H1N1-H275Y-infected mice. The treatment alleviated body weight loss and lung injury. Moreover, sodium baicalin exerted a clear inhibitory effect on NAs. The IC50 values of sodium baicalin against H1N1-H275Y and cells-expressing A/Anhui/1/2013-R294K (H7N9-R294K) NA protein (N9-R294K) were 214.4 μM and 216.3 μM. Direct interactions between sodium baicalin and NA were observed, and we simulated the interactions of sodium baicalin with N9-R294K and N9 near the active sites of OC-N9-R294K and OC-N9. The residues responsible for the sodium baicalin-N9-R294K and sodium baicalin-N9 interactions were the same, confirming that sodium baicalin exerts effects on wild-type and oseltamivir-resistant viral strains.

Cbf-14 (RLLRKFFRKLKKSV), a designed peptide derived from cathelicidin family AMP, has proven to be potent against drug-resistant bacteria. In the present study, we investigated the anti-cryptococcal activity of Cbf-14 in vitro and in a pulmonary infection mouse model. Sensitivity test indicated that Cbf-14 possessed effective antifungal activity against Cryptococcus neoformans with an MIC of 4–16 µg/ml, and killing experiments showed that fungicidal activity was achieved after only 4 h treatment with Cbf-14 at 4× MIC concentrations in vitro. Meanwhile, Cbf-14 was effective at prolonging the survival of infected mice when compared with controls, and significantly inhibited the secretion of pro-inflammatory cytokines TNF-α, IL-1β and IL-6, suggesting its anti-inflammatory activity against fungal infections. As a positively charged peptide, Cbf-14 was proven to neutralize the negative zeta potential of the fungal cell surface, disrupt the capsule polysaccharide of fungi, and further damage cell membrane integrity. These results were confirmed by flow cytometry analysis of the fluorescence intensity after PI staining, while cell membrane damage could be clearly observed by transmission electron microscopy after Cbf-14 (4× MIC) treatment for 1 h. In addition, Cbf-14 increased the IL-10 levels in cultured RAW 264.7 cells, which were stimulated by C. neoformans infection. The obtained data demonstrated that Cbf-14 could rapidly kill C. neoformans cells in vitro, effectively inhibit C. neoformans induced-infection in mice, and inhibit inflammation in vitro / vivo. Therefore, Cbf-14 could potentially be used for the treatment of fungal infections clinically.

Atherosclerosis attracts increasing global attention because of its morbidity and mortality. G004, as a synthetic sulfonylurea compound, has been confirmed to have anti-hyperglycaemia, anti-platelet and anti-thrombus effects. The aim of the present study was to investigate whether G004 suppress the onset and development of atherosclerosis and illuminate its probable mechanism of action. ApoE-/- mice that were fed a high-fat diet were randomly divided into five groups by weight; subsequently, they were treated with vehicle, G004, at different doses or atorvastatin once daily for 12weeks. Meanwhile, C57BL/6 mice with the same diet served as the normal controls. Then, the serum lipid profiles and histopathological damage to the liver, kidney, aortic arch and aortic root were analysed. The activation of endothelial nitric oxide synthase (eNOS) and levels of inflammatory markers were detected. Reverse cholesterol transport (RCT) was assessed in vivo by intraperitoneal injection of RAW264.7 cells that were radiolabelled with 3H-cholesterol. The results indicated that G004 ameliorated the serum lipid accumulation, atherosclerotic lesions and liver steatosis. Additionally, this compound increased the expression of SIRT1 and eNOS as well as the phosphorylation and deacetylation of eNOS in the aorta, alleviating the inflammatory state. RCT was promoted in ApoE-/- mice, which was accompanied by increased expression of SIRT1/LXRα/ABCA1/G1 in the liver, and similar results appeared in the cholesterol efflux assay in RAW264.7 cells. The results provide a strong rationale for G004 to be an efficient anti-atherosclerosis agent that improved vascular endothelial dysfunction by stimulating SIRT1/eNOS and promoted RCT by stimulating SIRT1/LXRα/ABCA1/G1.

Here, we investigated whether I 4 , which was initially developed as a hypoglycemic agent, possesses anti-atherosclerotic activity and attempted to elucidate the probable mechanism of action underlying this activity. ApoE −/− mice were fed a Western diet and simultaneously administered I 4 , glimepiride, or pioglitazone once daily for 12 weeks, and the atherosclerotic vascular lesions, lipid content, and expression levels of LOX-1, ICAM-1, VCAM-1 and Bax/Bcl-2 in mouse aortas were assessed. RAW264.7 macrophage-derived foam cells were obtained via ox-LDL stimulation to investigate the lipid-lowering, anti-atherosclerotic inflammation and anti-apoptotic effect of I 4 . The data indicated that I 4 significantly decreased the lipid accumulation in the circulation and tissue, especially for TG and FFA levels (p vs model group), alleviating the arterial and liver lesions induced by lipotoxicity. Its lipid-reducing effects may due to LOX-1and CD36 expression suppression. I 4 , at doses of 20 mg/kg and 10 mg/kg, significantly decreased serum IL-6, IL-1β, and TNF-α production and suppressed the expression of p-ERK, p-p38, VCAM-1 and ICAM-1 protein. I 4 attenuated atherosclerotic inflammation by blocking NF-κB nuclear translocation, suppressing MAPK/NF-κB signaling pathway and diminishing NF-κB-VCAM-1 promoter region binding. Additionally, I 4 suppressed p-p53 and cleaved-caspase-3 expression to inhibit foam cell apoptosis induced by ox-LDL uptake. Overall, I 4 exerts potent inhibitory effects on atherosclerosis onset and development.

Fibrotic diseases have become a major cause of death in the developed world. AdipoR1 agonists are potent inhibitors of fibrotic responses. Here, we focused on the in silico identification of novel AdipoR1 peptide agonists. A homology model was constructed to predict the 3D structure of AdipoR1. By docking to known active peptides, the putative active site of the model was further explored. A virtual screening study was then carried out with a set of manually designed peptides using molecular docking. Peptides with high docking scores were then evaluated for their anti-fibrotic properties. The data indicated that the novel peptide Pep70 significantly inhibited the proliferation of hepatic stellate cells (HSC) and NIH-3T3 cells (18.33% and 27.80%) and resulted in favouring cell-cycle arrest through increasing the accumulation of cells in the G0/G1 phase by 17.08% and 15.86%, thereby reducing the cell population in the G2/M phase by 11.25% and 15.95%, respectively. Additionally, Pep70 exhibited the most marked suppression on the expression of α-smooth muscle actin (α-SMA), collagen type I alpha1 (COL1A1) and TGF-β1. Therefore, the peptide Pep70 was ultimately identified as an inhibitor of fibrotic responses and as a potential AdipoR1 agonist.

Helicobacter pylori (H. pylori) infection is highly prevalent, and has developed antimicrobial resistance to virtually all existing antibiotics. Currently, treatment of H. pylori infection (involving proton pump inhibitors and broad-spectrum antibiotics) is suboptimal, with high failure rates. Thus, there is a pressing need to develop new anti-H. pylori therapies. Cbf-K

Non-small cell lung cancer (NSCLC) is one of the most common malignancies in the world. Although extensive studies showed that luteolin exhibited antitumor effects against NSCLC, the mechanism has not been fully established. In the present study, we found that luteolin significantly reduced the expression of absent in melanoma 2 (AIM2) at both mRNA and protein levels leading to the suppression of AIM2 inflammasome activation, which induced G2/M phase arrest and inhibited epithelial–mesenchymal transition (EMT) in NSCLC. Furthermore, the inhibitory effects of luteolin on NSCLC cells were abolished by the knockdown of AIM2. On the contrary, the antitumor effects of luteolin could be notably reversed by the overexpression of AIM2. In addition, luteolin reduced poly(dA:dT)-induced caspase-1 activation and IL-1β cleavage in NSCLC cells. These findings suggested that AIM2 was essential to luteolin-mediated antitumor effects. The antitumor effects of luteolin, which were closely associated with AIM2, were also confirmed in the A549 and H460 xenograft mouse models. Collectively, our study displayed that the antitumor effects of luteolin on NSCLC were AIM2 dependent and the downregulation of AIM2 might be an effective way for NSCLC treatment.

Introduction: Fibrotic disorders are a leading cause of morbidity and mortality; hence effective treatments are still vigorously sought. AdipoRs (AdipoR1 and Adipo2) are responsible for the antifibrotic effects of adiponectin (APN). APN exerts antifibrotic effects by binding to its receptors. APN concentration and AdipoR expression are closely associated with fibrotic disorders. Decreased AdipoR expression may reduce APN-AdipoR signaling, while the upregulation of AdipoR expression may restore the anti-fibrotic effects of APN. Loss of APN signaling exacerbates fibrosis in vivo and in vitro. Areas covered: We assess the relationship between APN and fibrotic disorders, the structure of receptors for APN and the pathways accounting for APN or its analogs blocking fibrotic disorders. This article also discusses designed APN products and their therapeutic prospects for fibrotic disorders. Expert opinion: AdipoRs have a critical role in blocking fibrosis. The development of small-molecule agonists toward this target represents a valid drug development pathway.

Antibiotic resistance is spreading faster than the development of new antibiotics into clinical practice. Currently, the design of antimicrobial peptides (AMPs), potential new antibacterial agents with rare antimicrobial resistance, is the available strategy to enhance the antimicrobial activity and lower the toxicity of AMPs. In this study, a peptide derived from hagfish intestinal peptide was designed and termed as HF-18 (GFFKKAWRKVKKAFRRVL). After antimicrobial/bactericidal test in vitro, we found that HF-18 exhibited a potent antimicrobial activity with MIC of only 4 μg/ml against drug-resistant Staphylococcus aureus (S. aureus). Meanwhile, it eliminated the test bacteria within 1 h, suggesting its rapid bactericidal effect. Importantly, this peptide had no obvious hemolytic activity and cytotoxicity to mammalian cells. Furthermore, its notable antimicrobial effects in vivo was confirmed again in S. aureus induced mouse bacteremia and skin wound infection, reflecting as the decrease in bacterial counts in mouse lung or skin (up to 1.9 or 3.5 log CFU respectively), and including the inhibitory activity on inflammatory cytokines secretion. The possible mechanisms underlying HF-18 against drug-resistant S. aureus may attribute that HF-18 neutralized the negative charge in S. aureus surface and then disrupted the integrity of cell membranes to enhance the permeation of bacterial membrane, showing as the increased uptake of NPN and PI and the obvious morphology changes of S. aureus. In addition, this peptide bound to bacterial genomic DNA to suppress the expression of Panton-Valentine leukocidin (pvl) and nuclease (nuc) genes, which play major roles in S. aureus virulence. The properties of HF-18 suggest a path towards developing antibacterial agents that has stronger antibacterial activity and greater security for clinical treatment of infection induced by S. aureus, especially drug-resistant S. aureus.

Colorectal cancer (CRC) is one of the most common malignancies, and multidrug resistance (MDR) reduces the efficiency of anticancer drugs. Therefore, the development of novel anticancer drugs that are highly active against CRC with MDR is urgently needed. Our previous study showed that 5-(3,4,5-trimethoxybenzoyl)-4-methyl-2-(p-tolyl) imidazol (BZML) is not a P-glycoprotein (P-gp) substrate and has a potent anticancer effect against paclitaxel -sensitive or -resistant non-small-cell lung cancer (NSCLC) in vitro and in vivo. In the present study, we found that BZML exhibited strong anticancer activity not only in sensitive CRC cells (SW480 and HCT-116 cells) but also in intrinsically drug-resistant CRC cells (Caco2 cells). In addition, by targeting the colchicine binding site, BZML inhibited tubulin polymerization, which induced G2/M phase arrest, and it caused DNA damage by directly targeting DNA or producing ROS. Further, BZML induced apoptosis through the time-dependent ROS-mediated mitochondrial apoptotic pathway in the CRC cells. Additionally, BZML inhibited P-gp-mediated drug efflux and enhanced the inhibition of the cell growth that had been induced by paclitaxel or doxorubicin in Caco2 cells. In summary, BZML is a multi-targeted anticancer drug that targets tubulin and DNA, and the mechanisms underlying its potent anticancer activity involve disrupting microtubule assembly, causing DNA damage, inducing cell cycle arrest and eventually activating the ROS-mediated mitochondrial apoptotic pathway in SW480, HCT-116 and Caco2 cells. Therefore, the novel compound BZML is a promising anticancer drug that has tremendous potential for CRC treatment, especially for the treatment of drug-resistant CRC.

Helicobacter pylori (H. pylori) infection is highly prevalent, and has developed antimicrobial resistance to virtually all existing antibiotics. Currently, treatment of H. pylori infection (involving proton pump inhibitors and broad-spectrum antibiotics) is suboptimal, with high failure rates. Thus, there is a pressing need to develop new anti-H. pylori therapies. Cbf-K16, a cathelicidin-like antimicrobial peptide, presented broad antimicrobial activity during our previous research. This study further evaluated the therapeutic potential and the mode of action underlying Cbf-K16 against clarithromycin- and amoxicillin-resistant H. pylori SS1. The MIC and MBC of Cbf-K16 against the tested H. pylori were 16 and 32 μg/ml, respectively, and its killing kinetics was time-dependent, reflecting the thorough elimination of drug-resistant bacteria within 24 h. This peptide also protected H. pylori-infected gastric epithelial cells (GES-1) from death by reducing the cell supernatant and intracellular bacterial counts by 1.9 and 2.9-log10 units, respectively. These data indicated the powerful antimicrobial effects of Cbf-K16 in vitro. Meanwhile, notable antimicrobial activity in the mouse gastritis model was observed, with decreasing bacterial counts by 3.9-log10 units in stomach tissues and Cbf-K16 could effectively suppress the secretion of inflammatory cytokine IL-8. For its mode of action, Cbf-K16 not only neutralized the negative potential and increased the membrane uptake of NPN and PI by 78.5% and 85.1%, respectively, but also bound to genomic DNA, which in turn downregulated the expression of adhesion genes (alpA and alpB) and virulence gene (cagA), indicating its effective activities on membrane disruption, DNA-binding and gene expression. The data above demonstrated that Cbf-K16 possessed effective antimicrobial and anti-inflammatory activities and downregulated the expression of adhesion- and cytotoxin-associated genes of drug-resistant H. pylori SS1, making it a potential candidate for anti-infective therapy.

A polysaccharide isolated from Strongylocentrotus nudus eggs (SEP) reportedly displays immune activity in vivo. Here, its effect and underlying mechanism in the treatment of pancreatic cancer were investigated. SEP obviously inhibited pancreatic cancer growth by activating NK cells in vitro/vivo via TLR4/MAPKs/NF-κB signaling pathway, The tumor inhibitory rate achieved to 44.5% and 50.8% at a dose of 40 mg/kg in Bxpc-3 and SW1990 nude mice, respectively. Moreover, SEP obviously augmented the Gemcitabine (GEM) antitumor effect by upregulating NKG2D, which improved the sensitivity of NK cells targeting to its ligand MICA; meanwhile, the antitumor inhibitory rate was 68.6% in BxPC-3 tumor-bearing mice. Moreover, SEP reversed GEM-induced apoptosis and atrophy in both spleen and bone marrow via suppressing ROS secretion in vivo. These results suggested that pancreatic cancer was effectively inhibited by SEP-enhanced NK cytotoxicity mediated primarily through TLR4/MAPKs/NF-κB signaling pathway, representing a potential immunotherapy candidate for the treatment of pancreatic cancer.

Influenza A virus is a leading cause of mortality in humans and poses a global health emergency due to its newly adapted and resistant strains. Thus, there is an urgency to develop novel anti-influenza drugs. Peptides are a type of biological molecule having a wide range of inhibitory effects against bacteria, fungi, viruses and cancer cells. The prospects of several peptides and their mechanisms of action have received significant attention. BF-30, a 30 amino acid residue peptide isolated from the venom of the snake, Bungarus fasciatus, is reported to have antibacterial and antitumor activities. Here, we demonstrated that the 50% cytotoxic concentration (CC50) of the peptide to MDCK cells is 67.7 μM. While BF-30 could inhibit the influenza virus strains H1N1, H3N2 and the oseltamivir-resistant strain H1N1, in vitro, with 50% effective concentration (EC50) of 5.2, 7.4 and 18.9 μM, respectively. In animal experiments, mice treated with BF-30 showed 50% survival at a dosage of 4 μM, with an approximately 2 log viral titer decrease in the lung. However, further studies showed that BF-30 worked on only the virus invasion stage, and inhibited the influenza virus infection by causing virion membrane fusion rather than interacting with hemagglutinin or neuraminidase. These results demonstrated that the peptide BF-30 exhibited an effective inhibitory activity against the influenza A virus and could be a promising candidate for influenza virus therapy.

Baicalin is a natural flavonoid glycoside which has potent anti-tumor and antioxidant activity in cancer cells. In the present study, we found that baicalin treatment significantly induced senescence in colon cancer cells. Furthermore, baicalin upregulated the expression of decidual protein induced by progesterone (DEPP) in HCT116 colon cancer cells, which accompanied with the activation of Ras/Raf/MEK/ERK and p16INK4A/Rb signaling pathways. Meanwhile, these phenomena also appeared under the anti-oxidation effect exerted by baicalin. In addition, ectopic expression of DEPP in HCT116 cells significantly induced the activity of senescence-associated β-galactosidase (SA-β-Gal) in tumor cells regulated by Ras/Raf/MEK/ERK signaling pathway. Knockdown of DEPP by RNA interference efficiently counteracted the baicalin-mediated growth inhibition, senescence and cell cycle arrest in cancer cells. Importantly, in a xenograft mouse model of human colon cancer, we further confirmed that baicalin treatment dramatically inhibited tumor growth, which was due to the induction of tumor cellular senescence via the upregulation of DEPP and the activation of Ras/Raf/MEK/ERK signaling in vivo. In addition to baicalin treatment, we found that the hypoxia-response protein DEPP functions as a positive regulator involving the regulations of Ras/Raf/MEK/ERK signaling pathway and inhibition of human colon cancer by other anti-oxidative drugs, such as curcumin and sulforaphane, resulting in tumor cellular senescence. These results collectively suggest that baicalin upregulates the expression of DEPP and activates its downstream Ras/Raf/MEK/ERK and p16INK4A/Rb pathways by acting as an antioxidant, leading to senescence in colon cancer cells.

1-[4-[2-(4-Bromobenzene-sulfonamino)ethyl]phenylsulfonyl]-3-(trans-4-methylcyclohexyl) urea (G004, CAS865483-06-3) is a synthetic sulfonylurea, incorporating the hypoglycemic active structure of glimepiride (CAS 93479-97-1) and anti-TXA2 receptor (TP) active structure of BM-531(CAS 284464-46-6). In this study, we evaluated the effect of G004 on hyperglycemia and dyslipidemia as well as diabetic nephropathy (DN) in db/db mice by gavage over 90 consecutive days of treatment. The fasting blood glucose (FBG), glucose, and insulin tolerance as well as dyslipidemia were effectively ameliorated in db/db mice treated with G004. Interestingly, renal histological results of db/db mice revealed that G004 markedly reversed the expansion of mesangial extracellular matrix (ECM), the early hallmark of DN. Indeed, G004 treatment downregulated the renal expressions of type 4 collagen (Col IV) and transforming growth factor-β1 (TGF-β1) in db/db mice. In addition, imbalance in expressions of matrix metalloproteinase-9 (MMP-9) and its tissue inhibitor-1 (TIMP-1) in db/db mice kidneys was observed. However, G004 increased and decreased the expressions of MMP-9 and TIMP-1, respectively. It is well known that TGF-β pathway signaling plays an essential role in hyperglycemia-induced cell protein synthesis. On the other hand, MMP/TIMP system is responsible for the breakdown and turnover of ECM. Thus, we speculate that G004 possibly attenuated ECM accumulation via remodeling the synthesis and degradation of ECM component Col IV through modulation in TGF-β1 and MMP-9/TIMP-1 expressions in kidneys of db/db mice. Results from this study provide a strong rationale for G004 to be an efficient glucose-controlling agent with significant reno-protective properties.

Broad spectrum activities, a unique mode of actions and rare resistant variants make antimicrobial peptide (AMP) a potential alternative to antibiotics. However, AMPs still have limitations in clinical development due to their physiological stability, toxicity and manufacturing costs. Cbf-14, derived from cathelicidin-BF has been proven to be effective against drug-resistant bacteria. Herein, a series of Cbf-14 mutants were designed to overcome these limitations. Design strategies included substitutions of lysine (Lys) or leucine (Leu) with similar residues such as ornithine (Orn) and norleucine (Ile), which are unnatural amino acids, to generate AMPs with enhanced therapeutic potential. Antimicrobial activity, hemolytic activity and cytotoxicity against mouse spleen cells of the peptide mutants were investigated. Among all of the mutants, Cbf-14-2 was regarded as the most potent candidate with the amino acid sequence of RLLR-Orn-FFR-Orn-LKKSV-NH2, which exhibited a superior antimicrobial activity with a minimum inhibitory concentration (MIC) of 4-32μg/ml. Meanwhile, Cbf-14-2 displayed low levels of hemolysis in sheep red blood cells (sRBCs) and negligible cytotoxicity against mouse spleen cells, suggesting low toxicity against mammalian cells. A circular dichroism (CD) study indicated that Cbf-14-2 has a higher alpha-helix content than Cbf-14 (68.3% vs 35.1%) in SDS, which may contribute to its superior activity. Time-killing curves showed Cbf-14-2 can eliminate all tested bacteria within 240min, suggesting its rapid bactericidal effect. Transmission electron microscopy (TEM), flow cytometry and calcein release assays revealed its excellent antimicrobial potency by inducing membrane permeation and disruption. In addition, Cbf-14-2 (10mg/kg) could significantly elevate the survival rate of clinical strain infected mice, with a survival rate of 70.0%. Taken together, the data suggest that Cbf-14-2 possesses effective antimicrobial activity against penicillin-resistant bacteria in vitro and in vitro, thus rendering it as a potential anti-infective agent in clinical settings.

Here, we investigated whether I

Cbf-14, a cationic peptide derived from a cathelin-like domain, was designed by inserting the highly α-helical sequence RLLR into an antibacterial sequence and deleting the inactive amino acids in Cbf-K16. Clinical penicillin-resistant isolates as well as NDM-1-carrying Escherichia coli and a correspondingly infected mice model were employed to evaluate Cbf-14 antibacterial activity. The results showed that Cbf-14 possessed potent antimicrobial effects with an MIC of 8-64 μg/ml, and killed almost all bacteria within 240 min. Cbf-14-treated mice achieved an 80% survival rate and approximate 2.5 log unit reduction in CFU in tissues; additionally, this peptide significantly suppressed the production of pro-inflammatory cytokines by the disaggregation of lipopolysaccharide (LPS), suggesting its anti-inflammatory effects. Furthermore, Cbf-14, concentration higher than 2 × MIC value, increased membrane uptake to NPN and PI dye by 96.2% and 63.7%, respectively, neutralised the negative zeta potential of LPS and bacteria surface, and induced 100% leakage of liposome-entrapped calcein and cytoplasmic membrane disruption of E. coli, indicating obvious membrane permeation. Finally, it bound to DNA and respectively evoked 85.0% and 63.3% inhibition of gene replication and protein expression of NDM-1 at sub-MIC concentration in E. coli BL21 (DE3)-NDM-1. These data indicated that Cbf-14 possessed effective antimicrobial activity against penicillin-resistant bacteria in vitro/vivo through membrane disruption, DNA binding, down-regulating NDM-1 expression by plasmid replication inhibition, and anti-inflammatory activity by LPS disaggregation, suggesting a potential anti-infective clinical agent.

I4 is a new synthetic anti-diabetes sulfonylurea compound. The aim of present study was to investigate the preventive effects and primary action mechanisms of I4 on platelet-mediated arterial thrombosis. Platelet aggregation and 5-hydroxytryptamine (5-HT) secretion ex vivo was detected. The time-to-occlusion (TTO), thrombus weight and content of von Willebrand factor (vWF) in rat model of electrical- and ferric chloride-induced vessel occlusion were determined. Meanwhile, a rat model of type 2 diabetes mellitus (T2DM) was established to evaluate the effect of I4 on levels of plasma p-selectin, 6-keto-prostaglandin F1a (6-keto-PGF1a), thromboxane B2 (TXB2), tissue-type plasminogen activator (t-PA) and type-1 plasminogen activator inhibitor (PAI-1). NO synthesis, NOS activity, adhesion of platelet toward endothelial cell and intercellular adhesion molecule-1 (ICAM-1) expression were examined. Results showed that I4 exhibited a higher inhibitory potency than Glimepiride on ADP-induced platelet aggregation and 5-HT release ex vivo. In addition, I4 reduced the thrombus weight and content of vWF and markedly prolonged TTO. Oral administration of I4 (1 � 10 mg/kg) inhibited p-selectin production, elevated the ratio of plasma 6-keto-PGF1a/TXB2 and t-PA/PAI-1 in T2DM rats. Furthermore, I4 significantly improved NO synthesis and NOS activity, lowered adhesion ratio of platelet toward endothelial cells and ICAM-1 expression on HUVECs. These observations suggest that I4 markedly improves platelet-mediated arterial thrombosis by inhibiting platelet activation and release reaction, ameliorating the endothelial dysfunction such as the suppression of vWF production and the reduction of the overexpression of ICAM-1, displayed its potential in alleviating diabetes-associated vascular complications.