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DNA damage response (DDR) pathways are crucial for the survival of cancer cells and are attractive targets for cancer therapy. Bloom syndrome protein (BLM) is a DNA helicase that performs important roles in DDR pathways. Our previous study discovered an effective new BLM inhibitor with a quinazolinone scaffold by a screening assay. Herein, to better understand the structure-activity relationship (SAR) and biological roles of the BLM inhibitor, a series of new derivatives were designed, synthesized, and evaluated based on this scaffold. Among them, compound 9h exhibited nanomolar inhibitory activity and binding affinity for BLM. 9h could effectively disrupt BLM recruitment to DNA in cells. Furthermore, 9h inhibited the proliferation of the colorectal cell line HCT116 by significantly triggering DNA damage in the telomere region and inducing apoptosis, especially in combination with a poly (ADP-ribose) polymerase (PARP) inhibitor. This result suggested a synthetic lethal effect between the BLM and PARP inhibitors in DDR pathways.

Nonalcoholic fatty liver disease (NAFLD)/nonalcoholic steatohepatitis (NASH) are global epidemic public health problems with pathogenesis incompletely understood. Hepatocyte excessive apoptosis is a significant symbol for NAFLD/NASH patients, and therefore anti-apoptosis therapy could be used for NAFLD/NASH treatment. Up-regulation of BCL-2 has been found to be closely related with anti-apoptosis. BCL-2 gene promoter region has a C-rich sequence, which can form i-motif structure and play important role in regulating gene transcription. In this study, after extensive screening and evaluation, we found that acridone derivative A22 could up-regulate BCL-2 transcription and translation in vitro and in cells through selective binding to and stabilizing BCL-2 gene promoter i-motif. Our further experiments showed that A22 could reduce hepatocyte apoptosis in NAFLD/NASH model possibly through up-regulating BCL-2 expression. A22 could reduce inflammation, endoplasmic reticulum stress and cirrhosis in high-fat diet-fed mice liver model. Our findings provide a potentially new approach of anti-apoptosis for NAFLD/NASH treatment, and A22 could be further developed as a lead compound for NAFLD/NASH therapy. Our present study first demonstrated that gene promoter i-motif could be targeted for gene up-regulation for extended treatment of other important diseases besides cancer.

Cancer metabolism is an attractive target of the therapeutic strategy for cancer. The present study identified bouchardatine (Bou) as a potent suppressor of rectal cancer growth by cycle-arresting independent of apoptosis. In cultured HCT-116 rectal cancer cells, Bou increased glucose uptake/oxidation and capacity of mitochondrial oxidation. These effects were associated with an upregulation of uncoupling protein 2 (UCP2) and the activation of its upstream Sirtuin 1 (SIRT1)/(Liver kinase B1) LKB1- (Adenosine monophosphate-activated protein kinase) AMPK axis. The pivotal role of UCP2 in the cancer-suppressing effect was demonstrated by overexpressing UCP2 in HCT-116 cells with similar metabolic effects to those produced by Bou. Interestingly, Bou activated peroxisome proliferators activated receptor γ coactivator 1α (PGC-1α) and recruited it to the promoter of UCP2 in HCT-116 cells along with deacetylation (thus activation) by SIRT1. The requirement of SIRT1 for the cancer-suppressing effect through the PGC-1α–UCP2 was confirmed by the reciprocal responses to Bou in HCT-116 with defected and overexpressed SIRT1. Whereas knockdown, mutation or pharmacological inhibition of SIRT1 all abolished Bou-induced deacetylation/activation of PGC-1α, the opposing effects were observed after overexpressing SIRT1. In mice, administration of Bou (50 mg/kg) also suppressed the growth of rectal cancer associated with increases the UCP2 expression and mitochondria capacity in the tumor. Collectively, our findings suggest that Bou has a therapeutic potential for the treatment of rectal cancer by disrupting the metabolic path of cancer cells via activating the PGC-1α-UCP2 axis with SIRT1 as its primary target.

The gut bacterium Akkermansia muciniphila has been increasingly recognized for its therapeutic potential in treating metabolic disorders, including obesity, diabetes, and metabolicdysfunction-associated fatty liver disease (MAFLD). However, its underlying mechanism involved in its well-known metabolic actions needs further evaluation. The present study explored the therapeutic effect and mechanism of A. muciniphila in intervening MAFLD by using a high-fat and high-cholesterol (HFC) diet induced obese mice model. Mice treated with A. muciniphila efficiently reversed MAFLD in the liver, such as hepatic steatosis, inflammatory, and liver injury. These therapeutic effects persisted after long-term drug withdrawal and were slightly weakened in the antibiotics-treated obese mice. A. muciniphila treatment efficiently increased mitochondrial oxidation and bile acid metabolism in the gut-liver axis, ameliorated oxidative stress-induced cell apoptosis in gut, leading to the reshaping of the gut microbiota composition. These metabolic improvements occurred with increased L-aspartate levels in the liver that transported from the gut. The administration of L-aspartate in vitro or in mice displayed the similar beneficial metabolic effects mentioned above and efficiently ameliorated MAFLD. Together, these data indicate that the anti-MAFLD activity of A. muciniphila correlated with lipid oxidation and improved gut–liver interactions through regulating the metabolism of L-aspartate. A. muciniphila could be a potential agent for clinical intervention in MAFLD.

Upregulation of platelet-derived growth factor receptor β (PDGFR-β) has been found to be associated with development of various types of cancers, which has become an attractive target for anti-tumor treatment. Previously, we have synthesized and studied an acridone derivative B19, which can selectively bind to and stabilize oncogene c-myc promoter i-motif, resulting in down-regulation of c-myc transcription and translation, however its effect on tumor cells apoptosis requires improvement. In the present study, we synthesized a variety of B19 derivatives containing a known anti-cancer fluorescent chromophore naphthalimide for the purpose of enhancing anti-cancer activity. After screening, we found that acridone-naphthalimide derivative WZZ02 could selectively stabilize PDGFR-β promoter G-quadruplex and destabilize its corresponding i-motif structure, without significant interaction to other oncogenes promoter G-quadruplex and i-motif. WZZ02 down-regulated PDGFR-β gene transcription and translation in a dose-dependent manner, possibly due to above interactions. WZZ02 could significantly inhibit cancer cell proliferation, and induce cell apoptosis and cycle arrest. WZZ02 exhibited tumor growth inhibition activity in MCF-7 xenograft tumor model, which could be due to its binding interactions with PDGFR-β promoter G-quadruplex and i-motif. Our results suggested that WZZ02 as a dual G-quadruplex/i-motif binder could be effective on both oncogene replication and transcription, which could become a promising lead compound for further development with improved potency and selectivity. The wide properties for the derivatives of 1,8-naphthalimide could facilitate further in-depth mechanistic studies of WZZ02 through various fluorescent physical and chemical methods, which could help to further understand the function of PDGFR-β gene promoter G-quadruplex and i-motif.

Novel mansonone F derivative MSN54 (9-bromo-2,3-diethylbenzo[de]chromene-7,8-dione) exhibited significant cytotoxicity against twelve human tumor cell lines in vitro, with particularly strong potency against HL-60/MX2 cell line resistant to Topo II poisons. MSN54 was found to have IC50 of 0.69 and 1.43 µM against HL-60 and HL-60/MX2 cells, respectively. The resistance index is 10 times lower than that of the positive control VP-16 (etoposide). Various biological assays confirmed that MSN54 acted as a Topo IIα specific non-intercalative catalytic inhibitor. Furthermore, MSN54 exhibited good antitumor efficacy and low toxicity at a dose of 5 mg/kg in A549 tumor xenograft models. Thus, compound MSN54 is a promising candidate for the development of novel antitumor agents.

Euphorhelipanes A (1) and B (2), two Euphorbia diterpenoids with a new 4-(5,5-dimethylheptan-2-yl)-2,7-dimethylbicyclo[4.3.0]nonane skeleton, were isolated from a 95% ethanol extract of the whole plants of Euphorbia helioscopia. Their structures were elucidated by spectroscopic data analysis, quantum chemical calculations, and single-crystal X-ray diffraction data. Compounds 1 and 2 represent the first examples of Euphorbia diterpenoids with a 5/6 fused carbon ring system, and their plausible biosynthetic pathways originating from jatrophanes are proposed. Compounds 1 and 2 showed a triglyceride-lowering effect in oleic-acid-stimulated HuH7 cells at concentrations of 1-50 μM.

Amyloid-β (Aβ) and tau protein are two crucial hallmarks in Alzheimer's disease (AD). Their aggregation forms are thought to be toxic to the neurons in the brain. A series of new 1,2,3,4-tetrahydro-1-acridone analogues were designed, synthesized, and evaluated as potential dual inhibitors for Aβ and tau aggregation. In vitro studies showed that compounds 25-30 (20 μM) with N-methylation of the quinolone ring effectively inhibited Aβ1-42 aggregation by 84.7%-99.5% and tau aggregation by 71.2%-101.8%. Their structure-activity relationships are discussed. In particular, 30 could permeate the blood-brain barrier, bind to Aβ1-42 and tau, inhibit Aβ1-42 β-sheets formation, and prevent tau aggregation in living cells.

Our previous studies reveal that indolizinoquinolinedione scaffold is a base to develop novel DNA topoisomerase IB (TOP1) catalytic inhibitors. In this work, twenty-three novel indolizinoquinolinedione derivatives were synthesized. TOP1-mediated relaxation, nicking and unwinding assays revealed that three fluorinated derivatives 26, 28 and 29, and one N,N-trans derivative 46 act as TOP1 catalytic inhibitors with higher TOP1 inhibition (++++) than camptothecin (+++) and without TOP1-mediated unwinding effect. MTT assay against five human cancer cell lines indicated that the highest cytotoxicity is 20 for CCRF-CEM cells, 25 for A549 and DU-145 cells, 26 for HCT116 cells, and 33 for Huh7 cells with GI50 values at nanomolar range. The drug-resistant cell assay indicated that compound 26 may mainly act to TOP1 in cells and are less of Pgp substrates. Flow cytometric analysis showed that compounds 26, 28 and 29 can obviously induce apoptosis of HCT116 cells. Moreover, the structure-activity relationship (SAR) of indolizinoquinolinedione derivatives was analyzed.

Background Telomeric repeat–containing RNA (TERRA) is a large non-coding RNA in mammalian cells, which forms an integral component of telomeric heterochromatin. TERRA can bind to an allosteric site of telomeric repeat factor 2 (TRF2), a key component of Shelterin that protect chromosome termini. Both TERRA and TRF2 have been recognized as promising new therapeutic targets for cancer treatment. Methods Our methods include FRET assay, SPR, CD, microscale thermophoresis (MST), enzyme-linked immunosorbent assay (ELISA), chromatin immunoprecipitation (ChIP), colony formation assays, Western blot, immunofluorescence, cell cycle arrest and apoptosis detection, and xCELLigence real-time cell analysis (RTCA). Results In our routine screening of small molecule libraries, we found that a Quindoline derivative, CK1-14 could bind to and stabilize TERRA G-quadruplex structure, which could bind more tightly with an allosteric site of a telomeric binding protein TRF2, resulting in dissociation of TRF2 from telomeric DNA. Further in cellular studies indicated that the above effect of CK1-14 on TERRA G-quadruplex could activate DNA-damage response and cause cell cycle arrest, resulting in inhibition of U2OS cell proliferation and causing cell apoptosis. Conclusions Our mechanistic studies indicated that interaction of CK1-14 with TERRA induces telomeric DNA-damage response in U2OS cancer cells through inhibition of TRF2. CK1-14 could be further developed as a promising lead compound targeting telomere for cancer treatment. General significance Our present study provides the first evidence that allosteric modulation of TRF2 by TERRA G-quadruplex with a binding ligand could become a promising new strategy for cancer treatment especially for ALT tumor cells.

Telomeric repeat factor 2 (known as TRF2 or TERF2) is a key component of telomere protection protein complex named as Shelterin. TRF2 helps the folding of telomere to form T-loop structure and the suppression of ATM-dependent DNA damage response activation. TRF2 has been recognized as a potentially new therapeutic target for cancer treatment. In our routine screening of small molecule libraries, we found that Curcusone C had significant effect in disrupting the binding between TRF2 and telomeric DNA, with potent antitumor activity against cancer cells. Our result showed that Curcusone C could bind with TRF2 without binding interaction with TRF1 (telomeric repeat factor 1) although these two proteins share high sequence homology, indicating that their binding conformations and biological functions in telomere could be different. Our mechanistic studies showed that Curcusone C bound with TRF2 possibly through its DNA binding site causing blockage of its interaction with telomeric DNA. Further in cellular studies indicated that the interaction of TRF2 with Curcusone C could activate DNA-damage response, inhibit tumor cell proliferation, and cause cell cycle arrest, resulting in tumor cell apoptosis. Our studies showed that Curcusone C could become a promising lead compound for further development for cancer treatment. Here, TRF2 was firstly identified as a target of Curcusone C. It is likely that the anti-cancer activity of some other terpenes and terpenoids are related with their possible effect for telomere protection proteins.

Background and purpose Promoting energy metabolism is known to provide therapeutic effects for obesity and associated metabolic disorders. The present study evaluated the therapeutic effects of the newly identified bouchardatine (Bou) on obesity-associated metabolic disorders and the molecular mechanisms of these effects. Experimental approach The molecular mode of action of Bou for its effects on lipid metabolism was first examined in 3T3-L1 adipocytes and HepG2 cells. This was followed by an evaluation of its metabolic effects in mice fed a high-fat diet for 16 weeks with Bou being administered in the last 5 weeks. Further mechanistic investigations were conducted in pertinent organs of the mice and relevant cell models. Key results In 3T3-L1 adipocytes, Bou reduced lipid content and increased sirtuin 1 (SIRT1) activity to facilitate liver kinase B1 (LKB1) activation of AMPK. Chronic administration of Bou (50 mg∙kg−1 every other day) in mice significantly attenuated high-fat diet-induced increases in body weight gain, dyslipidaemia and fatty liver without affecting food intake and no adverse effects were detected. These metabolic effects were associated with activation of the SIRT1–LKB1–AMPK signalling pathway in adipose tissue and liver. Of particular note, UCP1 expression and mitochondrial biogenesis were increased in both white and brown adipose tissues of Bou-treated mice. Incubation with Bou induced similar changes in primary brown adipocytes isolated from mice. Conclusions and implications Bou may have therapeutic potential for obesity-related metabolic diseases by increasing the capacity of energy expenditure in adipose tissues and liver through a mechanism involving the SIRT1–LKB1–AMPK axis.

For the development of novel anticancer agents, we designed and synthesized a total of 37 perimidine o-quinone derivatives containing the o-quinone group at the A or B ring and different substituents (alkyl groups, aryl groups or heterocycles) at the C ring of the compounds. The structure-activity relationships (SARs) were established based on the cytotoxicity data of compounds from the HL-60, Huh7, Hct116, and Hela cell lines. The cytotoxicity results showed that most compounds exhibited potent cytotoxicity. In particular, compound b-12 showed the best anti-proliferative activity (IC50 ≤ 1 μM) against four cancer cell lines and strong potency against the HL-60/MX2 (0.47 μM) cell line, which is resistant to Topo II poisons. Further studies showed that b-12 exhibited potent Topo IIα inhibitory activity (IC50 = 7.54 μM) compared with Topo I, which acted as a class of non-intercalative Topo IIα catalytic inhibitor by inhibiting the ATP binding site of Topo II. Cell apoptosis and cell cycle assays confirmed that b-12 could induce the apoptosis of Huh7 cells in a dose-dependent manner.

Homologous recombination repair (HRR), a crucial approach in DNA damage repair, is an attractive target in cancer therapy and drug design. The Bloom syndrome protein (BLM) is a 3′–5′ DNA helicase that performs an important role in HRR regulation. However, limited studies about BLM inhibitors and their biological effects have been reported. Here, we identified a class of isaindigotone derivatives as novel BLM inhibitors by synthesis, screening, and evaluating. Among them, compound 29 was found as an effective BLM inhibitor with a high binding affinity and good inhibitory effect on BLM. Cellular evaluation indicated that 29 effectively disrupted the recruitment of BLM at DNA double-strand break sites, promoted an accumulation of RAD51, and regulated the HRR process. Meanwhile, 29 significantly induced DNA damage responses, as well as apoptosis and proliferation arrest in cancer cells. Our finding provides a potential anticancer strategy based on interfering with BLM via small molecules.

Multiple G-quadruplex units in the 3΄-terminal overhang of human telomeric DNA can associate and form multimeric structures. The specific targeting of such distinctive higher-order G-quadruplexes might be a promising strategy for developing selective anticancer agents with fewer side effects. However, thus far, only a few molecules were found to selectively bind to telomeric multimeric G-quadruplexes, and their effects on cancer cells were unknown. In this study, a new triaryl-substituted imidazole derivative called IZNP-1 was synthesized and found to specifically bind to and strongly stabilize telomeric multimeric G-quadruplexes through intercalating into the pocket between the two quadruplex units. The pocket size might affect the binding behavior of IZNP-1. Further cellular studies indicated that IZNP-1 could provoke cell cycle arrest, apoptosis and senescence in Siha cancer cells, mainly because of telomeric DNA damage and telomere dysfunction induced by the interactions of IZNP-1 with telomeric G-quadruplexes. Notably, IZNP-1 had no effect on the transcriptional levels of several common oncogenes that have the potential to form monomeric G-quadruplex structures in their promoter regions. Such behavior differed from that of traditional telomeric G-quadruplex ligands. Accordingly, this work provides new insights for the development of selective anticancer drugs targeting telomeric multimeric G-quadruplexes.

At present, wt1, a Wilms’ tumor suppressor gene, is recognized as a critical regulator of tumorigenesis and a potential therapeutic target. WT1 shows the ability to regulate the transcription of bcl-2 by binding to a GC-rich region in the promoter, which can then fold into a special DNA secondary structure called the G-quadruplex. This function merits the exploration of the effect of a G-quadruplex ligand on the binding and subsequent regulation of WT1 on the bcl-2 promoter. In the present study, WT1 was found to bind to the double strand containing the G-quadruplex-forming sequence of the bcl-2 promoter. However, the G-quadruplex ligand SYUIQ-FM05 effectively blocked this binding by interacting with the GC-rich sequence. Our new findings are significant in the exploration of new strategies to block WT1's transcriptional regulation for cancer-cell treatment., Highlights • WT1 bound to double-stranded but not G-quadruplex structure in bcl-2′s promoter. • G-quadruplexes ligand SYUIQ-FM05 blocked the binding of WT1 to bcl-2. • SYUIQ-FM05's regulation effects depends on its interaction with GC-rich sequence.

Aberrant overexpression of heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a key feature in oncogenesis and progression of many human cancers. hnRNP K has been found to be a transcriptional activator to up-regulate c-myc gene transcription, a critical proto-oncogene for regulation of cell growth and differentiation. Therefore, down-regulation of c-myc transcription by inhibiting hnRNP K through disrupting its binding to c-myc gene promoter is a potential approach for cancer therapy. In the present study, we synthesized and screened a series of Quinoline derivatives and evaluated their binding affinity for hnRNP K. Among these derivatives, (E)-1-(4-methoxyphenyl)-3-(4-morpholino-6-nitroquinolin-2-yl)prop-2-en-1-one (compound 25) was determined to be the first-reported hnRNP K binding ligand with its KD values of 4.6 and 2.6 μM measured with SPR and MST, respectively. Subsequent evaluation showed that the binding of compound 25 to hnRNP K could disrupt its unfolding of c-myc promoter i-motif, resulting in down-regulation of c-myc transcription. Compound 25 showed a selective anti-proliferative effect on human cancer cell lines with IC50 values ranged from 1.36 to 3.59 μM. Compound 25 exhibited good tumor growth inhibition in a Hela xenograft tumor model, which might be related to its binding with hnRNP K. These findings illustrated that inhibition of DNA-binding protein hnRNP K by compound 25 could be a new and selective strategy of regulating oncogene transcription instead of targeting promoter DNA secondary structures such as G-quadruplexes or i-motifs.

Mulberry Diels-Alder-type adducts (MDAAs) are a group of structurally unique natural products biosynthetically derived from the intermolecular [4 + 2] cycloaddition of a dehydroprenylphenol and a chalcone. In the current study, ten MDAAs, including an undescribed one, inethermulberrofuran C, were isolated from the root bark of Morus alba. The anti-Alzheimer's disease (anti-AD) properties of these isolates were systematically screened for a series of potential targets (Aβ self-aggregation, tau aggregation, and ChEs) as well as the anti-neuroinflammatory and neuroprotective activities. Four compounds, mulberrofuran C, mulberrofuran K, mulberrofuran G, and isomulberrofuran G, turned out to be potent multi-targeted agents for AD. Among them, mulberrofuran K with a good blood-brain barrier (BBB) permeability (8.7 ± 0.3 × 10−6 cm/s) was selected as a promising candidate for further mechanism study in glutamate-induced HT22 cell model, which showed its neuroprotective ability on up-regulation of the glutathione (GSH) level and suppression of the reactive oxygen species (ROS) production.

The guanine-rich sequences are able to fold into G-quadruplexes in living cells, making these structures promising anti-cancer drug targets. In the current study, we identified a small molecule, Ber8, from a series of 9-substituted berberine derivatives and found that it could induce acute cell growth arrest and senescence in cancer cells, but not in normal fibroblasts. Further analysis revealed that the cell growth arrest was directly associated with apparent cell cycle arrest, cell senescence, and profound DNA damage at the telomere region. Significantly, our studies also provided evidence that Ber8 could stabilize endogenous telomeric G-quadruplexes structures in cells. Ber8 could then induce the delocalization of TRF1 and POT1 from the telomere accompanied by a rapid telomere uncapping. These results provide compelling insights into direct binding of telomeric G-quadruplexes and might contribute to the development of more selective, effective anticancer drugs.

In our previous work, indolizinoquinolinedione derivative 1 was identified as a Top1 catalytic inhibitor. Herein, a series of 6-substituted indolizinoquinolinedione derivatives were synthesized through modification of the parent compound 1. Top1 cleavage and relaxation assays indicate that none of these novel compounds act as classical Top1 poison, and that the compounds with alkylamino terminus at C-6 side chain, including 8, 11-16, 18-21, 25, 26 and 28-30, are the most potent Top1 catalytic inhibitors. Top1-mediated unwinding assay demonstrated that 14, 22 and 26 were Top1 catalytic inhibitors without Top1-mediated unwinding effect. Moreover, MTT results showed that compounds 26, 28-30 exhibit significant cytotoxicity against human leukemia HL-60 cells, and that compound 26 exerts potent cytotoxicity against A549 lung cancer cells at nanomolar range.

c-MYC is an important oncogene that is considered as an effective target for anticancer therapy. Regulation of this gene's transcription is one avenue for c-MYC-targeting drug design. Direct binding to a transcription factor and generating the intervention of a transcriptional programme appears to be an effective way to modulate gene transcription. NM23-H2 is a transcription factor for c-MYC and is proven to be related to the secondary structures in the promoter. Here, we first screened our small-molecule library for NM23-H2 binders and then sifted through the inhibitors that could target and interfere with the interaction process between NM23-H2 and the guanine-rich promoter sequence of c-MYC. As a result, a quinazolone derivative, SYSU-ID-01: , showed a significant interference effect towards NM23-H2 binding to the guanine-rich promoter DNA sequence. Further analyses of the compound-protein interaction and the protein-DNA interaction provided insight into the mode of action for SYSU-ID-01: . Cellular evaluation results showed that SYSU-ID-01: could abrogate NM23-H2 binding to the c-MYC promoter, resulting in downregulation of c-MYC transcription and dramatically suppressed HeLa cell growth. These findings provide a new way of c-MYC transcriptional control through interfering with NM23-H2 binding to guanine-rich promoter sequences by small molecules.

G-quadruplexes are four-stranded DNA structures formed from G-rich sequences that are built around tetrads of hydrogen-bonded guanine bases. Accumulating studies have revealed that G-quadruplex structures are formed in vivo and play important roles in biological processes such as DNA replication, transcription, recombination, epigenetic regulation, meiosis, antigenic variation, and maintenance of telomeres stability. Mounting evidence indicates that a variety of proteins are capable of binding selectively and tightly to G-quadruplex and play essential roles in G-quadruplex-mediated regulation processes. Some of these proteins promote the formation or/and stabilization of G-quadruplex, while some other proteins act to unwind G-quadruplex preferentially. From a drug discovery perspective, many of these G-quadruplex binding proteins and/or their complexes with G-quadruplexes are potential drug targets. Here, we present a general summary of reported G-quadruplex binding proteins and their biological functions, with focus on those of medicinal research significance. We elaborated the possibility for some of these G-quadruplex binding proteins and their complexes with G-quadruplexes as potential drug targets.

Novel topoisomerase II (Topo II) inhibitors have gained considerable interest for the development of anticancer agents. In this study, a series of carbazole derivatives containing chalcone analogs (CDCAs) were synthesized and investigated for their Topo II inhibition and cytotoxic activities. The results from Topo II mediated DNA relaxation assay showed that CDCAs could significantly inhibit the activity of Topo II, and the structure-activity relationship indicated the halogen substituent in phenyl ring play an important role in the activity. Further mechanism studies revealed that CDCAs function as non-intercalative Topo II catalytic inhibitors. Moreover, some CDCAs showed micromolar cytotoxic activities. The most potent compound 3h exhibited notable growth inhibition against four human cancer cell lines. Flow cytometric analysis revealed that compounds 3d and 3h arrested the HL-60 cells in sub G1 phase by induction of apoptosis. It was further confirmed by Annexin-V-FITC binding assay. Western blot analysis revealed that compound 3h induces apoptosis likely through the activation of caspase proteins.

Background Alzheimer's disease (AD) is a progressive neurodegenerative brain disorder that is characterized by dementia, cognitive impairment, and memory loss. Diverse factors are related to the development of AD, such as increased level of β-amyloid (Aβ), acetylcholine, metal ion deregulation, hyperphosphorylated tau protein, and oxidative stress. Methods The following methods were used: organic syntheses of 1H-phenanthro[9,10-d]imidazole derivatives, inhibition of self-mediated and metal-induced Aβ1–42 aggregation, inhibition studies for acetylcholinesterase and butyrylcholinesterase, anti-oxidation activity studies, CD, MTT assay, transmission electron microscopy, dot plot assay, gel electrophoresis, Western blot, and molecular docking studies. Results We synthesized and characterized a new type of 1H-phenanthro[9,10-d]imidazole derivatives as multifunctional agents for AD treatment. Our results showed that most of these derivatives exhibited strong Aβ aggregation inhibitory activity. Compound 9g had 74% Aβ1–42 aggregation inhibitory effect at 10 μM concentration with its IC50 value of 6.5 μM for self-induced Aβ1–42 aggregation. This compound also showed good inhibition of metal-mediated (Cu2 + and Fe2 +) and acetylcholinesterase-induced Aβ1–42 aggregation, as indicated by using thioflavin T assay, transmission electron microscopy, gel electrophoresis, and Western blot. Besides, compound 9g exhibited cholinesterase inhibitory activity, with its IC50 values of 0.86 μM and 0.51 μM for acetylcholinesterase and butyrylcholinesterase, respectively. In addition, compound 9g showed good anti-oxidation effect with oxygen radical absorbance capacity (ORAC) value of 2.29. Conclusions Compound 9g was found to be a potent multi-target-directed agent for Alzheimer's disease. General significance Compound 9g could become a lead compound for further development as a multi-target-directed agent for AD treatment.

Background Angiogenesis is thought to be important in tumorigenesis and tumor progress. Vascular endothelial growth factor (VEGF) is a pluripotent cytokine and angiogenic growth factor that plays crucial roles in embryonic development and tumor progression. In many types of cancer, VEGF is overexpressed and is generally associated with tumor progression and survival rate. The polypurine/polypyrimidine sequence located upstream of the promoter region in the human VEGF gene can form specific parallel G-quadruplex structures, raising the possibility for transcriptional control of VEGF through G-quadruplex ligands. Methods PCR stop assay, circular dichroism (CD) spectra, RNA extraction and RT-PCR, enzyme-linked immunosorbent assay (ELISA), luciferase Assays, cell scrape test, xCELLigence real-time cell analysis (RTCA), and chick embryo chorioallantoic membrane (CAM) assay. Results and conclusions We found that quindoline derivatives can interact with the G-rich DNA sequences of the VEGF promoter to stabilize this G-quadruplex and suppress the transcription and expression of the VEGF protein. We also demonstrated that these derivatives exhibit potential anti-angiogenic activity in chick embryos and antitumor activity, including the inhibition of cell proliferation and migration. General significance Our new findings have significances not only for understanding the mechanism of the G-quadruplex ligands mediating the VEGF transcription inhibition, but also for exploring a new anti-tumor strategy to blocking the transcription of VEGF to inhibit the angiogenesis in cancer cells.

Background Cellular nucleic acid binding protein (CNBP) has been implicated in vertebrate craniofacial development and in myotonic dystrophy type 2 (DM2) and sporadic inclusion body myositis (sIBM) human diseases by controlling cell proliferation and survival to mediate neural crest expansion. CNBP has been found to bind single-stranded nucleic acid and promote rearrangements of nucleic acid secondary structure in an ATP-independent manner, acting as a nucleic acid chaperone. Methods A variety of methods were used, including cell viability assays, wound-scratch assays, chemotaxis assays, invasion assays, circular dichroic (CD) spectroscopy, NMR spectroscopy, chromatin immunoprecipitation, expression and purification of recombinant human CNBP, electrophoretic mobility shift assay (EMSA), surface plasmon resonance (SPR), fluorescence resonance energy transfer (FRET) analyses, luciferase reporter assay, Western blotting, and isothermal titration calorimetry (ITC). Results Up-regulation of CNBP induced human fibrosarcoma cell death and suppressed fibrosarcoma cell motility and invasiveness. It was found that CNBP transcriptionally down-regulated the expression of heterogeneous ribonucleoprotein K (hnRNP K) through its conversion of a G-rich sequence into G-quadruplex in the promoter of hnRNP K. G-quadruplex stabilizing ligand tetra-(N-methyl-4-pyridyl) porphyrin (TMPyP4) could interact with and stabilize the G-quadruplex, resulting in downregulation of hnRNP K transcription. Conclusions CNBP overexpression caused increase of cell death and suppression of cell metastasis through its induction of G-quadruplex formation in the promoter of hnRNP K resulting in hnRNP K down-regulation. General significance The present result provided a new solution for controlling hnRNP K expression, which should shed light on new anticancer drug design and development.

Background Telomere is protected by its G-quadruplex, T-loop structure, telomerase, and binding protein complex. Protein POT1 (protection of telomeres 1) is one subunit of telomere binding protein complex Shelterin. POT1 acts as a regulator of telomerase-dependent telomere length, and it can help telomere to form D-loop structure to stabilize telomere. POT1 protects telomere ends from ATR-dependent DNA damage response as well. Methods Extensive methods were used, including CD, EMSA, ITC, PCR stop assay, luciferase reporter assay, quantitative real-time PCR, Western blot, chromatin immunoprecipitation (Ch-IP), cloning, expression and purification of proteins. Results We found a new G-rich 30-base-pair long sequence (P-pot1 G18) located from − 165 to − 136 base pairs upstream of the translation starting site of protein POT1. This sequence in the promoter region of pot1 gene formed G-quadruplex resulting in down-regulation of pot1 gene transcription. This G-rich sequence is close to a binding site “TCCC” for transcription factor hnRNP K (heterogeneous nuclear ribonucleoprotein K), and its conversion to G-quadruplex prevented the access of hnRNP K to this binding site. The binding of hnRNP K could up-regulate pot1 gene transcription. TMPyP4 (meso-tetra(N-methyl-4-pyridyl)porphine) has been widely used as G-quadruplex binding ligand, which stabilized the G-quadruplex in vitro and in cellulo, resulting in down-regulation of pot1 gene transcription. Conclusions This G-quadruplex might become a potentially new drug target for antitumor agents. General significance Our results first demonstrated that G-quadruplex formation can affect the binding of transcription factor to its nearby binding site, and thus making additional influence to gene transcription.

N'-(7-Fluoro-5-N-methyl-10H-indolo[3,2-b]quinolin-5-ium)-N,N-dimethylpropane-1,3-diamine iodide (SYUIQ-FM05) is a semi-synthetic derivative of cryptolepine which is from Cryptolepis sanguinolenta (Lindl.) Schlechter (Periplocaeae). This ligand inhibits telomerase activity by stabilizing the G-quadruplex structure and induces growth arrest in cancer cells.The anticancer activity of SYUIQ-FM05 via inhibiting c-kit transcription was investigated in leukemic cells.The cytotoxicity of SYUIQ-FM05 in K562 cells was evaluated using a cell viability assay and flow cytometry (FCM) at 0.4, 2.0, 10.0 and 20.0 nM. Under the same concentrations of SYUIQ-FM05 or 100 nM imatinib mesylate (IM), quantitative polymerase chain reaction (Q-PCR) investigated transcription of c-kit and bcl-2, and western blotting analyzed the expression levels of c-Kit, total mitogen-activated protein kinase kinases (MEKs), phospho-MEK (p-MEK), total extracellular regulated protein kinases (ERKs), phospho-ERK (p-ERK), Bcl-2 and Bax.SYUIQ-FM05 inhibited cellular growth with an IC(50) of 10.83 ± 0.05 nM in K562 cells. c-Kit transcription was suppressed 2.69-, 4.39-, 7.71- and 10.52-fold at 0.4, 2.0, 10.0 and 20.0 nM SYUIQ-FM05, respectively, which produced proportional loss of total c-Kit protein except IM. Both SYUIQ-FM05 and IM downregulated p-MEK and p-ERK. Furthermore, bcl-2 transcription was suppressed 1.58- and 1.86-fold at 10.0 and 20.0 nM SYUIQ-FM05, respectively, but 0.4 and 2.0 nM SYUIQ-FM05 had no effect. A decrease in Bcl-2 and an increase in Bax appeared in these treated cells.These findings demonstrate that SYUIQ-FM05 could induce apoptosis in a leukemic cell line through inhibiting c-kit transcription, which supports the anticancer potency of SYUIQ-FM05 in c-Kit-positive leukemic cells.

A series of 4-anilinoquinazoline derivatives were designed and synthesized as novel c-myc promoter G-quadruplex binding ligands. Subsequent biophysical and biochemical evaluation demonstrated that the introduction of aniline group at 4-position of quinazoline ring and two side chains with terminal amino group improved their binding affinity and stabilizing ability to G-quadruplex DNA. RT-PCR assay and Western blot showed that compound 7a could down-regulate transcription and expression of c-myc gene in Hela cells, which was consistent with the behavior of an effective G-quadruplex ligand targeting c-myc oncogene. More importantly, RTCA and colony formation assays indicated that 7a obviously inhibited Hela cells proliferation, without influence on normal primary cultured mouse mesangial cells. Flow cytometric assays suggested that 7a induced Hela cells to arrest in G0/G1 phase both in a time-dependent and dose-dependent manner.

A series of isaindigotone derivatives and analogues were designed, synthesized and evaluated as dual inhibitors of cholinesterases (ChEs) and self-induced β-amyloid (Aβ) aggregation. The synthetic compounds had IC50 values at micro or nano molar range for cholinesterase inhibition, and some compounds exhibited strong inhibitory activity for AChE and high selectivity for AChE over BuChE, which were much better than the isaindigotone derivatives previously reported by our group. Most of these compounds showed higher self-induced Aβ aggregation inhibitory activity than a reference compound curcumin. The structure–activity relationship studies revealed that the derivatives with higher inhibition activity on AChE also showed higher selectivity for AChE over BuChE. Compound 6c exhibiting excellent inhibition for both AChE and self-induced Aβ aggregation was further studied using CD, EM, molecular docking and kinetics.

A series of novel 7-alkoxyl substituted indolizinoquinoline-5,12-dione derivatives were synthesized. The cholinesterase inhibition assays indicated that most synthesized compounds exhibited good activity for acetylcholinesterase (AChE) and high selectivity index of AChE over butyrylcholinesterase (BuChE). Compound 12b exhibited the most potent AChE inhibitory activity with an IC(50) value of 0.068 µM and the highest selectivity index of 144. Kinetic study of AChE indicated that a mixed type of inhibition pattern existed for these indolizinoquinoline-5,12-dione derivatives. Molecular docking study indicated that compound 12b could bind to both the catalytically active site and the peripheral anionic site of AChE.

A series of 9-bromo-substituted indolizinoquinoline-5,12-dione derivatives was synthesized. Antimicrobial activity assessment indicates that compounds 1, 26, 27 and 28 exhibit strong activity against gram-positive bacterial strains, including Beta-hemolytic streptococcus CMCC32210, Staphylococcus aureus ATCC25923, Staphylococcus epidermidis ATCC12228, Enterococcus faecalis ATCC29212 and methicillin-resistant S. aureus ATCC43300 (MRSA). Compound 27 shows the best anti-MRSA activity with an MIC value of 0.031 μg/ml. To assess the mechanism of action, the inhibitory activities of compound 1 against DNA gyrase and DNA topoisomerase IV were also measured. The results indicate that compound 1 has strong inhibitory effects on the Escherichia coli DNA gyrase supercoiling activity and S. aureus Topo IV relaxing activity.

A series of mansonone F (MF) derivatives were designed and synthesized. These compounds were found to be strong inhibitors for topoisomerases, with much more significant inhibition for topoisomerase II rather than topoisomerase I. The best inhibitor showed 20 times stronger anti-topoisomerase II activity than a positive control Etoposide. The cytotoxic activity of these MF derivatives was evaluated against human cancer cell lines CNE-2 and Glc-82, which showed that these compounds were potent antitumor agents. The structure–activity relationships (SARs) study revealed that o-quinone group and pyran ring are important for their cytotoxic activity.

A new series of heterobivalent tacrine derivatives were designed, synthesized and evaluated as potential multi-functional anti-Alzheimer agents for their inhibitory activity on cholinesterases, antioxidant activity and self-induced β-amyloid (Aβ) aggregation. All these synthesized compounds had potent inhibition activity on acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) at nanomolar range. A Lineweaver-Burk plot and molecular modeling study showed that these compounds targeted both the catalytic active site (CAS) and the peripheral anionic site (PAS) of AChE. The compounds containing hydroxyl group showed potent peroxyl radical absorbance activity. In addition, compound 5j exhibited higher self-induced Aβ aggregation inhibitory activity than curcumin, which could become a multi-functional agent for further development for the treatment of AD.

A series of 3-substituted (5c-5f, 6c-6f) and 4-substituted (10a-10g) oxoisoaporphine derivatives were synthesized. It was found that all these synthetic compounds had IC50 values at micro or nano molar range for cholinesterase inhibition, and most of them could inhibit amyloid β (Aβ) self-induced aggregation with inhibition ratio from 31.8% to 57.6%. The structure-activity relationship studies revealed that the derivatives with higher selectivity on AChE also showed better inhibition on Aβ self-induced aggregation. The results from cell toxicity study indicated that most quaternary methiodide salts had higher IC50 values than the corresponding non-quaternary compounds. This study provided potentially important information for further development of oxoisoaporphine derivatives as lead compounds for the treatment of Alzheimer's disease.

A novel series of N,N'-bis-methylenedioxybenzyl-alkylenediamines 5a-5g have been designed, synthesized and evaluated as bivalent anti-Alzheimer's disease ligands. The enzyme inhibition assay results indicated that compounds 5e-5g inhibit both acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) in the micromolar range (IC(50), 2.76-4.24 µM for AChE and 3.02-5.14 µM for BuChE), which was in the same potential as the reference compound rivastigmine (IC(50), 5.50 µM for AChE and 1.60 µM for BuChE). It was found that compounds could bind simultaneously to the peripheral and catalytic sites of AChE. β-Amyloid (Aβ) aggregation inhibition assay results showed that compound 5e exhibited highest self-mediated Aβ fibril aggregation inhibition activity (40.3%) with a similar potential as curcumin (41.6%). It was also found that 5e-5g did not affect neuroblastoma cell viability at the concentration of 50 μM.

A series of novel indolizinophthalazine-5,12-dione derivatives were designed and synthesized by the reaction of 6,7-dichlorophthalazine-5,8-dione with active methylene reagents (AMR) and pyridine derivatives. Some of synthesized compounds exhibited significant in vitro antiproliferative activity at micromolar level toward four human tumor cell lines, including lung adenocarcinoma cell, large-cell lung carcinoma cell, breast carcinoma cell and ardriamycin-resistance breast carcinoma cell. The DNA topoisomerase IB inhibitory assay indicated that DNA topoisomerase IB might be a biological target of the synthesized compounds.

Mammalian thioredoxin reductase (TrxR), a ubiquitous selenocysteine containing oxidoreductase, catalyzes the NADPH-dependent reduction of oxidized thioredoxin (Trx). TrxR has been suggested as a potential target for anticancer drugs development for its overexpression in human tumors and its diverse functions in intracellular redox control, cell growth and apoptosis. Mansonone F (MF) compounds have been shown to exhibit antiproliferative effects, but their complex mechanisms are unknown. In the present study, we have investigated the effects of some synthesized MF analogues on TrxR and HeLa cells. The studies of the mode of inhibition and the interactions of IG3, one of the most potent MF analogues, with TrxR showed MF compounds could be partly reduced by the C-terminal selenolthiol active site, and possibly by the N-terminal dithiol motif and/or FAD domain of TrxR simultaneously, accompanied by redox cycling with the generation of superoxide anion radicals. In addition, MF analogues exhibited the potential to inhibit the growth of HeLa cells and reduce TrxR activity in cell lysates. The cell cycle was arrested in G2/M phase and apoptosis was induced in a dose-dependent manner. Furthermore, our results showed that IG3-treated HeLa cells induced the change of intracellular ROS. Taken together, the reported results here suggest that inhibition of TrxR by MF analogues provides a possible complex mechanism for explaining the anticancer activity of MF compounds.

G-quadruplexes are specialized secondary structures in nucleic acids that possess significant conformational polymorphisms. The precise G-quadruplex conformations in vivo and their relevance to biological functions remain controversial and unclear, especially for telomeric G-quadruplexes. Here, we report a novel single-chain variable fragment (scFv) antibody, D1, with high binding selectivity for parallel G-quadruplexes in vitro and in vivo. Genome-wide chromatin immunoprecipitation using D1 and deep-sequencing revealed the consensus sequence for parallel G-quadruplex formation, which is characterized by G-rich sequence with a short loop size (

G-quadruplex nucleic acids are four-stranded DNA or RNA secondary structures that are formed in guanine-rich sequences. These structures exhibit extensive structural polymorphism and play a pivotal role in the control of a variety of cellular processes. To date, diverse approaches for high-throughput identification of G-quadruplex structures have been successfully developed, but high-throughput methods for further characterization of their topologies are still lacking. In this study, we report a new tetra-arylimidazole probe psIZCM-1, which was found to display significant and distinctive changes in both the absorption and the fluorescence spectra in the presence of parallel G-quadruplexes but show insignificant changes upon interactions with anti-parallel G-quadruplexes or other non-quadruplex oligonucleotides. In view of this dual-output feature, we used psIZCM-1 to identify the parallel G-quadruplexes from a large set of 314 oligonucleotides (including 300 G-quadruplex-forming oligonucleotides and 14 non-quadruplex oligonucleotides) via a microplate reader and accordingly established a high-throughput method for the characterization of parallel G-quadruplex topologies. The accuracy of this method was greater than 95%, which was much higher than that of the commercial probe NMM. To make the approach more practical, we further combined psIZCM-1 with another G-quadruplex probe IZCM-7 to realize the high-throughput classification of parallel, anti-parallel G-quadruplexes and non-quadruplex structures.

Novel topoisomerase II (Topo II) inhibitors have gained considerable interest for the development of anticancer agents. In this study, a series of 1,3-benzoazolyl-substituted pyrrolo[2,3-b]pyrazine derivatives were designed, synthesized, and evaluated as potential Topo II catalytic inhibitors. It was found that some of derivatives had good antiproliferative activity on seven cancer cell lines, especially on HL-60/MX2, a cancer cell line derivative from HL-60 that is resistant to Topo II poison. Topo II mediated DNA relaxation assay results showed that derivatives could significantly inhibit the activity of Topo II, and the structure-activity relationship studies indicated the importance of the alkylamino side chain and the benzoazolyl group. Further mechanism studies revealed that derivatives function as Topo II nonintercalative catalytic inhibitors and may block the ATP binding site of Topo II. Moreover, flow cytometric analysis showed that this class of compounds could induce apoptosis of HL-60 cells.

Our recent study has shown that the natural product bouchardatine (1) can reduce the triglyceride (TG) content in 3T3-L1 adipocytes (EC50 ≈ 25 μM). Here, we synthesized two series of compounds by introducing amine side chains at the 5 or 8 position of 1 and evaluated the lipid-lowering activity of derivatives. It was found that some of the compounds had significant lipid-lowering effects, and the most active compound 3d showed better activity (EC50 = 0.017 μM) than 2 (EC50 = 0.086 μM), a compound reported by us. Further, the mechanism studies revealed that 3d blocked TG accumulation via activation of the LKB1-AMPK signaling pathway, efficiently down-regulating the expression of key regulators of adipogenesis/lipogenesis. Cell uptake assay and confocal imaging of 3d in cells indicated that compound 3d had favorable cell permeability. Our results suggest that 3d may be a promising agent for the treatment of obesity and related metabolic disorders.

Environments surrounding G-rich sequences remarkably affect the conformations of these structures. A proper evaluation system mimicking the crowded environment in a cell with macromolecules should be developed to perform structural and functional studies on G-quadruplexes. In this study, the topology and stability of a G-quadruplex formed by human telomeric repeat sequences were investigated in a macromolecule-crowded environment created by polyethylene glycol 200 (PEG200), tumor cell extract, and Xenopus laevis egg extract. The interactions between small molecules and telomeric G-quadruplexes were also evaluated in the different systems. The results suggested that the actual behavior of G-quadruplex structures in cells extract is quite different from that in the PEG crowding system, and proteins or other factors in extracts might play a very important role in G-quadruplex structures., Highlights • Cell-free system was constructed using HL60 cell extract. • Topologies and stability of G-quadruplexes were identified in different systems. • G-quadruplex’s ligands’ effects were evaluated in different systems.

Up-regulation of a disintegrin and metalloproteinase 10 (ADAM10) to prevent the formation of β-amyloid (Aβ) peptides might be a promising strategy to treat Alzheimer's disease (AD). RNA G-quadruplex motif within the 5'-UTR of the ADAM10 mRNA is an inhibitory element for ADAM10 translation. Thus, mitigation of the suppressive effect of this motif using an RNA G-quadruplex-forming G-rich sequence (QGRS) binder might be a new approach for AD therapy. Herein, a series of new methylquinolinium derivatives were synthesized and screened by surface plasmon resonance (SPR) and the dual-luciferase reporter assay. Among them, compound 24 showed selective affinity for the QGRS of ADAM10 and could strongly up-regulate the translation of it. Moreover, treatment with 24 led to a significant increase of the secretion of sAPPα, consequently decreasing the Aβ40 in cellular. These results illustrate that the interaction between the RNA QGRS and a small molecule may be a new molecular strategy to modulate the translation of ADAM10.

Bouchardatine (1), a naturally occurring β-indoloquinazoline alkaloid, was synthesized. For the first time, the lipid-lowering effect and mechanism of 1 was investigated in 3T3-L1 adipocytes. Our study showed that 1 could significantly reduce lipid accumulation without cytotoxicity and mainly inhibited early differentiation of adipocyte through proliferation inhibition and cell cycle arrested in dose-dependent manner. Furthermore, the inhibition of early differentiation was reflected by down-regulation of key regulators of adipogenesis/lipogenesis, including CCAAT enhancer binding proteins (C/EBPβ, C/EBPδ, C/EBPα), peroxisome proliferator-activated receptors γ (PPARγ) and sterol-regulatory element binding protein-1c (SREBP-1c), in both of mRNA and protein levels. Subsequently decreasing the protein levels of acetyl CoA carboxylase (ACC), fatty acid synthase (FAS), and stearyl coenzyme A desaturated enzyme 1 (SCD-1), the rate-limited metabolic enzymes of fatty acid synthesis, were also observed. Further studies revealed that 1 persistently activated adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK) during differentiation, suggesting that the AMPK may be an upstream mechanism for the effect of 1 on adipogenesis and lipogenesis. Our data suggest that 1 can be a candidate for the development of new therapeutic drugs against obesity and related metabolic disorders.

Natural curcumin (1), demethoxycurcumin (2) and bisdemethoxycurcumin (3) isolated from Curcuma longa (turmeric), and synthetic curcumin analogs (A(1-7), B(1-7), C(1-6) and D(1-7)) were evaluated in vitro for the alpha-glucosidase inhibitory activity via UV and circular dichroism (CD) spectroscopy. The results indicated that natural curcuminoid compound 3 showed a remarkable inhibitory effect with IC(50) of 23.0 microM, and the synthetic compounds A(2), B(2), C(2) and D(2) showed potent inhibitory effects with IC(50) of 2.8, 2.6, 1.6 and 8.2 microM, respectively. Kinetic study exhibited that the mechanism of alpha-glucosidase inhibition of both 3 and C(2) was non-competitive. The structure activity relationship revealed that the ortho dihydroxyl groups could form a more tight interaction with alpha-glucosidase to exert more potential inhibitory activities.

In this study, chitosan microspheres and sponges with uniform spherical and porousmorphologies were prepared by coiling the stretched chains of chitosan with addition of salt and choosingdifferent kinds of organic solvents as evaporation solvents. Cellulase was immobilized to the support by acovalent method. The enzyme exhibited a considerable affinity to the support, and the protein loading of145.5mgg − 1 support was fairly high. The immobilized cellulase had a higher K m than free cellulase andhad better stability with respect to pH, thermal stability, reuses and storage stability than free cellulase.  2005 Society of Chemical Industry Keywords: chitosan; microsphere; sponge; immobilized cellulase; ammonium sulfate INTRODUCTION Cellulosic material is an abundant renewable resourcethat can serve as substrate for the production of chem-icals and fuel ethanol by enzymatic conversion. 1 , 2 Cellulase can be used as the biocatalyst for cellu-lose hydrolysis, but its activity is rather lower thanthat of amylase. Unfavorable economic problemshave restricted the development of this process. Inorder to reduce the cost and raise the activity ofcellulase, immobilization of cellulase

Synergetic inhibitory effect of genistein (I2) and d -glucose (I1) on α-glucosidase has been studied with kinetics method. It was concluded that the inhibitory effect was much greater when I2 and I1 were both added to the reactant solution simultaneously than that they were added, respectively, which suggesting the inhibitors bind to the different sites of α-glucosidase at the same time, and demonstrating synergetic inhibition.

A novel series of benzo[a]phenazin derivatives bearing alkylamino side chains were designed, synthesized and evaluated for their topoisomerases inhibitory activity as well as cytotoxicity against four human cancer cell lines (HL-60, K-562, HeLa, and A549). These compounds were found to be dual inhibitors of topoisomerase (Topo) I and Topo II, and exhibited excellent antiproliferative activity, in particular against HL-60 cells with submicromolar IC50 values. Further mechanistic studies showed that this class of compounds acted as Topo I poisons by stabilizing the Topo I-DNA cleavage complexes and Topo II catalytic inhibitors by inhibiting the ATPase activity of hTopo II. Molecular docking studies revealed the binding modes of these compounds for Topo I and Topo II.

Background Accelerated proliferation of solid tumor and hematologic cancer cells is related to accelerated transcription of ribosomal DNA by the RNA polymerase I to produce elevated level of ribosomal RNA. Therefore, down-regulation of RNA polymerase I transcription in cancer cells is an important anticancer therapeutic strategy. Methods A variety of methods were used, including cloning, expression and purification of protein, electrophoretic mobility shift assay (EMSA), circular dichroic (CD) spectroscopy, CD-melting, isothermal titration calorimetry (ITC), chromatin immunoprecipitation (Ch-IP), RNA interference, RT-PCR, Western blot, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) cell assay. Results Our results showed that 2,4-disubstituted quinazoline derivative Sysu12d could down-regulate c-myc through stabilization of c-myc promoter G-quadruplex, resulting in down-regulation of nucleolin expression. Sysu12d could also disrupt nucleolin/G-quadruplex complex. Both of the above contributed to the down-regulation of ribosomal RNA synthesis, followed by activation of p53 and then cancer cell apoptosis. Conclusions These mechanistic studies set up the basis for further development of Sysu12d as a new type of lead compound for cancer treatment. General significance 2,4-Disubstituted quinazoline derivatives may have multi-functional effect for cancer treatment.