氮杂甾体吡啶类衍生物的合成及抗肿瘤活性

利用药物设计的拼合原理,以黄体酮为原料,经过氧化开环、闭环、还原及缩合反应等步骤,设计合成了9个目标化合物,其结构经IR、NMR和HR-MS确证。以阿比特龙和5-氟脲嘧啶为阳性对照,采用MTT法初步评价目标化合物对体外人前列腺癌细胞PC-3、人卵巢癌细胞Skov3、人恶性黑色瘤细胞A375和人正常前列腺上皮细胞RWPE-1的抗增殖活性。结果显示,2-氨基-4-吡啶-4-基-6-(4′-氮杂孕甾-3′-酮-17′-基)烟腈、2-氨基-4-呋喃-2-基-6-(4′-氮杂孕甾-3′-酮-17′-基)烟腈对PC-3和Skov3细胞抗增殖活性优于阳性对照药阿比特龙和5-氟脲嘧啶。特别是2-氨基-4-吡啶-4-基-6-(4′-氮杂孕甾-3′-酮-17′-基)烟腈对PC-3细胞的IC₅₀值为(2.73±0.80)μmol/L,明显优于阿比特龙,值得进一步研究。     

蒎烷基噻唑腙类衍生物的合成及抗肿瘤活性研究

以诺蒎酮为原料,经过加成、缩合、环化等反应,合成出蒎烷基噻唑腙衍生物4a～4n,采用FTIR、1HNMR、13CNMR和HRMS对化合物的结构进行了表征。考察了化合物4a～4n对人肝癌细胞(HepG2)、人多发性骨髓瘤细胞(RPMI-8226)、人肺癌细胞(A549)和乳腺癌细胞(MDA-MB-231)的抗肿瘤活性。实验结果表明,化合物4a的抗肿瘤活性最强,其对Hep G2、8226、A549和231细胞的IC50分别低至5.8、8.8、7.1和10.6μmol/L;化合物4c的抗肿瘤活性也较强,其IC50分别为8.9、8.7、7.3和9.7μmol/L。细胞凋亡和周期实验数据显示,当化合物4a浓度从0增加到40μmol/L时,A549细胞的总凋亡率从6.55%增加到47.20%,G2/M期的细胞数量从13.50%上升至51.72%。以上结果表明,化合物4a能够诱导A549细胞凋亡,并将细胞有丝分裂周期阻滞在G2/M期。     

异长叶烷基-喹唑啉-2-胺衍生物的合成与生物活性评价

以异长叶烷酮为原料,经缩合、环化等反应合成出6个喹唑啉-2-胺衍生物(Ⅲa～Ⅲf)。采用1HNMR、13CNMR、IR、HRMS等对其结构进行了表征。采用MTT法考察了化合物Ⅲa～Ⅲf对人肝癌细胞(Hep G2)和人脐静脉内皮细胞(HUVECs)的抑制活性。结果表明:化合物Ⅲa、Ⅲb、Ⅲd、Ⅲf[半抑制浓度(IC50)分别为8.58±0.5、44.52±0.9、57.18±0.8、32.83±0.6μmol/L]对Hep G2有一定的抗肿瘤活性。其中,4-(4?-氯苯基)-6,6,10,10-四甲基-5,6,6a,7,8,9,10,10a-八氢-6a,9-甲基苯并[h]喹唑啉-2-胺(Ⅲa)抗HepG2的活性最强;只有4-[4?–(二甲基氨基)苯基]-6,6,10,10-四甲基-5,6,6a,7,8,9,10,10a-八氢-6a,9-甲基苯并[h]喹唑啉-2-胺(Ⅲf)对HUVECs有抑制活性。同时,采用叶浸渍法考察了化合物Ⅲa～Ⅲf对桃蚜的杀虫活性,结果表明,化合物Ⅲa、Ⅲd[致死中浓度(LC50)=41.0073,37.4589 mg/L]对桃蚜具有较好的杀虫活性。     

Synthesis and Biological Evaluation of Novel Dehydroabietic Acid Derivatives Conjugated with Acyl-Thiourea Peptide Moiety as Antitumor Agents

A series of dehydroabietic acid (DHAA) acyl-thiourea derivatives were designed and synthesized as potent antitumor agents. The in vitro pharmacological screening results revealed that the target compounds exhibited potent cytotoxicity against HeLa, SK-OV-3 and MGC-803 tumor cell lines, while they showed lower cytotoxicity against HL-7702 normal human river cells. Compound 9n (IC₅₀ = 6.58 ± 1.11 μM) exhibited the best antitumor activity against the HeLa cell line and even displayed more potent inhibitory activity than commercial antitumor drug 5-FU (IC₅₀ = 36.58 ± 1.55 μM). The mechanism of representative compound 9n was then studied by acridine orange/ethidium bromide staining, Hoechst 33,258 staining, JC-1 mitochondrial membrane potential staining, TUNEL assay and flow cytometry, which illustrated that this compound could induce apoptosis in HeLa cells. Cell cycle analysis indicated that compound 9n mainly arrested HeLa cells in the S phase stage. Further investigation demonstrated that compound 9n induced apoptosis of HeLa cells through a mitochondrial pathway.     

Encapsulation of the Dinuclear Trithiolato-Bridged Arene Ruthenium Complex Diruthenium-1 in an Apoferritin Nanocage: Structure and Cytotoxicity

The effects of encapsulating the cytotoxic dinuclear trithiolato-bridged arene ruthenium complex [(η⁶-p-MeC₆H₄iPr)₂Ru₂(μ₂-S-p-C₆H₄tBu)₃]Cl (DiRu-1) within the apoferritin (AFt) nanocage were investigated. The DiRu-1-AFt nanocarrier was characterized by UV/Vis spectroscopy, ICP-MS, CD and X-ray crystallography. In contrast to previously reported Au- and Pt-based drug-loaded AFt carriers, we found no evidence of direct interactions between DiRu-1 and AFt. DiRu-1-AFt is cytotoxic toward immortalized murine BALB/c-3T3 fibroblasts transformed with SV40 virus (SVT2) and human epidermoid carcinoma A431 malignant cells, and exhibits moderate selectivity for these cancer cells over normal BALB/c-3T3 cells. DiRu-1-AFt triggers the production of reactive oxygen species, depolarization of mitochondrial membrane potential, and induces cell death via p53-mediated apoptosis. Comparison between our data and previous results suggests that the presence of specific interactions between a metal-based drug and AFt within the protein cage is not essential for drug encapsulation.     

Aging process of polyamidoamine dendrimers: Effect of pH and shaking in the fluorescence emission and aggregation-state

In the last years, it has been discovered and intensely studied the non-traditional intrinsic luminescence of polyamidoamine (PAMAM) dendrimers. Nevertheless, their aging process in aqueous suspension is scarcely studied, being unknown the causes of the changes in their luminescence properties. Hence, this work aims to characterize the amine-terminated (DG4.0) and carboxylic acid-terminated (DG4.5) PAMAM dendrimers of generations 4.0 and 4.5, respectively, through the aging process at three different pH conditions, stored with or without shaking. The UV–Vis absorption, the fluorescence emission, and the dendrimer-size distribution are studied for up to 16 days. In a different way than the already published works, this work demonstrates that there is no chemical change in dendrimers through the aging process, even though changes in fluorescence emission are observed. Besides, the changes in the agglomeration patterns of dendrimers are not related to the change in the fluorescence emission through aging. Moreover, large aggregates of DG4.5 are present in water and need to be disrupted by shaking before an in vivo administration.     

Strategies to Reduce the On-Target Platelet Toxicity of Bcl-xL Inhibitors: PROTACs,SNIPERs and Prodrug-Based Approaches

Apoptosis is a highly regulated cellular process. Aberration in apoptosis is a common characteristic of various disorders. Therefore, proteins involved in apoptosis are prime targets in multiple therapies. Bcl-x_L is an antiapoptotic protein. Compared to other antiapoptotic proteins, the expression of Bcl-x_L is common in solid tumors and, to an extent, in some leukemias and lymphomas. The overexpression of Bcl-x_L is also linked to survival and chemoresistance in cancer and senescent cells. Therefore, Bcl-x_L is a promising anticancer and senolytic target. Various nanomolar range Bcl-x_L inhibitors have been developed. ABT-263 was successfully identified as a Bcl-x_L/Bcl-2 dual inhibitor. But it failed in the clinical trial (phase-II) because of its on-target platelet toxicity, which also implies an essential role of Bcl-x_L protein in the survival of human platelets. Classical Bcl-x_L inhibitor designs utilize occupancy-driven pharmacology with typical shortcomings (such as dose-dependent off-target and on-target platelet toxicities). Hence, event-driven pharmacology-based approaches, such as proteolysis targeting chimeras (PROTACs) and SNIPERs (specific non-genetic IAP-based protein erasers) have been developed. The development of Bcl-x_L based PROTACs was expected, as 600 E3-ligases are available in humans, while some (such as cereblon (CRBN), von Hippel-Lindau (VHL)) are relatively less expressed in platelets. Therefore, E3 ligase ligand-based Bcl-x_L PROTACs (CRBN: XZ424, XZ739; VHL: DT2216, PZ703b, 753b) showed a significant improvement in platelet therapeutic index than their parent molecules ( ABT-263 : DT2216, PZ703b, 753b, XZ739, PZ15227; A1155463 : XZ424). Other than their distinctive pharmacology, PROTACs are molecularly large, which limits their cell permeability and plays a role in improving their cell selectivity. We also discuss prodrug-based approaches, such as antibody-drug conjugates ( ABBV-155 ), phosphate prodrugs ( APG-1252 ), dendrimer conjugate ( AZD0466 ), and glycosylated conjugates ( Nav-Gal ). Studies of in-vitro, in-vivo, structure-activity relationships, biophysical characterization, and status of preclinical/clinical inhibitors derived from these strategies are also discussed in the review.     

Polyethylene glycol‐polyethylenimine‐tetrachloroplatinum (IV): A novel conjugate with good abilities of antitumor and gene delivery

It is much importance to develop novel multifunctional delivery systems for the combination therapy of drug and gene. In this work, a novel conjugate, polyethylene glycol‐polyethylenimine‐tetrachloroplatinum (IV) (PEG‐PEI‐Pt), with good abilities of antitumor and gene delivery was proposed by combining PEG (Mw 3400 Da), low molecular weight PEI (Mw 800 Da), and tetrachloroplatinum (IV). The antitumoral and gene transfection activities of PEG‐PEI‐Pt were analyzed in many tumor (A549, A375, HepG‐2, HuH‐7, and B16 cells) and normal (COS‐7 cells) cell lines. Similar to cisplatin (one platinum anticancer drug), PEG‐PEI‐Pt showed much higher sensitivity in tumor cells than in normal cells. More importantly, PEG‐PEI‐Pt had a potential to treat drug‐resistant tumors. Almost no transfection efficiency was observed for PEI (Mw 800 Da) and PEG‐PEI. Very interestingly, PEG‐PEI‐Pt could condense plasmid DNA efficiently, and exhibited good transfection efficiency in B16, HepG‐2, A375 and COS‐7 cells, comparable to even higher than PEI 25 kDa. In addition, PEG‐PEI‐Pt could also effectively deliver siRNA into the cytoplasm of tumor cells. With the good antitumoral and gene delivery abilities, PEG‐PEI‐Pt may have a great potential for combination therapy of drug and gene. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2012     

Polyaminoacid Based Core@shell Nanocarriers of 5-Fluorouracil: Synthesis,Properties and Theranostics Application

Cancer is one of the most important health problems of our population, and one of the common anticancer treatments is chemotherapy. The disadvantages of chemotherapy are related to the drug’s toxic effects, which act on cancer cells and the healthy part of the body. The solution of the problem is drug encapsulation and drug targeting. The present study aimed to develop a novel method of preparing multifunctional 5-Fluorouracil (5-FU) nanocarriers and their in vitro characterization. 5-FU polyaminoacid-based core@shell nanocarriers were formed by encapsulation drug-loaded nanocores with polyaminoacids multilayer shell via layer-by-layer method. The size of prepared nanocarriers ranged between 80–200 nm. Biocompatibility of our nanocarriers as well as activity of the encapsulated drug were confirmed by MTT tests. Moreover, the ability to the real-time observation of developed nanocarriers and drug accumulation inside the target was confirmed by fluorine magnetic resonance imaging (¹⁹F-MRI).     

Novel Hybrid Compounds Containing Benzofuroxan and Aminothiazole Scaffolds: Synthesis and Evaluation of Their Anticancer Activity

A series of novel hybrid compounds containing benzofuroxan and 2-aminothiazole moieties are synthesized via aromatic nucleophilic substitution reaction. Possible reaction pathways have been considered quantum-chemically, which allowed us to suggest the most probable products. The quantum chemical results have been proved by X-ray data on one compound belonging to the synthesized series. It was shown that the introduction of substituents to both the thiazole and amine moieties of the compounds under study strongly influences their UV/Vis spectra. Initial substances and obtained hybrid compounds have been tested in vitro as anticancer agents. Target compounds showed selectivity towards M-HeLa tumor cell lines and were found to be more active than starting benzofuroxan and aminothiazoles. Furthermore, they are considerably less toxic to normal liver cells compared to Tamoxifen. The mechanism of action of the studied compounds can be associated with the induction of apoptosis, which proceeds along the mitochondrial pathway. Thus, new hybrids of benzofuroxan are promising candidates for further development as anticancer agents.     

Incorporation of theophylline in a chitosan/chondroitin sulfate hydrogel matrix: In vitro release studies and mechanical properties according to pH changes

Hydrogels based on natural polymers have been widely applied as vehicles for controlled drug release because of their advantages and interesting properties. In this study, a physical hydrogel based on chitosan and chondroitin sulfate (CS) was formed under mild conditions to act as a potential device for the controlled release of theophylline (TH). In vitro CS and TH release studies at pH 2 and 8 were performed. Under acid conditions (pH 2), the fraction of TH released (ca. 0.87) was higher than that of CS (ca. 0.13). On the other hand, under basic conditions, the fractions released of both substances were similar (ca. 0.57). In addition, the system presented in this work was able to sustain the TH release in a controlled way for 30 h. The variation of the pH affected the mechanical properties and contributed to form ordered regions within the hydrogel network, as observed through compression tests and wide‐angle X‐ray scattering analysis. The experimental data and discussion presented in this article will contribute to the development of a new vehicle for controlled TH release; this ensures the efficacy of the drug and reduced the number of daily doses administered. © 2012 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013     

Design and characterization of pH stimuli-responsive nanofiber drug delivery system: The promising targeted carriers for tumor therapy

New carrier platforms have been designed for an electrospun pyridinium calixarene nanofiber for controlled drug delivery. First, 5,11,17,23-tetra-tert-butyl-25,27-bis(3-aminomethyl-pyridineamido)-26,28-dihydroxycalix[4]arene (3-AMP) scaffold was produced by electrospinning. AMP scaffold was modified by human serum albumin (HSA), folic acid (FA), and glutathione (GSH). Doxorubicin (DOX) was loaded to surfaces of the AMP, AMP-HSA, AMP-HSA-FA, and AMP-HSA-GSH nanofibers by using DOX solution in different buffers with, 2.2, 4.0, 6.0, and 7.4 pH. The release studies DOX from four different nanofibers was also done in a various amount microenviroments by changing pH values. The loading and release amount of DOX was estimated from the calibration curve drawn at 480 and 560 nm of excitation and emission wavelengths by using a fluorescence spectrophotometer. The loading studies were confirmed by Fourier transforms infrared, atomic force microscopy, transmission electron microscopy, scanning electron microscope, and energy-dispersive X-ray (EDX) analysis.     

1,2,4-Oxadiazole-Based Bio-Isosteres of Benzamides: Synthesis,Biological Activity and Toxicity to Zebrafish Embryo

To discover new compounds with broad spectrum and high activity, we designed a series of novel benzamides containing 1,2,4-oxadiazole moiety by bioisosterism, and 28 benzamides derivatives with antifungal activity were synthesized. These compounds were evaluated against four fungi: Botrytis cinereal, FusaHum graminearum, Marssonina mali, and Thanatephorus cucumeris. The results indicated that most of the compounds displayed good fungicidal activities, especially against Botrytis cinereal. For example, 10a (84.4%), 10d (83.6%), 10e (83.3%), 10f (83.1%), 10i (83.3%), and 10l (83.6%) were better than pyraclostrobin (81.4%) at 100 mg/L. In addition, the acute toxicity of 10f to zebrafish embryo was 20.58 mg/L, which was classified as a low-toxicity compound.     

In Vivo Anti-Tumor Activity and Toxicological Evaluations of Perillaldehyde 8,9-Epoxide,a Derivative of Perillyl Alcohol

Recent studies have revealed the high cytotoxicity of p-menthane derivatives against human tumor cells. In this study, the substance perillaldehyde 8,9-epoxide, a p-menthane class derivative obtained from (S)-(−)-perillyl alcohol, was selected in order to assess antitumor activity against experimental sarcoma 180 tumors. Toxicological effects related to the liver, spleen, kidneys and hematology were evaluated in mice submitted to treatment. The tumor growth inhibition rate was 38.4%, 58.7%, 35.3%, 45.4% and 68.1% at doses of 100 and 200 mg/kg/day for perillaldehyde 8,9-epoxide, perillyl alcohol and 25 mg/kg/day for 5-FU intraperitoneal treatments, respectively. No toxicologically significant effect was found in liver and kidney parameters analyzed in Sarcoma 180-inoculated mice treated with perillaldehyde 8,9-epoxide. Histopathological analyses of the liver, spleen, and kidneys were free from any morphological changes in the organs of the animals treated with perillaldehyde 8,9-epoxide. In conclusion, the data suggest that perillaldehyde 8,9-epoxide possesses significant antitumor activity without systemic toxicity for the tested parameters. By comparison, there was no statistical difference for the antitumor activity between perillaldehyde 8,9-epoxide and perillyl alcohol.     

Aromatic Core Extension in the Series of N‐Cyclic Bay‐Substituted Perylene G‐Quadruplex Ligands: Increased Telomere Damage,Antitumor Activity,and Strong Selectivity for Neoplastic over Healthy Cells

Based on previous work on both perylene and coronene derivatives as G‐quadruplex binders, a novel chimeric compound was designed: N,N′‐bis[2‐(1‐piperidino)‐ethyl]‐1‐(1‐piperidinyl)‐6‐[2‐(1‐piperidino)‐ethyl]‐benzo[ghi]perylene‐3,4:9,10‐tetracarboxylic diimide (EMICORON), having one piperidinyl group bound to the perylene bay area (positions 1, 12 and 6, 7 of the aromatic core), sufficient to guarantee good selectivity, and an extended aromatic core able to increase the stacking interactions with the terminal tetrad of the G‐quadruplex. The obtained “chimera” molecule, EMICORON, rapidly triggers extensive DNA damage of telomeres, associated with the delocalization of telomeric protein protection of telomeres 1 (POT1), and efficiently limits the growth of both telomerase‐positive and ‐negative tumor cells. Notably, the biological effects of EMICORON are more potent than those of the previously described perylene derivative (PPL3C), and more interestingly, EMICORON appears to be detrimental to transformed and tumor cells, while normal fibroblasts expressing telomerase remain unaffected. These results identify a new promising G‐quadruplex ligand, structurally and biologically similar on one side to coronene and on the other side to a bay‐monosubstituted perylene, that warrants further studies.     

Development and Biochemical Characterization of Self-Immolative Linker Containing GnRH-III-Drug Conjugates

The human gonadotropin releasing hormone (GnRH-I) and its sea lamprey analogue GnRH-III specifically bind to GnRH receptors on cancer cells and can be used as targeting moieties for targeted tumor therapy. Considering that the selective release of drugs in cancer cells is of high relevance, we were encouraged to develop cleavable, self-immolative GnRH-III-drug conjugates which consist of a p-aminobenzyloxycarbonlyl (PABC) spacer between a cathepsin B-cleavable dipeptide (Val-Ala, Val-Cit) and the classical anticancer drugs daunorubicin (Dau) and paclitaxel (PTX). Alongside these compounds, non-cleavable GnRH-III-drug conjugates were also synthesized, and all compounds were analyzed for their antiproliferative activity. The cleavable GnRH-III bioconjugates revealed a growth inhibitory effect on GnRH receptor-expressing A2780 ovarian cancer cells, while their activity was reduced on Panc-1 pancreatic cancer cells exhibiting a lower GnRH receptor level. Moreover, the antiproliferative activity of the non-cleavable counterparts was strongly reduced. Additionally, the efficient cleavage of the Val-Ala linker and the subsequent release of the drugs could be verified by lysosomal degradation studies, while radioligand binding studies ensured that the GnRH-III-drug conjugates bound to the GnRH receptor with high affinity. Our results underline the high value of GnRH-III-based homing devices and the application of cathepsin B-cleavable linker systems for the development of small molecule drug conjugates (SMDCs).     

Polymer micellar aggregates of novel amphiphilic biodegradable graft copolymer composed of poly(aspartic acid) derivatives: Preparation,characterization, and effect of pH on aggregation

Novel amphiphilic biodegradable graft copolymer based on poly(aspartic acid) was prepared by attaching monomethoxy polyethylene glycol (mPEG) as hydrophiphic segment to poly(aspartic acid‐g‐octadecylamine) (PASP‐g‐ODA) as hydrophobic backbone. The chemical structures of amphiphilic copolymers were confirmed by FTIR and ¹H NMR spectroscopy. The polymeric micelles were prepared with solvent evaporation and their physicochemical properties in aqueous media were characterized by dynamic light scattering (DLS) and fluorescence spectroscopy. These micelles were confirmed to be pH‐sensitive by measuring optical transmittance of micelle solution and the size of micellar aggregates. The number average diameter of polymeric micelles prepared in medium at pH 2.5 was larger than that in neutral and basic medium and showed a bimodal size distribution because of the protonation of carboxyl groups in backbone. Furthermore, the polymeric micelle can load water‐insoluble drug (podophyllotoxin), and the drug release from micelles showed a pH‐dependency. © 2005 Wiley Periodicals, Inc. J Appl Polym Sci, 2006     

Self-Assembly of pH-Labile Polymer Nanoparticles for Paclitaxel Prodrug Delivery: Formulation,Characterization, and Evaluation

The efficacy of paclitaxel (PTX) is limited due to its poor solubility, poor bioavailability, and acquired drug resistance mechanisms. Designing paclitaxel prodrugs can improve its anticancer activity and enable formulation of nanoparticles. Overall, the aim of this work is to improve the potency of paclitaxel with prodrug synthesis, nanoparticle formation, and synergistic formulation with lapatinib. Specifically, we improve potency of paclitaxel by conjugating it to α-tocopherol (vitamin E) to produce a hydrophobic prodrug (Pro); this increase in potency is indicated by the 8-fold decrease in half maximal inhibitory concentration (IC₅₀) concentration in ovarian cancer cell line, OVCA-432, used as a model system. The efficacy of the paclitaxel prodrug was further enhanced by encapsulation into pH-labile nanoparticles using Flash NanoPrecipitation (FNP), a rapid, polymer directed self-assembly method. There was an 1100-fold decrease in IC₅₀ concentration upon formulating the prodrug into nanoparticles. Notably, the prodrug formulations were 5-fold more potent than paclitaxel nanoparticles. Finally, the cytotoxic effects were further enhanced by co-encapsulating the prodrug with lapatinib (LAP). Formulating the drug combination resulted in synergistic interactions as indicated by the combination index (CI) of 0.51. Overall, these results demonstrate this prodrug combined with nanoparticle formulation and combination therapy is a promising approach for enhancing paclitaxel potency.