Synthesis and discovery of Baylis-Hillman adducts as potent and selective thioredoxin reductase inhibitors for cancer treatment.

The selenoprotein thioredoxin reductase (TrxR) is of paramount importance in maintaining cellular redox homeostasis, and aberrant upregulation of TrxR is frequently observed in various cancers due to their elevated oxidative stress in cells. Thus, it seems promising and feasible to target the ablation of intracellular TrxR for the treatment of cancers. We report herein the design and synthesis of a series of Baylis-Hillman adducts, and identified a typical adduct that possesses the superior cytotoxicity against HepG2 cells over other types of cancer cells. The biological investigation shows the selected typical adduct selectively targets TrxR in HepG2 cells, which thereafter results in the collapse of intracellular redox homeostasis. Further mechanistic studies reveal that the selected typical adduct arrests the cell cycle in G1/G0 phase. Importantly, the malignant metastasis of HepG2 cells is significantly restrained by the selected typical adduct. With well-defined molecular target and mechanism of action, the selected typical adduct, even other Baylis-Hillman skeleton-bearing compounds, merits further development as candidate or ancillary agent for the treatment of various cancers.

Progress and perspective on hydrogen sulfide donors and their biomedical applications.

Following the discovery of nitric oxide (NO) and carbon monoxide (CO), hydrogen sulfide (H2 S) has been identified as the third gasotransmitter in humans. Increasing evidence have shown that H2 S is of preventive or therapeutic effects on diverse pathological complications. As a consequence, it is of great significance to develop suitable approaches of H2 S-based therapeutics for biomedical applications. H2 S-releasing agents (H2 S donors) play important roles in exploring and understanding the physiological functions of H2 S. More importantly, accumulating studies have validated the theranostic potential of H2 S donors in extensive repertoires of in vitro and in vivo disease models. Thus, it is imperative to summarize and update the literatures in this field. In this review, first, the background of H2 S on its chemical and biological aspects is concisely introduced. Second, the studies regarding the H2 S-releasing compounds are categorized and described, and accordingly, their H2 S-donating mechanisms, biological applications, and therapeutic values are also comprehensively delineated and discussed. Necessary comparisons between related H2 S donors are presented, and the drawbacks of many typical H2 S donors are analyzed and revealed. Finally, several critical challenges encountered in the development of multifunctional H2 S donors are discussed, and the direction of their future development as well as their biomedical applications is proposed. We expect that this review will reach extensive audiences across multiple disciplines and promote the innovation of H2 S biomedicine.

Structural Modification of Aminophenylarsenoxides Generates Candidates for Leukemia Treatment via Thioredoxin Reductase Inhibition.

Upregulation of the selenoprotein thioredoxin reductase (TrxR) is of pathological significance in maintaining tumor phenotypes. Thus, TrxR inhibitors are promising cancer therapeutic agents. We prepared different amino-substituted phenylarsine oxides and evaluated their cytotoxicity and inhibition of TrxR. Compared with our reported p-substituted molecule (8), the o-substituted molecule (10) shows improved efficacy (nearly a fourfold increase) to kill leukemia HL-60 cells. Although the compounds 8 and 10 display similar potency to inhibit the purified TrxR, the o-substitution 10 exhibits higher potency than the p-substitution 8 to inhibit the cellular TrxR activity. Molecular docking results demonstrate the favorable weak interactions of the o-amino group with the TrxR C-terminal active site. Efficient inhibition of TrxR consequently induces the oxidative stress-mediated apoptosis of cancer cells. Silence of the TrxR expression sensitizes the cells to the arsenic compound treatment, further supporting the critical involvement of TrxR in the cellular actions of compound 10.

Synthesis and biological evaluation of disulfides as anticancer agents with thioredoxin inhibition.

Altered redox homeostasis as a hallmark of cancer cells is exploited by cancer cells for growth and survival. The thioredoxin (Trx), an important regulator in maintaining the intracellular redox homeostasis, is cumulatively recognized as a promising target for the development of anticancer drugs. Herein, we synthesized 72 disulfides and evaluated theirinhibition for Trx and antitumor activity. First, we established an efficient and fast method to screen Trx inhibitors by using the probe NBL-SS that was developed by our group to detect Trx function in living cells. After an initial screening of the Trx inhibitory activity of these compounds, 8 compounds showed significant inhibition activity against Trx. We then evaluated the cytotoxicity of these 8 disulfides, compounds 68 and 69 displayed high cytotoxicity to HeLa cells, but less sensitive to normal cell lines. Next, we performed kinetic studies of both two disulfides, 68 had faster inhibition of Trx than 69. Further studies revealed that 68 led to the accumulation of reactive oxygen species and eventually induced apoptosis of Hela cells via inhibiting Trx. The establishment of a method for screening Trx inhibitors and the discovery of 68 with remarkable Trx inhibition provide support for the development of anticancer candidates with Trx inhibition.

Streptochlorin analogues as potential antifungal agents: Design, synthesis, antifungal activity and molecular docking study.

Streptochlorin is a small molecule of indole alkaloid isolated from marine Streptomyces sp., it is a promising lead compound due to its potent bioactivity in preventing many phytopathogens in our previous study, but further structural modifications are required to improve its antifungal activity. Our work in this paper focused on the replacement of oxazole ring in streptochlorin with the imidazole ring, to discover novel analogues. Based on this design strategy, three series of streptochlorin analogues were efficiently synthesized through sequential Vilsmeier-Haack reaction, Van Leusen imidazole synthesis and halogenation reaction. Some of the analogues displayed excellent activity in the primary assays, and this is highlighted by compounds 4g and 4i, the growth inhibition against Alternaria Leaf Spot and Rhizoctorzia solani under 50 µg/mL are 97.5% and 90.3%, respectively, even more active than those of streptochlorin, pimprinine and Osthole. Molecular docking models indicated that streptochlorin binds with Thermus thermophiles Leucyl-tRNA Synthetase in a similar mode to AN2690, offering a perspective on the mode of action study for antifungal activities of streptochlorin derivatives. Further study is still ongoing with the aim of discovering synthetic analogues, with improved antifungal activity and clear mode of action.

Generation of potent Nrf2 activators via tuning the electrophilicity and steric hindrance of vinyl sulfones for neuroprotection.

Oxidative stress is constantly involved in the etiopathogenesis of an ever-widening range of neurodegenerative diseases. As a consequence, effective repression of cellular oxidative stress to a redox homeostatic condition is a promising and feasible strategy to treat, or at least retard the progression of, such disorders. Nrf2, a primary orchestrator of cellular antioxidant response machine, is responsible for detoxifying and compensating for deleterious oxidative stress via transcriptional activation of a diverse array of antioxidant biomolecules. In the framework of our persistent interest in disclosing small molecules that interfere with cellular redox-regulating machinery, we report herein the synthesis, optimization, and biological assessment of 47 vinyl sulfone scaffold-bearing small molecules, most of which exhibit robust neuroprotective effect against H2O2-mediated lesions to PC12 cells. After initial screening, the most potent neuroprotective compounds 9b and 9c with marginal cytotoxicity were selected for the follow-up studies. Our results demonstrate that their neuroprotective effects are attributed to the up-regulation of a panel of antioxidant genes and corresponding gene products. Further mechanistic studies indicate that Nrf2 is indispensable for the cellular performances of 9b and 9c, arising from the fact that silence of Nrf2 gene drastically nullifies their protective action. Taken together, 9b and 9c discovered in this work merit further development as neuroprotective candidates for the treatment of oxidative stress-mediated pathological conditions.

Diversity-oriented synthesis and antifungal activities of novel pimprinine derivative bearing a 1,3,4-oxadiazole-5-thioether moiety.

Based on the strategy of diversity-oriented synthesis and the structures of natural product pimprinine and streptochlorin, two series of novel pimprinine derivatives containing 1,3,4-oxadiazole-5-thioether moieties were efficiently synthesized under the optimized reaction conditions. Biological assays conducted at Syngenta showed the designed derivatives displayed an altered pattern of biological activity, of which 5h was identified as the most promising compound with strong activity against Pythium dissimile and also a broad antifungal spectrum in primary screening. Further structural optimization of pimprinine and streptochlorin derivatives is well under way, aiming to discover synthetic analogues with improved antifungal activity. Two series of novel pimprinine derivatives containing 1,3,4-oxadiazole-5-thioether moieties were efficiently synthesized through diversity-oriented synthesis strategy under the optimized conditions. Biological assays showed the designed derivatives exhibited potential activity.

Small molecules regulating reactive oxygen species homeostasis for cancer therapy.

Elevated intracellular reactive oxygen species (ROS) and antioxidant defense systems have been recognized as one of the hallmarks of cancer cells. Compared with normal cells, cancer cells exhibit increased ROS to maintain their malignant phenotypes and are more dependent on the "redox adaptation" mechanism. Thus, there are two apparently contradictory but virtually complementary therapeutic strategies for the regulation of ROS to prevent or treat cancer. The first strategy, that is, chemoprevention, is to prevent or reduce intracellular ROS either by suppressing ROS production pathways or by employing antioxidants to enhance ROS clearance, which protects normal cells from malignant transformation and inhibits the early stage of tumorigenesis. The second strategy is the ROS-mediated anticancer therapy, which stimulates intracellular ROS to a toxicity threshold to activate ROS-induced cell death pathways. Therefore, targeting the regulation of intracellular ROS-related pathways by small-molecule candidates is considered to be a promising treatment for tumors. We herein first briefly introduce the source and regulation of ROS, and then focus on small molecules that regulate ROS-related pathways and show efficacy in cancer therapy from the perspective of pharmacophores. Finally, we discuss several challenges in developing cancer therapeutic agents based on ROS regulation and propose the direction of future development.

Sanguinarine as a new chemical entity of thioredoxin reductase inhibitor to elicit oxidative stress and promote tumor cell apoptosis.

The alteration of redox homeostasis is a hallmark of cancer cells. As a critical player in regulating cellular redox signaling, thioredoxin reductase (TrxR) enzymes are increasingly recognized as attractive targets for anticancer drug development. We reported herein the natural product sanguinarine (SAN) as a potent inhibitor of TrxR with a new chemical scaffold. Inhibition of TrxR leads to accumulation of the oxidized thioredoxin, elicits oxidative stress, and finally promotes apoptosis of cancer cells. Further synthesis of different model compounds of SAN demonstrated that the phenanthridinium unit is responsible for the TrxR inhibition. The core structure of SAN, e.g., the phenanthridinium moiety, is different from those of known TrxR inhibitors, and thus SAN is a new chemical entity of TrxR inhibitors and may serve a lead for further development. In addition, as the phenanthridinium scaffold is widely present in natural products, the disclosure of TrxR inhibition by such unit sheds light in understanding the pharmacological actions of these molecules.

Synthesis of Dithiolethiones and Identification of Potential Neuroprotective Agents via Activation of Nrf2-Driven Antioxidant Enzymes.

Oxidative stress is implicated in the pathogenesis of a wide variety of neurodegenerative disorders, and accordingly, dietary supplement of exogenous antioxidants or/and upregulation of the endogenous antioxidant defense system are promising for therapeutic intervention or chemoprevention of neurodegenerative diseases. Nrf2, a master regulator of the cellular antioxidant machinery, cardinally participates in the transcription of cytoprotective genes against oxidative/electrophilic stresses. Herein, we report the synthesis of 59 structurally diverse dithiolethiones and evaluation of their neuroprotection against 6-hydroxydopamine- or H2O2-induced oxidative damages in PC12 cells, a neuron-like rat pheochromocytoma cell line. Initial screening identified compounds 10 and 11 having low cytotoxicity but conferring remarkable protection on PC12 cells from oxidative-mediated damages. Further studies demonstrated that both compounds upregulated a battery of antioxidant genes as well as corresponding genes' products. Significantly, silence of Nrf2 expression abolishes cytoprotection of 10 and 11, indicating targeting Nrf2 activation is pivotal for their cellular functions. Taken together, the two lead compounds discovered here with potent neuroprotective functions against oxidative stress via Nrf2 activation merit further development as therapeutic or chemopreventive candidates for neurodegenerative disorders.

Design and synthesis of novel 7-[(N-substituted-thioureidopiperazinyl)-methyl]-camptothecin derivatives as potential cytotoxic agents.

As part of continuing our research on diverse C-7 derivatives of camptothecin (CPT), 16 CPT derivatives bearing piperazinyl-thiourea chemical scaffold and different substituent groups have been designed, synthesized and evaluated in vitro for cytotoxicity against five tumor cell lines (A-549, MDA-MB-231, MCF-7, KB and KBvin). As a result, all the synthesized compounds showed promising in vitro cytotoxic activity against the five tumor cell lines tested, and were more potent than irinotecan. Importantly, compounds 13 g (IC50 = 0.514 µM) and 13o (IC50 = 0.275 µM) possessed similar or better antiproliferative activity against the multidrug-resistant (MDR) KBvin subline than that of topotecan (IC50 = 0.511 µM) and merit further development as anticancer candidates for clinical trail. With these results in hand, we have a reason to conclude that incorporating piperazinyl-thiourea motifs into position-7 of camptothecin confers well cytotoxic activity against cancer cell lines, probably resulting in new anticancer drugs.

Design, synthesis and antineoplastic activity of novel 20(S)-acylthiourea derivatives of camptothecin.

For the purpose of advancing our research on diverse C-20 decorated derivatives of camptothecin (CPT), 46 new CPT acylthiourea derivatives were synthesized and evaluated in vitro for their cytotoxicity. All the compounds showed promising in vitro cytotoxicity against six tumor cell lines (Hep3B, MCF7, A549, MDA-MB-231, KB and KB-vin). Out of them, compound c20 possesses remarkable in vitro cytotoxic activity and is more potent than topotecan. Mechanistically, c20 not only induces cell cycle arrest and cell apoptosis in A549 cells, but also inhibits Topo I activity in the cell and cell-free system in a manner similar to that of topotecan. In both xenograft and primary HCC mouse models, c20 displays significant in vivo anti-cancer activity and is more potent than topotecan. In addition, the acute toxicity assay showed that c20 has no apparent toxicity to mouse liver, kidney and hemopoietic system of the FVB/N mice. Take together, these results indicated that compound c20 could be a potential anti-cancer candidate for further clinical trial.

Design, semisynthesis and potent cytotoxic activity of novel 10-fluorocamptothecin derivatives.

Fluorination is a well-known strategy for improving the bioavailability of bioactive molecules in the lead optimization phase of drug discovery projects. In an attempt to improve the antitumor activity of camptothecins (CPTs), novel 10-fluoro-CPT derivatives were designed, synthesized and evaluated for cytotoxicity against five human cancer cell lines (A-549, MDA-MB-231, KB, KB-VIN and MCF-7). All of the derivatives showed more potent in vitro cytotoxic activity than the clinical CPT-derived drug irinotecan against the tumor cell lines tested, and most of them showed comparable or superior potency to topotecan. Remarkably, compounds 16b (IC50, 67.0nM) and 19b (IC50, 99.2nM) displayed the highest cytotoxicity against the multidrug-resistant (MDR) KB-VIN cell line and merit further development as preclinical drug candidates for treating cancer, including MDR phenotype. Our study suggested that incorporation of a fluorine atom into position 10 of CPT is an effective method for discovering new potent CPT derivatives.

Design, synthesis, and cytotoxic activity of novel 7-substituted camptothecin derivatives incorporating piperazinyl-sulfonylamidine moieties.

In our continuing search for camptothecin (CPT)-derived antitumor drugs, novel 7-substituted CPT derivatives incorporating piperazinyl-sulfonylamidine moieties were designed, synthesized and evaluated for cytotoxicity against five tumor cell lines (A-549, MDA-MB-231, MCF-7, KB, and KB-VIN). All of the derivatives showed promising in vitro cytotoxic activity against the tested tumor cell lines, and were more potent than irinotecan. Remarkably, most of the compounds exhibited comparable cytotoxicity against the multidrug-resistant (MDR) KB-VIN and parental KB tumor cell lines, while irinotecan lost activity completely against KB-VIN. Especially, compounds 13r and 13p (IC50 0.38 and 0.85µM, respectively) displayed the greatest cytotoxicity against the MDR KB-VIN cell line and merit further development into preclinical and clinical drug candidates for treating cancer, including MDR phenotype.

Design, synthesis, cytotoxic activity and molecular docking studies of new 20(S)-sulfonylamidine camptothecin derivatives.

In an ongoing investigation of 20-sulfonylamidine derivatives (9, YQL-9a) of camptothecin (1) as potential anticancer agents directly and selectively inhibiting topoisomerase (Topo) I, the sulfonylamidine pharmacophore was held constant, and a camptothecin derivatives with various substitution patterns were synthesized. The new compounds were evaluated for antiproliferative activity against three human tumor cell lines, A-549, KB, and multidrug resistant (MDR) KB subline (KBvin). Several analogs showed comparable or superior antiproliferative activity compared to the clinically prescribed 1 and irinotecan (3). Significantly, the 20-sulfonylamidine derivatives exhibited comparable cytotoxicity against KBvin, while 1 and 3 were less active against this cell line. Among them, compound 15c displayed much better cytotoxic activity than the controls 1, 3, and 9. Novel key structural features related to the antiproliferative activities were identified by structure-activity relationship (SAR) analysis. In a molecular docking model, compounds 9 and 15c interacted with Topo I-DNA through a different binding mode from 1 and 3. The sulfonylamidine side chains of 9 and 15c could likely form direct hydrogen bonds with Topo I, while hydrophobic interaction with Topo I and π-π stacking with double strand DNA were also confirmed as binding driving forces. The results from docking models were consistent with the SAR conclusions. The introduction of bulky substituents at the 20-position contributed to the altered binding mode of the compound by allowing them to form new interactions with Topo I residues. The information obtained in this study will be helpful for the design of new derivatives of 1 with most promising anticancer activity.

Design and synthesis of novel PEG-conjugated 20(S)-camptothecin sulfonylamidine derivatives with potent in vitro antitumor activity via Cu-catalyzed three-component reaction.

In our continuing search for camptothecin (CPT)-derived antitumor drugs, novel structurally diverse PEG-based 20(S)-CPT sulfonylamidine derivatives were designed, synthesized via a Cu-multicomponent reaction (MCR), and evaluated for cytotoxicity against four human tumor cell lines (A-549, MDA-MB-231, KB, and KBvin). All of the derivatives showed promising in vitro cytotoxic activity against the tested tumor cell lines, and were more potent than irinotecan. Significantly, these derivatives exhibited comparable cytotoxicity against KBvin, while irinotecan was less active against this cell line. With a concise efficient synthesis and potent cytotoxic profiles, especially significant activity towards KBvin, these compounds merit further development as a new generation of CPT-derived PEG-conjugated drug candidates.

Design and synthesis of novel spin-labeled camptothecin derivatives as potent cytotoxic agents.

In our continuing search for natural product-based spin-labeled antitumor drugs, 20 novel spin-labeled camptothecin derivatives were synthesized via a Cu-catalyzed one pot reaction and evaluated for cytotoxicity against four human tumor cell lines (A-549, MDA-MB-231, KB, and KBvin). Eighteen of the target compounds (9a, 9b, 9d-9k, 9m-9t) exhibited significant in vitro antiproliferative activity against these four tested tumor cell lines. Compounds 9e and 9j (IC50 0.057 and 0.072µM, respectively) displayed the greatest cytotoxicity against the multidrug-resistant (MDR) KBvin cell line and merit further development into preclinical and clinical drug candidates for treating cancer including MDR phenotype.

Design and synthesis of new 7-(N-substituted-methyl)-camptothecin derivatives as potent cytotoxic agents.

A series of novel 7-(N-substituted-methyl)-camptothecin derivatives was designed, synthesized, and evaluated for in vitro cytotoxicity against four human tumor cell lines, A-549, MDA-MB-231, KB, and KBvin. All of the derivatives showed promising in vitro cytotoxic activity against the tested tumor cell lines, with IC50 values ranging from 0.0023 to 1.11 µM, and were as or more potent than topotecan. Compounds 9d, 9e, and 9r exhibited the highest antiproliferative activity among all prepared derivatives. Furthermore, all of the compounds were more potent than paclitaxel against the multidrug-resistant (MDR) KBvin subline. With a concise efficient synthesis and potent cytotoxic profiles, especially significant activity towards KBvin, compounds 9d, 9e, and 9r merit further development as a new generation of camptothecin-derived anticancer clinical trial candidates.