Pharmacological Effects of FTY720 and its Derivatives.

FTY720 is an analog of sphingosine-1-phosphate (S1P) derived from the ascomycete Cordyceps sinensis. As a new immunosuppressant, FTY720 is widely used to treat multiple sclerosis. FTY720 binds to the S1P receptor after phosphorylation, thereby exerting immunosuppressive effects. The nonphosphorylated form of FTY720 can induce cell apoptosis, enhance chemotherapy sensitivity, and inhibit tumor metastasis of multiple tumors by inhibiting SPHK1 (sphingosine kinase 1) and activating PP2A (protein phosphatase 2A) and various cell death pathways. FTY720 can induce neutrophil extracellular traps to neutralize and kill pathogens in vitro, thus exerting anti- infective effects. At present, a series of FTY720 derivatives, which have pharmacological effects such as anti-tumor and alleviating airway hyperresponsiveness, have been developed through structural modification. This article reviews the pharmacological effects of FTY720 and its derivatives.

An Immunomodulatory Peptide Dendrimer Inspired from Glatiramer Acetate.

Glatiramer acetate (GA) is a random polypeptide drug used to treat multiple sclerosis (MS), a chronic autoimmune disease. With the aim of identifying a precisely defined alternative to GA, we synthesized a library of peptide dendrimers with an amino acid composition similar to GA. We then challenged the dendrimers to trigger the release of the anti-inflammatory cytokine interleukin-1 receptor antagonist (IL-1Ra) from human monocytes, which is one of the effects of GA on immune cells. Several of the largest dendrimers tested were as active as GA. Detailed profiling of the best hit showed that this dendrimer induces the differentiation of monocytes towards an M2 (anti-inflammatory) state as GA does, however with a distinct immune marker profile. Our peptide dendrimer might serve as starting point to develop a well-defined immunomodulatory analog of GA.

Synthesis and Biological Evaluation of Celastrol Derivatives as Potential Immunosuppressive Agents.

Celastrol, a friedelane-type triterpenoid isolated from the genus Triperygium, possesses antitumor, anti-inflammatory, and immunosuppressive activities. A total of 42 celastrol derivatives (1a-1t, 2a-2l, and 3a-3j) were synthesized and evaluated for their immunosuppressive activities. Compounds 2a-2e showed immunosuppressive effects, with IC50 values ranging from 25 to 83 nM, and weak cytotoxicity (CC50 > 1 µM). Compound 2a, with a selectivity index value 31 times higher than that of celastrol, was selected as a lead compound. Further research showed that 2a exerted its immunosuppressive effects by inducing apoptosis and inhibiting cytokine secretion via Lck- and ZAP-70-mediated signaling pathways.

Local delivery of FTY720 in mesoporous bioactive glass improves bone regeneration by synergistically immunomodulating osteogenesis and osteoclastogenesis.

The addition of osteoimmunology drugs to bone repair materials is beneficial to bone regeneration by regulating the local immune microenvironment. Fingolimod (FTY720) has been reported to be an osteoimmunology drug that promotes osteogenesis. However, there is no ideal biomaterial for the sustained release of FTY720 in the bone defect areas. In the present work, FTY720 loaded mesoporous bioactive glass (FTY720@MBGs) was successfully prepared based on the mesoporous properties of MBGs and electrostatic attraction. FTY720 achieved a sustained release for 7 days. The in vitro study found that FTY720@MBGs could synergistically promote osteogenesis and inhibit osteoclastogenesis due to their ability to promote macrophages toward the M2 phenotype. The in vivo study confirmed that FTY720@MBGs could significantly improve bone regeneration. This study provides new strategies for designing smart cell-instructive biomaterials that can play a role in all bone healing processes from early inflammation to bone reconstruction.

Novel Coupled Molecules from Active Structural Motifs of Synthetic and Natural Origin as Immunosuppressants.

INTRODUCTION: Nitric oxide (NO) is an important mediator in the pathogenesis and control of immune system-related disorders and its levels are modulated by inducible NO synthase (iNOS). Oxidative stress is another pathological indication in majority of autoimmune disorders. The present study aims at the development of coupled molecules via selection of pharmacophores for both immunomodulatory and antioxidant activities through iNOS inhibition. METHODS: Variedly substituted coumarin moieties are coupled with naturally occurring phenols through an amide linkage and were predicted for activities using computer-based program PASS. The compounds predicted to have dual activities were synthesized. Docking studies were carried out against iNOS (PDB 1R35) and compounds having good docking score were evaluated for immunomodulatory and antioxidant activities. RESULTS: The synthesized compounds were found to be pure and were obtained in good yields. Compounds with maximum docking score (YR1a, YR2e, YR2c and YR4e) were selected for evaluation by in vitro models. Compounds YR2e and YR2c markedly inhibited the reduction of NBT dye and showed maximum % iNOS inhibition. In DPPH assay, compound YR4e was observed as the most potent antioxidant (EC50 0.33 µM/mL). Based on these studies, compounds YR2e and YR2c were selected for haemagglutination test. Compound YR2e was observed as the most active immunosuppressant with maximal inhibitory ability of iNOS and NBT reduction and lower HAT value of 3.5. CONCLUSION: Compound YR2e can be utilized as a pharmacological agent in the prevention or treatment of immunomodulatory diseases such as tumors, rheumatoid arthritis, ulcerative colitis, organ transplant and other autoimmune disorders.

Identification of U11snRNA as an endogenous agonist of TLR7-mediated immune pathogenesis.

The activation of innate immune receptors by pathogen-associated molecular patterns (PAMPs) is central to host defense against infections. On the other hand, these receptors are also activated by immunogenic damage-associated molecular patterns (DAMPs), typically released from dying cells, and the activation can evoke chronic inflammatory or autoimmune disorders. One of the best known receptors involved in the immune pathogenesis is Toll-like receptor 7 (TLR7), which recognizes RNA with single-stranded structure. However, the causative DAMP RNA(s) in the pathogenesis has yet to be identified. Here, we first developed a chemical compound, termed KN69, that suppresses autoimmunity in several established mouse models. A subsequent search for KN69-binding partners led to the identification of U11 small nuclear RNA (U11snRNA) as a candidate DAMP RNA involved in TLR7-induced autoimmunity. We then showed that U11snRNA robustly activated the TLR7 pathway in vitro and induced arthritis disease in vivo. We also found a correlation between high serum level of U11snRNA and autoimmune diseases in human subjects and established mouse models. Finally, by revealing the structural basis for U11snRNA's ability to activate TLR7, we developed more potent TLR7 agonists and TLR7 antagonists, which may offer new therapeutic approaches for autoimmunity or other immune-driven diseases. Thus, our study has revealed a hitherto unknown immune function of U11snRNA, providing insight into TLR7-mediated autoimmunity and its potential for further therapeutic applications.

Simplified immunosuppressive and neuroprotective agents based on gracilin A.

The architecture and bioactivity of natural products frequently serve as embarkation points for the exploration of biologically relevant chemical space. Total synthesis followed by derivative synthesis has historically enabled a deeper understanding of structure-activity relationships. However, synthetic strategies towards a natural product are not always guided by hypotheses regarding the structural features required for bioactivity. Here, we report an approach to natural product total synthesis that we term 'pharmacophore-directed retrosynthesis'. A hypothesized, pharmacophore of a natural product is selected as an early synthetic target and this dictates the retrosynthetic analysis. In an ideal application, sequential increases in the structural complexity of this minimal structure enable development of a structure-activity relationship profile throughout the course of the total synthesis effort. This approach enables the identification of simpler congeners retaining bioactivity at a much earlier stage of a synthetic effort, as demonstrated here for the spongiane diterpenoid, gracilin A, leading to simplified derivatives with potent neuroprotective and immunosuppressive activity.

Synthesis and antiproliferative activity of new mycophenolic acid conjugates with adenosine derivatives.

New conjugates of mycophenolic acid (MPA) and adenosine derivatives were synthesized and assessed as potential immunosuppressants on Jurkat cell line and peripheral blood mononuclear cells (PBMC) from healthy donors. As compared to MPA, all compounds were found to be more active against Jurkat cell line. The antiproliferative activities were compared with MPA and adenosine, in both 2',3'-O-isopropylidene protected and free hydroxyl groups possessing forms. The obtained results were also discussed in terms of selectivity index, defined as SI = IC50/EC50.

Synthesis, Immunosuppressive Properties, and Mechanism of Action of a New Isoxazole Derivative.

This work describes the synthesis of a new series of isoxazole derivatives, their immunosuppressive properties, and the mechanism of action of a representative compound. A new series of N'-substituted derivatives of 5-amino-N,3-dimethyl-1,2-oxazole-4-carbohydrazide (MM1⁻MM10) was synthesized in reaction of 5-amino-N,3-dimethyl-1,2-oxazole-4-carbohydrazide with relevant carbonyl compounds. The isoxazole derivatives were tested in several in vitro models using human cells. The compounds inhibited phytohemagglutinin A (PHA)-induced proliferation of peripheral blood mononuclear cells (PBMCs) to various degrees. The toxicity of the compounds with regard to a reference A549 cell line was also differential. 5-amino-N'-(2,4-dihydroxyphenyl)methylidene-N,3-dimethyl-1,2-oxazole-4-carbohydrazide (MM3) compound was selected for further investigation because of its lack of toxicity and because it had the strongest antiproliferative activity. The compound was shown to inhibit lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF α) production in human whole blood cell cultures. In the model of Jurkat cells, MM3 elicited strong increases in the expression of caspases, Fas, and NF-κB1, indicating that a proapoptotic action may account for its immunosuppressive action in the studied models.

Determination of the mitigating effect of colon-specific bioreversible codrugs of mycophenolic acid and aminosugars in an experimental colitis model in Wistar rats.

AIM: To design colon-targeted codrugs of mycophenolic acid (MPA) and aminosugars as a safer option to mycophenolate mofetil (MMF) in the management of inflammatory bowel disease. METHODS: Codrugs were synthesized by coupling MPA with aminosugars (D-glucosamine and D-galactosamine) using EDCI coupling. The structures were confirmed by infrared radiation, nuclear magnetic resonance, mass spectroscopy and elemental analysis. The release profile of codrugs was extensively studied in aqueous buffers, upper gastrointestinal homogenates, faecal matter and caecal homogenates (in vitro) and rat blood (in vitro). Anti-colitic activity was assessed in 2,4,6-trinitrobezenesulfonic acid-induced colitis in Wistar rats by the estimation of various demarcating parameters. Statistical evaluation was performed by applying one-way and two-way ANOVA when compared with the disease control. RESULTS: The prodrugs resisted activation in HCl buffer (pH 1.2) and stomach homogenates of rats with negligible hydrolysis in phosphate buffer (pH 7.4) and intestinal homogenates. Incubation with colon homogenates (in vitro) produced 76% to 89% release of MPA emphasizing colon-specific activation of codrugs and the release of MPA and aminosugars at the site of action. In the in vitro studies, the prodrug of MPA with D-glucosamine (MGLS) was selected which resulted in 68% release of MPA in blood. in vitro studies on MGLS revealed its colon-specific activation after a lag time of 8 h which could be ascribed to the hydrolytic action of N-acyl amidases found in the colon. The synthesized codrugs markedly diminished disease activity score and revived the disrupted architecture of the colon that was comparable to MMF but superior to MPA. CONCLUSION: The significant attenuating effect of prodrugs and individual aminosugars on colonic inflammation proved that the rationale of the codrug approach is valid.

Development of Glatopa® (Glatiramer Acetate): The First FDA-Approved Generic Disease-Modifying Therapy for Relapsing Forms of Multiple Sclerosis.

The multiple sclerosis (MS) treatment landscape in the United States has changed dramatically over the past decade. While many disease-modifying therapies (DMTs) have been approved by the US Food and Drug Administration (FDA) for the treatment of relapsing forms of MS, DMT costs continue to rise. The availability of generics and biosimilars in the MS-treatment landscape is unlikely to have a major impact on clinical benefit. However, their availability will provide alternative treatment options and potentially lower costs through competition, thus increasing the affordability of and access to these drugs. In April 2015, the first generic version of the complex drug glatiramer acetate (Glatopa® 20 mg/mL) injection was approved in the United States as a fully substitutable generic for all approved indications of the 20 mg/mL branded glatiramer acetate (Copaxone®) dosage form. Despite glatiramer acetate's complex nature-being a chemically synthesized (ie, nonbiologic) mixture of peptides-the approval occurred without conducting any clinical trials. Rather, extensive structural and functional characterization was performed to demonstrate therapeutic equivalence to the innovator drug. The approval of Glatopa signifies an important milestone in the US MS-treatment landscape, with the hope that the introduction of generic DMTs and eventually biosimilar DMTs will lead to future improvements in the affordability and access of these much-needed treatments for MS.

Binding Mode and Structure-Activity Relationships of ITE as an Aryl Hydrocarbon Receptor (AhR) Agonist.

Discovered as a modulator of the toxic response to environmental pollutants, aryl hydrocarbon receptor (AhR) has recently gained attention for its involvement in various physiological and pathological pathways. AhR is a ligand-dependent transcription factor activated by a large array of chemical compounds, which include metabolites of l-tryptophan (l-Trp) catabolism as endogenous ligands of the receptor. Among these, 2-(1'H-indole-3'-carbonyl)thiazole-4-carboxylic acid methyl ester (ITE) has attracted interest in the scientific community, being endowed with nontoxic, immunomodulatory, and anticancer AhR-mediated functions. So far, no information about the binding mode and interactions of ITE with AhR is available. In this study, we used docking and molecular dynamics to propose a putative binding mode of ITE into the ligand binding pocket of AhR. Mutagenesis studies were then instrumental in validating the proposed binding mode, identifying His 285 and Tyr 316 as important key residues for ligand-dependent receptor activation. Finally, a set of ITE analogues was synthesized and tested to further probe molecular interactions of ITE to AhR and characterize the relevance of specific functional groups in the chemical structure for receptor activity.

Synthesis and Functional Investigations of Computer Designed Novel Cladribine-Like Compounds for the Treatment of Multiple Sclerosis.

Cladribine (2-CdA) is used as an anti-cancer drug but is currently studied as a potential treatment for use in relapsing-remitting multiple sclerosis (MS). In this study, we computer designed, synthesized, and characterized two novel derivatives of 2-CdA, K1-5d and K2-4c, and investigated their underlying mechanism of beneficial effect using the CCRF-CEM and RAJI cell lines. For this purpose, we first determined their effect on MS and DNA damage and repair-related gene expression profiles using custom arrays along with 2-CdA treatment at non-toxic doses. Then, we determined whether cells underwent apoptosis after treatment with 2-CdA, K1-5d, and K2-4c in CCRF-CEM and RAJI cells, using the DNA fragmentation assay. It was found that both derivatives modulated the expression of the pathway-related genes that are important in inflammatory signaling, apoptosis, ATM/ATR, double-strand break repair, and the cell cycle. Furthermore, 2-CdA, K1-5d, and K2-4c significantly activated apoptosis in both cell lines. In summary, our data demonstrate that although both derivatives act as anti-inflammatory and apoptotic agents, inducing the accumulation of DNA strand breaks and activating the ultimate tumor suppressor p53 in T and B lymphocytes, the K1-5d derivative has shown more promising activities for further studies.

Synthesis and Immunosuppressive Activity of New Mycophenolic Acid Derivatives.

BACKGROUND: Immunosuppressive drugs are widely used to prevent and treat allograft rejection and autoimmune diseases. Mycophenolic acid (MPA) and its derivatives are currently one of the most prescribed immunosuppressive drugs; however, metabolic drawbacks and variable interand intrapatient responses limit their use. OBJECTIVE: In order to find out new safe and effective immunosuppressive compounds, we report here the synthesis and pharmacological evaluation of hybrid MPA derivatives containing the thalidomide/ phthalimide subunits. RESULTS: All compounds 3a-d exhibited an enhanced ability to reduce the levels of pro-inflammatory cytokines compared to the parental drugs MPA and thalidomide. The mixed lymphocyte reaction assay has demonstrated that compound 3d - (E)-(3-(1,3-dioxoisoindolin-2-yl)-2,6-dioxopiperidin-1- yl)methyl-6-(4-hydroxy-6-methoxy-7-methyl-3-oxo-1,3-dihydroisobenzofuran-5-yl)-4-methylhex-4- enoate - has superior activity compared to that of MPA. In addition, compound 3d was less cytotoxic against Jurkat cells than MPA and did not demonstrate in vivo genotoxic effect. CONCLUSION: All these data have shown that compound 3d is a promising lead compound useful in the immunosuppressive therapy.

Discovery, Total Synthesis and Key Structural Elements for the Immunosuppressive Activity of Cocosolide, a Symmetrical Glycosylated Macrolide Dimer from Marine Cyanobacteria.

A new dimeric macrolide xylopyranoside, cocosolide (1), was isolated from the marine cyanobacterium preliminarily identified as Symploca sp. from Guam. The structure was determined by a combination of NMR spectroscopy, HRMS, X-ray diffraction studies and Mosher's analysis of the base hydrolysis product. Its carbon skeleton closely resembles that of clavosolides A-D isolated from the sponge Myriastra clavosa, for which no bioactivity is known. We performed the first total synthesis of cocosolide (1) along with its [α,α]-anomer (26) and macrocyclic core (28), thus leading to the confirmation of the structure of natural 1. The convergent synthesis featured Wadsworth-Emmons cyclopropanation, Sakurai annulation, Yamaguchi macrocyclization/dimerization reaction, α-selective glycosidation and ß-selective glycosidation. Compounds 1 and 26 potently inhibited IL-2 production in both T-cell receptor dependent and independent manners. Full activity requires the presence of the sugar moiety as well as the intact dimeric structure. Cocosolide also suppressed the proliferation of anti-CD3-stimulated T-cells in a dose-dependent manner.

The Plant Resources, Structure Characteristics, Biological Activities and Synthesis of Pyranoflavonoids.

Pyranoflavonoids are mainly distributed in Leguminosae, Moraceae. As a potent drug candidate, pyranoflavonoids have attracted much attention in late years due to their impressive pharmaceutical activities. The structure characteristics, plant resources and pharmaceutical activities of pyranoflavonoids are reviewed in this paper. The fragmentation in mass spectrometry is also discussed for quick identification of these chemicals. Due to the low abundance of this flavonoid sub-class in plants, it is required to find an efficient way to synthesize them. Therefore, an overview of chemical synthesis and biological synthesis is presented in this paper. The information given in this paper will be helpful to know pyranoflavonoids and to be utilized in medicines and nutraceuticals.

A phenotypic drug discovery study on thienodiazepine derivatives as inhibitors of T cell proliferation induced by CD28 co-stimulation leads to the discovery of a first bromodomain inhibitor.

A phenotypic screening of thienodiazepines derived from a hit compound found through a binding assay targeting co-stimulatory molecules on T cells and antigen presenting cells successfully led to the discovery of a thienotriazolodiazepine compound (7f) possessing potent immunosuppressive activity. A chemical biology approach has succeeded in revealing that 7f is a first inhibitor of epigenetic bromodomain-containing proteins. 7f is expected to become an anti-cancer agent as well as an immunosuppressive agent.

(E)-1,3-diphenyl-1H-pyrazole derivatives containing O-benzyl oxime moiety as potential immunosuppressive agents: Design, synthesis, molecular docking and biological evaluation.

A series of novel (E)-1,3-diphenyl-1H-pyrazole derivatives containing O-benzyl oxime moiety were firstly synthesized and their immunosuppressive activities were evaluated. Among all the compounds, 4n exhibited the most potent inhibitory activity (IC50 = 1.18 µM for lymph node cells and IC50 = 0.28 µM for PI3Kγ), which was comparable to that of positive control. Moreover, selected compounds were tested for their inhibitory activities against IL-6 released in ConA-simulated mouse lymph node cells, 4n exhibited the most potent inhibitory ability. Furthermore, in order to study the preliminary mechanism of the compounds with potent inhibitory activity, the RT-PCR experiment was performed to assay the effect of selected compounds on mRNA expression of IL-6. Among them, compound 4n strongly inhibited the expression of IL-6 mRNA.

A Novel Synthetic Compound, YH-1118, Inhibited LPS-Induced Inflammatory Response by Suppressing IkappaB Kinase/NF-kappaB Pathway in Raw 264.7 Cells.

For the search of a potent first-in-class compound to inactivate macrophages responsible for inflammatory responses, in the present study, we investigated the anti-inflammatory effects of YH-1118, a novel synthetic compound, in a lipopolysaccharide (LPS)-stimulated mouse macrophage cell line, Raw 264.7. YH-1118 inhibited LPS-induced nitric oxide (NO) production and inducible NO synthase (iNOS) expression at both the protein and mRNA levels. The suppression of LPS-induced iNOS expression by YH-1118 was mediated via nuclear factor kappa B (NF-κB), but not activator protein-1 (AP-1) transcription factor. This was supported by the finding that YH-1118 attenuated the phosphorylation of inhibitor of κBα (IκBα) and nuclear translocation and DNA binding activity of NF-kappaB. Through the mechanisms that YH- 1118 inhibited the activation of IkappaB kinases (IKKs), upstream activators of NF-κB, or p38 MAPK, YH-1118 significantly suppressed LPS-induced production of pro-inflammatory cytokines, tumor necrosis factor-α, interleukin-1ß (IL-1ß), and IL-6 (p < 0.05). In conclusion, our results suggest that YH-1118 inhibits LPS-induced inflammatory responses by blocking IKK and NF-κB activation in macrophages, and may be a therapeutic candidate for the treatment of various inflammatory diseases.