Small Molecular Weight Antagonist Research Articles

Dynamics of the Human Y1-Receptor in the Inactive and APO States Investigated by Solid-State Nuclear Magnetic Resonance Spectroscopy and Molecular Dynamics Simulations

The Y1 receptor (Y1R) is part of the G protein-coupled receptor (GPCR) superfamily, which plays a crucial role in many biological signaling processes and is of high interest for medical applications. It is involved in many physiological and pathophysiological functions, related to obesity and cancer. Y1R is activated by neuropeptide Y (NPY), which is involved in several physiological processes including the stimulation of food intake. Here, we studied the receptor dynamics bound to UR-MK299, which is a small molecular weight antagonist, and in the apo form both embedded in POPC membranes at 37 °C. For investigation by nuclear magnetic resonance (NMR) spectroscopy the receptor was labeled uniformly with 13C. This allowed investigating its dynamics by measurement of order parameters. However, due to the uniform labeling the NMR order parameters are averaged over the whole receptor without any site resolution and therefore could only show general differences between the different states. To obtain information of individual segmental motions, we conducted molecular dynamics (MD) simulations of both states using the recently published crystal structure in presence of UR-MK299 (PDB entry code: 5ZBQ). For the apo state, the antagonist was removed and the receptor relaxed for several microseconds before analysis. For the UR-MK299 bound state the CHARMM force field of the antagonist was parametrized with Gaussian. The simulations were prepared following a newly developed general scheme for setup of GPCR MD simulations. The production runs were conducted for several microseconds and compared to the experimentally available data followed by a detailed analysis of the segmental motions. Aim of this analysis is to elucidate the complex structure function dynamics relationship of GPCRs combining state-of-the-art biophysical methods to obtain their time resolved motions.

CD22-Antagonists with nanomolar potency: The synergistic effect of hydrophobic groups at C-2 and C-9 of sialic acid scaffold

In earlier studies, we identified the C-9 amido derivative 1 (9-(4′-hydroxy-4-biphenyl)acetamido-9-deoxy-Neu5Gcα2-6GalOMP) and the C-9 amino derivative 2 (9-(4′-hydroxy-4-biphenyl)methylamino-9-deoxy-Neu5Gcα2-6GalOMP) have the most promising affinity for mouse CD22 and human CD22, respectively. Replacing the subterminal galactose residue (2-6Gal-OMP) of 1 with benzyl (5) or biphenylmethyl (6) as aglycone led to even higher potency for mCD22. In this study, both compounds showed improved potency and selectivity for CD22 (IC50 70nM) and 712-fold more selective for CD22 than for MAG. The corresponding derivatives of 2, compounds 8 and 9, showed comparable activity to 2 but lower potency and selectivity than 5 and 6. Although compounds 5–9 are simple and small molecular weight antagonists, they showed much high potency and selectivity than the corresponding compounds having α 2-6Gal linkage. Both biological and computational docking simulation studies suggest that the 2-6Gal-OMP residues of 1 and 2 are not critical for binding process and could be replaced with hydrophobic non-carbohydrate moieties. The data presented herein has significant implications for the design and discovery of next-generation CD22-antagonists.

The p53-MDM2/MDMX axis - A chemotype perspective.

The protein-protein interaction (PPI) of the tumor suppressor p53 and its negative regulator MDM2 consists of the most intense studied PPI with a group of small molecular weight antagonists described and many more disclosed in patent literature. Due to the Å-level structural insight into p53 interaction with MDM2 there is a reasonable understanding of the requirements of the molecules to bind. In contrast and despite the very close homology and 3-D similarity no potent MDMX antagonist has been disclosed up to date. The current review summarizes the different disclosed chemotypes for MDM2 including a discussion of the cocrystal structures. Structures and approaches to reconstitute functional p53 from mutated p53 are presented. Finally new screening methods and recent biotech deals based on p53 are discussed.

Organ-specific inhibition of metastatic colon carcinoma by CXCR3 antagonism

e14619 Background: Liver and lung metastases are the predominant cause of colorectal cancer (CRC) related mortality. Recent research has indicated that CXCR3/chemokines interactions that orchestrate hematopoetic cell movement are implicated in the metastatic process of malignant tumors, including that of CRC cells to lymph nodes. To date, however, the contribution of CXCR3 to liver and lung metastasis in CRC has not been addressed. Methods: To determine whether CXCR3 receptors regulate malignancy-related properties of CRC cells, we have used CXCR3-expressing CRC cell lines of human (HT29 cells) and murine (C26 cells) origins that enable the development of liver and lung metastases when injected into immunodeficient and immunocompetent mice, respectively, and assessed the effect of CXCR3 blockade using AMG487, a small molecular weight antagonist. Results: In vitro, activation of CXCR3 on human and mouse CRC cells by its cognate ligands induced migratory and growth responses, both activities being abrogated by AMG487. In vivo, systemic CXCR3 antagonism by preventive or curative treatments with AMG487 markedly inhibited metastasis of human and mouse CRC cells to the lung without affecting that to the liver. Also, we measured increased levels of CXCR3 and ligands expression within lung nodules compared to liver tumors. Conclusions: Altogether, our findings indicate that CXCR3 is a key mediator for lung metastasis of colon carcinoma and identify CXCR3-blocking approaches as promising strategies against established lung metastases. No significant financial relationships to disclose.

Organ-specific inhibition of metastatic colon carcinoma by CXCR3 antagonism

Liver and lung metastases are the predominant cause of colorectal cancer (CRC)-related mortality. Recent research has indicated that CXCR3/chemokines interactions that orchestrate haematopoetic cell movement are implicated in the metastatic process of malignant tumours, including that of CRC cells to lymph nodes. To date, however, the contribution of CXCR3 to liver and lung metastasis in CRC has not been addressed. To determine whether CXCR3 receptors regulate malignancy-related properties of CRC cells, we have used CXCR3-expressing CRC cell lines of human (HT29 cells) and murine (C26 cells) origins that enable the development of liver and lung metastases when injected into immunodeficient and immunocompetent mice, respectively, and assessed the effect of CXCR3 blockade using AMG487, a small molecular weight antagonist. In vitro, activation of CXCR3 on human and mouse CRC cells by its cognate ligands induced migratory and growth responses, both activities being abrogated by AMG487. In vivo, systemic CXCR3 antagonism by preventive or curative treatments with AMG487 markedly inhibited the implantation and the growth of human and mouse CRC cells within lung without affecting that in the liver. In addition, we measured increased levels of CXCR3 and ligands expression within lung nodules compared with liver tumours. Altogether, our findings indicate that activation of CXCR3 receptors by its cognate ligands facilitates the implantation and the progression of CRC cells within lung tissues and that inhibition of this axis decreases pulmonary metastasis of CRC in two murine tumour models.

288: A small molecular weight antagonist of CXCR3 reduces the severity of experimental idiopathic pneumonia syndrome and improves survival following allogeneic stem cell transplantation

BACKGROUND: Idiopathic pneumonia syndrome (IPS) is a frequently fatal complication after allogeneic stem cell transplantation (allo-SCT) The pathophysiology of IPS involves the secretion of inflammatory cytokines including IFNγ and TNFα along with the recruitment of donor T cells to the lung. CXCR3 is a chemokine receptor that is expressed on activated Th1/Tc1 T cell subsets and the expression of its ligands CXCL9 (Mig) and CXCL10 (IP-10) can be induced in a variety of cell types by IFNγ alone or in combination with TNFα.

Antagonism of CXCR3 Inhibits Lung Metastasis in a Murine Model of Metastatic Breast Cancer

Tumor cells aberrantly express chemokines and/or chemokine receptors, and some may promote tumor growth and metastasis. We examined the expression and function of chemokine receptor CXCR3 in a syngeneic murine model of metastatic breast cancer. By flow cytometry, CXCR3 was detected in all murine mammary tumor cell lines examined. All human breast cancer cell lines examined also expressed CXCR3, as did the immortalized but nontumorigenic MCF-10A cell line. Interaction of CXCR3 ligands, CXCL9, CXCL10, and CXCL11, with CXCR3 on the highly malignant murine mammary tumor cell line 66.1 resulted in intracellular calcium mobilization and chemotaxis in vitro. To test the hypothesis that tumor metastasis is facilitated by CXCR3 expressed by tumor cells, we employed a small molecular weight antagonist of CXCR3, AMG487. 66.1 tumor cells were pretreated with AMG487 prior to i.v. injection into immune-competent female mice. Antagonism of CXCR3 on 66.1 tumor cells inhibited experimental lung metastasis, and this antimetastatic activity was compromised in mice depleted of natural killer cells. Systemic administration of AMG487 also inhibited experimental lung metastasis. In contrast to the antimetastatic effect of AMG487, local growth of 66.1 mammary tumors was not affected by receptor antagonism. These studies indicate that murine mammary tumor cells express CXCR3 which facilitates the development of lung metastases. These studies also indicate for the first time that a small molecular weight antagonist of CXCR3 has the potential to inhibit tumor metastasis.

Inhibition of human immunodeficiency virus type 1 infection in macrophages by an alpha-v integrin blocking antibody

Macrophages are key cells for HIV infection and HIV spreading inside the organism. Macrophages cultured in vitro can be successfully infected after differentiation with cytokines such as macrophage colony stimulating factor (M-CSF). In the monocyte to macrophage differentiation process with M-CSF, αv-integrins are upregulated concomitantly with the capacity of HIV to generate a productive virus infection. In the present study we show that an anti-αv antibody, 17E6, inhibited HIV-1 infection of primary macrophages. The effect of 17E6 on HIV-1 BaL replication in acutely infected macrophages was dose-dependent, with a 50% effective concentration (EC 50) of 17 ± 2 μg/ml in the absence of cytotoxicity. Similarly, a monoclonal antibody targeting the αvβ6 integrin (14D9.F8) also inhibited HIV-1 BaL infection in this cell type. 17E6 reduced the detection of HIV-1 BaL proviral DNA in acutely infected macrophages, but was completely ineffective against HIV-1 BaL production in chronically infected macrophages, suggesting that 17E6 inhibited HIV infection at an early stage of the virus cycle. Finally, a small molecular weight antagonist of the αvβ6 integrin, EMD 409849, reduced HIV replication at subtoxic concentrations. Therefore, our results suggest that αv-containing integrins could play a role in HIV replication in macrophages and suggest that small-molecular-weight compounds might interfere with HIV replication in macrophages through the interaction with αv integrins.

Inhibition of the development of collagen‐induced arthritis in rhesus monkeys by a small molecular weight antagonist of CCR5

Collagen-induced arthritis (CIA) in the rhesus monkey is a nonhuman primate model of rheumatoid arthritis (RA). The close phylogenetic relationship between humans and the rhesus monkey makes this model useful for the preclinical safety and efficacy testing of new therapies that are inactive in animals more distinctly related to humans. In this study, we tested the therapeutic potential of a novel, small molecular weight antagonist of CCR5, SCH-X, in this model. CIA was induced in 10 rhesus monkeys. The animals were allocated to receive SCH-X or saline as the control (n = 5 in each group). Treatment was initiated on the day of CIA induction and continued for 45 days. Monkeys were monitored before and 63 days after CIA induction for macroscopic signs of clinical arthritis, such as soft-tissue swelling and body weight. Furthermore, markers of inflammation and joint degradation were monitored to follow the disease course. Only 2 of 5 animals in the SCH-X-treated group displayed prominent soft-tissue swelling, compared with all 5 saline-treated monkeys. In addition to the suppression of joint inflammation, treatment with SCH-X resulted in a reduction in joint destruction, as demonstrated by lower rates of urinary excretion of collagen crosslinks, with confirmation by histology. Whereas in all saline-treated monkeys, marked erosion of joint cartilage was observed, this was absent in 4 of the 5 SCH-X-treated monkeys. The systemic effects of treatment with SCH-X were a suppressed acute-phase reaction (reduction in C-reactive protein level) in the 3 treated monkeys with CIA that remained asymptomatic, and an altered antibody response toward type II collagen. The results suggest that the CCR5 antagonist SCH-X might have a strong clinical potential for treatment during periods of active inflammation, as seen in RA.

Inhibition of Immune Complex-Induced Inflammation by A small Molecular Weight Selectin Antagonist

The anti-inflammatory effect of a small molecular weight antagonist of P- and E-selectin-dependent cell adhesion was examined. The glycolipid sulphatide was shown to block the adherence of thrombin-activated rat platelets to HL-60 cells. This interaction is known to be dependent on P-selectin. The rat dermal reverse passive Arthus reaction was used to assess the effect of sulphatide on a neutrophil dependent inflammatory response. Sulphatide dosedependently blocked both the vascular permeability increase and cell infiltration after intraperitoneal administration. These results show that a small molecular weight compound which blocks P- and E-selectin dependent adhesion in vitro can effectively block the inflammation due to immune complex deposition. A compound with this type of profile may have therapeutic potential in the treatment of immune complex mediated diseases.