Multivalency plays a major role in biological processes and particularly in the relationship between pathogenic microorganisms and their host that involves protein-glycan recognition. These interactions occur during the first steps of infection, for specific recognition between host and bacteria, but also at different stages of the immune response. The search for high-affinity ligands for studying such interactions involves the combination of carbohydrate head groups with different scaffolds and linkers generating multivalent glycocompounds with controlled spatial and topology parameters. By interfering with pathogen adhesion, such glycocompounds including glycopolymers, glycoclusters, glycodendrimers and glyconanoparticles have the potential to improve or replace antibiotic treatments that are now subverted by resistance. Multivalent glycoconjugates have also been used for stimulating the innate and adaptive immune systems, for example with carbohydrate-based vaccines. Bacteria present on their surfaces natural multivalent glycoconjugates such as lipopolysaccharides and S-layers that can also be exploited or targeted in anti-infectious strategies.

Toll-Like Receptor 4 (TLR4) signal pathway plays an important role in initiating the innate immune response and its activation by bacterial endotoxin is responsible for chronic and acute inflammatory disorders that are becoming more and more frequent in developed countries. Modulation of the TLR4 pathway is a potential strategy to specifically target these pathologies. Among the diseases caused by TLR4 abnormal activation by bacterial endotoxin, sepsis is the most dangerous one because it is a life-threatening acute system inflammatory condition that still lacks specific pharmacological treatment. Here, we review molecules at a preclinical or clinical phase of development, that are active in inhibiting the TLR4-MyD88 and TLR4-TRIF pathways in animal models. These are low-molecular weight compounds of natural and synthetic origin that can be considered leads for drug development. The results of in vivo studies in the sepsis model and the mechanisms of action of drug leads are presented and critically discussed, evidencing the differences in treatment results from rodents to humans.

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In many Gram-negative bacteria, lipopolysaccharide (LPS) and its lipid A moiety are pivotal for bacterial survival. Depending on its structure, lipid A carries the toxic properties of the LPS and acts as a potent elicitor of the host innate immune system via the Toll-like receptor 4/myeloid differentiation factor 2 (TLR4/MD-2) receptor complex. It often causes a wide variety of biological effects ranging from a remarkable enhancement of the resistance to the infection to an uncontrolled and massive immune response resulting in sepsis and septic shock. Since the bioactivity of lipid A is strongly influenced by its primary structure, a broad range of chemical syntheses of lipid A derivatives have made an enormous contribution to the characterization of lipid A bioactivity, providing novel pharmacological targets for the development of new biomedical therapies. Here, we describe and discuss the chemical aspects regarding lipid A and its role in innate immunity, from the (bio)synthesis, isolation and characterization to the molecular recognition at the atomic level.

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Toll-like receptor 4 (TLR4), together with MD-2, binds bacterial endotoxins (E) with high affinity, triggering formation of the activated homodimer (E-MD-2-TLR4)2. Activated TLR4 induces intracellular signaling leading to activation of transcription factors that result in cytokine and chemokine production and initiation of inflammatory and immune responses. TLR4 also responds to endogenous ligands called danger associated molecular patterns (DAMPs). Increased sensitivity to infection and a variety of immune pathologies have been associated with either too little or too much TLR4 activation. We review here the molecular mechanisms of TLR4 activation (agonism) or inhibition (antagonism) by small organic molecules of both natural and synthetic origin. The role of co-receptors MD-2 and CD14 in the TLR4 modulation process is also discussed. Recent achievements in the field of chemical TLR4 modulation are reviewed, with special focus on non-classical TLR4 ligands with a chemical structure different from lipid A.

Neuropathic pain remains a prevalent clinical problem because it is often poorly responsive to the currently used analgesics, thus it is crucial the identification of new potential targets and drugs. Recent evidence indicated that microglial cells in the spinal cord are critically involved in the development and maintenance of neuropathic pain, with a pivotal role of toll-like receptor 4 (TLR4). Binding of an endogenous ligand to TLR4 might be considered an important step in the regulation of microglia activity in pain facilitation, suggesting that a mechanism aimed to inhibit such a binding could be effective against neuropathic pain. We have synthesized new ligands to TLR4 with antagonistic activity. In the present work we evaluated the efficacy of the most potent TLR4 antagonist synthesized by us (FP-1), administered in mice with painful neuropathy. The repeated treatment of neuropathic mice with FP-1 (5-10 mg/kg, i.p.) evoked a relief of both thermal hyperalgesia and mechanical allodynia, whereas the administration of the highest dose to TLR4 knockout neuropathic mice revealed that in the absence of TLR4 receptor, the compound lost its efficacy. As consequence of TLR4 binding, the repeated treatment with FP-1 prevented the activation of the transcription factor NF-kB and the TNFalpha overproduction in the spinal cord. Together, our findings support the previous evidence indicative for a contribution of glial TLR4 to the initiation of neuropathic pain, suggest it as potential innovative target to treat this debilitating disease, and propose FP-1 as lead compound for the development of new effective drugs.

Angiogenesis plays a key role in various physiologic and pathologic conditions, including tumor growth. Drm/gremlin, a member the Dan family of bone morphogenic protein (BMP) antagonists, is commonly thought to affect different processes during growth, differentiation, and development by heterodimerizing various BMPs. Here, we identify Drm/gremlin as a novel proangiogenic factor expressed by endothelium. Indeed, Drm/gremlin was purified to homogeneity from the conditioned medium of transformed endothelial cells using an endothelial-cell sprouting assay to follow protein isolation. Accordingly, recombinant Drm/gremlin stimulates endothelial-cell migration and invasion in fibrin and collagen gels, binds with high affinity to various endothelial cell types, and triggers tyrosine phosphorylation of intracellular signaling proteins. Also, Drm/gremlin induces neovascularization in the chick embryo chorioallantoic membrane. BMP4 does not affect Drm/gremlin interaction with endothelium, and both molecules exert a proangiogenic activity in vitro and in vivo when administered alone or in combination.