Multispot, label-free biodetection at a phantom plastic-water interface.

Recognizing and quantifying specific biomolecules in aqueous samples are constantly needed in research and diagnostic laboratories. As the typical detection procedures are rather lengthy and involve the use of labeled secondary antibodies or other agents to provide a signal, efforts have been made over the last 10 y to develop alternative label-free methods that enable direct detection. We propose and demonstrate an extremely simple, low-cost, label-free biodetector based on measuring the intensity of light reflected by the interface between a fluid sample and an amorphous fluoropolymer substrate having a refractive index very close to that of water and hosting various antibodies immobilized in spots. Under these index-matching conditions, the amount of light reflected by the interface allows straightforward quantification of the amount of antigen binding to each spot. Using antibodies targeting heterologous immunoglobulins and antigens commonly used as markers for diagnoses of hepatitis B and HIV, we demonstrate the limit of detection of a few picograms per square millimeter of surface-bound molecules. We also show that direct and real-time access to the amount of binding molecules allows the precise extrapolation of adhesion rates, from which the concentrations of antigens in solution can be estimated down to fractions of nanograms per milliliter.

Polyanionic drugs and viral oncogenesis: a novel approach to control infection, tumor-associated inflammation and angiogenesis.

Polyanionic macromolecules are extremely abundant both in the extracellular environment and inside the cell, where they are readily accessible to many proteins for interactions that play a variety of biological roles. Among polyanions, heparin, heparan sulfate proteoglycans (HSPGs) and glycosphingolipids (GSLs) are widely distributed in biological fluids, at the cell membrane and inside the cell, where they are implicated in several physiological and/or pathological processes such as infectious diseases, angiogenesis and tumor growth. At a molecular level, these processes are mainly mediated by microbial proteins, cytokines and receptors that exert their functions by binding to HSPGs and/or GSLs, suggesting the possibility to use polyanionic antagonists as efficient drugs for the treatment of infectious diseases and cancer. Polysulfated (PS) or polysulfonated (PSN) compounds are a heterogeneous group of natural, semi-synthetic or synthetic molecules whose prototypes are heparin and suramin. Different structural features confer to PS/PSN compounds the capacity to bind and inhibit the biological activities of those same heparin-binding proteins implicated in infectious diseases and cancer. In this review we will discuss the state of the art and the possible future development of polyanionic drugs in the treatment of infectious diseases and cancer.

Polysulfated/sulfonated compounds for the development of drugs at the crossroad of viral infection and oncogenesis.

Virus infection and oncogenesis are two tightly linked processes. Some viruses are endowed with a direct transforming capability and infection activates inflammation that, in turn, favours tumor progression. Also, both inflammation and tumor trigger (and are strongly dependent from) angiogenesis. Finally, some oncogenic viruses release "virokines" that contribute to the development of virus-associated tumors. At a molecular level, viral proteins, cytokines, receptors and adhesion molecules "cross-contribute" to the different processes and, amazingly, many of them bind to heparin and to heparan sulfate proteoglycans to exert their functions. Heparin-like polysulfated (PS) or polysulfonated (PSN) compounds are an heterogeneous group of natural or synthetic molecules whose prototypes are PS heparin and PSN suramin. They vary in their backbone structure, length, number/disposition of sulfated/sulfonated groups. Different combinations of these features confer to PS/PSN the capacity to bind with variable specificity to those heparin-binding proteins that "cross-contribute" to virus infection and tumor progression. Taken together, these considerations suggest that heparin-like PS/PSN antagonists may act as multitarget drugs that may control at once virus infection and tumor progression by targeting different proteins simultaneously. Here we discuss the possibility to exploit PS/PSN compounds for the development of drugs at the cross-road of viral infection and oncogenesis, taking in consideration the past efforts, possible drawbacks and future perspectives.

Heparin-mimicking sulfonic acid polymers as multitarget inhibitors of human immunodeficiency virus type 1 Tat and gp120 proteins.

Human immunodeficiency virus (HIV) Tat and gp120 intriguingly share the feature of being basic peptides that, once released by HIV(+) cells, bind to polyanionic heparan sulfate proteoglycans (HSPGs) on target uninfected cells, contributing to the onset of AIDS-associated pathologies. To identify multitarget anti-HIV prodrugs, we investigated the gp120 and Tat antagonist potentials of a series of polyanionic synthetic sulfonic acid polymers (SSAPs). Surface plasmon resonance revealed that SSAPs inhibit with a competitive mechanism of action the binding of Tat and gp120 to surface-immobilized heparin, an experimental condition that resembles binding to cellular HSPGs. Accordingly, SSAPs inhibited HSPG-dependent cell internalization and the transactivating activity of Tat. Little is known about the binding of free gp120 to target cells. Here, we identified two classes of gp120 receptors expressed on endothelial cells, one of which was consistent with an HSPG-binding, low-affinity/high-capacity receptor that is inhibited by free heparin. SSAPs inhibited the binding of free gp120 to endothelial cells, as well as its capacity to induce apoptosis in the same cells. In all the assays, poly(4-styrenesulfonic acid) (PSS) proved to be the most potent antagonist of Tat and gp120. Accordingly, PSS bound both proteins with high affinity. In conclusion, SSAPs represent an interesting class of compounds that bind both gp120 and Tat and inhibit their HSPG-dependent cell surface binding and pathological effects. As these activities contribute to both AIDS progression and associated pathologies, SSAPs can be considered prototypic molecules for the development of multitarget drugs for the treatment of HIV infection and AIDS-associated pathologies.