Peptides and Drug Delivery.

Peptides have been used as drugs to treat various health conditions, and they are also being developed as diagnostic agents. Due to their receptor selectivity, peptides have recently been utilized for drug delivery to target drug molecules to specific types of cells (i.e. cancer cells, immune cells) to lower the side effects of the drugs. In this case, the drug is conjugated to the carrier peptide for directing the drug to the target cells (e.g. cancer cells) with higher expression of a specific receptor that recognizes the carrier peptide. As a result, the drug is directed to the target diseased cells without affecting the normal cells. Peptides are also being developed for improving drug delivery through the intestinal mucosa barrier (IMB) and the blood-brain barrier (BBB). These peptides were derived from intercellular junction proteins such as occludins, claudins, and cadherins and improve drug delivery through the IMB and BBB via the paracellular pathways. It is hypothesized that the peptides modulate protein-protein interactions in the intercellular junctions of the IMB and BBB to increase the porosity of paracellular pathways of the barriers. These modulator peptides have been shown to enhance brain delivery of small molecules and medium-sized peptides as well as a large protein such as 65 kDa albumin. In the future, this method has the potential to improve oral and brain delivery of therapeutic and diagnostic peptides and proteins.

Conjugates of Cell Adhesion Peptides for Therapeutics and Diagnostics Against Cancer and Autoimmune Diseases.

Overexpressed cell-surface receptors are hallmarks of many disease states and are often used as markers for targeting diseased cells over healthy counterparts. Cell adhesion peptides, which are often derived from interacting regions of these receptor-ligand proteins, mimic surfaces of intact proteins and, thus, have been studied as targeting agents for various payloads to certain cell targets for cancers and autoimmune diseases. Because many cytotoxic agents in the free form are often harmful to healthy cells, the use of cell adhesion peptides in targeting their delivery to diseased cells has been studied to potentially reduce required effective doses and associated harmful side-effects. In this review, multiple cell adhesion peptides from extracellular matrix and ICAM proteins were used to selectively direct drug payloads, signal-inhibitor peptides, and diagnostic molecules, to diseased cells over normal counterparts. RGD constructs have been used to improve the selectivity and efficacy of diagnostic and drug-peptide conjugates against cancer cells. From this precedent, novel conjugates of antigenic and cell adhesion peptides, called Bifunctional Peptide Inhibitors (BPIs), have been designed to selectively regulate immune cells and suppress harmful inflammatory responses in autoimmune diseases. Similar peptide conjugations with imaging agents have delivered promising diagnostic methods in animal models of rheumatoid arthritis. BPIs have also been shown to generate immune tolerance and suppress autoimmune diseases in animal models of type-1 diabetes, rheumatoid arthritis, and multiple sclerosis. Collectively, these studies show the potential of cell adhesion peptides in improving the delivery of drugs and diagnostic agents to diseased cells in clinical settings.

Membrane inlet for mass spectrometric measurement of catalysis by enzymatic decarboxylases.

Membrane inlet mass spectrometry (MIMS) uses diffusion across a permeable membrane to detect in solution uncharged molecules of small molecular weight. We point out here the application of MIMS to determine catalytic properties of decarboxylases using as an example catalysis by oxalate decarboxylase (OxDC) from Bacillus subtilis. The decarboxylase activity generates carbon dioxide and formate from the nonoxidative reaction but is accompanied by a concomitant oxidase activity that consumes oxalate and oxygen and generates CO(2) and hydrogen peroxide. The application of MIMS in measuring catalysis by OxDC involves the real-time and continuous detection of oxygen and product CO(2) from the ion currents of their respective mass peaks. Steady-state catalytic constants for the decarboxylase activity obtained by measuring product CO(2) using MIMS are comparable to those acquired by the traditional endpoint assay based on the coupled reaction with formate dehydrogenase, and measuring consumption of O(2) using MIMS also estimates the oxidase activity. The use of isotope-labeled substrate ((13)C(2)-enriched oxalate) in MIMS provides a method to characterize the catalytic reaction in cell suspensions by detecting the mass peak for product (13)CO(2) (m/z 45), avoiding inaccuracies due to endogenous (12)CO(2).

Nitric oxide reversibly inhibits Bacillus subtilis oxalate decarboxylase.

Membrane inlet mass spectrometry (MIMS) has been employed to assay the catalytic activity of oxalate decarboxylase (OxDC), allowing us to demonstrate that nitric oxide (NO) reversibly inhibits the enzyme under dioxygen-depleted conditions. X-band EPR measurements do not provide any direct evidence for the interaction of NO with either of the Mn(II) centers in OxDC raising the possibility that there is a separate dioxygen-binding pocket in the enzyme.