Novel immunosupressant circular peptides from plants

Natural peptides of great number and diversity occur in all organisms from microbes to plants to man and exhibit biological activity against unrelated targets, thus providing researchers with starting points for drug discovery. Natural peptide libraries offer a unique way to screen the diversity of interaction of peptides with soluble cytosolic proteins and transmembrane receptors and are thus available for therapeutic applications. Cyclotides are an abundant and diverse group of ribosomal-synthesized plant peptides containing a cyclic cystine- knotted structure that confers them with remarkable stability. They are explored for their distribution in plants, although little is known about the individual peptide content of a single species. Therefore we propose to chemically analyse the crude extracts of several coffee-family plants from the tribe Psychotriae using a rapid peptidomics workflow utilising nano-LC-MS, peptide reconstruct with database identification and nESI-MS/MS as well as MALDI-TOF/TOF automated sequence analysis to determine their cyclotide content. Biologically, cyclotides are mainly explored for applications in agriculture and drug design; based on their predicted numbers, diversity and plasticity of the cyclotide scaffold, which is amenable to a wide range of amino acid substitutions, they are considered a combinatorial peptide library that is available for screening of therapeutic effects. Our main goal is to analyse their growth-inhibiting effects on primary cells of the human immune system (activated T- lymphocytes) using biological and immunological endpoints in cell-based test systems. Preliminary data promise an effect in a defined concentration range that is not due to cytotoxic effects, so that these results open new avenues to utilize native and synthetically-optimized plant cyclotides (i) to study the mechanism of these effects and (ii) to test their therapeutic potential in vivo in model systems of immune-related disorders and as immunosuppressant peptides.

This project has led to a crucial development in the treatment of multiple sclerosis (MS). We demonstrated in an animal model that, following treatment with a specially synthesized plant peptide (cyclotide), there is no further progression of the usual clinical signs of multiple sclerosis. The one-off oral administration of the active agent brought about a great improvement in symptoms. There were no further attacks of the disease. This could slow down the course of the disease in general.MS is a chronic inflammatory autoimmune disease of the central nervous system, in which the insulating myelin sheaths around the nerve fibres are destroyed. The disease progresses in the form of attacks or episodes and is currently incurable. An episode is defined by the occurrence of new symptoms or flare-ups of pre-existing ones. Each episode is associated with immediate or deferred deterioration in the patient's condition. The mechanism of inflammation in the nervous system is partially understood. Based on this knowledge, there are treatments to slow down progression of the disease but these have significant side effects, particularly in long-term therapy. It is estimated that around 2.5 million people are affected by MS worldwide, around 8,000 of these in Austria.The discovery now offers hope that the disease can be halted at a very early stage or, at the very least, its progression greatly retarded. As soon as functional neurological problems occur and an MRI (Magnetic Resonance Imaging) scan identifies early pathological changes in the central nervous system, the drug can be given as a basic therapy. In an animal model for MS, symptoms were considerably reduced by the oral administration of cyclotides. It is therefore possible that the drug could extend the interval between episodes or possibly prevent an onset of the disease.We have demonstrated that cyclotides are macrocyclic plant peptides that can be isolated from all the main plant families (e.g. coffee plants, cucurbits or even grasses and plants of the nightshade family) and therefore represent a large and wide-ranging group of natural substances. Due to the extraordinary stability of cyclotides, a drug obtained from them can be taken orally. Many of the current MS treatments in common use have to be administered intravenously.In addition we elucidated the mode of action of cyclotides in that they suppress the messenger substance interleukin-2 and hence the division of T cells, which act as "killer" or "helper" cells in the human immune system response. Hence, cyclotides could also possibly be used to treat other diseases characterized by an overactive, misdirected immune response, such as rheumatoid arthritis, for example.