The present invention relates to nanostructured conjugates, more specifically to nanostructured fusion proteins suitable for the selective delivery of their conjugated therapeutic agents to specific cell and tissue types. It also relates to nanoparticles comprising such nanostructured proteins and the therapeutic uses thereof.

Various biodegradable polyglutamate-amino acids comprising recurring units of the general formulae (I) and (II) are prepared. Such polymers are useful for variety of drug, biomolecule and imaging agent delivery applications.

The invention provides compositions useful as molecular probes. In particular, the invention provides activatable cell penetrating peptides comprising a fluorescence donor and a fluorescence acceptor. Exemplary fluorescence donors and fluorescence acceptors include compounds derived from cyanine. Also provided are ratiometric, multispectral, and excitation lifetime imaging methods for detecting the molecular probes provided herein.

Various biodegradable polyglutamate-amino acids comprising recurring units of the general formulae (I) and (II) are prepared. Such polymers are useful for variety of drug, biomolecule and imaging agent delivery applications.

Polyglutamates are well known to be highly biocompatible, biodegradable and multifunctional polymers, which have been already used as building blocks in polymer drug conjugates and polymeric micelles. Those systems have been applied to various medical applications ranging from therapy to molecular imaging. Furthermore a polyglutamic acid (PGA) paclitaxel conjugate has already entered clinical studies (Opaxio PGA-PTX conjugate currently in phase III of Clinical trials). In this context, a synthetic pathway to a plethora functional polyglutamates (homopolymers, block-co-polymers, tribocks) with well-defined structure, adjustable molecular weight (Mw) and low dispersity (D=Mw/Mn<1.2) applying the ring opening polymerization (ROP) of N-carboxyanhydrides (NCA) has been developed. Additionally, the acid moieties of the polyglutamates can be activated with 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium (DMTMM) and various functionalities can be easily introduced by post-polymerization modification yielding a set orthogonal reactive attachment sides. The reactive moieties, such as azides, maleimides, thiols, akynes (linear or cyclic) offer the opportunity of specific conjugation of the drugs, targeting moieties or markers. Besides introducing reactive groups the functionalization strategy was also used for PEGylation of PGA reducing charge induced interactions and therefore pharmacological properties, such as blood circulation time may be adjusted. In summary, a tool kit of various polyglutamates has been developed enabling the synthesis of a variety of polymer drug conjugates or polymer based imaging agents. The functional polymeric precursors developed will allow us to functionalize and therefore adjust the polymer properties to a desired application.

A generic structure for the peptides of the present invention includes A-X-B-C, where C is a cargo moiety, the B portion includes basic amino acids, X is a cleavable linker sequence, and the A portion includes acidic amino acids. The intact structure is not significantly taken up by cells; however, upon extracellular cleavage of X, the B-C portion is taken up, delivering the cargo to targeted cells. Cargo may be, for example, a contrast agent for diagnostic imaging, a chemotherapeutic drug, or a radiation-sensitizer for therapy. X may be cleaved extracellularly or intracellularly. The molecules of the present invention may be linear, cyclic, branched, or have a mixed structure.

The present invention relates to nanostructured conjugates, more specifically to nanostructured fusion proteins suitable for the selective delivery of their conjugated therapeutic agents to specific cell and tissue types. It also relates to nanoparticles comprising such nanostructured proteins and the therapeutic uses thereof.

An embodiment of the present invention has an object of providing single-macromolecule particles each having a more accurately controlled hydrodynamic diameter, an active molecular complex, a method for producing the single-macromolecule particles, and a method for imaging biological tissue. In order to attain the object, provided are single-macromolecule particles characterized by each being formed of a single macromolecule and by having a molecular weight distribution M.sub.w/M.sub.n of not more than 1.5. This makes it possible to obtain particles each having an accurately controlled hydrodynamic diameter. The single macromolecule is preferably a structure consisting of a single hydrophilic macromolecule A or a structure in which one or more side chains are bonded to a main chain, the main chain being the single hydrophilic macromolecule A, each of the one or more side chains being a hydrophilic macromolecule B.