Intracellular protein delivery is offering numerous possibilities in research and in

therapy. Aside gene therapy, protein delivery into living cells is one of the most

promising tools for the treatment of various so far immedicable diseases including

cancer. To develop a practicable protein delivery platform, a test system which allows

easy control of successful intracellular delivery is needed. Therefore a test system

based on two model proteins was established. A nuclear localization signal tagged

EGFP molecule is enabling fast control of cellular uptake and endosomal release.

The second model protein ß-galactosidase is evidencing that protein conformation is

not irreversible disturbed by modification with the carrier molecules. Protein

transduction technology is opening the door for a promising alternative to gene

therapy, as it is lacking of the potential malignant side effects of gene therapy. The

most limiting step in the development of a therapeutic drug remains the delivery

process. In the last decade, many techniques to deliver proteins into living cells were

developed. Although great efforts were made, so far no all-purpose technique is

available that addresses all critical steps, like efficient uptake, endo-lysosomal

escape, low toxicity, while maintaining enzymatic activity. Each method has got its

limitation, for example cell type dependence. Among the so far used carriers, the

most effective ones are cationic polymers like polyethylenimine. These carriers are

lacking of precise structure and often show high toxicity, dependent on the molecular

weight of the used polymer. In this thesis the properties of the three arm cationic

oligomer 386, which was previously designed for siRNA delivery was investigated in

regard of being applicable as a transduction carrier for protein delivery. This carrier

molecule, in contrast to other cationic polymers used for protein delivery, is of precise

structure, of low molecular weight and potentially degradable by proteases. The

transduction oligomer was covalently bound to the protein by a bioreversible bond.

Our results reveal that covalent coupling of the structure defined cationic oligomer

386 to a protein leads to a high efficient, serum insensitive and low toxic alternative to

established protein transduction technologies. For a general all-purpose delivery

system covalent coupling of the carrier to the cargo protein is indispensable. Protein

delivery requires special properties to the linker molecule. Therefore in this work a

new pH sensitive linker was developed which combines the advantages of click

reactions with the implementation of a traceless cleavable bond between two conjugated molecules. Three different click chemistries were performed which all are

compatible with the acid labile properties. A traceless cleavage may be a particularly

important feature in protein transduction strategies, to maintain full bioactivity of

enzymes and other proteins. The current example of 386 carrier-mediated cytosolic

delivery and subsequent nuclear import of released nls-EGFP demonstrates the

advantage of the traceless linker. To demonstrate that the modification does not

irreversibly affect structure and biological activity of proteins, 386-AzMMMan-ßgalactosidase

was delivered as a model enzyme. It exhibited cytosolic activity in the

transduced cells far higher than without shuttle. Aside from these encouraging

options for protein delivery and modification, the linker might have broader use in the

design of novel programmed, acid labile and biodegradable drug delivery systems.

Targeted therapeutics could, after delivery into acidic tumor areas or upon cellular

uptake into endosomes, be dismantled from their outer shell including targeting

ligands. Besides drug delivery, the linker may also be of interest for other

applications, such as reversible labeling of various biological and also chemical

molecules. The developed linking strategy