CELL-PENETRATING PEPTIDE FUSION PROTEINS - Fusion Protein Technologies for Biopharmaceuticals: Applications and Challenges - page 383

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CELL-PENETRATING PEPTIDE FUSION PROTEINS

A NDRES M UNOZ -A LARCON ,H ENRIK H ELMFORS ,K RISTIN K ARLSSON , AND U LO L ANGEL

Department of Neurochemistry, Stockholm University, Stockholm, Sweden

26.1 Introduction
26.2 Typical applications and indications
26.3 Technological aspects
26.4 Successes and failures in preclinical and clinical research
26.5 Alternatives/variants of this approach
26.6 Conclusions and future perspectives
Acknowledgments
References

of overcoming the challenge posed by the cellular mem-
branes that otherwise prohibit intracellular delivery.
In 1994, the first member of a new class of peptides—
cell-penetrating peptides (CPPs)—with membrane penetrat-
ingabilitywas discovered[1]. CPPs constitute aheterogeneous
group of relatively short cationic and/or amphipathic peptides,
up to 40 amino acids in length. They possess the ability to
gain entry to cells independent of membrane receptors [2] and
they exhibit no cell-type specificity [3]. Furthermore, they
possess a low cellular toxicity and the capacity to promote
intracellular delivery of covalently or noncovalently conju-
gated bioactive cargoes. Several CPPs have been shown to
enable delivery of biologically active proteins into cells when
synthesized as a recombinant fusion protein or covalently
conjugated to full-length proteins [4]. Subsequently, CPPs
have been employed as vectors for intracellular delivery of a
variety of molecules that have difficulties in entering cells by
themselves [5,6]. Although peptides derived from protein
interactionsitesgeneratemoreextensive contactswithproteins
than small hydrophobic molecules and have been successfully
delivered using CPPs, peptides are often only capable of
partially mimicking the activity of full-length proteins. As a
result, proteins, albeit harder to deliver, are inmanywaysmore
attractive as potential drugs. The concept of intracellular
delivery of CPP-fusion proteins with biological activity is
commonly referred to as protein transduction.

26.1 INTRODUCTION

Protein interactions regulate many of the fundamental cel-
lular processes such as gene expression and cellular signal-
ing. Consequently, molecules with protein modulating
properties provide an attractive source of potential thera-
peutic agents. Although small molecules remain the domi-
nating therapeutic paradigm, development of traditional
small molecule pharmaceuticals relies heavily on chance
discovery of molecules that possess both cell-permeable
properties and the ability to modulate biological processes.
Many such small molecules contain insufficient biochemical
information needed to obtain high specificity while avoiding
undesirable and possibly toxic side effects. On the other
hand, the structures of macromolecules, such as proteins and
peptides, are better able to contain the information required
to obtain therapeutic agents showing superior biological
activity, specificity, and low toxicity. However, full-length
proteins and larger peptides are difficult to produce and most
often inherently cell impermeable. Regardless, the promis-
ing aspects of macromolecular therapeutics have prompted
the development of a wide range of delivery vectors capable

26.2 TYPICAL APPLICATIONS
AND INDICATIONS

In order to address the challenges in chemotherapy, imaging,
diagnostics and mechanistic chemical biology novel strate-
gies, and agents that enable or enhance the passage of drugs