Delivery of Protein and Peptide Drugs in Cancer

The following sections are included:

• Contents

• Contributors

To effectively treat cancer, we have to be able to selectively attack the tumor and individual cancer cells, while effectively protect normal tissue from possible toxicity and other side effects of anti cancer drugs. It is not an easy task, since systemically administered drugs may rapidly metabolize in the blood or be cleared from the body before reaching the tumor cells. In addition, on its way to the target, it has to overcome multiple physiological barriers, such as irregularities in the tumor blood flow, the high interstitial pressure, and the absence of a lymphatic drainage in tumors (Campbell, Chapter 2). Many drugs have also been found to perform their action inside the cells which requires their intracellular delivery through low permeable cell membranes. All these obstacles are especially pronounced in the case of protein and peptide drugs, whose successful application needs effective means of drug delivery into tumors. This book will consider various problems associated with tumor delivery of protein and peptide drugs and some of the current strategies to solve these problems.

The following sections are included:

• Introduction

• Blood Vessels: Modulation of Normal and Pathologic Function
  • General features of blood vessels in biological systems
  • The tumor vasculature: Specialized features and angiogenesis

• Transport of Peptide and Protein Molecules across Tumor Capillary Networks
  • Barriers limiting drug transport
  • The interstitial matrix: MMPs, collagen, invasion and metastasis
  • Overcoming barriers: Exploiting tumor physiology for therapeutic gain

• Concluding Remarks

• References

The following sections are included:

• Introduction

• Theory Underlying the EPR Effect

• Anatomical and Pathophysiological Abnormalities Related to EPR Effect

• Augmentation of the EPR Effect in Solid Tumor by Influencing Vascular Mediators

• Relation of Fluid Dynamics in Cancer Tissue to the EPR Effect

• Implications for Delivery of Drugs to Tumors

• Conclusion

• References

The term PEGylation defines the modification of a protein, peptide or non-peptide molecule by the chemical linking of one or more poly(ethylene glycol) (PEG) chains. PEG is the polymer of choice for drug conjugation, being non-toxic, non-immunogenic, non-antigenic and highly soluble in water; all properties that are conferred on the conjugated drugs. The PEG derivatives have the following advantages: (1) a prolonged residence in the body due to a reduced kidney clearance, as a consequence of the increased molecular weight; (2) a decreased degradation by proteolytic enzymes, thanks to the shielding effect of the polymer chains; (3) reduction or elimination of protein immunogenicity.

Thanks to these favorable properties, PEGylation now plays an important role in enhancing the use and potential of peptides and proteins as therapeutic agents that, in the native form, often encounter severe limitations in their use. So far, several chemical strategies have been developed to perform the linking of the polymer to a biologically active molecule, and a wide range of methods of which a selection is reported here. A clear demonstration of the potential of PEGylation is the numbers of products that have already reached the market or that will soon be available. In addition to a review of PEGylation methods, this chapter also describes recent achievements in PEGylation of protein and peptide drugs.

The following sections are included:

• Introduction

• Enzymes and PEG-Enzymes in Cancer Therapy
  • Metabolite depleting enzymes
    • PEG-asparaginase (PEG-ASNase)
    • PEG-methioninase (PEG-METase)
    • PEG-arginine deiminase (PEG-ADI)
    • PEG-uricase (PEG-UK)

• PEG-Antibodies

• Conclusions

• References

The half-lives of proteins in vivo can be increased by covalently attaching the polymer polyethylene glycol (PEG) to them. This can significantly decrease their rate of clearance from plasma. The clearance rate is inversely proportional to the total molecular weight of the construct. This approach has been used to design cytokine constructs that can be administered once a week, rather than on a daily or every other day schedule. Two cytokines for which this approach appears to be successful are PEG-interferon-α_2a (PEG-IFN-α_2a; Pegasys™) and PEG-granulocyte colony-stimulating factor (PEG-G-CSF; SD/01). Both use large MW PEG (20–40 kDa) to give sufficiently long duration in vivo. With PEG-G-CSF conjugates, the in vivo efficacy is directly proportional to molecular weight, whereas the in vitro activity is inversely proportional, suggestion that overall duration of contact is more important than affinity of the interaction. Conjugates of a number of other cytokines have been prepared, but until recently, few have used the large molecular weight polymers. As this approach is likely to be taken in the future to make new PEG-cytokine constructs, thorough in vivo pharmacokinetic studies will be essential for their development and clinical use.

The following sections are included:

• Role of Proteins in Tumor Development
  • Proto-oncogenes
  • Tumor suppressor genes
  • Pro-angiogenic proteins/pro-metastatic proteins
  • Proteins involved in drug resistance

• RNA Interference
  • Mechanism of RNA interference
  • Structural requirements for siRNA

• Approaches for siRNA Delivery

• siRNA Delivery for Cancer Therapy
  • siRNA to silence oncogenes
  • siRNA to silence mutated tumor suppressor genes
  • siRNA to silence pro-angiogenic/pro-metastatic genes
  • siRNA to silence genes that increase tumor drug resistance

• Concluding Remarks

• References

The following sections are included:

• Proteins and Peptides as Cancer Therapeutics

• Stability, Delivery and Distribution Issues for Peptide and Protein Drugs

• Possible Solutions. Liposomes as Pharmaceutical Carriers

• Liposomal Proteins and Peptides for Cancer Treatment

• Tumor Cell Targeting Using Specific Ligand-Bearing Liposomes

• Transmembrane Delivery of Protein and Peptide Drugs into Tumor Cells

• References

The membrane-bound folate receptor (FR) is overexpressed on a wide range of human cancers, such as those originating in ovary, lung, breast, endometrium, kidney and brain. The vitamin folic acid (FA) is a high affinity ligand of the FR, which retains its receptor binding properties when conjugated to other molecules. Consequently, folate mediated targeting has been successfully applied for the delivery of a wide variety of payloads, including protein toxins, immunotherapeutic agents, adenoviral vectors and radio-imaging agents to FR-positive cancers. These folate-bearing delivery systems have exhibited major enhancements in cancer cell specificity and selectivity over their non-targeted counterparts. Hence, it is hopeful that this targeting strategy should lead to improvements in the safety and efficacy of anti cancer agents, resulting in an increased therapeutic advantage in the clinic. This chapter reviews the growing literature on folate-targeted drugs, with specific emphasis on the delivery of proteins and peptides to cancers.

The following sections are included:

• Introduction

• Transferrin (Tf)
  • Structure
  • Biological function

• Transferrin-Receptors (TfR)
  • Structure
  • Clathrin-mediated endocytosis
  • Regulation of expression
  • Tissue distribution

• In vivo Application of Proteins Drugs, General Considerations

• Generation of Conjugates
  • Chemical coupling
  • Recombinant production of conjugates

• Tf and AntiTFR Targeted Toxins
  • Ribosomal inhibitors
  • Ribonucleases
  • Diphtheria toxin and Pseudomonas exotoxin

• Conclusions

• References

Progress in the treatment of cancer has been slow over the past few decades, with the development of new therapeutics being hampered by the poor target specificity of small molecules and the impermeability of the plasma membrane to any molecule greater than 500 Da. Adenoviral, retroviral, and lipid-based delivery systems have all met with only limited clinical success secondary to problems with immunogenicity, prolonged gene expression, and significant long-term, occasionally lethal, side effects. The recent identification of a particular group of proteins with enhanced ability to cross the plasma membrane has led to the discovery of a class of protein domains with cell penetrating properties. The fusion of these protein transduction domain sequences with large, biologically active, macromolecular cargo is sufficient to induce their rapid transduction into virtually every type of mammalian cell. Moreover, this nascent technology appears to circumvent many of the problems associated with DNA and viral-based methods. Protein transduction-mediated delivery of previously unavailable large molecules, peptides and proteins, offers a unique opportunity to specifically modulate tumor cell biology in vivo.

The following sections are included:

• Introduction

• Natural Cyokines

• Cryptic Protein Fragments

• Antibodies
  • Monoclonal antibodies (mAb)
  • Recombinant antibodies

• Soluble VEGF-Rs (Traps)

• Small Peptides
  • Vascular targeting peptides
  • Function-blocking peptides

• Conclusions and Perspectives

• Acknowledgments

• References

The gastrointestinal lymphatic system is a specific transport pathway through which dietary lipids, fat-soluble vitamins, and water-insoluble peptide type molecules can gain access to the systemic circulation. Drugs transported by way of the gastrointestinal lymphatic system bypass the liver and avoid potential hepatic first-pass metabolism. Lymphatic delivery of immunomodulatory and protein and peptide drugs used in the treatment of cancer cell metastases presents an opportunity to maximize therapeutic benefit while minimizing general systemic drug exposure. Furthermore, lymphatic drug transport may promote drug incorporation into the body's lipid-handling system, thus offering the potential to manipulate drug distribution and residence time within the body and reduce the invasiveness of treatment for the patient. This chapter will discuss the potential utilization of lymphatic transport in enhancing the delivery of drugs, including proteins and peptides, to metastatic tumors.

Primary and metastatic brain tumors remain a significant health problem and affect between 130 000 and 185 000 people each year in the USA. Conventional treatment with surgical resection, irradiation, and cytotoxic chemotherapy continues to be inadequate, with a median survival of approximately 1 year for most patients. Advances in molecular pharmacology and the biology of the blood–brain barrier (BBB) are improving the ability of researchers to target therapeutic peptides, proteins, and small molecules to the brain and brain tumors. Simple methods to improve BBB penetration and brain delivery include chemical modification, glycosylation, and pegylation. Drug delivery vehicles such as nanoparticles and liposomes are also under study. These vehicles can be modified by pegylation or conjugation with a targeting vector to improve delivery across the BBB. Targeting vectors include natural ligands (e.g. epidermal growth factor) or monoclonal antibodies to receptors (e.g. transferrin, insulin, interleukin). Other vector-mediated delivery approaches involve the conjugation of a therapeutic peptide or protein with a targeting molecule that can induce transcytosis across BBB endothelial cells. The most commonly used vectors are peptidomimetic antibodies to endothelial receptors, such as the transferrin and insulin receptors.

The following sections are included:

• Cancer Gene Therapy
  • Identifying the target gene/pathway for cancer therapy
  • Selection of gene therapy vector

• Protein-Based Cancer Gene Therapy
  • Growth inhibition and apoptosis
    • Toxic proteins
    • Tumor suppressor proteins
    • Antiangiogenic proteins
  • Enhancement of the immune response

• Peptide-Based Cancer Gene Therapy
  • Tumor targeting and gene transduction

• Conclusions

• References

The following sections are included:

• Index