A potential plasmid-based therapy for cancer and viral diseases?

Protein transduction domains (PTD) are cell permeable protein components or peptides that allow the ferrying of larger proteins across the plasma membrane. The TAT protein of HIV has a PTD consisting of a 9 amino acid sequence – RKKRRQRRR (amino acid 49 – 57) (Beerens et al, 2006). Subsequently, investigators have shown that the peptide sequence of the TAT PTD fused to proteins mediated their delivery into the cells (Xia et al, 2001). Usually, such amino acids are cationic in nature. More potent PTDs have also been synthesized from peptides, e.g. PTD-4, PTD-5, MST-1, L-R9 and peptide-2.

Thus, PTD fusion proteins have therapeutic potential and they can be targeted towards cancer cells, viral infected cells or normal cells (for additional functions). For example, investigators have shown that restoration of RhoB (Rho family of GTPases) expression led to ovarian tumor regression (Couderc et al, 2008). Thus, a RhoB-PTD fusion protein may have therapeutic prospects. A phase I gene therapy trial by Yoo and colleagues involved the introduction of the E1A gene-liposome complex into patients with recurrent breast and head and neck cancers (Yoo et al, 2001). The E1A protein is involved in transcriptional repression, cellular differentiation and induction of apoptosis in cancer cells. Thus, E1A-PTD fusion proteins can be another therapeutic prospect in the treatment of cancers.

Zinc finger proteins (ZFP) have the potential to be the mainstay of cancer-based or viral-based therapies. They can be artificially engineered to bind a particular DNA sequence and activate or repress selected genes. They have a DNA-binding domain and either an activating or repressor domain (Gommans et al, 2005). Investigators have successfully engineered zinc finger transcriptional factors which repress the HIV Long-terminal repeats (LTR) promoters and inhibit HIV replication (Reynolds et al, 2003). Thus, PTD-ZFP fusion proteins form another attractive therapeutic prospect, in which such fusion proteins can be disseminated to cells to achieve transcriptional control.

The protein of therapeutic interest can be fused to the PTD, and the gene encoding the fusion protein can be placed into a plasmid. The N -terminus of the protein contains the secretory signal while the C-terminus contains the PTD domain.* Such plasmids can be introduced to white blood cells, which circulate throughout the body and are capable of homing in on cancer tissues or virus-infected cells. Plasmid delivery may be facilitated by PTD peptides, which are oligomers of the arginine-rich motif of the HIV TAT protein (Rudolph et al, 2003). The white blood cells will then produce the PTD-fusion proteins, which can be secreted and disseminated to target cells of interest, e.g. cancer cells, virus-infected cells, cells implicated in other diseases, or normal cells (to acquire additional functions).

*Edits: Added N-terminus to contain secretory signals and C-terminus to contain PTD domain

References
1) Beerens AMJ, Al Hadithy AFY, Rots MG, Haisma HJ. Protein transduction domains and their utility in gene therapy. At: http://dissertations.ub.rug.nl/FILES/faculties/science/2006/a.m.j.beerens/02_c2.pdf

2) Xia H, Mao Q, Davidson BL. The HIV Tat protein transduction domain improves the biodistribution of B-glucuronidase expressed from recombinant viral vectors. Nature Biotechnology 19, 640 – 644 (2001)
doi:10.1038/90242

3) Couderc B, Pradines A, Rafii A, Golzio M, Deviers A, Allal C, Berg D, Penary M, Teissie J, Favre G. In vivo restoration of RhoB expression leads to ovarian tumor regression. Cancer Gene Ther. 2008 Jul;15(7):456-64.

4) Yoo GH, Hung MC, Lopez-Berestein G, LaFollette S, Ensley JF, Carey M, Batson E, Reynolds TC, Murray JL. Phase I trial of intratumoral liposome E1A gene therapy in patients with recurrent breast and head and neck cancer. Clin Cancer Res. 2001 May;7(5):1237-45.

5) Gommans WM, Haisma HJ, Rots MG. Engineering zinc finger protein transcription factors: the therapeutic relevance of switching endogenous gene expression on or off at command. J Mol Biol. 2005 Dec 2;354(3):507-19.

6) Reynolds L, Ullman C, Moore M, Isalan M, West MJ, Clapham P, Klug A, Choo Y. Repression of the HIV-1 5′ LTR promoter and inhibition of HIV-1 replication by using engineered zinc-finger transcription factors.
Proc Natl Acad Sci U S A. 2003 Feb 18;100(4):1615-20.

7) Rudolph C, Plank C, Lausier J, Schillinger U, Müller RH, Rosenecker J. Oligomers of the arginine-rich motif of the HIV-1 TAT protein are capable of transferring plasmid DNA into cells. J Biol Chem. 2003 Mar 28;278(13):11411-8.