The TAT Protein Transduction Domain as an Intra-Articular Drug Delivery Technology

Objective Intra-articular drug delivery holds great promise for the treatment of joint diseases such as osteoarthritis. The objective of this study was to evaluate the TAT peptide transduction domain (TAT-PTD) as a potential intra-articular drug delivery technology for synovial joints. Design Experiments examined the ability of TAT conjugates to associate with primary chondrocytes and alter cellular function both in vitro and in vivo. Further experiments examined the ability of the TAT-PTD to bind to human osteoarthritic cartilage. Results The results show that the TAT-PTD associates with chondrocytes, is capable of delivering siRNA for chondrocyte gene knockdown, and that the recombinant enzyme TAT-Cre is capable of inducing in vivo genetic recombination within the knee joint in a reporter mouse model. Last, binding studies show that osteoarthritic cartilage preferentially uptakes the TAT-PTD from solution. Conclusions The results suggest that the TAT-PTD is a promising delivery strategy for intra-articular therapeutics.

The TAT Protein Transduction Domain as an Intra-articular Drug-Delivery Technology

ObjectiveIntra-articular drug delivery holds great promise for the treatment of joint diseases such as osteoarthritis. The objective of this study was to evaluate the TAT peptide transduction domain (TAT-PTD) as a potential intra-articular drug delivery technology for synovial joints.DesignExperiments examined the ability of TAT conjugates to associate with primary chondrocytes and alter cellular function both in vitro and in vivo. Further experiments examined the ability of the TAT-PTD to bind to human osteoarthritic cartilage.ResultsThe results show that the TAT-PTD associates with chondrocytes, is capable of delivering siRNA for chondrocyte gene knockdown, and that the recombinant enzyme TAT-Cre is capable of inducing in vivo genetic recombination within the knee joint in a reporter mouse model. Lastly, binding studies show that osteoarthritic cartilage preferentially uptakes the TAT-PTD from solution.ConclusionsThe results suggest that the TAT-PTD is a promising delivery strategy for intra-articular therapeutics.

Therapeutic effects of cell-permeant peptides that activate G proteins downstream of growth factors

In eukaryotes, receptor tyrosine kinases (RTKs) and trimeric G proteins are two major signaling hubs. Signal transduction via trimeric G proteins has long been believed to be triggered exclusively by G protein-coupled receptors (GPCRs). This paradigm has recently been challenged by several studies on a multimodular signal transducer, Gα-Interacting Vesicle associated protein (GIV/Girdin). We recently demonstrated that GIV’s C terminus (CT) serves as a platform for dynamic association of ligand-activated RTKs with Gαi, and for noncanonical transactivation of G proteins. However, exogenous manipulation of this platform has remained beyond reach. Here we developed cell-permeable GIV-CT peptides by fusing a TAT-peptide transduction domain (TAT-PTD) to the minimal modular elements of GIV that are necessary and sufficient for activation of Gi downstream of RTKs, and used them to engineer signaling networks and alter cell behavior. In the presence of an intact GEF motif, TAT-GIV-CT peptides enhanced diverse processes in which GIV’s GEF function has previously been implicated, e.g., 2D cell migration after scratch-wounding, invasion of cancer cells, and finally, myofibroblast activation and collagen production. Furthermore, topical application of TAT-GIV-CT peptides enhanced the complex, multireceptor-driven process of wound repair in mice in a GEF-dependent manner. Thus, TAT-GIV peptides provide a novel and versatile tool to manipulate Gαi activation downstream of growth factors in a diverse array of pathophysiologic conditions.

Targeting the Oligomerization of BCR/ABL by Membrane Permeable Competitive Peptides Inhibits the Proliferation of Philadelphia Chromosome Positive Leukemic Cells

The BCR/ABL fusion protein is the hallmark of Philadelphia Chromosome positive (Ph+) leukemia. The constitutive activation of the ABL-kinase in BCR/ABL cells induces the leukemic phenotype. Targeted inhibition of BCR/ABL by small molecule inhibitors reverses the transformation potential of BCR/ABL. Recently, we definitively proved that targeting the tetramerization of BCR/ABL mediated by the N-terminal coiled-coil domain (CC) using competitive peptides, representing the helix-2 of the CC, represents a valid therapeutic approach for treating Ph+ leukemia. To further develop competitive peptides for targeting BCR/ABL, we created a membrane permeable helix-2 peptide (MPH-2) by fusing the helix-2 peptide with a peptide transduction tag. In this study, we report that the MPH-2: (i) interacted with BCR/ABL in vivo; (ii) efficiently inhibited the autophosphorylation of BCR/ABL; (iii) suppressed the growth and viability of Ph+ leukemic cells; and (iv) was efficiently transduced into mononuclear cells (MNC) in an in vivo mouse model. This study provides the first evidence that an efficient peptide transduction system facilitates the employment of competitive peptides to target the oligomerization interface of BCR/ABL in vivo.