RGD Peptides and Monoclonal Antibodies, Antagonists of αv-Integrin, Enter the Cells by Independent Endocytic Pathways

Castel, Susanna; Pagan, Roser; Mitjans, Francesc; Piulats, Jaume; Goodman, Simon L.; Jonczyk, Alfred; Huber, Florian; Vilaró, Senén; Reina, Manuel

doi:10.1038/labinvest.3780375

Cited by 87 publications

(71 citation statements)

References 47 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The early internalization route of TEAM-VP in HUVEC cells is in the line with published reports on cycloRGDfK peptides, 27,28 thus indicating that a mitochondriotoxic part can be added without to put a damper on RGD binding and internalization through a V b 3 receptors. Distribution in lyso- somes has already been observed for other RGD-peptides like cycloRGDfK, 27,28 but in our study confocal microscopy clearly shows that TEAM-VP reaches mitochondria after its release from lysosomes without degradation. This is the first detailed study showing microscopic evidence that RGDcontaining peptide reaches its targeted intracellular compartment.…”

Section: Discussionsupporting

confidence: 74%

Targeted Vpr-derived peptides reach mitochondria to induce apoptosis of αVβ3-expressing endothelial cells

Borgne‐Sanchez¹,

Dupont²,

Langonné³

et al. 2006

Cell Death Differ

View full text Add to dashboard Cite

The HIV-1 encoded apoptogenic protein Vpr induces mitochondrial membrane permeabilization (MMP) via interactions with the voltage-dependent anion channel (VDAC) and the adenine nucleotide translocator (ANT). We have designed a peptide, TEAM-VP, composed of two functional domains, one a tumor blood vessel RGD-like 'homing' motif and the other an MMP-inducing sequence derived from Vpr. When added to isolated mitochondria, TEAM-VP interacts with ANT and VDAC, reduces oxygen consumption and overcomes Bcl-2 protection to cause inner and outer MMP. TEAM-VP specifically recognizes cell-surface expressed a V b 3 integrins, internalizes, temporarily localizes to lysosomes and progressively co-distributes with the mitochondrial compartment with no sign of lysosomal membrane permeabilization. Finally TEAM-VP reaches mitochondria of angiogenic endothelial cells to induce mitochondrial fission, dissipation of the mitochondrial transmembrane potential (DW m ), cytochrome c release and apoptosis hallmarks. Hence, this chimeric peptide constitutes the first example of a virus-derived mitochondriotoxic compound as a candidate to kill selectively tumor neo-endothelia. Within the integrin family, a V b 3 receptors are considered to be an important target for anticancer therapy being upregulated in certain invasive tumors.1 a V b 3 recognizes a specific consensus sequence, an RGD motif, which is present in a number of extracellular matrix proteins like vitronectin, fibronectin, fibrinogen and thrombospondin.2 RGD-containing peptides have been shown to reduce angiogenesis induced by cytokines and to mediate apoptosis in endothelial cells.3 It appears that RGD peptides that are constrained in a cyclic conformation, show an increased affinity for binding to integrins. 4 One example is the cyclic RGD-4C peptide, a potent binder of a 5 b 1 , a V b 3 and a V b 5 used to selectively deliver doxorubicin to tumor blood vessels 5 and to internalize the amphipatic peptide (KLAKLAK) 2 in endothelial cells resulting in cellular apoptosis.

show abstract

Section: Discussionsupporting

confidence: 74%

Targeted Vpr-derived peptides reach mitochondria to induce apoptosis of αVβ3-expressing endothelial cells

Borgne‐Sanchez¹,

Dupont²,

Langonné³

et al. 2006

Cell Death Differ

View full text Add to dashboard Cite

show abstract

“…51), there have been only a few reports in cardiomyocytes: fluid phase endocytosis of solutes (52), phagocytosis of yeast cells (53) and Trypanosoma cruzi (54), and clathrin-mediated endocytosis of the fluorescent marker lucifer yellow (55). In addition, there has been no report on integrin-mediated endocytosis in adult cardiomyocytes despite a few reports in other cell types: endocytosis of synthetic peptide with RGD motifs by M21 melanoma cells (39) and uptake of ECM protein-bound microbeads by retinal pigment epithelial cells that implicates integrin ␣ 5 ␤ 1 in ECM remodeling (28). In this context, our studies demonstrate an intracellular accumulation of Alexa Fluor 594-labeled RGD peptide in a time-dependent manner in cardiomyocytes.…”

Section: Discussionmentioning

confidence: 99%

“…Activation-Previous studies demonstrate that when peptides bearing an RGD motif are added to cells, they undergo internalization (39) and that the FIG. 5.…”

Section: Rgd Undergoes Endocytosis and Inhibitors Of Endocytosis Blocmentioning

confidence: 99%

RGD-containing Peptides Activate S6K1 through β3 Integrin in Adult Cardiac Muscle Cells

Balasubramanian

Kuppuswamy

2003

Journal of Biological Chemistry

View full text Add to dashboard Cite

The enzyme p70S6 kinase (S6K1) is critical for cell growth, and we have reported its activation during cardiac hypertrophy. Because cardiac hypertrophy also involves integrin activation, we analyzed whether integrins could contribute to S6K1 activation. Using adult feline cardiomyocytes, here we report that integrin-interacting Arg-Gly-Asp (RGD) peptides activate S6K1 as observed by band shifting, kinase activity and phosphorylation at Thr-389 and Thr-421/Ser-424 of S6K1, and S6 protein phosphorylation. Perturbation of specific integrin function with blocking antibodies and by overexpressing the ␤ 1A cytoplasmic tail revealed that ␤ 3 but not ␤ 1 integrin mediates the RGD-induced S6K1 activation. This activation is focal adhesion complex-independent and is accompanied by the activation of extracellular signal-regulated kinases 1/2 (ERK) and mammalian target of rapamycin (mTOR). Studies using specific inhibitors and dominant negative c-Raf expression in cardiomyocytes indicate that the S6K1 activation involves mTOR, MEK/ERK, and phosphatidylinositol 3-kinase pathways and is independent of protein kinase C and c-Raf. Finally, addition of fluorescent-labeled RGD peptide to cardiomyocytes exhibits its internalization and localization to the endocytic vesicles, and pretreatment of cardiomyocytes with endocytic inhibitors reduced the S6K1 activation. These data suggest that RGD interaction with ␤ 3 integrin and its subsequent endocytosis trigger specific signaling pathway(s) for S6K1 activation in cardiomyocytes and that this process may contribute to hypertrophic growth and remodeling of myocardium.

show abstract

“…Examples of ligands that are effectively internalized include viruses (19) and ␣ v -integrin-blocking mAb. In contrast, internalization of the cyclic synthetic arginine-glycine-aspartate motif (cRGD) peptides targeted for ␣ v ␤ 3 takes place by an integrin-independent fluid-phase endocytosis pathway (20). To assess the feasibility of using the integrin-targeting cp38C2 constructs for drug delivery, we evaluated the internalization of the cp38C2 variants.…”

Section: Resultsmentioning

confidence: 99%

Breaking the one antibody–one target axiom

Guo

Das

Mueller

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Studies at the interface of chemistry and biology have allowed us to develop an immunotherapeutic approach called chemically programmed antibodies (cpAbs), which combines the merits of traditional small-molecule drug design with immunotherapy. In this approach, a catalytic antibody catalyzes the covalent conjugation of a small molecule or peptide to the active site of the antibody, effectively recruiting the binding specificity of the conjugated molecule to the antibody. In essence, this technology provides the tools for breaking the ''one antibody-one target axiom'' of immunochemistry. Our studies in this area have focused on using the chemistry of the well studied aldolase catalytic antibodies of which mAb 38C2 is a member. Previously, we explored reversible assembly of cpAbs available through diketone chemistry. In this article, we explore a unique proadapter assembly strategy wherein an antibody 38C2-catalyzed transformation unveils a reactive tag that then reacts to form a stable covalent bond with the antibody. An integrin ␣v␤3 antagonist was synthesized with the designed proadapter and studied using human breast cancer cell lines MDA-MB-231 and MDA-MB-435. We demonstrate that this approach allows for (i) the effective assembly of cpAbs in vitro and in vivo, (ii) selective retargeting of 38C2 to integrin ␣v␤3 expressing breast cancer cell lines, (iii) intracellular delivery of cpAbs into cells, (iv) dramatically increased circulatory half-life, and (v) substantial enhancement of the therapeutic effect over the peptidomimetic itself in animal models of breast cancer metastasis. We believe that this technology possesses potential for the treatment and diagnosis of disease.catalytic antibody ͉ chemical programming ͉ combinatorial antibody libraries M onoclonal antibodies are a rapidly growing class of therapeutics for a wide variety of diseases (1, 2). Some of the advantages of antibodies include their relative lack of nonspecific toxicity, long half-life, and ease of access from patient-derived or synthetic combinatorial antibody libraries. For certain diseases, such as cancer, that antibodies can carry their own effector functions is of prime importance because the antibody specificity directs the killing function endemic to the effector domain, the Fc. It has always been axiomatic in immunochemistry that even though one may desire one or more of the advantageous properties common to all antibodies, due to their clonal nature, each task requires a different antibody. A solution to this problem, namely chemically programmed antibodies (cpAbs), has emerged at the interface of chemistry and biology: One can use different low-molecular-weight targeting agents (programming agents or adapters) to selectively target the same antibody to different sites for different uses (3). This strategy has the advantage that only a single antibody is required for a multiplicity of tasks, and it taps into the unlimited chemical diversity and the specificity that can be engendered by organic synthesis (4). The antibody provides ...

show abstract

RGD Peptides and Monoclonal Antibodies, Antagonists of αv-Integrin, Enter the Cells by Independent Endocytic Pathways

Cited by 87 publications

References 47 publications

Targeted Vpr-derived peptides reach mitochondria to induce apoptosis of αVβ3-expressing endothelial cells

Targeted Vpr-derived peptides reach mitochondria to induce apoptosis of αVβ3-expressing endothelial cells

RGD-containing Peptides Activate S6K1 through β3 Integrin in Adult Cardiac Muscle Cells

Breaking the one antibody–one target axiom

Contact Info

Product

Resources

About