Non-peptide Antagonist Research Articles

Kinin B1 Receptor Inhibition With BI113823 Reduces Inflammatory Response, Mitigates Organ Injury, and Improves Survival Among Rats With Severe Sepsis.

This study examined the therapeutic effects of an orally active nonpeptide kinin B1 receptor antagonist, BI113823, in a clinically relevant experimental model of polymicrobial sepsis in rats. Sepsis was induced by cecal ligation and puncture (CLP). Animals received treatment with either vehicle or BI113823. The experiment was terminated in the first set of animals 15 hours after CLP. Seven-day survival following CLP was determined in the second set of animals. Compared with vehicle treatment, administration of BI113823 reduced neutrophil and macrophage infiltration, reduced cytokine production, attenuated intestinal mucosal hyperpermeability, prevented hemodynamic derangement, and improved cardiac output. Furthermore, administration of BI113823 reduced inducible nitric oxide synthase expression and the injury score in the lung and attenuated nuclear factor ĸB activation and apoptosis in the liver. Treatment with BI113823 also reduced plasma levels of cardiac troponin, aspartate aminotransferase, alanine aminotransferase, urea, and lactate, as well as proteinuria. Finally, administration of BI113823 improved the 7-day survival rate following CLP in rats. Administration of BI113823 reduced systemic and tissue inflammatory responses, prevented hemodynamic derangement, attenuated multiorgan injury, and improved overall survival.

Development and characterization of Telmisartan self-microemulsifying drug delivery system for bioavailability enhancement

Telmisartan is a potent, long-lasting, nonpeptide antagonist of the angiotensin II type-1 (AT1) receptor that is indicated for the treatment of essential hypertension. It belongs to a class II drug in BCS classification i.e. low solubility and high permeability. One of the major problems with this drug is its low solubility in biological fluids which results into poor bioavailability after oral administration. The objective of present work was to develop a self microemulsifying drug delivery system (SMEDDS) to enhance the oral bioavailability of poorly water soluble Telmisartan. SMEDDS is a mixture of oil, surfactant, and cosurfactant, which are emulsified in aqueous medium under gentle digestive motility in the gastrointestinal tract. Psuedoternary phase diagrams were constructed to identify the efficient self-emulsifying region. A SMEDDS were further evaluated for its percentage transmittance, emulsification time, drug content, phase separation, globule size, zeta potential, pH, refractive index, X-ray diffraction, Differential scanning calorimetry and in vitro dissolution studies. Optimized formulation was also compared with marketed product (Telma 20) in male spraguedawley rats. The pharmacokinetic study exhibited 1.54 fold increase in the oral bioavailability of Telmisartan SMEDDS compared with the Marketed product.

Apelin receptors: From signaling to antidiabetic strategy

The G protein-coupled receptor APJ and its cognate ligand, apelin, are widely expressed throughout human body. They are implicated in different key physiological processes such as angiogenesis, cardiovascular functions, fluid homeostasis and energy metabolism regulation. On the other hand, this couple ligand–receptor is also involved in the development and progression of different pathologies including diabetes, obesity, cardiovascular disease and cancer. Recently, a new endogenous peptidic ligand of APJ, named Elabela/Toddler, has been identified and shown to play a crucial role in embryonic development. Whereas nothing is yet known regarding Elabela/Toddler functions in adulthood, apelin has been extensively described as a beneficial adipokine regarding to glucose and lipid metabolism and is endowed with anti-diabetic and anti-obesity properties. Indeed, there is a growing body of evidence supporting apelin signaling as a novel promising therapeutic target for metabolic disorders (obesity, type 2 diabetes). In this review, we provide an overview of the pharmacological properties of APJ and its endogenous ligands. We also report the activity of peptidic and non-peptidic agonists and antagonists targeting APJ described in the literature. Finally, we highlight the important role of this signaling pathway in the control of energy metabolism at the peripheral level and in the central nervous system in both physiological conditions and during obesity or diabetes.

A structural feature of the non-peptide ligand interactions with mice mu-opioid receptors.

By binding to and activating the G-protein coupled μ-, κ- and δ-opioid receptors in the central nervous system, opiates are known to induce analgesic and sedative effects. In particular, non-peptide opioid ligands are often used in clinical applications to induce these therapeutically beneficial effects, due to their superior pharmacokinetics and bioavailability in comparison to endogenous neuropeptides. However, since opioid alkaloids are highly addictive substances, it is necessary to understand the exact mechanisms of their actions, specifically the ligand-binding properties of the target receptors, in order to safely apply opiates for therapeutic purposes. Using an in silico molecular docking approach (AutoDock Vina) combined with two-step cluster analysis, we have computationally obtained the docking scores and the ligand-binding pockets of twelve representative non-peptide nonendogenous agonists and antagonists at the crystallographically identified μ-opioid receptor. Our study predicts the existence of two main binding sites that are congruently present in all opioid receptor types. Interestingly, in terms of the agonist or antagonist properties of the substances on the receptors, the clustering analysis suggests a relationship with the position of the ligand-binding pockets, particularly its depth within the receptor structure. Furthermore, the binding affinity of the substances is directly correlated to the proximity of the binding pockets to the extracellular space. In conclusion, the results provide further insights into the structural features of the functional pharmacology of opioid receptors, suggesting the importance of the binding position of non-peptide agonists and antagonists- specifically the distance and the level of exposure to the extracellular space- to their dissociation kinetics and subsequent potency.

P7, a novel antagonist of corticotropin releasing factor receptor type 1 (CRFR1) screened from phage display library

The corticotropin releasing factor (CRF) plays a central role in regulating the activities of hypothalamic-pituitary-adrenal (HPA) axis in the presence of a variety of stressful stimuli via binding to its type 1 receptors (CRFR1). Despite that many peptidic or non-peptidic antagonists of CRFR1 have been developed to serve as therapeutic tools to CRF-related pathologies, none of them have been utilized clinically. Targeting the extracellular domain 1 (EC1) of CRFR1, the CRF-binding site, represents a new strategy to inhibit the function of the receptor. However, no such agents have been identified up to now. Herein, by using an 87-amino acid fragment corresponding to the EC1 region as the bait, we screened the binding polypeptides from a phage display (Ph.D.-12) peptide library. After 3-round biopanning, positive clones were selected and the polypeptides carried by them were identified. 5 polypeptides were found to bind with the target specifically. Among them, the P7 exhibited the highest affinity. By evaluating the cAMP accumulation in the CRFR1 or CRFR2-expressing HEK293 cells, we demonstrated that P7 blocking the function of CRFR1, but not CRFR2. In addition, we also found that P7 and CRF act on CRFR1 competitively. Taken together, we reveal that P7, a novel polypeptide identified from phage display library, inhibits the function of CRFR1 effectively and specifically by binding at its EC1 domain. The new polypeptide might provide a promising agent for diagnostic or therapeutic utilities in CRF-related disorders.

Synthesis toward CRHR1 Antagonists through 2,7-Dimethylpyrazolo[1,5-α][1,3,5]triazin-4(3H)-one C-H Arylation.

A novel synthetic protocol for 8-aryl substituted pyrazolo[1,5-α][1,3,5]triazin-4(3H)-ones was developed employing Pd-catalyzed C-H arylation. The reaction yield was influenced by the presence of a phosphine ligand, pivalic acid, and base selection. With the use of 5-10 mol % catalyst, reactions of 2 with p- or m-substituted aryl bromides proceeded in moderate to good yields. Lower yields were observed with o-substituted aryl bromides. Using this method a precursor for MJL1-109-2, a known nonpeptide CRHR-1 antagonist, was successfully synthesized.

Convergent QSAR studies on a series of NK3 receptor antagonists for schizophrenia treatment

The dopamine hypothesis states that decreased dopaminergic neurotransmission reduces schizophrenia symptoms. Neurokinin-3 receptor (NK3) antagonists reduce dopamine release and have shown positive effects in pre-clinical and clinical trials. We employed 2D and 3D-QSAR analysis on a series of 40 non-peptide NK3 antagonists. Multivariate statistical analysis, PCA and HCA, were performed to rational training/test set splitting and PLS regression was employed to construct all QSAR models. We constructed one highly predictive CoMFA model (q2 = 0.810 and r2 = 0.929) and acceptable HQSAR and CoMSIA models (HQSAR q2 = 0.644 and r2 = 0.910; CoMSIA q2 = 0.691, r2 = 0.911). The three different techniques provided convergent physicochemical results. All models indicate cyclopropane, piperidine and di-chloro-phenyl ring attached to cyclopropane ring and also the amide group attached to the piperidine ring could play an important role in ligand–receptor interactions. These findings may contribute to develop potential NK3 receptor antagonists for schizophrenia.

The amide linker in nonpeptide neurotensin receptor ligands plays a key role in calcium signaling at the neurotensin receptor type 2.

Compounds acting via the GPCR neurotensin receptor type 2 (NTS2) display analgesia in relevant preclinical models. The amide bond in nonpeptide NTS1 antagonists plays a central role in receptor recognition and molecular conformation. Using NTS2 FLIPR and binding assays, we found that it is also a key molecular structure for binding and calcium mobilization at NTS2. We found that reversed amides display a shift from agonist to antagonist activity and provided examples of the first competitive nonpeptide antagonists observed in the NTS2 FLIPR assay. These compounds will be valuable tools for determining the role of calcium signaling in vitro to NTS2 mediated analgesia.

Peptide ligand recognition by G protein-coupled receptors.

The past few years have seen spectacular progress in the structure determination of G protein-coupled receptors (GPCRs). We now have structural representatives from classes A, B, C, and F. Within the rhodopsin-like class A, most structures belong to the α group, whereas fewer GPCR structures are available from the β, γ, and δ groups, which include peptide GPCRs such as the receptors for neurotensin (β group), opioids, chemokines (γ group), and protease-activated receptors (δ group). Structural information on peptide GPCRs is restricted to complexes with non-peptidic drug-like antagonists with the exception of the chemokine receptor CXCR4 that has been crystallized in the presence of a cyclic peptide antagonist. Notably, the neurotensin receptor 1 is to date the only peptide GPCR whose structure has been solved in the presence of a peptide agonist. Although limited in number, the current peptide GPCR structures reveal great diversity in shape and electrostatic properties of the ligand binding pockets, features that play key roles in the discrimination of ligands. Here, we review these aspects of peptide GPCRs in view of possible models for peptide agonist binding.

AT2 receptor activities and pathophysiological implications.

Although angiotensin II subtype-2 receptor (AT2R) was discovered over 2 decades ago, its contribution to physiology and pathophysiology is not fully elucidated. Current knowledge suggests that under normal physiologic conditions, AT2R counterbalances the effects of angiotensin II subtype-1 receptor (AT1R). A major obstacle for AT2R investigations was the lack of specific agonists. Most of the earlier AT2R studies were performed using the peptidic agonist, CG42112A, or the nonpeptidic antagonist PD123319. CGP42112A is nonspecific for AT2R and in higher concentrations can bind to AT1R. Recently, the development of specific nonpeptidic AT2R agonists boosted the efforts in identifying the therapeutic potentials for AT2R stimulation. Unlike AT1R, AT2R is involved in vasodilation by the release of bradykinin and nitric oxide, anti-inflammation, and healing from injury. Interestingly, the vasodilatory effects of AT2R stimulation were not associated with significant reduction in blood pressure. In the kidney, AT2R stimulation produced natriuresis, increased renal blood flow, and reduced tissue inflammation. In animal studies, enhanced AT2R function led to reduction of cardiac inflammation and fibrosis, and reduced the size of the infarcted area. Similarly, AT2R stimulation demonstrated protective effects in vasculature and brain.

Genetic depletion and pharmacological targeting of αv integrin in breast cancer cells impairs metastasis in zebrafish and mouse xenograft models

IntroductionIncreased expression of αv integrins is frequently associated with tumor cell adhesion, migration, invasion and metastasis, and correlates with poor prognosis in breast cancer. However, the mechanism by which αv integrins can enhance breast cancer progression is still largely unclear. The effects of therapeutic targeting of αv integrins in breast cancer also have yet to be investigated.MethodsWe knocked down αv integrin in MDA-MB-231 and MCF10A-M4 breast cancer cells, or treated these cells with the αv antagonist GLPG0187. The effects of αv integrin depletion on mesenchymal markers, transforming growth factor-β (TGF-β)/Smad signaling and TGF-β-induced target gene expression were analyzed in MDA-MB-231 cells by RNA analysis or Western blotting. The function of αv integrin on breast cancer cell migration was investigated by transwell assay in vitro, and its effect on breast cancer progression was assessed by both zebrafish and mouse xenografts in vivo. In the mouse model, GLPG0187 was administered separately, or in combination with the standard-of-care anti-resorptive agent zoledronate and the chemotherapeutic drug paclitaxel, to study the effects of combinational treatments on breast cancer metastasis.ResultsGenetic interference and pharmacological targeting of αv integrin with GLPG0187 in different breast cancer cell lines inhibited invasion and metastasis in the zebrafish or mouse xenograft model. Depletion of αv integrin in MDA-MB-231 cells inhibited the expression of mesenchymal markers and the TGF-β/Smad response. TGF-β induced αv integrin mRNA expression and αv integrin was required for TGF-β-induced breast cancer cell migration. Moreover, treatment of MDA-MB-231 cells with non-peptide RGD antagonist GLPG0187 decreased TGF-β signaling. In the mouse xenografts GLPG0187 inhibited the progression of bone metastasis. Maximum efficacy of inhibition of bone metastasis was achieved when GLPG0187 was combined with the standard-of-care metastatic breast cancer treatments.ConclusionThese findings show that αv integrin is required for efficient TGF-β/Smad signaling and TGF-β-induced breast cancer cell migration, and for maintaining a mesenchymal phenotype of the breast cancer cells. Our results also provide evidence that targeting αv integrin could be an effective therapeutic approach for treatment of breast cancer tumors and/or metastases that overexpress αv integrin.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-015-0537-8) contains supplementary material, which is available to authorized users.

Synthesis and biological evaluation of novel phenothiazine derivatives as non-peptide arginine vasopressin V2 receptor antagonists

Abstract A series of novel phenothiazine derivatives was synthesized and tested for arginine vasopressin receptor antagonist activity. They were synthesized as novel arginine vasopressin receptor antagonists from phenothiazine as a scaffold via successive acylation, reduction and acylation reactions. Their structures were characterized by 1 H NMR, 13 C NMR and HRMS, and biological activity was evaluated by in vitro and in vivo studies. The in vitro binding assay indicated that several compounds are potent selective V2 receptor antagonists. Compounds with promising binding affinity to V2 receptors were selected to conduct the in vivo diuretic studies on Sprague-Dawley rats. Among them, 1n , 1r , 1t and 1v exhibited excellent diuretic activity, especially 1r and 1v . Therefore, 1r and 1v are potent novel AVP V2 receptor antagonist candidates.

Procedure—economical enantioselective total syntheses of asperlicins C and E

We report a procedure—economical method for the highly enantioselective and protecting-group free total syntheses of nonpeptidal CCK antagonists asperlicins C and E. Starting from l-tryptophan, the synthesis of asperlicin C has been achieved in three steps, which features the low-valent titanium (LVT: TiCl4–Zn combination)-mediated reductive cyclization of o-nitrobenzamide to construct the (3H)-quinazolin-4-one moiety. This is the first employment of LVT for the synthesis of asperlicin C, which allowed accessing asperlicin C in >99% enantioselectivity. Asperlicin C was converted, in one-pot, into asperlicin E and 2,3-di-epi-asperlicin E by dimethyl dioxirane (DMDO)-mediated tandem reactions. The use of DMDO as a green, cheap, and easily available oxidant to replace the photochemical method renders the synthesis of asperlicin E experimentally convenient.

One-step (18)F labeling of non-peptidic bivalent integrin αvβ3 antagonist for cancer imaging.

A rapid one-step 18F labeling reaction with fluoridealuminum complex, which is based on chelation chemistry, has received a surge of interest for 18F radiolabeling of peptides. In this study, a non-peptidic bivalent integrin αvβ3 antagonist (bivalent-IA) was conjugated with 1,4,7-triazacyclononane-1,4-diiacetic acid (NODA). A novel 18F labeled radiotracer, 18F-bivalent-IA, was developed via one step 18F–AlF/NODA chelation reaction in aqueous phase with high radiochemical yield (65–75%, decay corrected) and good specific activity (750–850 mCi/μmol). The tumor integrin targeting efficiency and in vivo pharmacokinetic profile of 18F-bivalent-IA were evaluated in U-87 MG (integrin positive) and MDA-MB-231 (integrin negative) models by small-animal PET/CT scan followed by a biodistribution study. The PET/CT and ROI results showed high tumor uptake of 18F-bivalent-IA in U-87 MG tumor-bearing mice from 5 to 120 min p.i. with good contrast, and the U-87 MG tumor uptake values (6.35 ± 0.67%ID/g, at 1 h p.i.) were 6 times higher than those of MDA-MB-231 tumor (1.05 ± 0.12%ID/g, at 1 h p.i.) (P < 0.0001) which correlated with the integrin αvβ3 expression in tumor tissues confirmed by immunohistochemistry. Co-injection of the 18F-bivalent-IA with 6 nmol (6 μg) of nonradioactive bivalent-IA effectively blocked tumor uptake demonstrating the integrin αvβ3-specificity. In conclusion, the first 18F labeled non-peptidic bivalent integrin αvβ3 targeting radiotracer, 18F-bivalent-IA, was developed and proved to be a highly potent and specific PET radiopharmaceutical for noninvasive imaging of integrin αvβ3, which plays a critical role in tumor angiogenesis and metastasis.

Recent Progress in the Use of Glucagon and Glucagon Receptor Antago-nists in the Treatment of Diabetes Mellitus

Glucagon is an important pancreatic hormone, released into blood circulation by alpha cells of the islet of Langerhans. Glucagon induces gluconeogenesis and glycogenolysis in hepatocytes, leading to an increase in hepatic glucose production and subsequently hyperglycemia in susceptible individuals. Hyperglucagonemia is a constant feature in patients with T2DM. A number of bioactive agents that can block glucagon receptor have been identified. These glucagon receptor antagonists can reduce the hyperglycemia associated with exogenous glucagon administration in normal as well as diabetic subjects. Glucagon receptor antagonists include isoserine and beta-alanine derivatives, bicyclic 19-residue peptide BI-32169, Des-His1-[Glu9] glucagon amide and related compounds, 5-hydroxyalkyl-4-phenylpyridines, N-[3-cano-6- (1,1 dimethylpropyl)-4,5,6,7-tetrahydro-1-benzothien-2-yl]-2-ethylbutamide, Skyrin and NNC 250926. The absorption, dosage, catabolism, excretion and medicinal chemistry of these agents are the subject of this review. It emphasizes the role of glucagon in glucose homeostasis and how it could be applied as a novel tool for the management of diabetes mellitus by blocking its receptors with either monoclonal antibodies, peptide and non-peptide antagonists or gene knockout techniques.

ML-18 is a non-peptide bombesin receptor subtype-3 antagonist which inhibits lung cancer growth

Bombesin receptor subtype (BRS)-3 is a G protein coupled receptor (GPCR) for the bombesin (BB)-family of peptides. BRS-3 is an orphan GPCR and little is known of its physiological role due to the lack of specific agonists and antagonists. PD168368 is a nonpeptide antagonist for the neuromedin B (NMB) receptor (R) whereas PD176252 is a nonpeptide antagonist for the gastrin releasing peptide (GRP) R and NMBR but not BRS-3. Here nonpeptide analogs of PD176252 e.g. the S-enantiomer ML-18, and the R-enantiomer, EMY-98, were investigated as BRS-3 antagonists using lung cancer cells. ML-18 and EMY-98 inhibited specific 125I-BA1 (DTyr-Gln-Trp-Ala-Val-βAla-His-Phe-Nle-NH2)BB6-14 binding to NCI-H1299 lung cancer cells stably transfected with BRS-3 with IC50 values of 4.8 and >100μM, respectively. In contrast, ML-18 bound with lower affinity to the GRPR and NMBR with IC50 values of 16 and >100μM, respectively. ML-18 (16μM), but not its enantiomer EMY-98, inhibited the ability of 10nM BA1 to elevate cytosolic Ca2+ in a reversible manner using lung cancer cells loaded with FURA2-AM. ML-18 (16μM), but not EMY-98, inhibited the ability of 100nM BA1 to cause tyrosine phosphorylation of the EGFR and ERK in lung cancer cells. ML-18 but not EMY-98 inhibited the proliferation of lung cancer cells. The results indicate that ML-18 is a nonpeptide BRS-3 antagonist that should serve as a template to improve potency and selectivity.

International Union of Basic and Clinical Pharmacology. XCII. Urotensin II, urotensin II-related peptide, and their receptor: from structure to function.

Urotensin II (UII) is a cyclic neuropeptide that was first isolated from the urophysis of teleost fish on the basis of its ability to contract the hindgut. Subsequently, UII was characterized in tetrapods including humans. Phylogenetic studies and synteny analysis indicate that UII and its paralogous peptide urotensin II-related peptide (URP) belong to the somatostatin/cortistatin superfamily. In mammals, the UII and URP genes are primarily expressed in cholinergic neurons of the brainstem and spinal cord. UII and URP mRNAs are also present in various organs notably in the cardiovascular, renal, and endocrine systems. UII and URP activate a common G protein-coupled receptor, called UT, that exhibits relatively high sequence identity with somatostatin, opioid, and galanin receptors. The UT gene is widely expressed in the central nervous system (CNS) and in peripheral tissues including the retina, heart, vascular bed, lung, kidney, adrenal medulla, and skeletal muscle. Structure-activity relationship studies and NMR conformational analysis have led to the rational design of a number of peptidic and nonpeptidic UT agonists and antagonists. Consistent with the wide distribution of UT, UII has now been shown to exert a large array of biologic activities, in particular in the CNS, the cardiovascular system, and the kidney. Here, we review the current knowledge concerning the pleiotropic actions of UII and discusses the possible use of antagonists for future therapeutic applications.

Effect of small molecule vasopressin V1a and V2 receptor antagonists on brain edema formation and secondary brain damage following traumatic brain injury in mice.

The attenuation of brain edema is a major therapeutic target after traumatic brain injury (TBI). Vasopressin (AVP) is well known to play a major role in the regulation of brain water content and vasoendothelial functions and to be involved in brain edema formation. Therefore, the aim of the current study was to analyze the antiedematous efficacy of a clinically relevant, nonpeptidic AVP V1a and V2 receptor antagonists. C57Bl6 mice were subjected to controlled cortical impact (CCI) and V1a or V2 receptors were inhibited by using the highly selective antagonists SR-49059 or SR-121463A either by systemic (intraperitoneal, IP) or intracerebroventricular (ICV) application. After 24 h, brain edema, intracranial pressure (ICP), and contusion volume were assessed. Systemically applied AVP receptor antagonists could not reduce secondary lesion growth. In contrast, ICV administration of AVP V1a receptor antagonist decreased brain edema formation by 68%, diminished post-traumatic increase of ICP by 46%, and reduced secondary contusion expansion by 43% 24 h after CCI. The ICV inhibition of V2 receptors resulted in significant reduction of post-traumatic brain edema by 41% 24 h after CCI, but failed to show further influence on ICP and lesion growth. Hence, centrally applied vasopressin V1a receptor antagonists may be used to reduce brain edema formation after TBI.

K3036.58 in the μ opioid (MOP) receptor is important in conferring selectivity for covalent binding of β-funaltrexamine (β-FNA)

β-funaltrexamine (β-FNA) is an irreversible μ opioid (MOP) receptor antagonist and a reversible agonist of κ opioid (KOP) receptor. β-FNA binds covalently to the MOP receptor at Lys2335.39, which is conserved among opioid receptors. Molecular docking of β-FNA showed that K3036.58 in the MOP receptor and E2976.58 in the KOP receptor played distinct roles in positioning β-FNA. K3036.58E MOP receptor and E2976.58K KOP receptor mutants were generated. The mutations did not affect β-FNA affinity or efficacy. K3036.58E mutation in the MOP receptor greatly reduced covalent binding of [3H]β-FNA; however, E2976.58K did not enable the KOP receptor to bind irreversibly to β-FNA. Molecular modeling demonstrated that the ε-amino group of K3036.58 in the MOP receptor interacted with CO of the acetate group of β-FNA to facilitate covalent bond formation with Lys2335.39. Replacement of K3036.58 with Glu in the MOP receptor resulted in repulsion between the COOH of Glu and the CO of β-FNA and increased the distance between K2335.39 and the fumarate group, making it impossible for covalent bond formation. These findings will be helpful for design of selective non-peptide MOP receptor antagonists.

Selective in vivo imaging of syngeneic, spontaneous, and xenograft tumors using a novel tumor cell-specific hsp70 peptide-based probe.

Although in vivo targeting of tumors using fluorescently labeled probes has greatly gained in importance over the last few years, most of the clinically applied reagents lack tumor cell specificity. Our novel tumor cell-penetrating peptide-based probe (TPP) recognizes an epitope of Hsp70 that is exclusively present on the cell surface of a broad variety of human and mouse tumors and metastases, but not on normal tissues. Because of the rapid turnover rate of membrane Hsp70, fluorescently labeled TPP is continuously internalized into syngeneic, spontaneous, chemically/genetically induced and xenograft tumors following intravenous administration, thereby enabling site-specific labeling of primary tumors and metastases. In contrast with the commercially available nonpeptide small molecule αvβ3-integrin antagonist IntegriSense, TPP exhibits a significantly higher tumor-to-background contrast and stronger tumor-specific signal intensity in all tested tumor models. Moreover, in contrast with IntegriSense, TPP reliably differentiates between tumor cells and cells of the tumor microenvironment, such as tumor-associated macrophages and fibroblasts, which were found to be membrane-Hsp70 negative. Therefore, TPP provides a useful tool for multimodal imaging of tumors and metastases that might help to improve our understanding of tumorigenesis and allow the establishment of improved diagnostic procedures and more accurate therapeutic monitoring. TPP might also be a promising platform for tumor-specific drug delivery and other Hsp70-based targeted therapies.