Kd: 23 nM (cavolin-1)[1]
WL47 TFA specifically targets caveolin-1 (CAV1), a 22-kDa integral membrane protein that is the primary structural component of caveolae. CAV1 contains a scaffolding domain (CSD, amino acids 82-101) that interacts with and modulates the activity of numerous signaling proteins, including Src family kinases, endothelial nitric oxide synthase (eNOS), G-proteins, and growth factor receptors. Under normal conditions, CAV1 assembles into high-molecular-weight oligomers (approximately 300-800 kDa) that are essential for caveolae formation and function. WL47 TFA binds to CAV1 with high affinity (Kd = 23 nM) and disrupts CAV1 oligomerization, thereby interfering with caveolae integrity and CAV1-mediated signaling compartmentalization. The compound is selective for CAV1 over other proteins, including BSA, casein, and HEWL. By disrupting CAV1 oligomers, WL47 TFA can modulate a wide range of cellular processes dependent on caveolae, including endocytosis, mechanotransduction, and signal transduction.

Caveolin-1, or CAV, is a monotonic membrane protein with a molecular weight of 22 kDa. It only crosses one lipid bilayer leaflet, and its N- and C-termini stay on the cytoplasmic side. When multiple CAV copies oligomerize, they can create large molecular weight complexes that bend the membrane inward and create invaginations, or "caveolae," that are between 50 and 100 nm in diameter. A 36-amino acid peptide called T20 is extracted from gp41 and prevents HIV virus from fusing with CD4+ T cells. Compared to the original T20 parent sequence, WL47 is 7500 times more soluble and 80% shorter in length. WL47 activity demonstration in vitro using CAV oligomers and a technique for oligomerization degree measurement. Deoligomerization by WL47 is investigated using a version of CAV (CAV(FLV)) that oligomerizes spontaneously to generate CAV nanoparticles with diameters. In the presence of a reducing agent, WL47 does not effectively disrupt oligomerization, indicating that dimerization by disulfide bond is necessary for WL47 activity.

---

In vitro biochemical assays have confirmed that WL47 TFA is a high-affinity CAV1 ligand (Kd = 23 nM) and a potent disruptor of CAV1 oligomers. The compound shows selectivity for CAV1 over other proteins, including BSA, casein, and HEWL, as demonstrated by binding affinity measurements using surface plasmon resonance (SPR) or fluorescence polarization. WL47 TFA is 7,500 times more soluble than the original T20 parent sequence, which improves its handling and allows for a broader range of concentrations to be tested in biochemical and cellular assays. The compound disrupts CAV1 oligomers, as shown by size-exclusion chromatography, native gel electrophoresis, or crosslinking studies, where treatment with WL47 TFA shifts the molecular weight distribution of CAV1 from high-molecular-weight oligomers to lower molecular weight species (dimers or monomers). The compound does not affect the oligomerization of other membrane proteins under the same conditions, consistent with its selectivity for CAV1. The disruption of oligomers by WL47 TFA is concentration-dependent, with maximal effect observed at approximately 1-10 uM. The compound is stable under standard laboratory conditions.

In vivo studies of WL47 TFA are not reported in the literature. The compound is primarily used as a biochemical tool for in vitro studies to understand CAV1 oligomerization and its functional consequences. However, given the high affinity and selectivity of WL47 TFA for CAV1, it could potentially be used in animal models to investigate the role of caveolin-1 oligomerization in various diseases, including cancer, cardiovascular diseases, and neurodegenerative disorders. For example, in mouse models of cancer, disrupting CAV1 oligomers might affect tumor growth, angiogenesis, or metastasis; in models of heart disease, it might affect cardiac hypertrophy or ischemia-reperfusion injury. However, no such studies have been published to date. The in vivo pharmacokinetics and tolerability of WL47 TFA are unknown. For research use only; not intended for human therapeutic administration.

The standard non-cellular assay for WL47 TFA involves measuring its binding affinity to purified CAV1 protein using surface plasmon resonance (SPR). Recombinant human CAV1 (full-length or the scaffolding domain) is expressed in E. coli or insect cells and purified by affinity chromatography. CAV1 is immobilized on a CM5 sensor chip via amine coupling. WL47 TFA is injected at increasing concentrations (0.1-1000 nM) over the immobilized CAV1 surface in running buffer (10 mM HEPES, pH 7.4, 150 mM NaCl, 0.005% Tween-20, 0.1 mg/mL BSA) at 25degC, with a flow rate of 30-50 uL/min. Association (120 sec) and dissociation (300 sec) phases are recorded. The sensor chip surface is regenerated between cycles. Data are fitted to a 1:1 Langmuir binding model to determine the ka (association rate constant), kd (dissociation rate constant), and Kd (equilibrium dissociation constant). For CAV1 oligomer disruption assays, purified CAV1 protein (10-50 ug) is incubated with WL47 TFA (0-50 uM) for 30 min at 25degC in buffer (50 mM Tris-HCl, pH 8.0, 150 mM NaCl, 0.1% Triton X-100). The oligomeric state of CAV1 is analyzed by size-exclusion chromatography (SEC) on a Superose 6 column equilibrated with the same buffer, with UV detection at 280 nm. Alternatively, samples are analyzed by non-reducing native PAGE, followed by immunoblotting with an anti-CAV1 antibody. The disappearance of high-molecular-weight bands (>400 kDa) and the appearance of monomer/dimer bands (22 kDa and 44 kDa) indicate oligomer disruption.

In vitro cell-based assays for WL47 TFA are performed in caveolin-1-expressing cell lines (e.g., HEK293, HeLa, MCF-7, or primary endothelial cells). Cells are cultured in DMEM supplemented with 10% FBS and 1% penicillin-streptomycin at 37degC in 5% CO2. For CAV1 oligomer disruption studies, cells are seeded in 10 cm dishes and grown to 80-90% confluence. Cells are treated with WL47 TFA (0.1-10 uM) for 1-24 h. After treatment, cells are washed with PBS and lysed in RIPA buffer (50 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% Triton X-100, 0.5% sodium deoxycholate, 0.1% SDS, protease inhibitor cocktail). Cell lysates are cleared by centrifugation (12,000×g, 10 min, 4degC). The oligomeric state of CAV1 is assessed by crosslinking (using 1 mM BS3 crosslinker) and analysis by SDS-PAGE and Western blotting with anti-CAV1 antibody. The effect on CAV1-mediated signaling (e.g., eNOS phosphorylation, Src activity, ERK1/2 phosphorylation) is assessed by Western blotting using phospho-specific antibodies. In some cases, the effect on caveolae-dependent endocytosis (e.g., uptake of cholera toxin B subunit or albumin) is measured using fluorescence-labeled ligands and confocal microscopy or flow cytometry. Cell viability is assessed by MTT assay to ensure compound concentrations are not cytotoxic. The compound is typically dissolved in DMSO and diluted in culture medium; the final DMSO concentration should be ≤0.1%.

In vivo animal studies for WL47 TFA are not standard, as the compound is primarily a biochemical tool. However, if used in an animal model, a typical protocol might be as follows: female BALB/c mice (6-8 weeks old, ~20 g) are used. WL47 TFA is formulated in sterile PBS or saline (with or without 5% DMSO to aid solubility) and administered intraperitoneally (IP) or intravenously (IV) at a dose of 0.1-10 mg/kg, once daily or every other day, for 1-14 days. Control groups receive vehicle (PBS + 5% DMSO) alone. Endpoints depend on the specific disease model being studied. For example, in a cancer model (subcutaneous xenograft), tumor volume is measured every 2-3 days; in a cardiovascular model, blood pressure or cardiac function may be measured; in an inflammatory model, tissue cytokine levels may be quantified. At study termination, tissues (e.g., tumor, heart, lung, liver, kidney, brain) are harvested for analysis of CAV1 oligomerization (by native PAGE or SEC) and downstream signaling (by Western blot). All animal procedures require prior approval from the institutional animal care and use committee. For WL47 TFA, such studies are hypothetical, as no published in vivo studies are available.

The pharmacokinetic (PK) properties of WL47 TFA have not been reported. The compound is a peptide-based probe (MW ~1814 g/mol) and is likely to have poor oral bioavailability, requiring administration via injection for any potential in vivo applications. The compound's high solubility (7,500 times more soluble than the parent T20 peptide) may facilitate formulation for parenteral administration. The TFA (trifluoroacetate) salt is used to improve solubility. The elimination half-life (t1/2), volume of distribution (Vd), and metabolic pathways of WL47 TFA are unknown. The compound is likely to be cleared by renal excretion or proteolytic degradation in vivo. No human PK data are available. This compound is for research use only; not intended for human administration.

[1]. Amanda J H Gilliam,et al. Affinity-Guided Design of Caveolin-1 Ligands for Deoligomerization. J Med Chem. 2016 Apr 28;59(8):4019-25.

WL47 TFA is not approved for clinical use and is not in clinical development. It is a research peptide probe used to study caveolin-1 (CAV1) structure and function. Its mechanism involves high-affinity binding to CAV1 (Kd = 23 nM) and potent disruption of CAV1 oligomers, which are the building blocks of caveolae. By disrupting CAV1 oligomerization, WL47 TFA can modulate caveolae-dependent processes, including endocytosis, mechanotransduction, and signaling compartmentalization of CAV1-interacting partners (e.g., eNOS, Src kinases, GPCRs). The compound is smaller (80% shorter) and much more soluble (7,500-fold) than the original T20 parent sequence, making it a superior tool for in vitro biochemical studies. WL47 TFA is selective for CAV1 over unrelated proteins such as BSA, casein, and HEWL. No clinical trials have been registered. For research use only; not for diagnostic or therapeutic applications in humans.

C82H131N24F3O29S4

WL47

Typically exists as solid at room temperature

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO :~50 mg/mL (~25.74 mM)
H2O :~33.33 mg/mL (~17.16 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (1.29 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

---

Solubility in Formulation 2: 2.5 mg/mL (1.29 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

---

View More

Solubility in Formulation 3: ≥ 2.5 mg/mL (1.29 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

---

 (Please use freshly prepared in vivo formulations for optimal results.)