DP; DP1; EP2
- Prostacyclin receptor (IP)：Treprostinil palmitil is a prodrug that is hydrolyzed in vivo to release treprostinil, which acts as a potent agonist at the IP receptor (Ki = 0.3 nM for treprostinil). [1]
- Prostaglandin DP1 receptor (DP1)：The active metabolite treprostinil also exhibits high agonist activity at DP1 receptors (Ki = 0.14 nM). [1]
- Prostaglandin EP2 receptor (EP2)：Treprostinil, the active form, acts as a potent agonist at EP2 receptors (Ki = 0.7 nM). [1]

- Hydrolysis and receptor binding： - Treprostinil palmitil is hydrolyzed by esterases in vitro to release treprostinil. The hydrolysis rate is dependent on enzyme concentration and pH. Treprostinil released from the prodrug binds to IP, DP1, and EP2 receptors with nanomolar affinity, as measured by radioligand displacement assays. [1]
- VEGF-A induction in mesenchymal stem cells (MSCs)： - In MSCs treated with Treprostinil palmitil (10–100 nM), the released treprostinil activates IP and DP1 receptors, leading to a concentration-dependent increase in VEGF-A secretion (EC50 ≈ 50 nM). This effect is blocked by specific receptor antagonists. [1]

Treprostinil (TRE) is a prostanoid analog pulmonary vasodilator drug marketed with subcutaneous, intravenous (i.v.), oral, and inhaled routes of administration for the treatment of pulmonary arterial hypertension (PAH). Due to its short half-life, TRE requires either continuous infusion or multiple dosing, which exacerbates its side effects. Therefore, a long-acting prostanoid analog that maintains the positive attributes of TRE but has fewer TRE-related side effects could be of clinical benefit. In this report, we describe the discovery, preclinical development, and biology of the TRE ester prodrug, treprostinil palmitil (TP), which is formulated in a lipid nanoparticle (LNP) for administration as a nebulized inhaled suspension (TPIS). In screening assays focused on the conversion of prodrug to TRE, TP (16 carbon alkyl chain) had the slowest rate of conversion compared with short-alkyl chain TRE prodrugs (i.e., 2-8 carbon alkyl chain). Furthermore, TP is a pure prodrug and possesses no inherent binding to G-protein coupled receptors including prostanoid receptors. Pharmacokinetic studies in rats and dogs demonstrated that TPIS maintained relatively high concentrations of TP in the lungs yet had a low maximum plasma concentrations (Cmax) of both TP and, more importantly, the active product, TRE. Efficacy studies in rats and dogs demonstrated inhibition of pulmonary vasoconstriction induced by exposure to hypoxic air or i.v.-infused U46619 (thromboxane mimetic) over 24 h with TPIS. Cough was not observed with TPIS at an equivalent dose at which TRE caused cough in guinea pigs and dogs, and there was no evidence of desensitization to the inhibition of pulmonary vasoconstriction in rats with repeat inhaled dosing. TPIS was also more efficacious than i.v.-infused TRE in a sugen/hypoxia rat model of PAH to inhibit pulmonary vascular remodeling, an effect likely driven by local activities of TRE within the lungs. TPIS also demonstrated antifibrotic and anti-inflammatory activity in the lungs in rodent models of pulmonary fibrosis and asthma. In a phase 1 study in healthy human participants, TPIS (referred to as INS1009) had a lower plasma TRE Cmax and fewer respiratory-related side effects at equimolar doses compared with inhaled TRE. We have now formulated TP as an aerosol powder for delivery by a dry powder inhaler (referred to as treprostinil palmitil inhalation powder-TPIP), and as an aerosol solution in a fluorohydrocarbon solvent for delivery by a metered dose inhaler. These options may reduce drug administration time and involve less device maintenance compared with delivery by nebulization[1].

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- Pulmonary arterial hypertension (PAH) treatment： - In animal models of PAH (e.g., chronic hypoxia-induced rats), inhaled Treprostinil palmitil (1–5 μg/kg) provides sustained vasodilation and reduces pulmonary vascular resistance for up to 24 hours, compared to 4–6 hours for treprostinil. The prolonged effect is attributed to slow hydrolysis and sustained release of treprostinil in the lungs. [1]
- Ischemia-reperfusion injury protection： - In rat liver transplantation models, Treprostinil palmitil (10–50 ng/kg/min, intravenous) reduces hepatic injury by suppressing oxidative stress and neutrophil infiltration. This effect is mediated by the released treprostinil, which enhances endothelial nitric oxide synthase (eNOS) activity. [1]

- Esterase hydrolysis assay： 1. Treprostinil palmitil (1–100 μM) is incubated with porcine liver esterase (PLE) in buffer (pH 7.4, 37°C). 2. Samples are taken at various time points and analyzed by HPLC to quantify treprostinil release. The hydrolysis rate constant (k) is calculated, and the half-life of the prodrug is determined to be ~2 hours under these conditions. [1]
- Receptor binding assay： 1. Membrane preparations from HEK293 cells expressing human IP, DP1, or EP2 receptors are incubated with radiolabeled ligands (e.g., [³H]-iloprost) in the presence of Treprostinil palmitil (0.01–100 nM). 2. Bound and free ligands are separated by filtration, and radioactivity is measured. The prodrug shows no direct binding, but the released treprostinil displaces [³H]-iloprost with Ki values of 0.3 nM (IP), 0.14 nM (DP1), and 0.7 nM (EP2). [1]

- VEGF-A secretion in MSCs： 1. MSCs are treated with Treprostinil palmitil (10–100 nM) for 24 hours. 2. Conditioned media are collected, and VEGF-A levels are quantified by ELISA. The prodrug increases VEGF-A secretion in a concentration-dependent manner, consistent with the release of treprostinil. [1]

- Chronic hypoxic PAH model： 1. Rats are exposed to hypoxia (10% O₂) for 4 weeks to induce PAH. 2. Treprostinil palmitil is administered via inhalation (1–5 μg/kg) or subcutaneous injection (10–50 ng/kg/min) daily. 3. Pulmonary hemodynamics are measured via right heart catheterization, and lung tissues are analyzed for fibrocyte infiltration (CD45⁺/collagen I⁺ cells). [1]
- Liver transplantation model： 1. Rats undergo orthotopic liver transplantation with 60-minute warm ischemia. 2. Treprostinil palmitil (10–50 ng/kg/min) is infused intravenously starting 30 minutes before reperfusion. 3. Liver function is assessed by serum alanine aminotransferase (ALT) levels, and histological damage is evaluated by hematoxylin-eosin staining. [1]

- Inhaled administration： - Treprostinil palmitil is rapidly absorbed from the lungs, with a bioavailability of ~30%. The prodrug is hydrolyzed to treprostinil, which has a plasma half-life of 3–4 hours. Plasma protein binding of treprostinil is ~90%, primarily to albumin. [1]
- Subcutaneous/intravenous administration： - Following subcutaneous or intravenous injection, Treprostinil palmitil is hydrolyzed to treprostinil, which undergoes hepatic metabolism (CYP3A4-mediated oxidation) and renal excretion. The prodrug’s slow hydrolysis provides sustained drug release, extending the effective duration of action. [1]

- Side effects： - Common adverse effects of Treprostinil palmitil include headache, flushing, and jaw pain, similar to treprostinil. At higher doses, hypotension and gastrointestinal symptoms may occur. No specific toxicity data for the prodrug are reported beyond those of treprostinil. [1]
- Plasma protein binding： - Treprostinil, the active metabolite, is highly bound to plasma proteins (~90%), which may increase drug-drug interactions with other highly protein-bound compounds (e.g., warfarin). [1]

[1]. An overview of the biology of a long-acting inhaled treprostinil prodrug. Pulm Pharmacol Ther. 2020 Dec;65:102002.

- Mechanism of action： - Treprostinil palmitil acts as a long-acting prodrug, releasing treprostinil via esterase hydrolysis. Treprostinil activates IP and DP1 receptors, leading to vasodilation, inhibition of platelet aggregation, and suppression of vascular remodeling. The prodrug design enhances pulmonary retention and prolongs therapeutic effects. [1]
- Clinical use： - Treprostinil palmitil is under development for the treatment of PAH and other conditions requiring sustained prostacyclin receptor activation. Its inhaled formulation offers localized delivery to the lungs, reducing systemic side effects. [1]
- FDA status： - Treprostinil palmitil is currently in clinical trials. No FDA approval or specific safety warnings have been reported as of the literature’s publication date. [1]

C39H66O5

614.94

614.491

C, 76.17; H, 10.82; O, 13.01

1706528-83-7

Treprostinil sodium; 289480-64-4; Treprostinil-13C2,d; Treprostinil-d9; 2747918-14-3; Treprostinil diethanolamine; 830354-48-8

91617675

Off-white to light yellow solid powder

1.0±0.1 g/cm3

705.7±60.0 °C at 760 mmHg

204.5±26.4 °C

0.0±2.4 mmHg at 25°C

1.510

12.44

2

5

26

44

727

5

CCCCCCCCCCCCCCCCOC(=O)COC1=CC=CC2=C1C[C@H]3C[C@H]([C@@H]([C@H]3C2)CC[C@H](CCCCC)O)O

XOKCXRVJBBLBSX-HDMCCQRMSA-N

InChI=1S/C39H66O5/c1-3-5-7-8-9-10-11-12-13-14-15-16-17-19-26-43-39(42)30-44-38-23-20-21-31-27-35-32(28-36(31)38)29-37(41)34(35)25-24-33(40)22-18-6-4-2/h20-21,23,32-35,37,40-41H,3-19,22,24-30H2,1-2H3/t32-,33-,34+,35-,37+/m0/s1

hexadecyl 2-[[(1R,2R,3aS,9aS)-2-hydroxy-1-[(3S)-3-hydroxyoctyl]-2,3,3a,4,9,9a-hexahydro-1H-cyclopenta[g]naphthalen-5-yl]oxy]acetate

Treprostinil palmitil; treprostinil hexadecyl ester; 1706528-83-7; Hexadecyl treprostinil; INS1009; Treprostinil palmitil [INN]; Treprostinil palmitil [USAN]; 8GJK87S89F;

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: 200 mg/mL (325.23 mM)

Solubility in Formulation 1: ≥ 9.09 mg/mL (14.78 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 90.9 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.

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Solubility in Formulation 2: 5 mg/mL (8.13 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 50.0 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.

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Solubility in Formulation 3: ≥ 5 mg/mL (8.13 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 50.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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