G protein-coupled receptor kinase 2 (GRK2) and GRK3 inhibitor. For GRK2 inhibition in biochemical assays, IC₅₀ values of 35 nM, 54 nM, and 290 nM have been reported for CMPD101, with values varying due to slightly different assay conditions [2]
GRK3 may be inhibited with similar potency: using identical assay conditions, the IC₅₀ value of CMPD101 was 32 nM for GRK3 [2]
Notably, even at CMPD101 concentrations up to 125 μM, no inhibition of GRK1 or GRK5 was observed in biochemical assays [2]

The HEK-B2 cell line's plasma membrane retains the β2AR-GFP protein fusion while CMPD101 (100 μM; first 20 min) inhibits the internalization of β2AR and significantly lowers the formation of protein-coated vesicles induced by isopropanol inducer. CMPD101 (3-30 μM; first 30 min) produces strong Ser375 phosphorylation, which is partially inhibited by 3 μM Cmpd101 for 30 min. 30 μM Cmpd101 totally replenishes cell death. 1]. prevents MOPr from being phosphorylated on the residues of Thr370, Thr376, and Thr379 [2]. CMPD101 (3-30 μM; 30 min ago) elicits baseline ERK1/2 phosphorylation in HEK 293 cells expressing HA-MOPrs Already added, however DAMGO-induced ERK1/2 and Elk-1 activation is unaffected by the compound[2].

---

CMPD101 (5 μM and 50 μM) concentration-dependently inhibited electric field stimulation (EFS)-induced, frequency-dependent contractions of human prostate strips [2]
CMPD101 (50 μM) inhibited concentration-dependent contractions of human prostate strips induced by the α₁-adrenoceptor agonists noradrenaline and phenylephrine, as well as by the non-adrenergic agonists endothelin-1 and the thromboxane A₂ analogue U46619. It reduced the maximum response (Eₘₐₓ) but did not change the EC₅₀ values for these agonists [2]
CMPD101 (50 μM) did not affect the phosphorylation state of the myosin light chain (MLC, serine 19) or the Rho kinase substrate MYPT-1 (threonine 696) in human prostate tissues, suggesting its inhibitory effect on contraction is not mediated through direct inhibition of MLC kinase or Rho kinase [2]
CMPD101 (50 μM) did not significantly alter contractions induced by high molar KCl (80 mM) in human prostate strips, indicating its effect is not due to non-specific inhibition of voltage-gated calcium channels or the downstream contractile machinery [2]
CMPD101 (50 μM) did not change the phosphorylation state of β₂-adrenoceptors (at serine 346 or serine 355) in human prostate tissues [2]
CMPD101 (50 μM) did not alter relaxations of pre-contracted human prostate strips induced by the β₂-adrenoceptor agonists isoproterenol or salbutamol [2]
Western blot analyses and peroxidase/fluorescence stainings suggested expression of GRK2 and GRK3 proteins in the stromal smooth muscle cells of human prostate tissues [2]

Biochemical assays using purified enzymes were performed to determine the inhibitory activity of CMPD101 against GRK2 and GRK3, reporting IC₅₀ values in the nanomolar range (35 nM, 54 nM, 290 nM for GRK2; 32 nM for GRK3). These assays confirmed selectivity over GRK1 and GRK5 at concentrations up to 125 μM [2]
Off-target inhibition assays were conducted using pure enzymes. In these biochemical assays, 1 μM CMPD101 was reported to inhibit Rho kinase 2 activity by 47%, while inhibition of PKC, JNK, and MLCK was less than 20% or 10%, respectively [2]
Another study reported the IC₅₀ of CMPD101 for Rho kinase inhibition to be higher than 2 μM [2]

Western Blot Analysis[2]
Cell Types: HEK 293 cells stably expressing HA-tagged rat MOPr
Tested Concentrations: 3 μM, 30 μM
Incubation Duration: 30 minutes ago
Experimental Results: No effect on DAMGO-induced p-ERK1/2 and p- Elk-1 expression.

---

The effect of CMPD101 on β₂-adrenoceptor desensitization was studied in HEK293 cells. CMPD101 concentration-dependently inhibited β₂-adrenoceptor desensitization in a range of 3-30 μM [2]
In a subsequent study using cultured cells and rat brain slices, GRK2-mediated μ-opioid receptor phosphorylation was only partially inhibited by 3 μM CMPD101, but fully inhibited by 30 μM CMPD101. Concomitant analyses excluded that this was due to off-target inhibition such as Rho kinase 2 or SGK1 [2]

One experiment mentioned involves the isolated perfused rat heart. Rat hearts were perfused and treated with isoproterenol to induce βAR desensitization. The effect of CMPD-101 on preventing this desensitization was evaluated, likely by measuring functional parameters like contractility or cAMP levels.

The manuscript compared the nanomolar IC₅₀ values of CMPD101 for GRK2/3 inhibition under biochemical assay conditions with micromolar EC₅₀ values in intact cells (e.g., EC₅₀ values of 3–30 μM for inhibiting β₂-adrenergic receptor desensitization in HEK293 cells). This difference is attributed to the fact that the compound must cross barriers such as cell membranes in cellular or tissue systems [2].

In a broad selection screening, CMPD-101 exhibited low activity against HERG ion channels (9% inhibition at 1 µM and 31% inhibition at 10 µM), indicating a low potential risk of cardiovascular toxicity associated with HERG blockade. Its IC50 for CYP3A4 inhibition is >10 µM, suggesting a low likelihood of CYP3A4-mediated drug interactions at relevant concentrations.

[1]. Design, Synthesis, and Evaluation of the Highly Selective and Potent G-Protein-Coupled Receptor Kinase 2 (GRK2) Inhibitor for the Potential Treatment of Heart Failure. J Med Chem. 2017 Aug 24;60(16):6942-6990.

[2]. Inhibition of prostatic smooth muscle contraction by the inhibitor of G protein-coupled receptor kinase 2/3, CMPD101.Eur J Pharmacol. 2018 Jul 15;831:9-19.

CMPD101 has been described as the first commercially available small molecule inhibitor believed to be specific for GRK2/3[2]. This study proposes that CMPD101 can inhibit human prostate smooth muscle contraction induced by α₁-adrenergic, neurogenic (EFS), and non-adrenergic (endothelin-1, thromboxane A₂) stimulation. This broad-spectrum inhibitory effect contrasts with α₁ receptor blockers that target only adrenergic contraction[2]. The potential mechanism by which CMPD101 relaxes prostate smooth muscle appears to be independent of GRK inhibition, β₂-adrenergic receptor phosphorylation, and Rho kinase or MLCK inhibition, suggesting that the compound has unknown properties[2]. From a translational medicine perspective, CMPD101 may induce urodynamic effects in vivo and, due to its simultaneous action on both adrenergic and non-adrenergic contractile pathways, may be an ideal candidate for treating benign prostatic hyperplasia (BPH)-related lower urinary tract symptoms (LUTS). However, its application may be limited by the common side effects of kinase inhibitors and the lack of tolerability data [2].

C24H21F3N6O

466.458354711533

466.172

C, 61.80; H, 4.54; F, 12.22; N, 18.02; O, 3.43

865608-11-3

11677079

White to off-white solid powder

1.3±0.1 g/cm3

1.622

3.3

2

8

7

34

659

0

O=C(C1C=C(NCC2N(C)C(C3C=CN=CC=3)=NN=2)C=CC=1)NCC1C(C(F)(F)F)=CC=CC=1

WFOVEDJTASPCIR-UHFFFAOYSA-N

InChI=1S/C24H21F3N6O/c1-33-21(31-32-22(33)16-9-11-28-12-10-16)15-29-19-7-4-6-17(13-19)23(34)30-14-18-5-2-3-8-20(18)24(25,26)27/h2-13,29H,14-15H2,1H3,(H,30,34)

3-[[[4-Methyl-5-(4-pyridinyl)-4H-1,2,4-triazol-3-yl]methyl]amino]-N-[[2-(trifluoromethyl)phenyl]methyl]benzamide

Takeda101; CMPD101; Takeda-101; Takeda 101; CMPD 101; CMPD-101.

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

DMSO : ~250 mg/mL (~535.95 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (4.46 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 20.8 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: ≥ 2.08 mg/mL (4.46 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in 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 20.8 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: ≥ 2.08 mg/mL (4.46 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 20.8 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.)