LMPTP INHIBITOR 1 hydrochloride is a selective inhibitor of low molecular weight protein tyrosine phosphatase ( LMPTP ), with an IC 50 of 0.8 μM for LMPTP-A.

LMPTPA:0.8 μM

LMPTP INHIBITOR 1 (hydrochloride) 是一种对低分子量蛋白质酪氨酸磷酸酶具有选择性抑制作用的抑制剂，对LMPTP-A的IC 50值为0.8 μM。LMPTP INHIBITOR 1 对LMPTP-A的抑制效果较之于LMPTP-B更为强大。LMPTP抑制剂1还在人类HepG2肝细胞中，通过胰岛素刺激后增强了HepG2 IR的磷酸化[1]。

LMPTP inhibitor 1 具有口服生物可用性，0.03% w/w 的处理约产生 680 nM 的平均血清浓度，而0.05% w/w 的处理则产生 >3 μM；该物质还能逆转肥胖小鼠的糖尿病。LMPTP inhibitor 1（0.05% w/w）抑制 LMPTP 活性，显著改善糖尿病 DIO 小鼠的葡萄糖耐受性并降低空腹胰岛素水平，但不影响体重[1]。

Phosphatase assays are performed in buffer containing 50 mM Bis-Tris, pH 6.0, 1 mM DTT and 0.01% Triton X-100 at 37°C. For assays conducted with 3-O-methylfluorescein phosphate (OMFP) as substrate, fluorescence is monitored continuously at λex = 485 and λem = 525 nm. For assays conducted with para-nitrophenyl phosphate (pNPP) as substrate, the reaction is stopped by addition of 2X reaction volume of 1 M NaOH, and absorbance is measured at 405 nm. IC50 values are determined from plots of LMPTP inhibitor 1 concentration versus percentage of enzyme activity. For inhibitor selectivity assays, each PTP is incubated with either 0.4 mM OMFP or 5 mM pNPP in the presence of 40 μM LMPTP inhibitor 1 or DMSO. Equal units of enzyme activity, comparable to the activity of 10 nM human LMPTP-A, are used. For the inhibitor reversibility assay, 50 nM human LMPTP-A is pre-incubated with 10 μM LMPTP inhibitor 1 or DMSO for 5 min. The enzyme is diluted 100X in phosphatase assay buffer containing 0.4 mM OMFP and fluorescence is measured at the indicated time points [1].

Human HepG2 cells are cultured in Eagle's Minimal Essential Medium (ATCC) containing 10% fetal bovine serum (FBS), 100 U/mL penicillin and 100 μg/mL streptomycin. The absence of Mycoplasma contamination in HepG2 cultures is confirmed using the Lonza MycoAlert Mycoplasma Detection Kit. Cells are treated with 10 μM LMPTP inhibitor 1 in serum-starvation media (0.1% FBS) overnight, following which cells are stimulated with 10 nM bovine insulin for 5 min at 37°C. For detection of IR tyrosine phosphorylation by immunoprecipitation/Western blotting, cells are lysed in radioimmunoprecipitation assay buffer containing 1 mM phenylmethylsulfonyl fluoride, 10 μg/mL aprotinin/leupeptin, 10 mM sodium orthovanadate, 5 mM sodium fluoride, and 2 mM sodium pyrophosphate, and the IR is immunoprecipitated using the anti-IRβ Ab. IR tyrosine phosphorylation of immunoprecipitates is determined by Western blotting with the anti-pIR/pIGFR-Y1162/Y1163 Ab [1].

LMPTP inhibitor 1 is administered to male B6 or Acp1fl/fl albumin-Cre+ DIO mice at 0.05% w/w in high-fat diet (HFD) rodent chow. Control groups consist of male B6 or Acp1fl/fl albumin-Cre+ littermate mice administered HFD rodent chow alone. Mice are allowed food and water ad libitum and weighed daily. Randomization is not used in these experiments; rather littermate mice are assigned to treatment or control groups in a manner to maintain similar mean body weights between the 2 groups at the start of the study. Insulin-induced liver IR phosphorylation, IPGTT, and fasting insulin levels are assessed after treatment. Diabetic (displaying overnight [13 hr] fasting blood glucose levels ≥140 mg/dL) B6 DIO mice are used in experiments to assess IPGTT and fasting insulin levels [1].