Lipid kinase activity

IC50 values were measured using a standard lipid kinase activity with PI as a substrate. The differences were that (i) 100 μM cold ATP was used instead of 10 μM, (ii) the DMSO concentration was 1% rather than 2%, and (iii) [γ-33P]ATP was used instead of [γ-32P]ATP. The TLC plates were quantified using a phosphorimager screen. The reported IC50 values were determined by non-linear regression analysis on the basis of at least three independent experiments repeated across multiple preparations of recombinant protein.

Cell lines

PC3 cells

Preparation method

The solubility of this compound in DMSO is >68.7mg/mL. General tips for obtaining a higher concentration: Please warm the tube at 37℃ for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below -20℃ for several months.

Reaction Conditions

0.2, 2 and 20 μM; 24 ~ 72 hrs

Applications

In PC3 cells, TGX-221 treatment (0.2, 2, and 20 μM) inhibited cell proliferation, and significantly reduced the activity of the p110β PI3K isoform.

Animal models

FeCl3-induced arterial thrombosis in mice

Dosage form

0.3 + 0.3, 1 + 1, 3 + 3 mg/kg + mg/kg/hr; i.v.

Applications

At the doses of 1 + 1 (49 % ± 13.9%) and 3 + 3 (88 % ± 10.6%), TGX-221 improved integrated blood flow over 30 mins in a mouse model. In addition, Tail bleeding time (BT) (sec) increased with TGX-221 doses of 3 + 3 (median 1560) and 1 + 1 (1305). In all TGX-221 groups, mean renal BT (sec) also increased.

Other notes

Please test the solubility of all compounds indoor, and the actual solubility may slightly differ with the theoretical value. This is caused by an experimental system error and it is normal.

TGX-221 is a potent inhibitor of (phosphatidylinositol 3-kinases) PI3K which specifically inhibits PI3K -p110β isoform with IC50 value of 8.5 nM [1].

In J774.2 macrophage cells, TGX-221 has been demonstrated to reduce insulin-induced phosphorylation of Ser473 of protein kinase B (PKB). While in in CHO-IR and 3T3-L1 cells, TGX-221 has no effect on PKB phosphprylation [1].

In vivo, TGX-221 significantly improved blood flow in FeCl3-induced arterial thrombosis as well as increased tail and renal bleeding times in mice. In addition, TGX-221 has revealed to disrupt CFRs in a Folts model of arterial thrombosis in male Sprague-Dawley rats [2].

Reference:
[1] Chaussade C1, Rewcastle GW, Kendall JD, Denny WA, Cho K, Gr nning LM, Chong ML, Anagnostou SH,Jackson SP, Daniele N, Shepherd PR.  Evidence for functional redundancy of class IA PI3K isoforms in insulin signalling. Biochem J. 2007 Jun 15;404(3):449-58.
[2] Bird JE1, Smith PL, Bostwick JS, Shipkova P, Schumacher WA.  Bleeding response induced by anti-thrombotic doses of a phosphoinositide 3-kinase (PI3K)-β inhibitor in mice. Thromb Res. 2011 Jun;127(6):560-4.