Fig. 1. Alexidine dihydrochloride is a selective inhibitor of PTPMT1. A, structure of alexidine dihydrochloride. B, inhibition of selected phosphatases by alexidine dihydrochloride. PTPMT1, VHR phosphatase, ��-Ppase, … Having established alexidine dihydrochloride as Nutlin-3a 675576-98-4 an inhibitor of PTPMT1, we then investigated whether the inhibition was selective toward this protein phosphatase. Hence, the capacity of the compound to inhibit a variety of related enzymes, including a protein serine/threonine phosphatase, �� protein phosphatase; the classical, tyrosine-specific PTP, T-cell protein tyrosine phosphatase; and the dual-specificity PTP, VH1-related phosphatase and PTEN, under optimal buffer and pH conditions for each enzyme, was assessed. Importantly, none of these protein phosphatases was appreciably inhibited by alexidine dihydrochloride (Fig.

1). The result with PTEN was particularly noteworthy because the sequence of the catalytic motif of the active site of PTEN formed the basis for the bioinformatics screen that led to the discovery of PTPMT1, and the two enzymes differ in this sequence by just two amino acids (Pagliarini et al., 2004). The Dibiguanide Structure of Alexidine Dihydrochloride is Important for Its Inhibition of PTPMT1. Having established alexidine dihydrochloride as a selective inhibitor of PTPMT1, we were keen to determine the structural features of the compound that contribute to this inhibition. The striking duplication in the structure of alexidine dihydrochloride prompted us to investigate whether this binate aspect contributed to the ability of the compound to inhibit PTPMT1, and whether other biguanide and dibiguanide compounds also inhibited PTPMT1.

Interestingly, we found that the dibiguanide compound chlorhexidine dihydrochloride significantly inhibited PTPMT1, albeit with a log-fold higher IC50 compared with alexidine dihydrochloride; the IC50 determined for the chlorhexidine analog was 19.7 �� 3.3 ��M (Fig. 2). In contrast, a monobiguanide compound comprising half of the alexidine dihydrochloride structure inhibited PTPMT1 very weakly with an IC50 of 207 �� 43 ��M and an accompanying Hill coefficient close to 1 (Fig. 2). In addition, two other monobiguanide compounds, the type II diabetes drugs metformin and phenformin, did not significantly inhibit PTPMT1 (Fig. 2).

Thus, the dibiguanide structure conferred a significantly higher level of potency of inhibition of PTPMT1 compared Batimastat with the monobiguanide structure, and the dibiguanide structure appeared to be important for the cooperativity observed in the Hill slope with alexidine dihydrochloride. Fig. 2. The dibiguanide structure of alexidine dihydrochloride is important for its inhibition of PTPMT1. A, inhibition of PTPMT1 by various biguanide and dibiguanide compounds using O-MFP as the substrate. … Inhibition of PTPMT1 by Alexidine Dihydrochloride Is Predominantly Uncompetitive.