Affinity-Enhanced Luminescent Re(I)- and Ru(II)-Based Inhibitors of the Cysteine Protease Cathepsin L.

Two new Re(I)- and Ru(II)-based inhibitors were synthesized with the formulas [Re(phen)(CO)3(1)](OTf) (7; phen = 1,10-phenanthroline, OTf = trifluoromethanesulfonate) and [Ru(bpy)2(2)](Cl)2 (8; bpy = 2,2'-bipyridine), where 1 and 2 are the analogues of CLIK-148, an epoxysuccinyl-based cysteine cathepsin L inhibitor (CTSL). Compounds 7 and 8 were characterized using various spectroscopic techniques and elemental analysis; 7 and 8 both show exceptionally long excited state lifetimes. Re(I)-based complex 7 inhibits CTSL in the low nanomolar range, affording a greater than 16-fold enhancement of potency relative to the free inhibitor 1 with a second-order rate constant of 211000 ± 42000 M-1 s-1. Irreversible ligation of 7 to papain, a model of CTSL, was analyzed with mass spectroscopy, and the major peak shown at 24283 au corresponds to that of papain-1-Re(CO)3(phen). Compound 7 was well tolerated by DU-145 prostate cancer cells, with toxicity evident only at high concentrations. Treatment of DU-145 cells with 7 followed by imaging via confocal microscopy showed substantial intracellular fluorescence that can be blocked by the known CTSL inhibitor CLIK-148, consistent with the ability of 7 to label CTSL in living cells. Overall this study reveals that a Re(I) complex can be attached to an enzyme inhibitor and enhance potency and selectivity for a medicinally important target, while at the same time allowing new avenues for tracking and quantification due to long excited state lifetimes and non-native element composition.

Total Synthesis of Tryprostatin B: Synthesis and Asymmetric Phase-Transfer-Catalyzed Reaction of Prenylated Gramine Salt.

A concise and efficient total synthesis of microtubule inhibitor tryprostatin B (1) is described. The key step is the preparation of a diprenylated gramine salt 9a. In this step, the prenyl group is incorporated at the 2-position of the indole moiety by direct lithiation of the Boc-protected gramine. We also developed and optimized the asymmetric phase-transfer-catalyzed reaction with salt 9a to provide the C2-prenyl tryptophan intermediate 2 resulting in 93% enantiomeric excess (ee) and 65% yield. The total synthesis of 1 is done in six steps with 35% overall yield.

Caging the uncageable: using metal complex release for photochemical control over irreversible inhibition.

Photochemical control over irreversible inhibition was shown using Ru(ii)-caged inhibitors of cathepsin L. Levels of control were dependent on where the Ru(ii) complex was attached to the organic inhibitor, reaching >10 : 1 with optimal placement. A new strategy for photoreleasing Ru(ii) fragments from inhibitor-enzyme conjugates is also reported.