Thapsigargin

We then further examined the effect of imidazolin-4-one 3 on human neural stemcells in culture in order to establish whether it was also more potent than Cl-amidine in live cells. Cell death was induced in human neural cells by treatment with 5 μM thapsigargin, and their pre-treatment with different concentrations of imidazolin-4-one 3 and Cl-amidine was examined for their ability to antagonise cell death (Fig. 4). Thapsigargin-induced cell death was not altered by Cl-amidine at or below concentrations of 1 mM, and a significant, though partial, increase in cell survival was observed only at concentrations above 10 mM (Fig. 4). In marked contrast, imidazolin-4-one 3 maximally restored cell survival at 100 nM, and was highly effective at all the concentrations tested (up to 1 mM). The greater difference in potency between imidazolin-4-one 3 and Cl-amidine in the cell assay as compared to the enzymatic assay in protein extracts may be at least in part due to a higher concentration of the imidazolin-4-one 3 in the cell, given its greater lipophilicity.

Thapsigargin. The Mediterranean umbelliferous plant Thapsia garganica is the source of the sesquiterpene lactone thapsigargin (1, Table 1), which, in addition to tunicamycin and brefeldin A, is one of the most widely used and recognized ER stress inducers, albeit through a different mechanism of action. From a biochemical point of view, thapsigargin is an irreversible SERCA inhibitor (1, Table 1), thus having a profound impact upon Ca²⁺ homeostasis.

Therefore, we hypothesized that salvianolic acid A and salvianolic acid C may alter intracellular Ca²⁺ via an unknown target. To validate this hypothesis, we used thapsigargin and cyclopiazonic acid as the probe molecules. Thapsigargin is a non-competitive inhibitor of sarco/endoplasmic reticulum Ca²⁺ATPase (SERCA), while cyclopiazonic acid is a specific inhibitor of SERCA. Both inhibitors can raise the intracellular calcium concentration by blocking the ability of the cell to pump calcium into the sarcoplasmic and endoplasmic reticula.

Thapsigargin is a potent and selective inhibitor of sarco/endoplasmic reticulumCa²⁺ ATPases (SERCAs). As such, thapsigargin is capable of severely unbalancing cellular Ca²⁺ concentrations, often leading to disrupted cell function and growth, and apoptosis of the affected cell. Significantly, this has led to the development of a thapsigargin-derived prodrug for the treatment of prostate cancer.When tested in vivo, the prodrug was selectively cytotoxic to prostate tumours, whilst displaying minimal host toxicity. Following the recent publication of the first total synthesis of thapsigargin, and with a growing understanding of its structure–activity relationship (SAR), highly active analogues of the natural product with simplified structures are increasingly within reach of the synthetic chemist. Furthermore, owing to the difficulties in cultivating Thapsia (from which thapsigargin is harvested in relatively small quantities), total synthesis appears attractive as a means of obtaining thapsigargin and related analogues.