Structure–Activity Relationships of Potent, Targeted Covalent Inhibitors That Abolish Both the Transamidation and GTP Binding Activities of Human Tissue Transglutaminase

Abstract

Human tissue transglutaminase (hTG2) is a multifunctional enzyme. It is primarily known for its calcium-dependent transamidation activity that leads to formation of an isopeptide bond between glutamine and lysine residues found on the surface of proteins, but it is also a GTP binding protein. Overexpression and unregulated hTG2 activity have been associated with numerous human diseases, including cancer stem cell survival and metastatic phenotype. Herein, we present a series of targeted covalent inhibitors (TCIs) based on our previously reported Cbz-Lys scaffold. From this structure-activity relationship (SAR) study, novel irreversible inhibitors were identified that block the transamidation activity of hTG2 and allosterically abolish its GTP binding ability with a high degree of selectivity and efficiency (k/K > 10 M min). One optimized inhibitor (VA4) was also shown to inhibit epidermal cancer stem cell invasion with an EC of 3.9 μM, representing a significant improvement over our previously reported "hit" NC9.

Benefits of Combined All-Trans Retinoic Acid and Arsenic Trioxide Treatment of Acute Promyelocytic Leukemia Cells and Further Enhancement by Inhibition of Atypically Expressed Transglutaminase 2

“…NC9 is a TG2-selective irreversible inhibitor that blocks the enzyme active site and locks TG2 in an open conformation, thereby abrogating GTP binding as well. [11,17]. Our results showed that inhibition of TG2 by NC9 in ATRA + ATO combined treatment significantly reduces not just reactive oxygen species (ROS) but also proinflammatory cytokines and chemokine production that significantly increases the severity of DS.…”

“…The pro-apoptotic effects of TG2 are associated with the calcium-dependent protein-protein cross-linking activity at high calcium levels. In its survival-promoting attribution, TG2 most often participates as a protein-protein interaction partner in GTP-bound form [17,18]. In earlier studies, others tested a novel irreversible TG2 inhibitor NC9 to identify agents that can selectively target cancer stem cells in tumors [13,19,20].…”

A Precision Strategy to Cure Renal Cell Carcinoma by Targeting Transglutaminase 2

“…The Keillor group recently reported the TGase 2 inhibitor VA4, optimized from hit compound NC9. This inhibitor blocks the transamidation activity of TGase 2, as well as its GTP binding ability, through an allosteric mechanism [72]. VA4 was reported to be the best lead compound among the sulfonamide and piperazinyl amide derivatives designed to interact with the active site of TGase 2 [72].…”

“…This inhibitor blocks the transamidation activity of TGase 2, as well as its GTP binding ability, through an allosteric mechanism [72]. VA4 was reported to be the best lead compound among the sulfonamide and piperazinyl amide derivatives designed to interact with the active site of TGase 2 [72]. We investigated whether these specific irreversible inhibitors for TGase 2 might also block TGase 2-p53 binding and p53 depletion in RCC cells.…”

Transglutaminase 2 takes center stage as a cancer cell survival factor and therapy target

“…Although direct measurements of TG2 intracellular conformation have not been performed in unperturbed gastric cancer cells, indirect data suggests that TG2 exists in the closed GTP bound form in these cells where it is expected to drive cancer cell survival signaling (Figure A). This suggests that TG2 inhibitors, which suppress GTP binding by indirectly disordering the TG2 GTP binding site, may be a useful treatment for gastric cancer.…”