Kinetic characterization of thiosemicarbazones as cysteine protease inhibitors and their potential use as therapeutic agents against metastatic cancer and Chagas' disease.




Chavarria Nolasco, Gustavo E.

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Human cathepsins L and K are cysteine proteases that play important roles in physiological and pathological processes, such as cancer metastasis, bone resorption and neurodegenerative diseases. This research has focused on the evaluation of synthetic thiosemicarbazones that could inhibit their proteolytic activities with the objective of preventing cancer metastasis. In addition, cruzain, a cathepsin L-like cysteine protease found in Trypanosoma cruzi, is a validated target for pivotal roles in the parasitic invasion in Chagas' disease, a condition that could be fatal if not treated. Currently, there is no effective treatment against the disease, which is rapidly extending to non-endemic areas in the United States and Europe. More than 150 synthetic thiosemicarbazones (obtained through a collaborative study) were tested against cathepsins L, K and cruzain. This work presents preliminary in vitro analysis of these compounds in order to characterize their potency and mode of inhibition. A number of potent inhibitors was found for each enzyme. A smaller subset of thiosemicarbazones were found to be selective. Results showed that compound 1, one of the most potent inhibitors in this library is a slow binding, slowly reversible, competitive inhibitor of these targets. Furthermore, 1 was able to delay and partially inhibit the activation of procathepsin K under acidic conditions. Similar results were found with 3-bromo-3'-hydroxybenzophenone thiosemicarbazone (8). Compounds 8 and 1 inhibited the in vitro type I collagenase activity of cathepsin L in a time-dependent manner and type IV collagenase activity of cathepsin K. Analogs, 1, 8, 156, 157, and 168 were also used in cell studies. These compounds were able to delay cell migration and cell invasion in MDA-MB-231 cells, a type of carcinoma breast cancer cell line. It was determined that cell invasion and cell migration were inhibited in a concentration dependent manner. Lastly, analog 17 was also found to be a slow reversible, slow binding, competitive inhibitor of cruzain. This compound was also able to inhibit the collagenase activity of recombinant cruzain when human type I collagen was used as a substrate. Kinetic studies and molecular modeling indicate the best thiosemicarbazone inhibitors form a reversible covalent bond with each enzyme.



Cathepsin L inhibitor., Cathepsin K inhibitor., Cruzain inhibitor., Enzyme inhibition., Thiosemicarbazone., Cell invasion., MDA-MB-231 breast cancer cells., Slow-binding inhibitor., Collagenase inhibition.


Chavarria, Gustavo. "Synthesis and biochemical evaluation of a series of non-peptidic thiochromanone thiosemicarbazone analogues as cathepsin L and B inhibitors." ACS Med. Chem. Lett. 3 (2012): 450–53.
Chavarria, Gustavo. "Functionalized benzophenone, thiophene, pyridine, and fluorene thiosemicarbazone derivatives as inhibitors of cathepsin L." Bioorganic & Medicinal Chemistry Letters 20 (2010): 6610-15.
Chavarria, Gustavo. "Design, synthesis, and biological evaluation of potent thiosemicarbazone based cathepsin L inhibitors." Bioorganic & Medicinal Chemistry Letters 20 (2010): 1415-19.