Ubiquitin-Specific Inhibitors for the Treatment of ... - ACS Publications

Nov 9, 2017 - Publication date: August 17, 2017. Priority Application: US 62/294,583. Priority date: February 12, 2016. Inventors: Guerin, D. J.; Bair...
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Ubiquitin-Specific Inhibitors for the Treatment of Cancers, Autoimmune, and Infectious Diseases Robert B. Kargbo* Usona Institute, 277 Granada Drive, San Luis Obispo, California 93401-7337, United States Patent Application Title: Patent Application Number: Priority Application: Inventors: Applicant(s): Disease Area: Summary:

Thienopyrazine Carboxamides as Ubiquitin-Specific Protease Inhibitors WO 2017139779 A1 Publication date: August 17, 2017 US 62/294,583 Priority date: February 12, 2016 Guerin, D. J.; Bair, K, W.; Caravella, J. A.; Ioannidis, S.; Lancia, D. R., Jr.; Li, H.; Mischke, S.; Ng, P.; Yee, R. D.; Schiller, S. E. R.; Shelekhin, T.; Wang, Z. Forma Therapeutics, Inc. Cancer, inflammation, autoimmune, and infectious diseases Biological Target: Ubiquitin USP28 and/or USP25 Ubiquitin is a protein consisting of 76 amino acids and is highly conserved throughout the tissues of most eukaryotic organisms. The addition of ubiquitin to protein substrates tags them for important biological processes such as degradation via proteasome or lysosomes, cellular activity, immune response, DNA repair, and so forth. In order to recycle ubiquitin molecules or edit polyubiquitin chains, deubiquitinating enzymes (DUBs) hydrolyze ubiquitin bound proteins and inhibit the proteasome degradation pathway. The cysteine proteases ubiquitin-specific proteases (USP25 and USP28) are paralogues of each other and have three distinct domains: an N-terminal, a pair of ubiquitin-interacting motifs (UIM), and a ubiquitin-specific protease (USP). USP25 and USP28 have several important cellular functions; the major role involves the regulation of the stability of cellular proteins. USP25 serves as a tumor-promoting factor and regulates cellular responses related to inflammation, autoimmune disease, infectious diseases, and so forth. However, USP28 serves to stabilize many oncoproteins and as a tumor-promoting factor. Many cancer types, including breast cancer, acute myeloid leukemia (AML), colorectal cancer, and ovarian cancer, involve amplification, deletions, and mutations of USP28. In addition, USP28 controls the stability of MYC, a transcription factor of genes involved in cell growth, proliferation, and apoptosis. There are studies that showed the knockdown of USP28 gene has led to a decrease of MYC protein and the inhibition of growth in human cell lines in vitro. Knockout of USP28 in mice has shown that its activity is not required for normal development and reproduction. Consequently, inhibition of USP25 and USP28 with small molecule inhibitors has potential in the treatment of cancers, inflammatory diseases, infectious diseases, autoimmune diseases, and so forth. The compounds of formula I in this patent application are described to possess inhibitory activity for USP25 and USP28 and may provide potential treatment for many of the above disorders, particularly in cancers, autoimmune diseases, and infectious diseases.

Important Compound Classes:

Received: October 28, 2017

© XXXX American Chemical Society

A

DOI: 10.1021/acsmedchemlett.7b00449 ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters

Patent Highlight

Key Structures:

Biological Assay:

Ubiquitin-Rhodamine 110 assay for USP28 Activity: The final volume of the assay buffer was 20 μL containing 20 mM Tris-HCl, 2 mM CaCl2, 2 mM BME (2-mercaptoethanol), 0.01% Prionex, and 0.01% Triton X-100. Enzyme USP28, construct USP28 (USP28-5(1−1077)-TEV-6*His), substrate (Ub-Rh110; Ubiquitin-Rhodamine 110), and fluorescence were read on Envision (excitation at 485 nm and emission at 535 nm) or on the PheraSTAR. Ubiquitin-Rhodamine 110 assay for USP25 Activity: The final volume of the assay buffer was 9 μL containing 20 mM Tris-HCl, 3 mM BME (2-mercaptoethanol), 0.03% BGG, and 0.01% Triton X-100. Enzyme USP25, construct USP25-His6, substrate (Ub-Rh110; Ubiquitin-Rhodamine 110), and fluorescence were read on the Envision or on the PheraSTAR.

Biological Data:

In the Table below; ++++ indicates IC50 < 0.2 μM, +++ indicates IC50 between 0.2 and 2 μM, and ++ indicates IC50 between 2 and 10 μM.

Claims:

48 Total claims 26 Composition of matter claims 22 Method of use claims

Recent Review Articles:

Kitazawa, S.; Yagi-Utsumi, M.; Kato, K.; Kitahara, R. Molecules 2017, 22, 1414. Ciana, A.; Achilli, C.; Minetti, G. Cell. Physiol. Biochem. 2017, 42, 1139. Morreale, F. E.; Testa, A.; Chaugule, V. K.; Bortoluzzi, A.; Ciulli, A.; Walden, H. J. Med. Chem. 2017, 60, 8183. Chen, H.; Wu, G.; Gao, S.; Guo, R.; Zhao, Z.; Yuan, H.; Liu, S.; Wu, J.; Lu, X.; Yuan, X.; Yu, Z.; Zu, X.; Xie, N.; Yang, N.; Hu, Z.; Sun, Q.; Zhang, W. J. Med. Chem. 2017, 60, 6828. 5. Hrdinka, M.; Gyrd-Hansen, M. Molecular Cell 2017, 68, 265.



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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Notes

The author declares no competing financial interest.

B

DOI: 10.1021/acsmedchemlett.7b00449 ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX