TY - JOUR
T1 - Hydrolysis-Resistant Ester-Based Linkers for Development of Activity-Based NIR Bioluminescence Probes
AU - Yadav, Anuj K.
AU - Zhao, Zhenxiang
AU - Weng, Yourong
AU - Gardner, Sarah H.
AU - Brady, Catharine J.
AU - Pichardo Peguero, Oliver D.
AU - Chan, Jefferson
N1 - C.J.B. acknowledges the Chemistry-Biology Interface Training Grant (T32-GM136629) and previous support from the Robert C. and Carolyn J. Springborn Graduate Fellowship. SHG thanks the Cancer Center at Illinois for a Graduate Student Cancer Scholarship. J.C. thanks the Helen Corley Petit Scholar Program and the Camille and Henry Dreyfus Foundation. Major funding for the 500 MHz Bruker CryoProbe was provided by the Roy J. Carver Charitable Trust (Muscatine, Iowa; Grant No. 15-4521) to the School of Chemical Sciences NMR Lab. The Q-Tof Ultima mass spectrometer was purchased in part with a grant from the National Science Foundation, Division of Biological Infrastructure (DBI-0100085). We also acknowledge Dr. Iwona Dobrucka and the Molecular Imaging Laboratory at the Beckman Institute for use of the IVIS imaging system. This work was supported by the National Institutes of Health (R35GM133581) and in part by a grant awarded to J.C. from the Chemistry Discovery Fund established by Ving & May Lee.
This work was supported by the National Institutes of Health (R35GM133581) and in part by a grant awarded to J.C. from the Chemistry Discovery Fund established by Ving & May Lee.
C.J.B. acknowledges the Chemistry-Biology Interface Training Grant (T32-GM136629) and previous support from the Robert C. and Carolyn J. Springborn Graduate Fellowship. SHG thanks the Cancer Center at Illinois for a Graduate Student Cancer Scholarship. J.C. thanks the Helen Corley Petit Scholar Program and the Camille and Henry Dreyfus Foundation. Major funding for the 500 MHz Bruker CryoProbe was provided by the Roy J. Carver Charitable Trust (Muscatine, Iowa; Grant No. 15-4521) to the School of Chemical Sciences NMR Lab. The Q-Tof Ultima mass spectrometer was purchased in part with a grant from the National Science Foundation, Division of Biological Infrastructure (DBI-0100085). We also acknowledge Dr. Iwona Dobrucka and the Molecular Imaging Laboratory at the Beckman Institute for use of the IVIS imaging system.
PY - 2023/1/18
Y1 - 2023/1/18
N2 - Activity-based sensing (ABS) probes equipped with a NIR bioluminescence readout are promising chemical tools to study cancer biomarkers owing to their high sensitivity and deep tissue compatibility. Despite the demand, there is a dearth of such probes because NIR substrates (e.g., BL660 (a NIR luciferin analog)) are not equipped with an appropriate attachment site for ABS trigger installation. For instance, our attempts to mask the carboxylic acid moiety with standard self-immolative benzyl linkers resulted in significant background signals owing to undesirable ester hydrolysis. In this study, we overcame this longstanding challenge by rationally designing a new hydrolysis-resistant ester-based linker featuring an isopropyl shielding arm. Compared to the parent, the new design is 140.5-fold and 67.8-fold more resistant toward spontaneous and esterase-mediated hydrolysis, respectively. Likewise, we observed minimal cleavage of the ester moiety when incubated with a panel of enzymes possessing ester-hydrolyzing activity. These impressive in vitro results were corroborated through a series of key experiments in live cells. Further, we showcased the utility of this technology by developing the first NIR bioluminescent probe for nitroreductase (NTR) activity and applied it to visualize elevated NTR expression in oxygen deficient lung cancer cells and in a murine model of non-small cell lung cancer. The ability to monitor the activity of this key biomarker in a deep tissue context is critical because it is associated with tumor hypoxia, which in turn is linked to drug resistance and aggressive cancer phenotypes.
AB - Activity-based sensing (ABS) probes equipped with a NIR bioluminescence readout are promising chemical tools to study cancer biomarkers owing to their high sensitivity and deep tissue compatibility. Despite the demand, there is a dearth of such probes because NIR substrates (e.g., BL660 (a NIR luciferin analog)) are not equipped with an appropriate attachment site for ABS trigger installation. For instance, our attempts to mask the carboxylic acid moiety with standard self-immolative benzyl linkers resulted in significant background signals owing to undesirable ester hydrolysis. In this study, we overcame this longstanding challenge by rationally designing a new hydrolysis-resistant ester-based linker featuring an isopropyl shielding arm. Compared to the parent, the new design is 140.5-fold and 67.8-fold more resistant toward spontaneous and esterase-mediated hydrolysis, respectively. Likewise, we observed minimal cleavage of the ester moiety when incubated with a panel of enzymes possessing ester-hydrolyzing activity. These impressive in vitro results were corroborated through a series of key experiments in live cells. Further, we showcased the utility of this technology by developing the first NIR bioluminescent probe for nitroreductase (NTR) activity and applied it to visualize elevated NTR expression in oxygen deficient lung cancer cells and in a murine model of non-small cell lung cancer. The ability to monitor the activity of this key biomarker in a deep tissue context is critical because it is associated with tumor hypoxia, which in turn is linked to drug resistance and aggressive cancer phenotypes.
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U2 - 10.1021/jacs.2c12984
DO - 10.1021/jacs.2c12984
M3 - Article
C2 - 36603103
AN - SCOPUS:85145987444
SN - 0002-7863
VL - 145
SP - 1460
EP - 1469
JO - Journal of the American Chemical Society
JF - Journal of the American Chemical Society
IS - 2
ER -