Wang, W. , Vellaisamy, K. , Li, G. , Wu, C. , Ko, C. N. , Leung, C. H Wang, W., Vellaisamy, K., Li, G., Wu, C., Ko, C. N., Leung, C. H., & Ma, D. L. (2017). Development of a long-lived luminescence probe for visualizing β-galactosidase in ovarian carcinoma cells. Analytical Chemistry, 89(21), 11679-11684. Jingtao Zhang

Introduction Ovarian cancer: 161,000 death The seventh-most common cancer Physical examination Blood test Ultrasound Rossing, M. A., Wicklund, K. G., Cushing-Haugen, K. L., & Weiss, N. S. (2010). Predictive value of symptoms for early detection of ovarian cancer. Journal of the National Cancer Institute, 102(4), 222-229. Ovarian cancer's early stages (I/II) are difficult to diagnose because most symptoms are nonspecific and thus of little use in diagnosis; as a result, it is rarely diagnosed until it spreads and advances to later stages (III/IV).

Background -------Screening CA125 Β-Gal (High sensitivity) Low specificity Worse specificity May not increase at early stage[1] strong and positive correlations CA125 is a protein as a biomarker for some cancers. It may also be elevated in other cancers, including endometrial cancer, fallopian tube cancer, lung cancer, breast cancer and gastrointestinal cancer. Β-Gal is a glycoside hydrolase enzyme that catalyzes the hydrolysis of β-galactosides. Chatterjee, S. K., Bhattacharya, M., & Barlow, J. J. (1979). Glycosyltransferase and glycosidase activities in ovarian cancer patients. Cancer research, 39(6 Part 1), 1943-1951.

Background Background ------- β-Gal Colorimetric assays, β-Gal has also been demonstrated as an important biomarker for cell senescence and primary ovarian cancer. Therefore, methods for the sensitive detection of β-gal could aid in the monitoring of ovarian cancer development. Colorimetric assays, Immunostaining assays, Histochemical methods. Low sensitivity, Tedious sample preparation, High cost.

Background -------Luminescence imaging Tung, C. H., Zeng, Q., Shah, K., Kim, D. E., Schellingerhout, D., & Weissleder, R. (2004). In vivo imaging of β-galactosidase activity using far red fluorescent switch. Cancer research, 64(5), 1579-1583. Kamiya, M., Asanuma, D., Kuranaga, E., Takeishi, A., Sakabe, M., Miura, M., ... & Urano, Y. (2011). β-Galactosidase fluorescence probe with improved cellular accumulation based on a spirocyclized rhodol scaffold. Journal of the American Chemical Society, 133(33), 12960-12963. Sakabe, M., Asanuma, D., Kamiya, M., Iwatate, R. J., Hanaoka, K., Terai, T., ... & Urano, Y. (2012). Rational design of highly sensitive fluorescence probes for protease and glycosidase based on precisely controlled spirocyclization. Journal of the American Chemical Society, 135(1), 409-414.

Background Fluorescent probes are limited by -------Luminescence imaging Fluorescent probes are limited by the high endogenous fluorescence of biological samples. Asanuma, D., Sakabe, M., Kamiya, M., Yamamoto, K., Hiratake, J., & Ogawa, M., et al. (2015). Sensitive β-galactosidase-targeting fluorescence probe for visualizing small peritoneal metastatic tumours in vivo. Nature Communications, 6, 6463.

Background Background -------phosphorescence https://commons.wikimedia.org/wiki/File:Electronic_Processes_Involving_Light.png limit interference between excitation and emission

Results & Discussion -------characterization Structural Confirmation Stability Exam Luminescent Spectra(TRES) High-resolution Mass Spectra Confocal

Results & Discussion -------Stability large Stokes shift

Results & Discussion Results & Discussion -------Spectra selectivity of complex 1 for β-gal To obtain the optimal performance of the probe

Results & Discussion Results & Discussion -------TRES A good linear relationship

Results & Discussion -------High-resolution Mass Results & Discussion

Results & Discussion Results & Discussion MLCT -------Mechanism 3 MLCT Complex 2 is nonemissive due to nonradiative interactions with the solvent leading to luminescence quenching. However, β-gal cleaves the galactose moiety of complex 1 to generate the dealkylated product 2, which binds to β-gal. Upon binding to β-gal, the complex is protected from the bulk solvent and hence becomes emissive via the triplet metal-to-ligand charge-transfer (3MLCT) excited state. synthetically prepared complex 2 showed little luminescence

Results & Discussion Results & Discussion -------Mechanism Typical pathways for cyclo-metalated Ir (III) complexes photochemistry You, Y., & Nam, W. (2012). Photofunctional triplet excited states of cyclometalated Ir (III) complexes: beyond electroluminescence. Chemical Society Reviews, 41(21), 7061-7084.

Results & Discussion -------Confocal Results & Discussion

Conclusion -------Innovation 1. First phosphorescent probe for ovarian cancer; 2. Good compatibility with reasonable stability in PBS; 3. Sensitivity Asanuma, D., Sakabe, M., Kamiya, M., Yamamoto, K., Hiratake, J., Ogawa, M., ... & Urano, Y. (2015). Sensitive β-galactosidase-targeting fluorescence probe for visualizing small peritoneal metastatic tumours in vivo. Nature communications, 6.

Conclusion Conclusion -------Shortcomings BT474 Maximum Excitation Wavelength

Conclusion -------Shortcomings They did not preclude the possibility that complex 2 may also bind to other proteins.

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