Triazole-derivatized near-infrared cyanine dyes enable local functional fluorescent imaging of ocular inflammation

Chloe N. Thomas; Nada Alfahad; Nicholas Capewell; Jamie Cowley; Eleanor Hickman; Antonio Fernandez; Neale Harrison; Omar S. Qureshi; Naomi Bennett; Nicholas M. Barnes; Andrew D. Dick; Colin J. Chu; Xiaoxuan Liu; Alastair K. Denniston; Marc Vendrell; Lisa J. Hill

doi:10.1016/j.bios.2022.114623

Triazole-derivatized near-infrared cyanine dyes enable local functional fluorescent imaging of ocular inflammation

Chloe N. Thomas, Nada Alfahad, Nicholas Capewell, Jamie Cowley, Eleanor Hickman, Antonio Fernandez, Neale Harrison, Omar S. Qureshi, Naomi Bennett, Nicholas M. Barnes, Andrew D. Dick, Colin J. Chu, Xiaoxuan Liu, Alastair K. Denniston, Marc Vendrell, Lisa J. Hill

Research output: Contribution to journal › Article › peer-review

Abstract / Description of output

Near-infrared (NIR) chemical fluorophores are promising tools for in-vivo imaging in real time but often succumb to rapid photodegradation. Indocyanine green (ICG) is the only NIR dye with regulatory approval for ocular imaging in humans; however, ICG, when employed for applications such as labelling immune cells, has limited sensitivity and does not allow precise detection of specific inflammatory events, for example leukocyte recruitment during uveitic flare-ups. We investigated the potential use of photostable novel triazole NIR cyanine
(TNC) dyes for detecting and characterising activated T-cell activity within the eye. Three TNC dyes were evaluated for ocular cytotoxicity in-vitro using a MTT assay and optimised concentrations for intraocular detection within ex-vivo porcine eyes after topical application or intracameral injections of the dyes. TNC labelled
T-cell tracking experiments and mechanistic studies were also performed in-vitro. TNC-1 and TNC-2 dyes exhibited greater fluorescence intensity than ICG at 10 μM, whereas TNC-3 was only detectable at 100 μM within the porcine eye. TNC dyes did not demonstrate any ocular cell toxicity at working concentrations of 10 μM.
CD4+T-cells labelled with TNC-1 or TNC-2 were detected within the porcine eye, with TNC-1 being brighter than TNC-2. Detection of TNC-1 and TNC-2 into CD4+T-cells was prevented by prior incubation with dynole 34–2 (50 μM), suggesting active uptake of these dyes via dynamin-dependent processes. The present study provides evi-dence that TNC dyes are suitable to detect activated CD4+T-cells within the eye with potential as a diagnostic marker for ocular inflammatory diseases.

Original language	English
Article number	114623
Number of pages	11
Journal	Biosensors and Bioelectronics
Volume	216
DOIs	https://doi.org/10.1016/j.bios.2022.114623
Publication status	Published - 13 Aug 2022

Keywords / Materials (for Non-textual outputs)

near-infrared
fluorophores
cyanine
uveitis
optical coherence tomography
leukocytes

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10.1016/j.bios.2022.114623

1-s2.0-S0956566322006637-main
This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
Final published version, 8.76 MBLicence: CC BY

https://linkinghub.elsevier.com/retrieve/pii/S0956566322006637

Dynamic Activatable Fluorophores
Vendrell Escobar, M.
EU government bodies
1/06/18 → 31/05/23
Project: Research

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Thomas, C. N., Alfahad, N., Capewell, N., Cowley, J., Hickman, E., Fernandez, A., Harrison, N., Qureshi, O. S., Bennett, N., Barnes, N. M., Dick, A. D., Chu, C. J., Liu, X., Denniston, A. K., Vendrell, M., & Hill, L. J. (2022). Triazole-derivatized near-infrared cyanine dyes enable local functional fluorescent imaging of ocular inflammation. Biosensors and Bioelectronics, 216, Article 114623. https://doi.org/10.1016/j.bios.2022.114623

@article{21c64180816c4594ba45d0ce6def4f1e,

title = "Triazole-derivatized near-infrared cyanine dyes enable local functional fluorescent imaging of ocular inflammation",

abstract = "Near-infrared (NIR) chemical fluorophores are promising tools for in-vivo imaging in real time but often succumb to rapid photodegradation. Indocyanine green (ICG) is the only NIR dye with regulatory approval for ocular imaging in humans; however, ICG, when employed for applications such as labelling immune cells, has limited sensitivity and does not allow precise detection of specific inflammatory events, for example leukocyte recruitment during uveitic flare-ups. We investigated the potential use of photostable novel triazole NIR cyanine(TNC) dyes for detecting and characterising activated T-cell activity within the eye. Three TNC dyes were evaluated for ocular cytotoxicity in-vitro using a MTT assay and optimised concentrations for intraocular detection within ex-vivo porcine eyes after topical application or intracameral injections of the dyes. TNC labelledT-cell tracking experiments and mechanistic studies were also performed in-vitro. TNC-1 and TNC-2 dyes exhibited greater fluorescence intensity than ICG at 10 μM, whereas TNC-3 was only detectable at 100 μM within the porcine eye. TNC dyes did not demonstrate any ocular cell toxicity at working concentrations of 10 μM. CD4+T-cells labelled with TNC-1 or TNC-2 were detected within the porcine eye, with TNC-1 being brighter than TNC-2. Detection of TNC-1 and TNC-2 into CD4+T-cells was prevented by prior incubation with dynole 34–2 (50 μM), suggesting active uptake of these dyes via dynamin-dependent processes. The present study provides evi-dence that TNC dyes are suitable to detect activated CD4+T-cells within the eye with potential as a diagnostic marker for ocular inflammatory diseases.",

keywords = "near-infrared, fluorophores, cyanine, uveitis, optical coherence tomography, leukocytes",

author = "Thomas, {Chloe N.} and Nada Alfahad and Nicholas Capewell and Jamie Cowley and Eleanor Hickman and Antonio Fernandez and Neale Harrison and Qureshi, {Omar S.} and Naomi Bennett and Barnes, {Nicholas M.} and Dick, {Andrew D.} and Chu, {Colin J.} and Xiaoxuan Liu and Denniston, {Alastair K.} and Marc Vendrell and Hill, {Lisa J.}",

note = "Funding Information: Chloe N Thomas and Lisa J Hill acknowledge funding from a Research Development Fund from the College of Medical and Dental Sciences at University of Birmingham . Antonio Fernandez acknowledges funding from Fundaci{\'o}n Seneca for a Saavedra Fajardo Grant ( 21124/SF/19 ). Marc Vendrell acknowledges funding from an ERC Consolidator Grant (DYNAFLUORS, 771443 ). Publisher Copyright: {\textcopyright} 2022 The Authors",

year = "2022",

month = aug,

day = "13",

doi = "10.1016/j.bios.2022.114623",

language = "English",

volume = "216",

journal = "Biosensors and Bioelectronics",

issn = "0956-5663",

publisher = "Elsevier",

}

Thomas, CN, Alfahad, N, Capewell, N, Cowley, J, Hickman, E, Fernandez, A, Harrison, N, Qureshi, OS, Bennett, N, Barnes, NM, Dick, AD, Chu, CJ, Liu, X, Denniston, AK, Vendrell, M & Hill, LJ 2022, 'Triazole-derivatized near-infrared cyanine dyes enable local functional fluorescent imaging of ocular inflammation', Biosensors and Bioelectronics, vol. 216, 114623. https://doi.org/10.1016/j.bios.2022.114623

TY - JOUR

T1 - Triazole-derivatized near-infrared cyanine dyes enable local functional fluorescent imaging of ocular inflammation

AU - Thomas, Chloe N.

AU - Alfahad, Nada

AU - Capewell, Nicholas

AU - Cowley, Jamie

AU - Hickman, Eleanor

AU - Fernandez, Antonio

AU - Harrison, Neale

AU - Qureshi, Omar S.

AU - Bennett, Naomi

AU - Barnes, Nicholas M.

AU - Dick, Andrew D.

AU - Chu, Colin J.

AU - Liu, Xiaoxuan

AU - Denniston, Alastair K.

AU - Vendrell, Marc

AU - Hill, Lisa J.

N1 - Funding Information: Chloe N Thomas and Lisa J Hill acknowledge funding from a Research Development Fund from the College of Medical and Dental Sciences at University of Birmingham . Antonio Fernandez acknowledges funding from Fundación Seneca for a Saavedra Fajardo Grant ( 21124/SF/19 ). Marc Vendrell acknowledges funding from an ERC Consolidator Grant (DYNAFLUORS, 771443 ). Publisher Copyright: © 2022 The Authors

PY - 2022/8/13

Y1 - 2022/8/13

N2 - Near-infrared (NIR) chemical fluorophores are promising tools for in-vivo imaging in real time but often succumb to rapid photodegradation. Indocyanine green (ICG) is the only NIR dye with regulatory approval for ocular imaging in humans; however, ICG, when employed for applications such as labelling immune cells, has limited sensitivity and does not allow precise detection of specific inflammatory events, for example leukocyte recruitment during uveitic flare-ups. We investigated the potential use of photostable novel triazole NIR cyanine(TNC) dyes for detecting and characterising activated T-cell activity within the eye. Three TNC dyes were evaluated for ocular cytotoxicity in-vitro using a MTT assay and optimised concentrations for intraocular detection within ex-vivo porcine eyes after topical application or intracameral injections of the dyes. TNC labelledT-cell tracking experiments and mechanistic studies were also performed in-vitro. TNC-1 and TNC-2 dyes exhibited greater fluorescence intensity than ICG at 10 μM, whereas TNC-3 was only detectable at 100 μM within the porcine eye. TNC dyes did not demonstrate any ocular cell toxicity at working concentrations of 10 μM. CD4+T-cells labelled with TNC-1 or TNC-2 were detected within the porcine eye, with TNC-1 being brighter than TNC-2. Detection of TNC-1 and TNC-2 into CD4+T-cells was prevented by prior incubation with dynole 34–2 (50 μM), suggesting active uptake of these dyes via dynamin-dependent processes. The present study provides evi-dence that TNC dyes are suitable to detect activated CD4+T-cells within the eye with potential as a diagnostic marker for ocular inflammatory diseases.

AB - Near-infrared (NIR) chemical fluorophores are promising tools for in-vivo imaging in real time but often succumb to rapid photodegradation. Indocyanine green (ICG) is the only NIR dye with regulatory approval for ocular imaging in humans; however, ICG, when employed for applications such as labelling immune cells, has limited sensitivity and does not allow precise detection of specific inflammatory events, for example leukocyte recruitment during uveitic flare-ups. We investigated the potential use of photostable novel triazole NIR cyanine(TNC) dyes for detecting and characterising activated T-cell activity within the eye. Three TNC dyes were evaluated for ocular cytotoxicity in-vitro using a MTT assay and optimised concentrations for intraocular detection within ex-vivo porcine eyes after topical application or intracameral injections of the dyes. TNC labelledT-cell tracking experiments and mechanistic studies were also performed in-vitro. TNC-1 and TNC-2 dyes exhibited greater fluorescence intensity than ICG at 10 μM, whereas TNC-3 was only detectable at 100 μM within the porcine eye. TNC dyes did not demonstrate any ocular cell toxicity at working concentrations of 10 μM. CD4+T-cells labelled with TNC-1 or TNC-2 were detected within the porcine eye, with TNC-1 being brighter than TNC-2. Detection of TNC-1 and TNC-2 into CD4+T-cells was prevented by prior incubation with dynole 34–2 (50 μM), suggesting active uptake of these dyes via dynamin-dependent processes. The present study provides evi-dence that TNC dyes are suitable to detect activated CD4+T-cells within the eye with potential as a diagnostic marker for ocular inflammatory diseases.

KW - near-infrared

KW - fluorophores

KW - cyanine

KW - uveitis

KW - optical coherence tomography

KW - leukocytes

U2 - 10.1016/j.bios.2022.114623

DO - 10.1016/j.bios.2022.114623

M3 - Article

SN - 0956-5663

VL - 216

JO - Biosensors and Bioelectronics

JF - Biosensors and Bioelectronics

M1 - 114623

ER -

Triazole-derivatized near-infrared cyanine dyes enable local functional fluorescent imaging of ocular inflammation

Abstract / Description of output

Keywords / Materials (for Non-textual outputs)

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Dynamic Activatable Fluorophores

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