Rapid Ex Vivo Visualization of Laryngeal Tumors Using a Sprayable pH-Activatable Fluorophore
10 affiliations
- 1Department of Otorhinolaryngology and Head and Neck Surgery, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands
- 2Department of Surgery, Leiden University Medical Center, 2333 ZA Leiden, The Netherlands
- 3Department of Otorhinolaryngology and Head and Neck Surgery, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands
- 4Department of Pathology, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands
- 5Centre for Human Drug Research, 2333 AL Leiden, The Netherlands
- 6Leiden Academic Center for Drug Research, Leiden University, 2300 RA Leiden, The Netherlands
- 7Department of Surgical Oncology and Gastrointestinal Surgery, Erasmus MC Cancer Institute, University Medical Center Rotterdam, Rotterdam, The Netherlands
- 8Gordon Center for Medical Imaging, Department of Radiology, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114 USA
- 9Center for Inflammation Imaging, Department of Radiology, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114 USA
- 10Department of Chemistry, Center of Diagnostics and Therapeutics, Georgia State University, Atlanta, GA 30303 USA
Abstract
Introduction: Achieving clear resection margins (> 5 mm) is critical during laryngeal and hypopharyngeal tumor resection, as positive margins are associated with reduced disease-specific survival. Near-infrared (NIR) fluorescence imaging has emerged as a valuable tool for intraoperative margin assessment by enabling real-time visualization of residual tumor tissue. However, current fluorescent contrast agents rely on intravenous administration, often requiring dosing hours to days before surgery. Also, targeting tumor-specific biomarkers with heterogeneous expression limits applicability. To address these limitations, we evaluated the feasibility of a topically applied pH-activatable fluorophore (PH10), which selectively fluoresces under acidic conditions. This ex vivo proof-of-concept study investigates whether topical PH10 can visualize laryngeal and hypopharyngeal tumors.
Methods: Freshly resected laryngeal and hypopharyngeal squamous cell carcinoma (SCC) specimens were obtained from ten patients and topically sprayed with PH10 immediately after surgery. Following a 2-min incubation period, NIR fluorescence images were acquired before and after three sequential saline washes. Fluorescence intensity was quantified as the tumor-to-background ratio (TBR); tumor localization was confirmed on histopathology.
Results: Laryngeal tumors demonstrated tumor-specific fluorescence upon PH10 application, with significantly higher signal intensity in tumor tissue compared to adjacent normal mucosa (P < 0.05). The median TBR after the first wash was 2.3 (interquartile range 1.7–3.0), exceeding the clinically relevant threshold of 1.5 in seven of seven laryngeal SCC cases; however, this threshold was not reached in either of the two hypopharyngeal carcinoma cases included.
Conclusions: This proof-of-concept study suggests that PH10, a sprayable pH-activatable fluorophore, can enable rapid and high-contrast visualization of laryngeal tumors, although this was not observed in the small number of hypopharyngeal cases evaluated. Given its topical application and rapid activation, PH10 may hold promise for intraoperative margin assessment, yet these preliminary findings, based on a small and heterogeneous sample, require validation in larger cohorts, including direct correlation with deep resection margins, before clinical utility can be established.
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BibTeX
@article{linders2026rapid,
title = {{Rapid Ex Vivo Visualization of Laryngeal Tumors Using a Sprayable pH-Activatable Fluorophore}},
author = {Daan G. J. Linders and Sebastiaan W. R. Dalmeijer and Bo E. Zweedijk and Hamed Abbasi and Danielle Boldewijn and Sjors A. Koppes and Bernd Kremer and Jacobus Burggraaf and Denise E. Hilling and Joost R. van der Vorst and Jinhui Ser and Satoshi Kashiwagi and Maged Henary and Jeroen C. Jansen and Alexander L. Vahrmeijer and Hak Soo Choi and Stijn Keereweer},
journal = {Molecular Diagnosis \& Therapy},
year = {2026},
month = sep,
doi = {10.1007/s40291-026-00878-x},
url = {https://doi.org/10.1007/s40291-026-00878-x},
note = {CC BY-NC 4.0},
}
RIS
TY - JOUR
TI - Rapid Ex Vivo Visualization of Laryngeal Tumors Using a Sprayable pH-Activatable Fluorophore
AU - Daan G. J. Linders
AU - Sebastiaan W. R. Dalmeijer
AU - Bo E. Zweedijk
AU - Hamed Abbasi
AU - Danielle Boldewijn
AU - Sjors A. Koppes
AU - Bernd Kremer
AU - Jacobus Burggraaf
AU - Denise E. Hilling
AU - Joost R. van der Vorst
AU - Jinhui Ser
AU - Satoshi Kashiwagi
AU - Maged Henary
AU - Jeroen C. Jansen
AU - Alexander L. Vahrmeijer
AU - Hak Soo Choi
AU - Stijn Keereweer
PY - 2026
DA - 2026/09/11
JO - Molecular Diagnosis & Therapy
DO - 10.1007/s40291-026-00878-x
UR - https://doi.org/10.1007/s40291-026-00878-x
AB - Introduction: Achieving clear resection margins (> 5 mm) is critical during laryngeal and hypopharyngeal tumor resection, as positive margins are associated with reduced disease-specific survival. Near-infrared (NIR) fluorescence imaging has emerged as a valuable tool for intraoperative margin assessment by enabling real-time visualization of residual tumor tissue. However, current fluorescent contrast agents rely on intravenous administration, often requiring dosing hours to days before surgery. Also, targeting tumor-specific biomarkers with heterogeneous expression limits applicability. To address these limitations, we evaluated the feasibility of a topically applied pH-activatable fluorophore (PH10), which selectively fluoresces under acidic conditions. This ex vivo proof-of-concept study investigates whether topical PH10 can visualize laryngeal and hypopharyngeal tumors. Methods: Freshly resected laryngeal and hypopharyngeal squamous cell carcinoma (SCC) specimens were obtained from ten patients and topically sprayed with PH10 immediately after surgery. Following a 2-min incubation period, NIR fluorescence images were acquired before and after three sequential saline washes. Fluorescence intensity was quantified as the tumor-to-background ratio (TBR); tumor localization was confirmed on histopathology. Results: Laryngeal tumors demonstrated tumor-specific fluorescence upon PH10 application, with significantly higher signal intensity in tumor tissue compared to adjacent normal mucosa (P < 0.05). The median TBR after the first wash was 2.3 (interquartile range 1.7–3.0), exceeding the clinically relevant threshold of 1.5 in seven of seven laryngeal SCC cases; however, this threshold was not reached in either of the two hypopharyngeal carcinoma cases included. Conclusions: This proof-of-concept study suggests that PH10, a sprayable pH-activatable fluorophore, can enable rapid and high-contrast visualization of laryngeal tumors, although this was not observed in the small number of hypopharyngeal cases evaluated. Given its topical application and rapid activation, PH10 may hold promise for intraoperative margin assessment, yet these preliminary findings, based on a small and heterogeneous sample, require validation in larger cohorts, including direct correlation with deep resection margins, before clinical utility can be established.
SN - 1177-1062
SN - 1179-2000
ER -