Ex Vivo Human Tissue Functions as a Testing Platform for the Evaluation of a Nerve-Specific Fluorophore.

SIGNIFICANCE

Selecting a nerve-specific lead fluorescent agent for translation in fluorescence-guided surgery is time-consuming and expensive. Preclinical fluorescent agent studies rely primarily on animal models, which are a critical component of preclinical testing, but these models may not predict fluorophore performance in human tissues.

AIM

The primary aim of this study was to evaluate and compare two preclinical models to test tissue-specific fluorophores based on discarded human tissues. The secondary aim was to use these models to determine the ability of a molecularly targeted fluorophore, LGW16-03, to label ex vivo human nerve tissues.

APPROACH

Patients undergoing standard-of-care transtibial or transfemoral amputation were consented and randomized to topical or systemic administration of LGW16-03 following amputation. After probe administration, nerves and background tissues were surgically resected and imaged to determine nerve fluorescence signal-to-background tissue ratio (SBR) and signal-to-noise ratio (SNR) metrics. Analysis of variance (ANOVA) determined statistical differences in metric means between administration cohorts and background tissue groups. Receiver operating characteristic (ROC) curve-derived statistics quantified the discriminatory performance of LGW16-03 fluorescence for labeling nerve tissues.

RESULTS

Tissue samples from 18 patients were analyzed. Mean nerve-to-adipose SBR was greater than nerve-to-muscle SBR (p = 0.001), but mean nerve-to-adipose SNR was not statistically different from mean nerve-to-muscle SNR (p = 0.069). Neither SBR nor SNR means were statistically different between fluorophore administration cohorts (p ≥ 0.448). When administration cohorts were combined, nerve-to-adipose SBR was greater than nerve-to-muscle SBR (mean ± standard deviation; 4.2 ± 2.9 vs. 1.8 ± 1.9; p < 0.001), but SNRs for nerve-to-adipose and nerve-to-muscle were not significantly different (5.1 ± 4.0 vs. 3.1 ± 3.4; p = 0.055). ROC curve-derived statistics to quantify LGW16-03 nerve labeling performance varied widely between patients, with sensitivities and specificities ranging from 0.2-99.9% and 0.4-100.0%.

CONCLUSION

Systemic and topical administration of LGW16-03 yielded similar fluorescence labeling of nerve tissues. Both administration approaches provided nerve-specific contrast similar to that observed in preclinical animal models. Fluorescence contrast was generally higher for nerve-to-adipose versus nerve-to-muscle. Ex vivo human tissue models provide safe evaluation of fluorophores in the preclinical phase and can aid in the selection of lead agents prior to first-in-human trials.