Introduction

Locally advanced esophageal cancer (EC) is treated with neoadjuvant chemo(radio)therapy (nCRT) followed by surgery, but only a modest percentage of patients (16-43%) achieves a pathological complete response post-nCRT. The addition of immune checkpoint inhibition (ICI) targeting programmed death-1 or its ligand (PD-1/PD-L1) is explored to increase response rates, yet adequate patient selection methods are lacking. PD-L1 expression’s predictive value in biopsies is inconsistent due to intra- and intertumoral heterogeneity, and the influence of nCRT on ICI response is unclear. Novel imaging methods are crucial to understand PD-L1 heterogeneity in order to ultimately select patients for ICI.

Aims & Methods

This ongoing study aims to assess the safety and feasibility of the innovative technique ultrasound guided quantitative fluorescence molecular endoscopy (US-qFME) using the fluorescently labelled ICI drug durvalumab-680LT before and after nCRT in EC patients to visualize, for the first time, PD-L1 drug distribution from the macroscopic to the cellular level. A durvalumab-680LT dose-optimization was performed in which up to now fifteen EC patients scheduled for nCRT were included in either the control (not receiving durvalumab-680LT), 4.5 mg, or 15 mg cohort. The next five patients will receive 25 mg durvalumab-680LT, after which the most optimal dose group will be expanded with 16 patients. US-qFME procedures were performed both before and after nCRT, consisting of in vivo fluorescence signal visualization, quantification by mucosal and ultrasound-guided spectroscopy measurements of healthy esophageal tissue, tumor and/or lymph nodes, and collection of biopsies. Subsequently, ex vivo analyses were performed for quantification of fluorescence signals, correlation to histology and visualization of tissue drug distribution and drug target cells.

Results

Higher fluorescence signals were visualized in vivo in tumor compared to healthy tissue. Ex vivo quantification of the fluorescence signals revealed higher median fluorescence signal intensities in tumor biopsies compared to healthy esophageal tissue pre-nCRT in both the 4.5 mg and 15 mg dose cohorts with a broad range in signals in the tumor (94.8 a.u. [41.0-131.7] vs. 31.0 a.u. [24.6-38.1] (p=0.125), and 103.9 a.u. [40.7-197.6] vs. 27.1 a.u. [25.8-40.8] (p=0.0625), respectively). Similar results were found during both mucosal and ultrasound-guided spectroscopy with higher signals in tumor tissue both in vivo and ex vivo compared to healthy tissue. The signals in healthy tissues of patients receiving the tracer were comparable to both tumor and healthy tissue in the control cohort (26.2 a.u. [21.5-37.6] and 11.7 a.u. [11.3-19.0]). An SDS-PAGE experiment confirmed the tracer’s integrity and stability in the mucosa. During fluorescence microscopy, only a dose of 15 mg durvalumab-680LT was sufficient to visualize membrane-bound durvalumab. Analyses post-nCRT are still ongoing. No (serious) adverse events occurred.

Conclusion

Durvalumab-680LT administration was safe and specifically targeted the tumor tissue as higher signals were found in tumor than in healthy tissue. In addition, the broad range in signals in the tumor tissue indicates, as expected, an inter- and intra-patient difference in PD-L1 expression. These preliminary findings show the potential of the novel combination of durvalumab-680LT and US-qFME to guide future patient selection for ICI therapy. Further analyses are ongoing, including correlation to PD-L1 staining and fluorescence microscopy to visualize drug distribution and potential drug-target interactions.

Supplementary Infographic ↗