Folate receptor-α targeted near-infrared fluorescence imaging in high-risk endometrial cancer patients: a tissue microarray and clinical feasibility study

Abstract

Objective: Detection and resection of all malignant lesions is pivotal in staging and cytoreductive surgery (CRS) of endometrial cancer (EC). Intraoperative EC detection could be enhanced using OTL-38, a fluorescent-labelled folate receptor-α (FRα) targeted imaging agent. The objectives of this study were to investigate which subgroups of high-risk EC patients express FRα and assess feasibility of intraoperative EC detection using OTL-38.

Results: FRα expression on TMA was significantly correlated with tumor type (p < 0.01). Eighty-two percent of serous and clear cell carcinomas showed FRα expression. Four patients were enrolled in the clinical study. Using fluorescence imaging all omental (n = 3) and lymph node (LN) metastases (n = 16) could be clearly identified, including one otherwise undetected omental metastasis. However, false-positive fluorescence was identified in 17/50 non-metastatic LNs, caused by OTL-38 targeting of FRβ, expressed by tumor-associated activated macrophages.

Conclusions: This study describes high FRα expression in serous and clear cell EC and demonstrates the first experience of intraoperative FRα-targeted tumor detection in patients with these subtypes of EC. Although all metastases could be clearly identified using OTL-38, the role of tumor-associated macrophages should be further evaluated.

Methods: Immunohistochemical (IHC) staining of FRα expression was performed on tissue micro arrays (TMA) of 116 patients with high-risk EC features. Patients with either serous or clear cell EC, planned for staging or CRS, were eligible for inclusion in the clinical study and received an intravenous dose of 0.0125 mg/kg OTL-38, 2-3 hours prior to surgery. Resected lesions, identified by standard-of-care and/or fluorescence imaging, were histopathologically assessed for FRα and tumor status.

Keywords: FRα; biomarker; targeting; tumor-specific imaging.

Figure 1. Representative examples of FRα expression status in endometrioid and clear cell EC

Shown are tissue cores with respectively weak, moderate and strong intensity of FRα staining in endometrioid (A) and clear cell carcinoma (B). Magnification: 10×.

Figure 2. Fluorescence imaging of a primary uterine serous adenocarcinoma and of the metastatic lymph nodes

(A) Intraoperative identification of para-aortic, metastatic lymph nodes (dashed arrow), located beneath a layer of overlying tissue (patient #1). The normal arrow indicates the fluorescence signal arising from the uterus. (B) Ex vivo fluorescence imaging of the resected para-aortal lymph nodes (patient #1), that show a clear fluorescence signal. (C) Ex vivo fluorescence imaging of the bisected uterus (patient #1). The fluorescence signal, detected during surgery, appears to be mainly arising from normal adjacent background uterine tissue instead of the primary tumor.

Figure 3. Histopathological evaluation of resected lesions

(A) Immunohistochemical staining for FRα of a uterine tumor shows FRα expression in cancer cells (black arrow, patient #1) and in normal epithelial cells (dashed arrow). Magnification: 5×. (B) Immunohistochemical staining for FRα of non-malignant adenomyosis tissue of the uterus (patient #2). Magnification: 5×. (C) Immunohistochemical staining for FRα of a clinically suspect and fluorescent lymph node shows positive FRα expression in the lymph node follicels (patient #1). The FRα expression correlates with the presence of tumor cells. Magnification: 0.5×. (D) Immunohistochemical staining for FRα of a fluorescent omental lesion, that contained tumor cells, shows positive FRα expression (patient #4). Magnification: 2×.

Figure 4. Histopathological evaluation of a false-positive lymph node

Shown are a NIR fluorescence image, haematoxylin & eosin (HE) staining, and FRα and FRβ staining of a (fluorescent) LN that did not contain tumor cells. Fluorescence is mainly seen in the sinuses and not in the follicles. The magnified images (HE, FRα and FRβ) show the lack of FRα staining, while the sinuses show a positive FRβ staining. Magnifications: 1× and 10×.