In this retrospective pilot study, the feasibility of the epithelial cell adhesion molecule (EpCAM) as an imaging target for lymph node (LN) metastatic disease of urothelial cell carcinoma (UCC) of the bladder was investigated. LN metastases and LNs without metastases of patients who underwent pelvic lymph node dissection because of muscle invasive bladder cancer (MIBC) were used. Primary tumors of the same patients were used from cystectomy specimen, transurethral resections, and biopsies. A pathologist, blinded to clinical data, scored EpCAM immunoreactivity. This method determines a total immunostaining score, which is the product of a proportion score and an intensity score. EpCAM expression was observed in 19/20 (95%) LNs with UCC metastases and in 11/12 (92%) of the primary tumors. EpCAM expression was absent in 14/14 (100%) LNs without metastases. Median EpCAM expression (TIS) in LN metastases was 5 (IQR 2.0–8.0) and in the primary tumors 6 (IQR 2.3–11.0). Based on the absence of staining in LNs without metastases, EpCAM show high tumor distinctiveness. EpCAM seems to be a feasible imaging target in LN metastases of UCC of the bladder. Pre- and perioperative visualization of these metastases will improve disease staging and improve the complete resection of LN metastases in MIBC.

SignificanceShortwave-infrared (SWIR) imaging is reported to yield better contrast in fluorescence-guided surgery than near-infrared (NIR) imaging, due to a reduction in scattering. This benefit of SWIR was shown in animal studies, however not yet in clinical studies with patient samples.AimWe investigate the potential benefit of SWIR to NIR imaging in clinical samples containing cetuximab-IRDye800CW in fluorescence-guided surgery.ApproachThe potential of the epidermal growth factor-targeted NIR dye cetuximab-IRDye800CW in the shortwave range was examined by recording the absorption and emission spectrum. An ex vivo comparison of NIR and SWIR images using clinical tumor samples of patients with penile squamous cell carcinoma (PSCC) and head and neck squamous cell carcinoma (HNSCC) containing cetuximab-IRDye800CW was performed. The comparison was based on the tumor-to-background ratio and an adapted contrast-to-noise ratio (aCNR) using the standard of care pathology tissue assessment as the golden standard.ResultsBased on the emission spectrum, cetuximab-IRDye800CW can be detected in the SWIR range. In clinical PSCC samples, overall SWIR imaging was found to perform similarly to NIR imaging (NIR imaging is better than SWIR in the 2/7 criteria examined, and SWIR is better than NIR in the 3/7 criteria). However, when inspecting HNSCC data, NIR is better than SWIR in nearly all (5/7) examined criteria. This difference seems to originate from background autofluorescence overwhelming the off-peak SWIR fluorescence signal in HNSCC tissue.ConclusionSWIR imaging using the targeted tracer cetuximab-IRDye800CW currently does not provide additional benefit over NIR imaging in ex vivo clinical samples. Background fluorescence in the SWIR region, resulting in a higher background signal, limits SWIR imaging in HNSCC samples. However, SWIR shows potential in increasing the contrast of tumor borders in PSCC samples, as shown by a higher aCNR over a line.

Background: In this study we investigated the expression of vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR) and prostate-specific membrane antigen (PSMA) to analyze their potency as targets for the detection of lymph node (LN) metastases of urothelial carcinoma of the bladder. Methods: Antigen expression was determined in 40 samples with urothelial carcinoma and compared to 17 matched samples without metastases by immunohistochemistry. The total immunostaining score (TIS 0–12) was determined as the product of a proportion score (PS 0–4) and intensity score (IS 0–3). Results: VEGF expression was high in primary tumor and LN metastases (median TIS 8 in both) and VEGF expression was also seen in LNs without metastases (median TIS 6). EGFR expression was low in primary tumor and LN metastases (median TIS 3 and 2 respectively) and absent in LNs without metastases. PSMA expression was low in samples with urothelial carcinoma (median TIS 2). Conclusion: VEGF shows moderate to high expression levels in both primary tumors and LN metastases and could be a candidate as a target agent for imaging modalities of urothelial carcinoma. EGFR and PSMA do show low staining levels in tumor tissue with urothelial carcinoma and do not seem suitable as target agents. Trial registration: The Medical Ethics Review Board of the University Medical Center Groningen approved this study on 14 December 2017 (METc UMCG 2017/639). Trial registration number (UMCG Research Register): 201700868.

Imaging modalities using tumor-directed monoclonal antibodies may be of value to improve the pre- and intraoperative detection and resection of lymph node (LN) metastatic disease in penile squamous cell carcinoma (PSCC). We investigated the expression of prostate-specific membrane antigen (PSMA), vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR) and epithelial cell adhesion molecule (EpCAM) to analyze their potency for diagnostic applications. Antigen expression was determined in primary tumors and LNs with and without metastases of 22 patients with PSCC. The total immunostaining score (TIS, 0–12) was determined as the product of a proportion score (PS, 0–4) and an intensity score (IS, 0–3). EGFR and VEGF expression were high in primary tumor (median TIS 8) and LN metastases (median TIS 6 and 8, respectively). No EGFR expression was seen in LNs without metastases. However, LNs without metastases did show VEGF expression (median TIS 6). No EpCAM or PSMA expression was seen in PSCC. This study shows that VEGF and EGFR expression is moderate to high in LN metastases of PSCC. Both VEGF and EGFR warrant further clinical evaluation to determine their value as a target for pre- and intraoperative imaging modalities in the detection of LN metastases in PSCC.

Background In this study we investigated the expression of vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR) and prostate-specific membrane antigen (PSMA) to analyze their potency as targets for the detection of lymph node (LN) metastases of urothelial carcinoma of the bladder. Methods Antigen expression was determined in 40 samples with urothelial carcinoma and compared to 17 matched samples without metastases by immunohistochemistry. The total immunostaining score (TIS, 0–12) was determined as the product of a proportion score (PS, 0–4) and intensity score (IS, 0–3). Results VEGF expression was high in primary tumor and LN metastases (median TIS 8 in both) and VEGF expression was also seen in LNs without metastases (median TIS 6). EGFR expression was low in primary tumor and LN metastases (median TIS 3 and 2 respectively) and absent in LNs without metastases. PSMA expression was low in samples with urothelial carcinoma (median TIS 2). Conclusion VEGF shows moderate to high expression levels in both primary tumors en LN metastases and could be a candidate as a target agent for imaging modalities of urothelial carcinoma. EGFR and PSMA do show low staining levels in tumor tissue with urothelial carcinoma and do not seem suitable as target agents. Trial registration: The Medical Ethics Review Board of the University Medical Center Groningen approved this study on 14 December 2017 (METc UMCG 2017/639). Trial registration number (UMCG Research Register): 201700868.