Tumor-Associated Lymphangiogenesis in the Development of Conjunctival Squamous Cell Carcinoma

Purpose To analyze whether tumor-associated lymphangiogenesis accompanies the development from premalignant conjunctival intraepithelial neoplasia (CIN) into invasive squamous cell carcinoma (SCC) of the conjunctiva and to study its association with prognosis and other tumor characteristics. Design Case-controlled, matched-pair cohort study. Participants Twenty patients with invasive SCC were closely matched with 20 patients with high-grade CIN and 20 patients with low-grade CIN regarding tumor size, tumor location, tumor extension, and patients' age. Methods Proliferating lymphatic vessels were identified using lymphatic vascular endothelial hyaluronan receptor-1 and podoplanin as specific lymphatic endothelial markers and Ki-67 as proliferation marker. Baseline tumor characteristics included tumor size, tumor-to-limbus distance, tumor-to-fornix distance, 2009 TNM classification, tumor cell type, mitotic rate, and Ki-67 proliferation index. Kaplan–Meier and Cox regression analyses of recurrence-free survival were performed. Main Outcome Measures Lymphatic vascular density (LVD) and relative lymphatic vascular area (RLVA) of proliferating lymphatic vessels within the tumor mass (intratumoral) and within an area ≤500 μm from the tumor border (peritumoral), and its association with tumor characteristics and recurrence-free survival. Results Intratumoral and peritumoral proliferating lymphatic vessels could be detected in all of the 60 conjunctival tumor samples. Invasive SCC revealed significantly higher values of intratumoral and peritumoral LVD and RLVA of proliferating lymphatics than high-grade or low-grade CIN ( P ≤0.001). Higher intratumoral lymphatic densities were significantly associated with larger tumor size ( P= 0.001), lower tumor-to-limbus distance ( P= 0.002), lower tumor-to-fornix distance ( P= 0.003), and higher TNM categories ( P< 0.001). Recurrence-free survival rates decreased significantly with higher intratumoral lymphatic densities ( P< 0.001). By multivariate Cox regression, large tumor size (hazard ratio 1.68, P= 0.002) and high intratumoral lymphatic density (hazard ratio 1.10, P= 0.046) were significant prognostic predictors of local recurrence. Conclusions Development of conjunctival SCC from premalignant lesions is accompanied by the outgrowth of new conjunctival lymphatic vessels. This active tumor-associated lymphangiogenesis seems to be associated with an increased risk of local recurrence in patients with CIN and conjunctival