Abstract 53: Increased expression of L-selectin (CD62L) in high grade bladder cancer: a potential biomarker for lymph node metastasis

Abstract

Abstract Introduction and Objectives: L-selectin (CD62L) is a vascular adhesion molecule which is primarily expressed on leucocytes. Its principal function is to direct leucocyte migration and homing to lymph nodes (LNs). In a gene expression microarray study of laser captured micro-dissected high grade muscle invasive bladder cancer (MIBC) vs. low grade bladder cancer (LGBC) from human specimens, we found CD62L to be the highest differentially expressed gene. We sought to examine the differential expression of CD62L in MIBCs and its clinical relevance. Methods: Unfixed fresh and formalin fixed paraffin-embedded human bladder cancer specimens and serum samples were obtained from the UCHC tumor bank. Tumor cells were isolated from 5-10 µm sections of frozen tumor tissue (3 MIBC and 3 LGBC) by laser capture microdissection followed by RNA isolation. Preparation of cRNA and hybridization to Agilent Human GE 4x44K v2 microarray chips was performed by GenUS Biosystems (Northbrook, IL). qPCR was used to validate the level of CD62L transcripts. Immunohistochemistry and ELISA were performed to evaluate the CD62L protein localization and expression level. Flow cytometry was used to identify the relative number of cells expressing CD62L in fresh tumor tissue. In silico studies were performed using the Oncomine Database. Results: Two-hundred fifty genes were differentially expressed (≥ 2-fold) between the groups with 86 overexpressed in MIBC compared to LGBC. The most highly elevated transcript (34-fold) was L-selectin. The increased expression of CD62L transcript in high grade MIBC vs. LGBC specimens was validated by qPCR. Using 12 human bladder tumor samples representing high grade MIBC and LGBC, we found a uniformly higher expression level of CD62L in MIBC samples. Further, CD62L localization was seen in foci of metastatic tumor cells in LN specimens from patients with high grade MIBC and known nodal involvement. Up-regulated expression of CD62L was also observed by flow cytometry of freshly isolated tumor cells from biopsies of high grade MIBC vs. LGBC patients. Circulating CD62L levels were also found to be higher (640 ± 75 vs. 558 ± 34 ng/ml) in serum samples from patients with high grade metastatic vs. high grade non-metastatic MIBC. In addition, in silico analysis of Oncomine Microarray Database showed a significant correlation between CD62L expression and tumor aggressiveness and clinical outcomes. Conclusion: These findings suggest that elevated CD62L expression may serve as a marker of either occult metastatic disease or the propensity of a tumor to metastasize. Citation Format: Dharamainder Choudhary, Poornima Hegde, Shilpa Choudhary, Kevin Claffey, Pramod Srivastava, Carol C. Pilbeam, John Arthur Taylor. Increased expression of L-selectin (CD62L) in high grade bladder cancer: a potential biomarker for lymph node metastasis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 53. doi:10.1158/1538-7445.AM2014-53