Unique Cell Surface Expression of Receptor Tyrosine Kinase ROR1 in Human B-Cell Chronic Lymphocytic Leukemia

Abstract

Purpose: Gene expression profiling identified receptor tyrosine kinase ROR1, an embryonic protein involved in organogenesis, as a signature gene in B-cell chronic lymphocytic leukemia (B-CLL). To assess the suitability of ROR1 as a cell surface antigen for targeted therapy of B-CLL, we carried out a comprehensive analysis of ROR1protein expression. Experimental Design: Peripheral blood mononuclear cells, sera, and other adult tissues from B-CLL patients and healthy donors were analyzed qualitatively and quantitatively for ROR1 protein expression by flow cytometry, cell surface biotinylation,Western blotting, and ELISA. Results: ROR1 protein is selectively expressed on the surface of B-CLL cells, whereas normal B cells, other normal blood cells, and normal adult tissues do not express cell surface ROR1. Moreover, cell surface expression of ROR1 is uniform and constitutive, i.e., independent of anatomic niches, independent of biological and clinical heterogeneity of B-CLL, independent of B-cell activation, and found at similar levels in all B-CLL samples tested. The antibody binding capacity of B-CLL cell surface ROR1was determined to be in the range of 10 3 to 10 4 molecules per cell. A portion of B-CLL cell surface ROR1 was actively internalized upon antibody binding. Soluble ROR1protein was detectable in sera of <25% of B-CLL patients and a similar fraction of healthy donors at concentrations below 200 ng/mL. Conclusions: The restricted, uniform, and constitutive cell surface expression of ROR1protein in B-CLL provides a strong incentive for the development of targeted therapeutics such as monoclonal antibodies.

Mass spectrometry-based identification of a naturally presented receptor tyrosine kinase-like orphan receptor 1-derived epitope recognized by CD8 + cytotoxic T cells

“…3,4 In agreement with these studies, we identified the HLA-B*07:02-restricted ROR1 peptide NPRYPNYMF as a component of the HLA ligandome of CLL samples (Figure 1 A,B and Online Supplementary Table S1). In contrast, the ROR1 peptide could not be found in the HLA ligandome of normal cells (data not shown).…”

“…It has been demonstrated that ROR1 messenger Ribonucleic Acid (mRNA) is highly expressed in CLL cells (Online Supplementary Figure S1) and a uniform expression of ROR1 protein is detected on their cell surface. 3,4 In contrast, ROR1 expression was not found in other blood cells or on bone marrow-derived cells, except on a small subset of precursor B cells.…”

RNAi screen for rapid therapeutic target identification in leukemia patients

“…Other targets identified in patients with ALL were ROR1, N-RAS, and CSF1R. Interestingly, overexpression of ROR1 has been observed in several studies of lymphocytic malignancies (30,31). Also, we identified JAK2 and EPHA5 as targets in patients with CMML, and FLT1, EPHA4, PDGFR␣/␤, PTK2B, FYN, and JAK1/3 in patients with AML.…”

Atomic force microscopy studies on cellular elastic and viscoelastic properties

“…However, if researchers are to better examine the role of cellular elastic and viscoelastic properties in vivo, primary cancer cells as well as their normal counterparts must be isolated for tests. Recent studies of primary cancerous B lymphocytes have shown that the receptor tyrosine kinase-like orphan receptor 1 (ROR1) is selectively expressed on the surface of B-cell chronic lymphocytic leukemia and on some B-cell lymphomas (including mantle cell lymphoma, marginal zone lymphoma, and follicular lymphoma), whereas normal B lymphocytes and other normal cells do not express ROR1 (Baskar et al, 2008;Bicocca et al, 2012). Hence, ROR1 is a suitable marker for isolating cancerous B lymphocytes from lymphoma patients.…”