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

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Clinical Cancer Research 14, 396-404, January 15, 2008. doi:

10.1158/1078-0432.CCR-07-1823

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

Chronic Lymphocytic Leukemia

Sivasubramanian Baskar1, Ka Yin Kwong1, Hofer1, M. Levy1,

G. Kennedy1, Elinor Lee3, Louis M. Staudt2, Wyndham H. 2, Wiestner3

and Christoph Rader1

Authors' Affiliations: 1 Experimental Transplantation and Immunology Branch and

2 Metabolism Branch, Center for Cancer Research, National Cancer Institute, and

3 Hematology Branch, National Heart, Lung and Blood Institute, NIH, Bethesda,

land

Requests for reprints: Christoph Rader, Experimental Transplantation and

Immunology Branch, Center for Cancer Research, National Cancer Institute, NIH,

9000 Rockville Pike, Building 10 CRC, Room 3-3150, Bethesda, MD 20892-1203.

Phone: 301-451-2235; Fax: 301-480-3431; E-mail: raderc@... .

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 ROR1 protein 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 ROR1 was determined to be in the

range of 103 to 104 molecules per cell. A portion of B-CLL cell surface ROR1 was

actively internalized upon antibody binding. Soluble ROR1 protein 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 ROR1 protein in B-CLL provides a strong incentive for the development of

targeted therapeutics such as monoclonal antibodies.