A homogeneous preparation of a urinary glycoprotein has been isolated from urine of patients with malignant melanoma and advanced adenocarcinomas of colon and lung. This molecule, Mr 30 kDa, is homologous to EDC1, a proteinase inhibitor antigenically related to plasma inter-alpha-trypsin inhibitor (IATI) originally isolated from the urine of a leukemic patient, E.D. The newly isolated EDC1 inhibits cellular proliferation of a Burkitt's lymphoma cell line, Raji, growing in serum-free medium supplemented with insulin, transferrin, selenium, and linoleic acid. This concentration-dependent inhibitory effect was monitored in terms of change in cell number and 3H-thymidine incorporation. The growth of cells treated with approximately 3.3 pmol EDC1/ml was 50% that of the control group by both assays. EDC1 was not cytotoxic to the cells because the EDC1-treated cells excluded trypan blue and resumed normal growth after removal of EDC1. In addition, EDC1 treatment of Raji cells prelabeled with 3H-labeled DNA did not release more radioactivity into the conditioned medium than the untreated labeled cells. EDC1 did not affect the growth of Hs2B2, a B-lymphoblast cell line, and Hs294T, a human malignant melanoma cell line. Equimolar and larger quantities of other proteinase inhibitors with inhibitory profiles similar to that of EDC1 (alpha-1 proteinase inhibitor, soybean trypsin inhibitor, lima bean trypsin inhibitor, and turkey ovomucoid) did not affect the growth of Raji cells. Raji cells have an absolute requirement of transferrin as a nutrient and require insulin to modulate the expression of transferrin receptors. The cells also synthesize interleukin-1 as an autocrine growth stimulator. EDC1 did not form a detectable complex with transferrin, insulin, or any autocrine factor synthesized by the cells.