Targeted cytotoxic effect of anti-JL1 immunotoxin against a human leukemic cell line and its clinical implications

Abstract

We have previously reported the identification of a unique thymocyte-specific surface molecule, JL1, which was detected using the monoclonal antibody (mAb), anti-JL1. Interestingly, JL1 was shown to be expressed in most leukemias, irrespective of their immunophenotype, and subpopulations of normal bone marrow (BM) mononuclear cells (MNCs). Here we investigated the potential usefulness of the anti-JL1 mAb as a therapeutic tool for leukemia. We demonstrated that the proliferation of cultured human leukemia cells was dramatically inhibited in vitro by anti-JL1 mAb conjugated with the polypeptide toxin, gelonin, but not by gelonin alone. We then systematically investigated the reactivity of the anti-JL1 mAb against normal human tissues to evaluate possible side effects along with various hematopoietic and nonhematopoietic tumor cell lines. All of 33 types of normal tissues except thymus and subpopulation of BM MNCs were clearly devoid of JL1 expression. Among tumor cell lines, all the nonhematopoietic cell lines tested were negative for JL1 expression, while some hematopoietic cell lines contained JL1 antigen. Collectively, the results showed the cytotoxic effects of anti-JL1-based immunotoxin against JL1-positive leukemic cells, sparing most normal tissues other than thymocytes and some BM MNCs. Therefore, we strongly suggest that gelonin-conjugated anti-JL1 mAb immunotoxin could be developed as a potential immunotherapeutic agent in the treatment of various types of JL1-positive acute leukemias.

Fig. 1a–c.

JL1 expression on the surface of Molt-4 cells (a) and U937 cells (b). c Dose response profiles of JL1-gelonin IT in Molt-4 (JL1-positive) cells and U937 (JL1-negative) cells. Starting at a density of 10⁵/ml, cells were incubated for 96 h at 37°C with JL1-gelonin IT and unconjugated JL1 mAb with free gelonin. The treatments are given in parentheses. Trypan blue was added and live cells were counted under a microscope

Fig. 2.

In vitro cytotoxicity of 17 nM JL1-gelonin IT in Molt-4 cells (JL1-positive) and U937 cells (JL1-negative). Cell viability was determined by trypan blue exclusion. The ordinate indicates the percentage of live cells after incubation for 72 h at 37°C in the presence of 17 nM JL1-gelonin IT

Fig. 3a, b.

Competition assay with free JL1 mAb. Starting at a density of 10⁵/ml, Molt-4 cells were incubated for 72 or 96 h at 37°C in the presence of only JL1-Gel IT (a) or JL1-Gel IT with free JL1 mAb (1.2 μM) (b). Trypan blue was added and live cells were counted under a microscope

Fig. 4.

No effect of JL1-Gel IT on the clonogenic activity of early myeloid cells in normal BM MNCs. The results are shown as the number of CFU-GM, CFU-M, and CFU-E per 2.0×10⁵ BM MNCs plated from the corresponding samples. All values are presented as the means±SD from four independent experiments