In the present study, immunotoxins (ITs) containing ricin A chain (RA) and anti-human T leukemia monoclonal antibodies SN1 and SN2 were used with or without α-interferon (IFN) and/or daunorubicin (DNR) for in vivo tumor suppression. SN1 and SN2 are directed toward two unique human T-leukemia-associated cell surface antigens, TALLA and GP37, respectively. As the tumor model, we used nude mice bearing ascitic tumors of Ichikawa, a human T acute lymphoblastic leukemia cell line. In initial studies, we investigated the effect of the IT injection schedule on the efficacy of ITs in the in vivo suppression of the ascitic tumors. Four doses of 20 μg each of SN1-RA and SN2-RA completely suppress the tumor growth in 100% of the treated mice when the IT treatment is initiated either 1 or 2 days after tumor inoculation of 1.6 × 107 Ichikawa cells into the mice. Subsequently, we investigated the potentiating effects of IFN and DNR on the in vivo antitumor activity of ITs. To this end, we chose to initiate the treatment 4 days after the tumor inoculation when IT treatment alone is only partially effective. ITs (10 μg each of SN1-RA and SN2-RA) plus IFN (2 × 105 IU) or ITs plus IFN plus DNR (5 μg) completely suppress tumor growth in 100% of the treated mice while similar treatment with any one of the three agents is only partially effective. Similar treatment with ITs plus DNR or IFN plus DNR results in complete suppression of tumor growth in 80% of the treated mice. These results were reproducible in a repeated experiment. To gain information about the mechanisms involving the IFN potentiation of IT activity, we carried out several experiments. The cell surface expression of TALLA and GP37 was slightly augmented by the in vitro incubation of Ichikawa cells with IFN as measured by fluorescence-activated cell sorter analysis. The degree of the increase in either TALLA or GP37 was significantly smaller than that of HLA class I antigens in the same experiment. In in vitro experiments, IFN did not show any significant cytotoxic activity against Ichikawa cells or augment the cytotoxic activity of ITs against Ichikawa cells. On the other hand, injections of IFN into nude mice augmented activity of macrophages and NK cells; however, Ichikawa leukemia cells were rather resistant to the NK cell lysis. These results collectively indicate that IFN potentiates the in vivo antitumor activity of our anti-human T leukemia ITs primarily by host-mediated effector mechanisms but not by direct action of IFN on leukemia cells. Particularly, the activation of macrophages by IFN in the tumor-bearing nude mice appears to be the major factor in the potentiation of the in vivo antitumor activity of the ITs in the present study. Furthermore, the present results clearly demonstrate the superior therapeutic efficacy of combined therapy, especially with ITs plus IFN, to therapy with ITs, IFN, or DNR alone.


This work was supported by USPHS Grant PO1 CA42683 awarded by the National Cancer Institute, Department of Health and Human Services.

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