Leukemic blasts from patients with acute phase chronic myeloid leukemia and refractory acute myeloid leukemia are highly resistant to many cytotoxic drugs. To overcome mult-drug resistance, we engineered a diptheria fusion protein by fusing human interleukin3 to a truncated form of diptheria toxin (DT). This DT-IL3 protein was more cytotoxic to IL-3R bearing human myeloid leukemia cell lines than IL-3R negative cell lines. Furthermore, exposure of monnuclear cells to DT-IL3 reduced the number of cells capable of forming colonies in semi-solid media. The sensitivity to DT-IL3 of leukemic progenitors from a number of acute phase patients suggests that this agent could have therapeutic potential for some patients with CML. To further investigate the ability of DT-IL3 to selectively inhibit the leukemic cell growth, the effects of DT-1L3 on the growth and induction of apoptosis in oncogene-transformed TF-1 human erthroleukemia cells were determined. Treatment of cytokine-dependent parental TF-1 cells with DT-IL3 results in the induction of apoptotic cell death after 2-3 days as determined by DNA laddering and Annexin V/PI binding. TF-1 cells were transformed to independence of exogenous cytokines after transfection with activated signal transduction molecules (Ha-Ras, Src, Raf, MEK1. PI3K. Akt, BCR-ABL and GM-CSF). Interestingly, the cell line which constitutive expresses autocrine GM-CSF as well as the oncogene-transformed cells, remained sensitve to the effects of DT-IL3 which indicated that cells which had by passed IL-3 signal transduction that retained IL-3 receptor expression were still sensitive to DT-IL3. The ICMs for DTIL3 remained relatively constant with less than 3-fold variation. In contrast, the cytokineindependent TF-1 cells were resistant to cell death induced by certain other chemotherapeutic agents such as all Irans retinoic acid (ATRA) as the IC5(J increases from 1 M.M in the cytokine-dependent cells to 50 μM in the oncogene-transformed cells. Similar results were observed with cis RA. Overexpression of the Bcl-2 oncogene in oncogenetransformed cells did not prevent the cells from the apoptotic-inducing effects of DT-IL3. The oncogene and autocrine transformed cell lines formed tumors in immunocompromised mice whereas the parental cytokine-dependent line does not. The ability of DT-II.3 treatment to reduce leukemic tumor load is being determined. These studies indicate DTIL3 may be an effective adjuvant in the treatment of certain leukemias which are resistant to differentiation-inducing agents such as ATRA.
|Issue number||11 PART I|
|Publication status||Published - 2000|
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