mTOR Inhibitors Induce Apoptosis and Inhibit Growth of Primary Adult Human ALL in Xenograft and Tissue Culture Models.

Abstract

While children with precursor B cell acute lymphoblastic leukemia (ALL) are often cured, adults with ALL usually succumb to their disease. Thus, the development of novel therapeutic agents is paramount. mTOR inhibitors (MTI) are a class of signal transduction inhibitors developed as immunosuppressive agents. We have previously shown the ability of MTI to inhibit growth and induce apoptosis in precursor B ALL cell lines and primary murine pre-B leukemia. To determine if this finding could be translated to clinical therapy, we explored if MTIs would be similarly effective in three primary human adult ALL samples. We have tested the MTIs rapamycin and CCI-779 in these models with similar results, but the results utilizing CCI-779 are presented below. Stromal culture. ALL blasts were maintained in vitro on irradiated bone marrow stromal cells. Cells normally can be maintained for several weeks in these conditions. Cells were either untreated or treated with CCI-779 at 100ng/ml. Long-term cultures were assessed for the effect of MTI on cell proliferation and short term (48hr) cultures were assessed for induction of apoptosis. Treated cells showed a dramatic decrease in cell proliferation (6–24 fold compared to untreated) and a 4–5 fold increase in apoptotic cells as detected by Annexin-V compared to untreated cells. NOD/SCID xenografts. To further evaluate the effect of MTI on ALL cells, patient samples were engrafted into NOD/SCID animals for analysis. Robust engraftment, expansion and repopulation in secondary hosts of ALL cells was seen in 78% of tested samples, demonstrating greater than 70% ALL in peripheral blood, bone marrow, and spleen in the majority of the mice. There was greater than 10 fold expansion of disease within the mouse. Engraftment was detected by flow cytometry for human CD19+/CD45+ cells. Engrafted animals with established disease (>5% peripheral blasts) were either not treated or treated with the MTI CCI-779. Currently we have treated 45 mice engrafted from three separate patient samples. Untreated animals continued to show expansion of human ALL cells. In dramatic contrast, animals treated with CCI-779 as a single agent showed a 4–30 fold decrease in peripheral blood blasts and a decrease in splenomegaly (p<.02). Our data show that both in vitro and in an in vivo model of established ALL, MTIs decrease proliferation of lymphoblasts and promote apoptosis of ALL cells. These results suggest that the mTOR signaling pathway is necessary for the survival of ALL cells and that MTIs should be analyzed as therapeutic agents for the therapy of ALL.