Expression of Midkine(MDK) and Sensitivity to Anti-Leukemic Drugs in Pediatric Acute Lymphoblastic Leukemia (ALL).

Midkine (MDK), a heparin-binding growth factor, is highly expressed in pediatric ALL, neuroblastomas, and various solid tumors. Previous gene expression profiling studies conducted in our laboratory on a cohort of 220 children with B precursor ALL treated on Children’s Oncology Group (COG) clinical trials have shown that higher levels of MDK expression in leukemic blasts are strongly associated with continuous complete remission (CCR) and that MDK is a powerful prognostic predictor of outcome (Wilcoxon ranksum test, p=0.000001), particularly in patients with central nervous system (CNS) involvement (Wilcoxon ranksum test, p =0.009). To explore the role of MDK in hematopoiesis and leukemogenesis and to further explore MDK as a potential novel target for ALL therapy, we examined MDK mRNA expression in three stages of normal B cell development in healthy controls and compared it to expression levels in leukemic blasts from children with B-ALL. Quantitative RT-PCR assays showed that MDK mRNA levels in CD19+ cells from peripheral blood mononuclear cells (PBMNC) were significantly higher than those in CD34+ or CD19+ cells from bone marrow (BM) (Mann Whitney test, p=0.0025 and 0.0018, respectively). ALL leukemic blasts samples expressed significantly higher levels of MDK than those in CD19+ cell in the BM of healthy controls (Mann Whitney test, p=0.0043). Similar results were obtained by ELISA detection of MDK protein in plasma of ALL patients and healthy controls. We have also investigated the relationship of MDK expression and the chemo-sensitivity of ALL cell lines. Using a lentiviral system, we have successfully over-expressed or knocked down MDK gene expression in the REH cell line containing the t(12;21). Unexpectedly, MTS assays revealed that MDK expression is inversely associated with sensitivity to L-asparaginase and doxorubicin, but not to vincristine and cytarabine, in REH cells. Higher levels of MDK were associated with chemo-resistance. Finally, we tested the effects of MDK expression on proliferation and apoptosis in REH cells and determined that although MDK expression was not associated with REH cell proliferation in vitro, it protected REH cells from doxorubicin-induced apoptotic cell death. Taken together, these results suggest that expression levels of MDK in ALL is inversely associated with in vitro drug sensitivity to specific anti-leukemic agents; consistent with observations in solid tumor models. It has been suggested that ₄β¹- and ₆β¹-integrins bind directly to MDK and are involved in MK-dependent cell migration. Further studies of the role of MDK signaling in vivo in the complex marrow/adhesion microenvironment and the effects on ALL survival and chemo-sensitivity and resistance are in progress.