Role of Connective Tissue Growth Factor (CTGF) in Survival and Chemosensitivity of Acute Lymphoblastic Leukemia

Abstract 2593

Connective tissue growth factor (CTGF/CCN2) is a member of the CCN family of proteins involved in extracellular matrix production, tumor cell proliferation, adhesion, migration, and metastasis. Recent studies have shown that CTGF expression is elevated in 75% of acute lymphoblastic leukemia (Br J Haematol, 2007; 138(6):740–8), and that increased expression of CTGF is associated with inferior outcome in B-ALL (Blood, 2007; 109(7):3080–3). In this study, we characterized the functional role and downstream signaling pathways of CTGF in ALL cells. First, we utilized lentiviral shRNA to knock-down CTGF in RS4;11 and REH ALL cells expressing high levels of CTGF mRNA (479.3±37.2 and 57.3±5.9 copies per 100 copies of ABL1, respectively). Silencing of CTGF (CTGF-knockdown, CTGF-kd) resulted in significant suppression of leukemia cell growth (57% in RS4;11 and by 70% in REH) compared to control vector. CTGF knockdown moderately reduced adhesion of RS4;11 to fibronectin (27%±0.1%). In the in vitro culture system, CTGF knockdown significantly enhanced growth inhibition and apoptosis induction after 48 hour exposure to chemotherapy agents (annexinV(+): Vincristine 25.8±3.5%, Vincristine/CTGF-kd 42.6±2.8%; Dexamethasone 66.3±1.8%, Dexamethasone/CTGF-kd 99.3±0.6%; Methotrexate, 17.4±0.6%, Methotrexate/CTGF-kd 39.5±3.9). Analysis of signaling pathways showed that CTGF down-regulation inhibits Src phosphorylation at Tyr416. Remarkably, phosphorylation of AKT at Ser473, and of mTOR downstream targets S6 Ribosomal Protein and 4E-BP1 were significantly inhibited in CTGF-knockdown RS4;11 cells, concomitantly with upregulation of expression of AKT targets Bim and p27. No changes in the levels of apoptotic regulators cIAP1 and Bcl-xL were found. This data suggest that CTGF regulates growth and chemosensitivity of ALL cells through Src and AKT/mTOR signaling. We previously reported that an anti-CTGF monoclonal antibody significantly extended median survival of mice implanted with xenografts derived from a primary CTGF expressing ALL sample in NOD/SCID mice. We are now investigating the effects of combining anti-CTGF treatment with cytotoxic chemotherapy in this model. Blocking CTGF signaling may represent a useful adjunct to cytotoxic therapies in acute lymphoblastic leukemia.