An increase in the number of stem cells in blood following mobilization is required to enhance engraftment after highdose chemotherapy and improve transplantation outcome. Therefore, an approach that improves stem cell mobilization is essential. The interaction between CXCL12 and its receptor, CXCR4, is involved in the retention of stem cells in the bone marrow. Therefore, blocking CXCR4 may result in mobilization of hematopoietic progenitor and stem cells. We have found that the CXCR4 antagonist known as 4F-benzoyl-TN14003 (T-140) can induce mobilization of hematopoietic stem cells and progenitors within a few hours posttreatment in a dose-dependent manner. Furthermore, although T-140 can also increase the number of white blood cells (WBC) in blood, including monocytes, B cells, and T cells, it had no effect on mobilizing natural killer cells. T-140 was found to efficiently synergize with granulocyte colony-stimulating factor (G-CSF) in its ability to mobilize WBC and progenitors, as well as to induce a 660-fold increase in the number of erythroblasts in peripheral blood. Comparison between the CXCR4 antagonists T-140 and AMD3100 showed that T-140 with or without G-CSF was significantly more potent in its ability to mobilize hematopoietic stem cells and progenitors into blood. These results demonstrate that different CXCR4 antagonists may have different therapeutic potentials.

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The chemokine receptor CXCR4 and its ligand CXCL12 is overexpressed in the majority of tumors and is critically involved in the development and metastasis of these tumors. CXCR4 is expressed in malignant tumor cells whereas its ligand SDF-1 (CXCL12) is expressed mainly by cancer associated fibroblasts (CAF). Similarly to CXCR4, the chemokine CCL20 is overexpressed in variety of tumors; however its role and regulation in tumors is not fully clear. Here, we show that the chemokine receptor CXCR4 stimulates the production of the chemokine CCL20 and that CCL20 stimulates the proliferation and adhesion to collagen of various tumor cells. Furthermore, overexpression of CCL20 in tumor cells promotes growth and adhesion in vitro and increased tumor growth and invasiveness in vivo. Moreover, neutralizing antibodies to CCL20 inhibit the in vivo growth of tumors that either overexpress CXCR4 or CCL20 or naturally express CCL20. These results reveal a role for CCL20 in CXCR4-dependent and -independent tumor growth and suggest a therapeutic potential for CCL20 and CCR6 antagonists in the treatment of CXCR4- and CCL20-dependent malignancies.

Familial dysautonomia (FD) is a genetic disease characterized by primary autonomic dysfunction including parasympathetic hypersensitivity. Breath-holding spells (BHS) are believed to be caused by autonomic dysregulation mediated via the vagus nerve and increased in patients with a family history of BHS. Details and understanding of its pathophysiology are lacking. In this retrospective study of patients with FD, the incidence of BHS was higher at 53.3%, compared with previous studies in normal children. Laughter as a precipitating factor for BHS has not been previously reported in FD and occurred in 10% of patients in this study. Lower lung volumes, chronic lung disease, chronic CO2 retention, and inadequate autonomic compensation occur in those with FD leading to a higher incidence and severity of BHS when crying or laughing. Thus, FD may be a good model for understanding manifestations of the autonomic nervous system dysfunction and contribute to our knowledge of BHS mechanisms.

3857 Poster Board III-793 The chemokine receptor CXCR4 and its ligand CXCL12 are involved in the development and progression of a diverse number of hematological malignancies, including leukemia, lymphoma and multiple myeloma (MM). Binding CXCL12 to CXCR4 activates a variety of intracellular signal transduction pathways and effector molecules that regulate cell chemotaxis, adhesion, survival, apoptosis and proliferation. It was previously shown that CXCR4 signaling can directly induce caspase-independent cell apoptosis through the interaction with the HIV gp120 envelope protein. In the present study we investigated the effect of CXCR4 specific antagonists 4F-benzoyl-TN14003 (T140) and AMD3100 on the survival and proliferation of different human hematological cancer cells. Here, we demonstrate that T140, but not AMD3100, exhibits preferential cytotoxicity towards malignant cells of hematopoietic origin, as compared to primary normal cells or solid prostate and breast tumor cells. The in vitro treatment with T140, but not with AMD3100, significantly decreased the number of viable chronic myeloid leukemia K562 cells, acute T cell leukemia Jurkat cells, acute promyelocytic leukemia NB4 and HL60 cells, and four different MM cell lines (U266, NCI-H929, RPMI8226 and ARH77), demonstrating the highest sensitivity to T140 (p<0.01). Notably, T140 inhibited the growth of freshly isolated leukemia and MM cells obtained from consenting patients. T140 inhibits the growth of MM and leukemic cells by inducing their apoptotic cell death. The apoptotic changes in the cells were associated with morphological changes, phosphatidylserine externalization, sub-G1 arrest, DNA double-stranded breaks, decrease in mitochondrial membrane potential, release of cytochrome c, and caspase 3 activation. The important role of CXCR4 in T140-mediated cell death was confirmed by demonstrating that CXCR4 over-expression in NB4 and K562 cells increased their sensitivity to T140. Furthermore, pretreatment of NB4 and HL60 cells with AMD3100 abolishes the effect of T140 on these cells, indicating the involvement of CXCR4 in T140-induced apoptosis. Importantly, the combination with novel anti-myeloma agent bortezomib significantly augments anti-myeloma activity of T140. The anti leukemic and MM effect of T140 was confirmed in xenograft in vivo tumor models. Subcutaneous (s.c.) or intra-peritoneal (i.p.) injections of T140 (100 or 300 mcg/mouse) significantly reduced, in a dose-dependent manner, the tumor size in immuno-deficient mice that were previously inoculated s.c. with human acute leukemia cells NB4 or MM cells RPMI8226 (p<0.01). Tumors from animals treated with T140 had smaller sizes and weights, larger necrotic areas and high apoptotic scores. Taken together, these data support the unique anti-cancer effect of T140 in hematological malignancies and indicate the potential therapeutic role of T140 in MM and leukemia patients. Disclosures: No relevant conflicts of interest to declare.