B cell neoplasms present heterogeneous patterns of lymphoid organ involvement, which may be a result of the differential expression of chemokine receptors. We found that chemokine receptor (CCR)7, CXC chemokine receptor (CXCR)4, or CXCR5, the main chemokine receptors that mediate B cell entry into secondary lymphoid tissues and their homing to T cell and B cell zones therein, were highly expressed in B malignancies with widespread involvement of lymph nodes. Conversely, those pathologies with little or no nodular dissemination showed no expression to very low levels of CCR7 and CXCR5 and low to moderate levels of CXCR4. These findings provide evidence for the role of CCR7, CXCR4, and CXCR5 in determining the pattern of lymphoid organ involvement of B tumors. Functional studies were performed on B malignancies expressing different levels of CCR7, CXCR5, and CXCR4. Multiple myeloma (MM) cells did not express CCR7 nor CXCR5 and did not migrate in response to their ligands; a moderate expression of CXCR4 on MM cells was accompanied by a migratory response to its ligand, CXCL12. By contrast, cells from B cell chronic lymphocytic leukemia (B-CLL) expressed the highest levels of these chemokine receptors and efficiently migrated in response to all ligands of CCR7, CXCR4, and CXCR5. In addition, the migration index of B-CLL cells in response to both of the CCR7 ligands correlated with the presence of clinical lymphadenopathy, thus indicating that the high expression of functional chemokine receptors justifies the widespread character of B-CLL, representing a clinical target for the control of tumor cell dissemination.

The activation of the endothelial cells by extravascular stimuli is the key event in the extravasation of circulating leukocytes to target tissues. L-selectin, a member of the selectin family, is constitutively expressed by white cells, and is the molecule involved in the initial binding of leukocytes to activated endothelium. After activation, leukocytes rapidly release L-selectin from the cell surface, -suggesting that the functional activity of this molecule is controlled in large part by its appearance and disappearance from cell surface. We have studied in a neutrophil-activated endothelial cell binding assay, the effect of different antiinflammatory drugs (steroidal and nonsteroidal) in the L-selectin-mediated interaction of neutrophils with activated endothelial cells. Some nonsteroidal antiinflammatory drugs (NSAIDs), such as indomethacin, diclofenac, ketoprofen, and aspirin, but not steroids, strongly inhibited the neutrophil-endothelial cell attachment. Furthermore, we also investigated the underlying mechanism of this functional effect. The expression of L-selectin on the neutrophil surface rapidly decreased in the presence of different NSAIDs, in a dose-and timedependent manner, whereas no changes in the expression of other adhesion molecules such as CD11a, CD11b, CD31, or ICAM-3 (CD50) were observed. Interestingly, studies in vivo on healthy volunteers treated with physiological doses of indomethacin showed a significant decrease of L-selectin neutrophil expression. Only diclofenac induced an upregulation of CD11b expression, suggesting an activating effect on neutrophils. No enzyme release was observed upon treatment of neutrophils with different NSAIDs, indicating a lack of degranulatory activity of NSAIDs, with the exception of diclofenac. The downregulation of L-selectin expression was due to the rapid cleavage and shedding of the membrane L-selectin, as determined by both immunoprecipitation from "2I-labeled neutrophils, and quantitative estimation in cell-free supernatants. These results suggest that NSAIDs exert a specific action on adhesion receptor expression in neutrophils, which might account, at least in part, for the antiinflammatory activities of NSAIDs. (J. Clin. Invest. 1995.

To date, chronic lymphocytic leukemia (CLL) remains incurable with current treatments, which include the monoclonal antibodies (mAbs) rituximab and alemtuzumab. The efficacy of rituximab is modest when used as single agent, and alemtuzumab induces severe immunosuppression. To develop more potent and specific therapies, we propose the CC chemokine receptor 7 (CCR7) as an attractive target molecule to treat CLL, as it not only fulfills the requirements of a high-surface expression and a good level of tissue specificity, but it also plays a crucial role in mediating the migration of the tumor cells to lymph nodes (LNs) and thus, in the development of clinical lymphadenopathy. In the current work, murine anti-human CCR7 mAb mediated a potent, complement-dependent cytotoxicity (CDC) against CLL cells while sparing normal T lymphocytes from the same patients. The sensitivity to CDC was related to the antigenic density of CCR7. Moreover, these mAb blocked the in vitro migration of CLL cells in response to CC chemokine ligand 19 (CCL19), one of the physiological ligands of CCR7. Conversely, CLL cells were poorly lysed through antibody-dependent, cell-mediated cytotoxicity (ADCC), probably as a result of the murine origin and the isotype of the anti-CCR7 mAb used. Molecular engineering techniques will allow us to obtain chimeric or humanized anti-CCR7 mAb to reach the best clinical response for this common and yet incurable leukemia.

Endothelial cells (EC) play a key role in the inflammatory response, both by the production of proinflammatory cytokines and by their interaction with leukocytes. Molecular genetic analysis has demonstrated that functional NF-kappa B sites are involved in the transcription of interleukin-6 (IL-6), IL-8, and granulocyte-macrophage colony-stimulating factor (GM-CSF) genes in response to inflammatory mediators. Thus, we have explored the effect of two inhibitors of the NF-kappa B activation, pyrrolidine dithiocarbamate (PDTC) and N-acetylcysteine (NAC), on the production of these cytokines by EC. Both PDTC and NAC inhibited, in a dose-dependent manner, the synthesis of IL-6, IL-8, and GM-CSF induced by tumor necrosis factor (TNF)-alpha or bacterial lipopolysaccharides (LPS) in human umbilical vein endothelial cells (HUVEC). PDTC appeared to prevent IL-6, IL-8, and GM-CSF gene transcription, as it blocked the induction of specific mRNA by TNF-alpha or LPS. The TNF-alpha mediated transcriptional activation of a chloramphenicol acetyltransferase (CAT) plasmid containing three copies of the -72 kappa B binding site from the IL-6 promoter was abrogated by PDTC. According to transfection experiments, electrophoretic mobility shift assays (EMSA) demonstrated that the antioxidant prevented the induction of NF-kappa B DNA-binding activity by TNF-alpha. Under the same conditions, PDTC by itself or in combination with TNF-alpha, enhanced the DNA-binding activity of AP-1, as well as c-fos and c-jun mRNA levels. Altogether, these results indicate that the antioxidant PDTC specifically inhibits the transcription of IL-6, IL-8, and GM-CSF genes through the inhibition of the NF-kappa B activation, while increasing the expression of AP-1. Our data make evident the antiinflammatory and immunoregulatory potential of the pharmacological inhibition of the NF-kappa B activation. In addition, PDTC and related molecules may be a useful tool to explore the expression of genes involved in the inflammatory response.