title: The CCL2/CCR2 Axis Affects Transmigration and Proliferation but Not Resistance to Chemotherapy of Acute Myeloid Leukemia Cells.
abstract: Acute myeloid leukemia (AML) has a high mortality rate despite chemotherapy and transplantation. Both CXCR4/SDF-1 and VLA-4/VCAM1 axes are involved in leukemia protection but little is known about the role of CCL2/CCR2 in AML biology and protection against chemotherapy. We measured CCR2 expression in AML cell lines and primary AML cells by flow cytometry (FCM), real time PCR (RT-PCR) and western blot (WB). CCL2 production was quantified by solid phase ELISA in peripheral blood (PB) and bone marrow (BM) serum. We measured chemotaxis in a transwell system with different concentrations of CCL2/CCR2 blockers; cell cycle with BrDU and propidium iodide and proliferation with yellow tetrazolium MTT. We determined synergy in in vitro cell apoptosis combining chemotherapy and CCL2/CCR2 blockade. Finally, we performed chemoprotection studies in an in vivo mouse model. Of 35 patients, 23 (65%) expressed CCR2 by FCM in PB. Two cell lines expressed high levels of CCR2 (THP-1 and murine AML). RT-PCR and WB confirmed CCR2 production. CCL2 solid phase ELISA showed significantly lower levels of CCL2 in PB and BM compared to normal controls. Chemotaxis experiments confirmed a dose-dependent migration in AML primary cells expressing CCR2 and THP-1 cells. A significant inhibition of transmigration was seen after CCL2/CCR2 blockade. Proliferation of CCR2+ AML cell lines was slightly increased (1.4-fold) after axis stimulation. We observed a non-significant increase in phase S THP-1 cells exposed to CCL2 and a concomitant decrease of cells in G1. The chemotherapy studies did not show a protective effect of CCL2 on cytarabine-induced apoptosis or synergy with chemotherapy after CCL2/CCR2 blockade both in vitro and in vivo. In conclusion, CCL2/CCR2 axis is expressed in the majority of monocytoid AML blasts. The axis is involved in cell trafficking and proliferation but no in vitro and in vivo chemotherapy protective effect was seen.
authors.affiliation: Hematology Oncology Department, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile.
authors.forename: Patricia
authors.initials: P
authors.lastname: Macanas-Pirard
authors.affiliation: Hematology Oncology Department, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile.
authors.forename: Thomas
authors.initials: T
authors.lastname: Quezada
authors.affiliation: Hematology Oncology Department, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile.
authors.forename: Leonardo
authors.initials: L
authors.lastname: Navarrete
authors.affiliation: Hematology Oncology Department, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile.
authors.forename: Richard
authors.initials: R
authors.lastname: Broekhuizen
authors.affiliation: Hematology Oncology Department, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile.
authors.forename: Andrea
authors.initials: A
authors.lastname: Leisewitz
authors.affiliation: Hematology Oncology Department, School of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile.
authors.forename: Bruno
authors.initials: B
authors.lastname: Nervi
authors.affiliation: Institute of Medicine, Faculty of Medicine, Universidad Austral de Chile, Valdivia, Región de los Ríos, Chile.
authors.forename: Pablo A
authors.initials: PA
authors.lastname: Ramírez
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D017209:Apoptosis; D001853:Bone Marrow / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D018932:Chemokine CCL2 / Q000378:metabolism*; D003114:Colony-Forming Units Assay; D019008:Drug Resistance, Neoplasm; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D015973:Gene Expression Regulation, Leukemic; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000188:drug therapy* / Q000378:metabolism; D015473:Leukemia, Promyelocytic, Acute / Q000188:drug therapy* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008875:Middle Aged; D009000:Monocytes / Q000166:cytology; D054390:Receptors, CCR2 / Q000378:metabolism*; D020298:U937 Cells; D055815:Young Adult
chemical_list: C111835:CCL2 protein, human; C515334:CCR2 protein, human; D018932:Chemokine CCL2; D054390:Receptors, CCR2