Limits...
CXCL12-induced VLA-4 activation is impaired in trisomy 12 chronic lymphocytic leukemia cells: a role for CCL21.

Ganghammer S, Hutterer E, Hinterseer E, Brachtl G, Asslaber D, Krenn PW, Girbl T, Berghammer P, Geisberger R, Egle A, Zucchetto A, Kruschinski A, Gattei V, Chigaev A, Greil R, Hartmann TN - Oncotarget (2015)

Bottom Line: Here, we demonstrate that high CD49d expression in tri12 CLL is accompanied by decreased CXCR4 expression.Concordantly, in real-time kinetic analyses we found CCL21 but not CXCL12 being capable to induce inside-out VLA-4 conformational changes in this CLL subgroup.Our results provide novel insights into the peculiar clinico-biological behaviour of tri12 CLL and emphasize its specific chemokine and integrin utilization during pathophysiologically and therapeutically relevant interactions with the microenvironment.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Immunological and Molecular Cancer Research, 3rd Medical Department with Hematology, Medical Oncology, Hemostaseology, Infectious Diseases and Rheumatology, Oncologic Center, Paracelsus Medical University Salzburg, Austria.

ABSTRACT
Homing to distinct lymphoid organs enables chronic lymphocytic leukemia (CLL) cells to receive pro-survival and proliferative signals. Cytogenetic aberrations can significantly affect CLL cell compartmentalization. Trisomy 12 (tri12) defines a CLL subgroup with specific clinical features and increased levels of the negative prognostic marker CD49d, the α4-subunit of the integrin VLA-4, which is a key regulator of CLL cell homing to bone marrow (BM). Chemokine-induced inside-out VLA-4 activation, particularly via the CXCL12-CXCR4 axis, increases the arrest of various cell types on VCAM-1 presenting endothelium. Here, we demonstrate that high CD49d expression in tri12 CLL is accompanied by decreased CXCR4 expression. Dissecting functional consequences of these alterations, we observed that tri12 CLL cell homing to murine BM is not affected by CXCR4-CXCL12 blockage using AMD3100 or olaptesed pegol/NOX-A12. In line, CCL21-CCR7 rather than CXCL12-CXCR4 interactions triggered VLA-4-mediated arrests of tri12 CLL cells to VCAM-1 under blood flow conditions. Concordantly, in real-time kinetic analyses we found CCL21 but not CXCL12 being capable to induce inside-out VLA-4 conformational changes in this CLL subgroup. Our results provide novel insights into the peculiar clinico-biological behaviour of tri12 CLL and emphasize its specific chemokine and integrin utilization during pathophysiologically and therapeutically relevant interactions with the microenvironment.

No MeSH data available.


Related in: MedlinePlus

VLA-4 expressed on tri12 CLL cells is not in a pre-activated conformationBinding of HUTS-21 mAb plotted versus LDV concentration. No tri12 and tri12 CLL cells were incubated with the indicated LDV concentrations in presence of an excess of HUTS-21 mAb. The fit to the data was done using the sigmoidal dose-response equation with variable slope using GraphPad Prism software. The EC50 values calculated for both subgroups indicate a resting state of the VLA-4 integrin [25]. A representative experiment of 2-3 independent experiments in the tri12 (n = 3) and no tri12 (n = 2) cohort is shown. Each point represents the mean ± SD of triplicates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4494922&req=5

Figure 5: VLA-4 expressed on tri12 CLL cells is not in a pre-activated conformationBinding of HUTS-21 mAb plotted versus LDV concentration. No tri12 and tri12 CLL cells were incubated with the indicated LDV concentrations in presence of an excess of HUTS-21 mAb. The fit to the data was done using the sigmoidal dose-response equation with variable slope using GraphPad Prism software. The EC50 values calculated for both subgroups indicate a resting state of the VLA-4 integrin [25]. A representative experiment of 2-3 independent experiments in the tri12 (n = 3) and no tri12 (n = 2) cohort is shown. Each point represents the mean ± SD of triplicates.

Mentions: Based on our observation of the strong basal interactions of unstimulated tri12 CLL cells with the VCAM-1 substrate under flow, we hypothesized a pre-activated high affinity VLA-4 conformation on these cells. Therefore, we used the conformationally sensitive anti-beta1 (CD29) antibody HUTS-21 that detects the exposed hybrid domain of the beta1 subunit. In the presence of VLA-4 ligand occupancy, absolute rates of HUTS-21 binding to activated cells are faster than the binding rates to resting cells, thus reporting the high affinity state of VLA-4 [25]. For VLA-4 ligation, we used the small molecule LDV, a peptide derived from the conserved high affinity LDV sequence of the VLA-4 binding site [21]. However, the binding kinetics of no tri12 and tri12 cells to the HUTS-21 antibody were comparable, indicating that tri12 CLL do not express pre-activated VLA-4 (Figure 5).


CXCL12-induced VLA-4 activation is impaired in trisomy 12 chronic lymphocytic leukemia cells: a role for CCL21.

Ganghammer S, Hutterer E, Hinterseer E, Brachtl G, Asslaber D, Krenn PW, Girbl T, Berghammer P, Geisberger R, Egle A, Zucchetto A, Kruschinski A, Gattei V, Chigaev A, Greil R, Hartmann TN - Oncotarget (2015)

VLA-4 expressed on tri12 CLL cells is not in a pre-activated conformationBinding of HUTS-21 mAb plotted versus LDV concentration. No tri12 and tri12 CLL cells were incubated with the indicated LDV concentrations in presence of an excess of HUTS-21 mAb. The fit to the data was done using the sigmoidal dose-response equation with variable slope using GraphPad Prism software. The EC50 values calculated for both subgroups indicate a resting state of the VLA-4 integrin [25]. A representative experiment of 2-3 independent experiments in the tri12 (n = 3) and no tri12 (n = 2) cohort is shown. Each point represents the mean ± SD of triplicates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494922&req=5

Figure 5: VLA-4 expressed on tri12 CLL cells is not in a pre-activated conformationBinding of HUTS-21 mAb plotted versus LDV concentration. No tri12 and tri12 CLL cells were incubated with the indicated LDV concentrations in presence of an excess of HUTS-21 mAb. The fit to the data was done using the sigmoidal dose-response equation with variable slope using GraphPad Prism software. The EC50 values calculated for both subgroups indicate a resting state of the VLA-4 integrin [25]. A representative experiment of 2-3 independent experiments in the tri12 (n = 3) and no tri12 (n = 2) cohort is shown. Each point represents the mean ± SD of triplicates.
Mentions: Based on our observation of the strong basal interactions of unstimulated tri12 CLL cells with the VCAM-1 substrate under flow, we hypothesized a pre-activated high affinity VLA-4 conformation on these cells. Therefore, we used the conformationally sensitive anti-beta1 (CD29) antibody HUTS-21 that detects the exposed hybrid domain of the beta1 subunit. In the presence of VLA-4 ligand occupancy, absolute rates of HUTS-21 binding to activated cells are faster than the binding rates to resting cells, thus reporting the high affinity state of VLA-4 [25]. For VLA-4 ligation, we used the small molecule LDV, a peptide derived from the conserved high affinity LDV sequence of the VLA-4 binding site [21]. However, the binding kinetics of no tri12 and tri12 cells to the HUTS-21 antibody were comparable, indicating that tri12 CLL do not express pre-activated VLA-4 (Figure 5).

Bottom Line: Here, we demonstrate that high CD49d expression in tri12 CLL is accompanied by decreased CXCR4 expression.Concordantly, in real-time kinetic analyses we found CCL21 but not CXCL12 being capable to induce inside-out VLA-4 conformational changes in this CLL subgroup.Our results provide novel insights into the peculiar clinico-biological behaviour of tri12 CLL and emphasize its specific chemokine and integrin utilization during pathophysiologically and therapeutically relevant interactions with the microenvironment.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Immunological and Molecular Cancer Research, 3rd Medical Department with Hematology, Medical Oncology, Hemostaseology, Infectious Diseases and Rheumatology, Oncologic Center, Paracelsus Medical University Salzburg, Austria.

ABSTRACT
Homing to distinct lymphoid organs enables chronic lymphocytic leukemia (CLL) cells to receive pro-survival and proliferative signals. Cytogenetic aberrations can significantly affect CLL cell compartmentalization. Trisomy 12 (tri12) defines a CLL subgroup with specific clinical features and increased levels of the negative prognostic marker CD49d, the α4-subunit of the integrin VLA-4, which is a key regulator of CLL cell homing to bone marrow (BM). Chemokine-induced inside-out VLA-4 activation, particularly via the CXCL12-CXCR4 axis, increases the arrest of various cell types on VCAM-1 presenting endothelium. Here, we demonstrate that high CD49d expression in tri12 CLL is accompanied by decreased CXCR4 expression. Dissecting functional consequences of these alterations, we observed that tri12 CLL cell homing to murine BM is not affected by CXCR4-CXCL12 blockage using AMD3100 or olaptesed pegol/NOX-A12. In line, CCL21-CCR7 rather than CXCL12-CXCR4 interactions triggered VLA-4-mediated arrests of tri12 CLL cells to VCAM-1 under blood flow conditions. Concordantly, in real-time kinetic analyses we found CCL21 but not CXCL12 being capable to induce inside-out VLA-4 conformational changes in this CLL subgroup. Our results provide novel insights into the peculiar clinico-biological behaviour of tri12 CLL and emphasize its specific chemokine and integrin utilization during pathophysiologically and therapeutically relevant interactions with the microenvironment.

No MeSH data available.


Related in: MedlinePlus