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Kaucka M.,Masaryk University | Krejci P.,Masaryk University | Krejci P.,Academy of Sciences of the Czech Republic | Krejci P.,Cedars Sinai Medical Center | And 11 more authors.
Acta Physiologica | Year: 2011

Aim: In this study, we analysed the post-translational modification of receptor tyrosine kinase-like orphan receptor (Ror1). Ror1 is highly upregulated in B cells of patients with chronic lymphocytic leukaemia (CLL). Molecularly, Ror1 acts as the Wnt receptor in the non-canonical Wnt pathway. Methods: The level of Ror1 glycosylation in HEK293 cells and in primary human CLL cells was analysed by treatment of inhibitors interfering with different steps of glycosylation process and by direct treatment of cell lysates with N-glycosidase. Ror1 ubiquitination was determined by ubiquitination assay. Functional consequences of post-translational modifications were analysed by immunohistochemistry and by analysis of cell surface proteins. Differences in Ror1 glycosylation were confirmed by analysis of 14 samples of B cells from CLL patients. Results: We demonstrate that Ror1 is extensively modified by N-linked glycosylation. Glycosylation produces several variants of Ror1 with electrophoretic migration of approx. 100, 115 and 130kDa. Inhibition of glycosylation interferes with cell surface localization of the 130-kDa variant of Ror1 and prevents Ror1-induced formation of filopodia. Moreover, we show that 130-kDa Ror1 is mono-ubiquitinated. Furthermore, individual CLL patients show striking differences in the electrophoretic migration of Ror1, which correspond to the level of glycosylation. Conclusion: Our data show that Ror1 undergoes complex post-translational modifications by glycosylation and mono-ubiquitination. These modifications regulate Ror1 localization and signalling, and are highly variable among individual CLL patients. These may suggest that Ror1 signals only in a subset of CLL patients despite Ror1 levels are ubiquitously high in all CLL patients. © 2011 The Authors. Acta Physiologica © 2011 Scandinavian Physiological Society.


Kaucka M.,Institute of Experimental Biology | Plevova K.,CEITEC Central European Institute of Technology | Plevova K.,University Hospital Brno and Medical Faculty | Pavlova S.,CEITEC Central European Institute of Technology | And 25 more authors.
Cancer Research | Year: 2013

The planar cell polarity (PCP) pathway is a conserved pathway that regulates cell migration and polarity in various contexts. Here we show that key PCP pathway components such as Vangl2, Celsr1, Prickle1, FZD3, FZD7, Dvl2, Dvl3, and casein kinase 1 (CK1)-e are upregulated in B lymphocytes of patients with chronic lymphocytic leukemia (CLL). Elevated levels of PCP proteins accumulate in advanced stages of the disease. Here, we show that PCP pathway is required for the migration and transendothelial invasion of CLL cells and that patients with high expression of PCP genes, FZD3, FZD7, and PRICKLE1, have a less favorable clinical prognosis. Our findings establish that the PCP pathway acts as an important regulator of CLL cell migration and invasion. PCP proteins represent an important class of molecules regulating pathogenic interaction of CLL cells with their microenvironment. © 2013 American Association for Cancer Research.


PubMed | University Hospital Brno and Medical Faculty and Masaryk University
Type: Journal Article | Journal: British journal of haematology | Year: 2016

The canonical Wnt pathway, dependent on -catenin-controlled transcription, is the most explored Wnt pathway, known to drive the malignant transformation of multiple cell types. Several reports have suggested that this pathway also participates in chronic lymphocytic leukaemia (CLL) pathogenesis. To get a better insight into the role of the Wnt/-catenin pathway in CLL we analysed in detail the expression of the most overexpressed Wnt ligand, encoded by the WNT3 gene, in a well-defined cohort of 137 CLL patients. Our analysis demonstrated that (i) untreated patients with more aggressive disease (with a notable exception of patients with 11q deletion) express less WNT3, (ii) WNT3 declines with disease progression in a significant proportion of patients and (iii) low WNT3 was identified as a strong independent marker indicating shorter treatment-free survival in CLL patients with IGHV mutation. Interestingly, CLL-related lymphoid cell lines, but not stromal cells, failed to respond to the ligand-induced activation of the Wnt/-catenin pathway. This opens the possibility that CLL cells use Wnt-3 to communicate with the cells in the microenvironment. We thus propose that the Wnt/-catenin pathway plays a more complex role in CLL pathogenesis than previously anticipated.


PubMed | University Hospital Brno and Medical Faculty, Azienda Ospedaliera Pugliese Ciaccio, University of Barcelona and UOC Ematologia
Type: | Journal: American journal of hematology | Year: 2017

Rai and Binet staging systems are important to predict the outcome of patients with CLL but do not reflect the biologic diversity of the disease nor predict response to therapy, which ultimately shape patients outcome. We devised a biomarkers-only CLL prognostic system, based on the two most important prognostic parameters in CLL (i.e. IGHV mutational status and FISH cytogenetics), separating three different risk groups: (1) low-risk (mutated IGHV+no adverse FISH cytogenetics [del(17p), del(11q)]); (2) intermediate-risk (either mutated IGHV or adverse FISH cytogenetics); and (3) high-risk (unmutated IGHV+adverse FISH cytogenetics. In 524 unselected subjects with CLL, the 10-year overall survival was 82% (95% CI 76-88%), 52% (45-62%), and 27% (17-42%) for the low-, intermediate-, and high-risk groups, respectively. Patients with low-risk comprised around 50% of the series and had a life expectancy comparable to the general population. The prognostic model was fully validated in two independent cohorts, including 417 patients representative of general CLL population and 337 patients with Binet stage A CLL. The model had a similar discriminatory value as the CLL-IPI. Moreover, it applied to all patients with CLL independently of age, and separated patients with different risk within Rai or Binet clinical stages. The biomarkers-only CLL prognostic system presented here simplifies the CLL-IPI and could be useful in daily practice and to stratify patients in clinical trials. This article is protected by copyright. All rights reserved.

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