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Research Laboratory on Chronic Lymphocytic Leukemia

MEMBERS

  • Pablo Oppezzo, PhD (Head)
  • Pablo Morande, PhD (Post-doctoral position)
  • Sandra Sernbo, PhD (Post-doctoral position)
  • Cecilia Abreu, PhD (Technical Assistant)
  • Agustín Correa, PhD (Principal technical assistant)
  • Claudia Ortega, PhD (Technical Assistant)
  • Florencia Palacios, PhD (Technical Assistant)
  • Daniel Prieto, MSc (PhD student)
  • Noé Seija, BSc (MSc. student)

 

RESEARCH

Our work is focus in the haematology area, tumoral immunology and the recombinant antibodies production. It lies on the interface between biochemistry and the molecular and cell biology fields, which in combination with protein expression approaches constitute the core of our experimental designs. From its beginning in 2007, our group has had a double mission at the IP Montevideo: to develop original lines of research, and to set up a technological core facility for the expression of recombinant proteins (Recombinant Protein Platform ).

The group leader (P. Oppezzo) has background in the area of tumoral immunology and recombinant proteins production. Immuno-haematological B cell malignancies, adaptive immunity, as well as recombinant antibody production has been Oppezzo’s main investigation area for the last 10 years.

Our work concentrates in the study of Chronic Lymphocytic Leukemia (CLL) as a biological model. This haematopoietic B-cell disease follows an extremely variable course and despite the fact that treatments often induce remissions, most patients relapse and CLL remains incurable.

The dissection of the molecular basis of the interactions between cancer cells and their microenvironment is leading to the development of new treatment modalities which are aimed at manipulating the communication of tumor cells with their milieu. In this regard, CLL is an instructive example of how these relationships influence the natural history of a disease.

 

Research lines

The dissection of the molecular basis of CLL progression focusing in the interactions between leukemic B cells and their microenvironment is becoming one of our main scientific interests. In this regard two research lines have been pursued since the establishment of our group in 2007.

Role of microenvironment interactions in CLL progression

At present, many questions remain unsolved concerning the role of the microenvironment interactions in the progression of tumoral diseases. We think that CLL represents an excellent model to study these relationships between the leukemic B lymphocyte and their milieu. A detailed characterization of proliferative tumoral subsets that exist in this leukemia may shed light on the association between lymphoid tumours progression and malignant transformation. We first demonstrated that in contrast to normal circulating B-lymphocytes, in progressive CLL cases, the leukemic cells express high levels of an active AID enzyme (Oppezzo et al, Blood, 2003) and (Oppezzo et al, Blood, 2005). These results and those from other groups suggest that, over-expression of AID could play an important role in CLL disease progression. In the last years, our group has described and characterized one of the proliferative tumor subsets in this leukemia. This subpopulation express the mutagenic enzyme AID by Activation-Induced Cytidine Deaminase and is associated with expression of tumor anti-apoptotic and cell proliferation markers (Palacios and Moreno et al, Blood, 2010). We also demonstrated that the proliferative pool in Unmutated CLL patients keep activated the PI3K signalling pathway. Our results show that this activation is triggered by up regulation of the microRNA Mir-22 which in turn down regulates the tumour suppressor PTEN molecule (Palacios et al., Leukemia 2014).

Development of new prognostic and therapeutic tools in CLL

This research line is outlined by our double profile as a research/facility group. Concerning the development of prognostic markers in CLL, we previously described that the expression ratio of Lipoprotein Lipase (LPL) and metalloprotease ADAM29 is an important additional marker for the prognosis of CLL (Oppezzo et al, Blood, 2005). This data was confirmed by several groups working in CLL in the consecutive years and at present, the prognostic marker LPL is used as one of the strongest prognostic factor in a comparative analysis of RNA-based markers in CLL disease (Kaderi et al., Haematologica, 2011). Despite the usefulness of LPL for CLL prognosis, its functional role and the molecular mechanism regulating its expression remain unsolved as yet. Our recent works in this area demonstrate that an epigenetic mechanism, triggered by the microenvironment, is responsible for anomalous expression of LPL in Um CLL patients (Moreno and Abreu et al., Leukemia 2013). This results lead to speculate that LPL expression on the cellular membrane of CLL B-cells could affect their biological behavior, by favoring cell spreading, and intracellular signalling in an activated tumoral microenvironment. (Abreu et al., Leukemia & Lymphoma, 2013).

In the context of therapeutics tools related with cancer, our group is recently focused on the generation of new therapeutics molecules named Artificial Binding Proteins (Affitins). Compared with classical therapeutics antibodies Affitins are able to maintain high affinity constants even when their molecular weight remains small. This could be very useful in lymphoid neoplasms, in order to gain access into solid tissues as secondary lymphoid organs, where leukemic cells receive pro-survival signals acquiring favorable proliferative conditions. In this line, a new generation of combinatorial protein engineering technologies has been recently set up in our laboratory. The results in this line has been allowed to propose the use of Affitins as versatile selective glycosidase inhibitors and, potentially, as enzymatic inhibitors in general, that could be envisaged for futures tumor therapy strategies (Correa et al., Plos One, 2014).

 

Networking

Production of recombinant antibodies as well as new protein scaffolds targeting tumor antigens constitutes a very useful tool to evaluate different prognostic and/or therapeutic molecules in cancer. To develop new therapeutic and prognostic methods in CLL it is mandatory to constitute a CLL network, that engages a continuous and coordinate work between our group (focused in the CLL biology) and different medical groups (specialized in the management of this disease). To initiate this, in the last years our group has become a reference laboratory that performs the molecular analysis of the immunoglobulin VH genes (IgVH) in CLL. The establishment of this standard procedure as a routine laboratory practice allowed us to start a strong collaboration with clinical hematologic groups of Hospital Maciel and Hospital de Clínicas in Montevideo and with the clinical hematologic group of Academy of Medicine in Buenos Aires , Argentine. These collaborations resulted in the foundation of the first LatinAmerican CLL group (LAG-CLL) with the participation of different laboratories of Argentine, Brasil and Uruguay. The consolidation of this network was recently achieved after obtaining the funds supported by CYTED. Oppezzo’s lab is the principal coordinator of this program (2011-2014) devoted to join efforts from the principal experts in lymphoproliferative disorders in the Iberoamerican region and to consolidate the regional CLL groups. Presently, a number of successful events have been achieved including workshops and student training. In this context, the first international CLL meeting was carried out in November 15th to 17th, 2013 in Punta del Este, receiving 285 participants. (http://www.clliberoamericangroup.com)

EDUCATION-COURSES-CONGRESS

  1. First Iberoamerican meeting on Chronic Lymphocytic Leukemia. Date and place: 15-17 November 2013, Montevideo, Uruguay. Participants: 285
  2. First LatinAmerican Workshop on prognosis markers in CLL: “Fluorescence in situ hibridization (FISH) as prognosis marker in CLL”. Date and place: 26-29 May 2014, Buenos Aires, Argentina. Participants: 41
  3. Second LatinAmerican Workshop on prognosis markers in CLL: “Cytometry approaches in the prognosis of CLL”. Date and place: 16-18 November 2014, Florianopolis, Brazil. Participants: 46
  4. Third LatinAmerican Workshop on prognosis markers in CLL: “Analysis of mutational profile of immunoglobulin VH genes in CLL. Date and place: 20-22 May 2015, Montevideo, Uruguya. Participants: 42

GRANTS

  •  Fondo María Viña – Dr. Pablo Oppezzo – “Development of Artificial Binding Proteins (Affitins) to evaluate new prognosis and treatment strategies in Chronic Lymphocytic Leukemia”– 2015-2017 – ANII, Uruguay
  • CSIC, I+D2014 – Dr. Pablo Oppezzo – “Implicancias de la expresión anómala de la enzima mutagénica  AID en la progresión de la Leucemia Linfoide Crónica” – 2014-2017 –Comisión sectorial de investigación científica de la Universidad de la República, Uruguay.
  • Fondo Clemente Estable – Dra. Cecilia Abreu – “Estudios genómicos del perfil de metilación del ADN en una población tumoral leucémica sobre-expresando la enzima AID” – 2013-2014 – ANII, Uruguay.
  • Fondo Clemente Estable – Dr. Pablo Oppezzo – “Implicancias de la expresión anómala de la enzima mutagénica AID en los procesos leucémicos: Desarrollo de un modelo tumoral” – 2013-2015 – ANII, Uruguay
  • Fondo María Viñas – Dr. Pablo Oppezzo – “Expresión de la Lipoproteína Lipasa en las células B de la Leucemia Linfoide Crónica (LLC): Hacia el desarrollo de un nuevo marcador pronóstico” – 2013-2015 – ANII, Uruguay
  • Fondo CYTED – Dr. Pablo Oppezzo – “Red-iberoamericana de Leucemia Linfoide Crónica: hacia el desarrollo de nuevos marcadores pronósticos” – 2011-2014 – CYTED.
  • Proyectos Transversales IPMont – Dr. Pablo Oppezzo –. “Genomic landscape of the methylation pattern and the microRNAs/mRNAs expression in progressive patients with Chronic Lymphocytic Leukemia” – 2013-2014 – Institut Pasteur de Montevideo, Uruguay.
  • Fondo Lady Tata – Dr. Pablo Oppezzo – “Characterisation of the proliferating pool in CLL.  Is AID expression a marker of this subpopulation?” – 2008-2011 – Lady Tata Foundation, United Kingdom.

PUBLICATIONS

  1. Montamat-Sicotte D, Litzler LC, Abreu C, Safavi S, Zahn A, Orthwein A, Müschen M, Oppezzo P, Muñoz DP, Di Noia JM. HSP90 inhibitors decrease AID levels and activity in mice and in human cells. Eur J Immunol. 2015 Aug;45(8):2365-76. doi: 10.1002/eji.201545462.
  2. Morande PE, Borge M, Abreu C, Galletti J, Zanetti SR, Nannini P, Bezares RF, Pantano S, Dighiero G, Oppezzo P, Gamberale R, Giordano M. Surface localization of high-mobility group nucleosome-binding protein 2 on leukemic B cells from patients with chronic lymphocytic leukemia is related to secondary autoimmune hemolytic anemia. Leuk Lymphoma. 2015 Apr;56(4):1115-22. doi: 10.3109/10428194.2014.957205.
  3. Palacios F, Prieto D, Abreu C, Ruiz S, Morande P, Fernández-Calero T, Libisch G, Landoni AI, Oppezzo P. Dissecting chronic lymphocytic leukemia microenvironment signals in patients with unmutated disease: microRNA-22 regulates phosphatase and tensin homolog/AKT/FOXO1 pathway in proliferative leukemic cells. Leuk Lymphoma. 2015 May;56(5):1560-5. doi: 10.3109/10428194.2014.990900.
  4. Palacios F, Abreu C, Prieto D, Morande P, Ruiz S, Fernández-Calero T, Naya H, Libisch G, Robello C, Landoni AI, Gabus R, Dighiero G, Oppezzo P. Activation of the PI3K/AKT pathway by microRNA-22 results in CLL B-cell proliferation Leukemia. 2014 Jan;29(1):115.
  5. Fischer S, Echeverría N, Moratorio G, Landoni AI, Dighiero G, Cristina J, Oppezzo P, Moreno P. Human endogenous retrovirus np9 gene is over expressed in chronic lymphocytic leukemia patients. Leuk Res Rep. 2014 Jul 25;3(2):70-2. doi: 10.1016/j.
  6. Borge M, Lenicov FR, Nannini PR, De Los Ríos Alicandú MM, Podaza E, Ceballos A, Grecco HF, Cabrejo M, Bezares RF, Morande PE, Oppezzo P, Giordano M, Gamberale R. The expression of sphingosine-1 phosphate receptor-1 in chronic lymphocytic leukemia cells is impaired by tumor microenvironmental signals and enhanced by piceatannol and R406 J Immunol. 2014 Sep 15;193(6):316.
  7. Libisch MG, Casás M, Chiribao M, Moreno P, Cayota A, Osinaga E, Oppezzo P, Robello C. GALNT11 as a new molecular marker in chronic lymphocytic leukemia. Gene. 2014 Jan 1;533(1):270.
  8. Oppezzo P, Dighiero G. Role of the B-cell receptor and the microenvironment in chronic lymphocytic leukemia’. Blood Cancer J. 2013 Sep 20;3:e149. doi: 10.1038/bcj.2013.45.
  9. Montamat-Sicotte D, Palacios F, Di Noia JM* and Oppezzo P*. Origins and consequences of AID expression in lymphoid neoplasms. Current Immunology Reviews, 2013; 9:75-85 .
  10. Almejun MB, Cols M, Zelazko M, Oleastro M, Cerutti A, Oppezzo P, Cunningham-Rundles C, Danielian S. Naturally occurring mutation affecting the MyD88-binding site of TNFRSF13B impairs triggering of class switch recombination. Eur J Immunol. 2013 Mar; 43(3):805-14. doi: 10.1002/eji.201242945. Impact Factor: 4.97
  11.  Moreno P, Abreu C, Borge M, Palacios F, Morande P, Pegazzano M, Bianchi S, Landoni AI, Agrelo R, Giordano M, Dighiero G, Gamberale R, Oppezzo P. Lipoprotein lipase expression in unmutated CLL patients is the consequence of a demethylation process induced by the microenvironment. Leukemia. 2013 Mar; 27(3):721-5. doi:1038/leu.2012.212.
  12. Rego N, Bianchi S, Moreno P, Persson H, Kvist A, Pena A, Oppezzo P, Naya H, Rovira C, Dighiero G, Pritsch O. Search for an aetiological virus candidate in chronic lymphocytic leukaemia by extensive transcriptome analysis. Br J Haematol. 2012 Jun; 1 57(6):709-17. doi: 10.1111/j.1365-2141.2012.09116.x.
  13. Hubert P, Heitzmann A, Viel S, Nicolas A, Sastre-Garau X, Oppezzo P, Pritsch O, Osinaga E, Amigorena S. Antibody-dependent cell cytotoxicity synapses form in mice during tumor-specific antibody immunotherapy. Cancer Res. 2011 Aug 1; 71(15):5134-43. doi: 10.1158/0008-5472.CAN-10-4222.
  14. Nannini PR, Borge M, Mikolaitis VC, Abreu C, Morande PE, Zanetti SR, Oppezzo P, Palacios F, Ledesma I, Bezares RF, Giordano M, Gamberale R. CCR4 expression in a case of cutaneous Richter’s transformation of chronic lymphocytic leukemia (CLL) to diffuse large B-cell lymphoma (DLBCL) and in CLL patients with no skin. Eur J Haematol. 2011 Jul;87(1):80-6. doi:10.1111/j.1600-0609.2011.01613.x.
  15.  Bianchi S, Moreno P, Landoni AI, Naya H, Oppezzo P, Dighiero G, Gabús R, Pritsch O. Immunoglobulin heavy chain V-D-J gene rearrangement and mutational status in Uruguayan patients with chronic lymphocytic leukemia. Leuk Lymphoma. 2010 Nov; 51(11):2070-8. doi: 10.3109/10428194.2010.522283.
  16.  Palacios F, Moreno P, Morande P, Abreu C, Correa A, Porro V, Landoni AI, Gabus R, Giordano M, Dighiero G, Pritsch O, Oppezzo P. High expression of AID and active class switch recombination might account for a more aggressive disease in unmutated CLL patients: link with an activated microenvironment in CLL disease. 2010 Jun 3; 115(22):4488-96. doi: 10.1182/blood-2009-12-257758.
  17. Galletti J, Cañones C, Morande P, Borge M, Oppezzo P, Geffner J, Bezares R, Gamberale R, Giordano M. Chronic lymphocytic leukemia cells bind and present the erythrocyte protein band 3: possible role as initiators of autoimmune hemolytic J Immunol. 2008 Sep 1; 181(5):3674-83.

Books

Book Chapter:

F. Palacios, C. Abreu, P. Moreno, M. Giordano, R. Gamberale and P. Oppezzo. Microenvironment Interactions in Chronic Lymphocytic Leukemia: A Delicate Equilibrium Linking the Quiescent and the Proliferative Pool. In the book “Chronic Lymphocytic Leukemia” edited by Pablo Oppezzo, ISBN 978-953-307-881-6, InTech, February 2, 2012

Book Edition:

“Chronic Lymphocytic Leukemia” edited by Pablo Oppezzo, ISBN 978-953-307-881-6, InTech, February 2, 2012

CONTACT

Dr Pablo Oppezzo

e-mail:poppezzo@pasteur.edu.uy