Sub theme 2.2
Molecular determinants of therapy response and outcome in childhood cancer


Goals of research: general outline
Scientific achievements
Future plans: special goals and approach
Running projects
Associated staff

Workgroup leaders   Department
Dr.  M.L.  den  Boer   Pediatrics
drs.  M.M.  van den  Heuvel-Eibrink   Pediatrics Hematology
dr.  J.P.P.  Meijerink   Pediatrics
MD, PhD  C.M.  Zwaan   Pediatrics Oncology

Goals of research: general outline

The pediatric oncology research program has the following aims:

  • Developing clinically and biologically relevant classification of childhood cancers, especially leukemia. This is done by genome-wide screening techniques on DNA level (array CGH), gene expression profiling and microRNA profiling and protein profiling of serum, cerebrospinal fluid and leukemic cells.
  • Identification of (epi)genetic abnormalities involved in  the development of genetic subtypes of leukemia and solid tumors including brain tumors in children. Especially the role of gene silencing of tumor suppressor genes by methylation is analysed.
  • Identification of molecules associated with chemotherapy resistance. New resistance genes are identified by comparing gene expression profiles of therapy resistant and sensitive patients which is confirmed by other techniques. New strategies to modulate the effects of these resistance genes are developed by functional studies.
  • Identification of new therapeutic targets and development of new treatment strategies for children with cancer. New targets are identified and confirmed by techniques described above in specific subgroups of patients. This is followed by validation studies to define the effect of inhibition of the target genes, a.o. by RNA interference. The efficacy of new drugs directed against these targets is tested in preclinical models.
  • Following this preclinical phase, new drugs are tested in clinical phase I and II studies in children. The Department of Pediatric Oncology of the Erasmus MC-Sophia Childrens Hospital is part of the European ITCC (Innovative Therapies for Children with Cancer) network to perform these studies. The research laboratory of Pediatric Oncology is responsible for the preclinical studies in especially leukemia in this consortium and plays a leading role in the clinical phase I/II studies.
  • The clinical part of the program includes also: (1) Phase III studies in pediatric oncology; (2) Improvements in supportive care, which are essential because of the intensive treatment; (3) Late effects of treatment and quality of life, which are important because of the improved cure rate (70%) of childhood cancer.

The ultimate goal of the research program is to develop targeted therapies for children with cancer. This should lead to more effective and less toxic treatment strategies. The program is mainly directed to children with hematological malignancies but also to brain tumors and solid tumors.


Scientific achievements

The research program has a preclinical and a clinical part. The preclinical part is based on 5 working groups and the clinical part on 2 groups. Here the 5 working groups related to molecular medicine will be described: 

 

1.     Molecular markers and drugable targets in pediatric acute leukemia (working group leader dr ML den Boer)

 

MiRNAs contribute to proliferation and differentiation by repressing gene expression at post-transcriptional level, i.e. translation from RNA to proteins. Several miRNAs co-localize with known genetic abnormalities in leukemia and we recently discovered new miRNAs whose expression differed between genetic subclasses of ALL. In the last years we have shown gene expression profiles for the different genetic subclasses of ALL. Using genome-wide gene expression analysis we recently identified a new genetic subclass that encomprises about 15% of all children with ALL and which is associated with a very poor outcome. In the last few years we have detected gene signatures associated with resistance to glucocorticoids and asparaginase. Our laboratory is the key-laboratory for the preclinical analysis of new drugs in childhood leukemia in the European network of the ITCC (Innovative Therapies for Children with Cancer).

 

2.     T-cell acute lymphoblastic leukemia (working group leader dr JPP Meijerink)

 

About half of children with T-ALL are characterized by specific chromosomal translocations affecting T-cell receptor genes in the vicinity of proto-oncogenes leading to activation of these genes. The significance of these abnormalities for therapy resistance and clinical outcome has been determined in the last 5 years by us and others. Cells from the other half of children with T-ALL without known genetic abnormalities have been analyzed by gene expression arrays and array-CGH for detection of new genetic abnormalities. We have discovered several new recurrent genetic abnormalities in childhood T-ALL; subsequent functional knock in and –out studies showed how these abnormalities play a role in the development of T-ALL.

 

3.     Infant acute lymphoblastic leukemia (dr RW Stam)

 

About 80% of ALL in infants (diagnosed before the age of 1 year) is characterized by a chromosomal translocation involving the MLL gene at chromosome 11q23 while only 1% of children above one year carry this abnormality. Infant ALL has a very poor prognosis (40-50% survival). In the last years we chararterized the drug resistance profile of MLL rearranged ALL cells. Based upon this, a large international collaborative treatment protocol for infant ALL was developed and coordinated by our group, that resulted in a significant improvement in survival for these infants. Also, FLT3 overexpression was detected and shown to be a suitable treatment target both in in vitro and in vivo models. This has resulted in a phase I/II study with FLT3 inhibitors in MLL rrearranged ALL starting in 2009. 

 

4.     Myeloid malignancies (working group leaders dr. C.M. Zwaan, Dr M.M. van den Heuvel-Eibrink)

 

This working group aims to identify molecular determinants for therapy response and outcome and to identify new therapeutic targets and develop new therapies for children with myeloid malignancies, myelodysplastic syndrome, acquired bone marrow failure and Down Syndrome leukemias. In the past years, gene expression profiling were analysed of ~ 300 children with AML. The potential of these gene expression profilies for classification of AML, as well as identification of new leukemia subgroups was established. New molecular-genetic aberrations in MLL-rearranged pediatric AML were detected. The differential impact of the various translocation partners on outcome was analyzed in a large international study. The molecular background of normal karyotype AML has been partly unraveled. The role of mutation independent constitutive FLT3 expression in JMML, the occurrence of B-RAF gene mutations in JMML, and the diagnostic value of TCRVbeta analysis were established.

 

5.     Epigenetics in childhood cancer (Dr M. van Noesel and Dr E. Michiels)

 

Pediatric solid tumors and brain tumors are the result of developmental defects during organ formation. The organ specific gene expression regulation leading to normal development is under tight epigenetic control of the genome. This research program aims at studying the disruption of the epigenetic regulation of genes in pediatric solid tumors. 1. Special focus is aimed at the structure of the chromatin in relation to gene expression as a central epigenetic control mechanism. 2.A second focus is on post-translational regulation of gene expression by small interfering RNA molecules or microRNA (miRNA). 3. Since most epigenetic aberrations can be corrected in vitro and in vivo by specific compounds, understanding epigenetic dysregulation of pediatric tumors can lead to the introduction of targetted drugs in the treatment of patients. Pre-clinical studies have determined the most effective drug combination and doses. Expression of the genome as determined by gene expression arrays before and after drug treatment renders the molecular effectiveness of epigenetic drugs in vitro.


Future plans: special goals and approach

1.     Molecular markers and drugable targets in pediatric acute leukemia (working group leader dr ML den Boer)

 

In the coming 5 years we will identify which (novel) miRNA encoding genes contribute to leukemogenesis and response to therapy in the different genetic subclasses of leukemia by studying expression as well as function.

In the coming years we will study the clinical application of these expression signatures in comparison to the diagnostic techniques currently in use. In about one quarter of ALL cases the underlying genetic abnormality is unknown. We will characterize the genetic and molecular defects in this novel subtype aiming to point to drugable targets. We will perform functional studies to analyze these novel signaling pathways that are disturbed in drug resistant ALL cells to develop new strategies to circumvent drug resistance. Example of this is the interference with glucose metabolism in glucocorticoid resistant cells. Using new generation proteomic techniques we are identifying aberrantly expressed proteins in plasma, cerebrospinal fluid and cellular lysates of leukemic patients. Studies in the coming 5 years aim to detect and further analyse these proteins and will also focus on activating mutations and functional analysis of kinase and phosphatase encoded genes. Many studies will be performed in this network in the forthcoming years.

 

2.     T-cell acute lymphoblastic leukemia (working group leader dr JPP Meijerink)

 

We will further investigate whether specific abnormalities in pathways that are involved in normal T-cell development are important for the pathogenesis and maintenance of T-ALL. Based upon the findings, it will be analyzed whether classification and stratification of T-ALL cases can be improved with possible better risk-adapted therapies. A mouse model will be established in which the role of the newly discovered genetic abnormalities and pathways in T-cell leukemogenesis can be studied in vivo.

 

3.     Infant acute lymphoblastic leukemia (dr RW Stam)

 

Other aims of the program of the coming 5 years are to characterize this malignancy more accurately in order to gain insights in the biology of this disease. identify genes that are characteristically expressed in the different genetic subtypes of infant MLL rearranged ALL by comparing gene expression profiling of a large group of infants with t(4;11), t(;11) and t(9;11). The expression of the identified genes will be modulated to study their role in leukemic cell survival and proliferation, followed by studying the potential of selected genes to serve as therapeutic target. Studies will be performed to further understand the biology of MLL translocation driven leukemogenesis. Other studies will investigate underlying mechanisms of therapy resistance, especially to glucocorticoids and asparaginase. Finally, the epigenetic character of this malignancy will be studied in relation to leukemogenesis and to exploiting these features to develop more specific therapeutic strategies using epigenetic drugs. Presently, we shall extend our studies in MLL-rearranged infant ALL by initiating studies focusing on the role of the bone marrow microenvironment and the chemo-protective interactions of MLL-rearranged ALL cells with the bone marrow stromal niche.

 

4.     Myeloid malignancies (working group leaders dr. C.M. Zwaan, Dr M.M. van den Heuvel-Eibrink)

 

Further studies in this will include kinome analysis by reverse phase protein arrays and flow cytometry. The prognostic significance of newly identified abnormalities will be studied. Validation studies will include expression levels, protein expression, and knock-down experiments to study tumor-cell dependency. A special target group that will be studied are children with Down syndrome leukemias. National and international studies will be perfomed by our group to identify which genetic aberrations drive the progression from transient myeloproliferative disease occurring in newborns to true AML later in life in these children. These studies include kinome analysis, array-CGH and gene expression profiling, as well studying the role of micro-RNAs. This project also includes various clinical studies, such as the prognostic significance of cytogenetic abnormalities in Down AML, and a screening study for transient myeloproliferative disease (TMD) in newborns in the Netherlands.Children with Down syndrome also have an increased risk of developing ALL, which is characterized by a lack of well-known genetic aberrations when compared with sporadic ALL. In fact, 50% of patients have a normal karyotype. We therefore aim at identifying the genetic aberrations in this disease, using techniques as mentioned for Down AML. Other projects will identify molecular diagnostic tools and drugable targets in myelodysplastic syndrome (MDS) and juvenile myelomonocytic leukemia (JMML). New projects are planned on the role of micro-RNAs in pediatric AML, as well as signal transduction abnormalities in pediatric AML, using reverse phase protein arrays and flowcytometric screening of aberrant signal transduction pathways.

 

5.     Epigenetics in childhood cancer (Dr M. van Noesel and Dr E. Michiels)

 

Specific aims for research for the coming 5 years are focused on genome wide DNA methylation patterns of pediatric neuroblastoma. DNA methylation induces gene silencing and methylation patterns are tumor specific. Genome wide analysis of methylation by using agilent CpG-island methylation arrays can determine genes and pathways disrupted or silenced in neuroblastomas and enables further classification and prognostification of neuroblastoma tumors based on methylation patterns. We aim to study the effectiveness of epigenetic drugs (DNA demethylating agents and HDAC inhibitors) in vitro in neuroblastoma cell lines. Neuroblastoma tumors and cell lines will be used as a model for future research of additional pediatric solid tumors. Other projects aim to identify causative and prognostic genetic mutations in brain tumors, especially medulloblastomas, ependymomas and low grade gliomas. This has been done by proteomic techniques as well as genome wide analyses. New studies will focus on abnormally expressed miRNAs and will study the functional role of these in medulloblastomas and ependymomas.


Most recent publications
  1. Holleman A/Cheok MH, den Boer ML, Yang W, Veerman AJ, Kazemier KM, Pei D, Cheng C, Pui CH, Relling MV, Janka-Schaub GE, Pieters R/Evans WE. Gene-expression patterns in drug-resistant acute lymphoblastic leukemia cells and response to treatment.  N Engl J Med 2004;351:533-42. (IF 50.0)
  2. Pieters R, Schrappe M, de Lorenzo P, Hann I, de Rossi G, Felice M, Hovi L, Leblance T, Szczepanski T, Ferster A, Janka G, Rubnitz J, Silverman L, Stary J, Campbell M, Li CH, Mann G, Suppiah R, Biondi A, Vora A, Valsecchi MG. A treatment protocol for infants younger than 1 year with acute lymphoblastic leukaemia (Interfant-99) : an observational study and a multicentre randomised trial. Lancet 2007; 370:240-250. (IF 28.4)
  3. Boer den ML, van Slegtenhorst M, De Menezes RX, Cheok MH, Buijs-Gladdines JGCAM, Peters STCJM, van Zutven LJCM, Beverloo HB, van der Spek PJ, Escherich G, Horstmann MA, Janka/Schaub GE, Kamps WA, Evans WE, Pieters R. A subtype of childhood acute lymphoblastic leukaemia with poor treatment outcome± a genome-wide classification study. Lancet Oncol 2009;10:125-134. (IF 13.2)
  4. Lugthart S/Cheok MH, den Boer ML, Yang W, Holleman A, Cheng C, Pui CH, Relling MV, Janka-Schaub GE, Pieters R/Evans WE. Identification of genes associated with chemotherapy crossresistance and treatment response in childhood acute lymphoblastic leukemia. Cancer Cell 2005;7:375-386. (IF 24.9)
  5. Armstrong SA, Staunton JE, Silverman LB, Pieters R, den Boer ML, Minden MD, Sallan SE, Lander ES, Golub TR, Korsmeyer SJ. MLL translocations specify a distinct gene expression profile, distinguishing a unique leukemia. Nature Genetics 2002;30:41-47. (IF 30.2)
  6. Armstrong SA, Kung AL, Mabon ME, Silverman LB, Stam RW, den Boer ML, Pieters R, Kersey JH, Sallan SE, Fletcher JA, Golub TR, Griffin JD, Korsmeyer SJ. Inhibition of FLT3 in MLL: Validation of a therapeutic target identified by gene expression based classification. Cancer Cell 2003:3:173-183. (IF 24.9)
  7. Lahortiga I, de Keersmaecker K, van Vlierberghe P, Graux C, Cauwelier B, Lambert F, Mentens N, Beverloo HB, Pieters R, Speleman F, Odero MD, Bauters M, Froyen G, Marynen P, Vandenberghe P, Wlodarska I, Meijerink JPP, Cools J. Duplication of the MYB oncogene in T cell acute lymphoblastic leukemia. Nat Genet 2007; 39:593-595. (IF 30.2)
  8. Boer den ML, Harms DO, Pieters R, Kazemier KM, Göbel U, Körholz D, Graubner U, Haas RJ, Jorch N, Spaar HJ, Kaspers GJL, Kamps WA, van der Does-van den Berg A, van Wering ER, Veerman AJP, Janka-Schaub GE. Patient stratification based on prednisolone-vincristine-asparaginase resistance profiles in children with acute lymphoblastic leukemia. J Clin Oncol 2003;17:3262-3268. (IF 17.1)
  9. Wei G, Twomey D, Lamb J, Schlis K, Agarwal J, Stam RW, Opferman JT, Sallan SE, den Boer ML, Pieters R, Golub TR, Armstrong SA. Gene expression-based chemical genomics identifies rapamycin as a modulator of MCL1 and glucocorticoid resistance. Cancer Cell 2006;10:331-342. (IF 24.9)
  10. Viseur le C, Hotfilder M, Bomken S, Wilson K, Röttgers S, Schrauder A, Rosemann A, Irving J, Stam RW, Shultz J, Harbott J, Jürgens H, Schrappe M, Pieters R, Vormoor J. In childhood acute lymphoblastic leukemia, blasts at different stages of immunophenotipic maturation have stem cell properties. Cancer Cell 2008;14:47-58. (IF 24.9).

People associated with sub theme 2.2: Molecular determinants of therapy response and outcome in childhood cancer
Name Position Department
A.M.   Aalbers PhD-candidate Pediatrics Oncology
MD  F.  Akbari Moqadam Postdoc/junior researcher Pediatrics Oncology
I.M.  van  Baarsen PhD-candidate Pediatrics Oncology
A.M.  BECHAKRA PhD-candidate Neurology
Dr.  A.  Beishuizen Senior researcher not being a workgroup leader
Dr.  A.  von  Bergh Senior researcher not being a workgroup leader
Dr.  M.L.  den  Boer Workgroup leader within the School Pediatrics
Drs.  J.M.  de  Bont PhD-candidate Pediatrics
drs.  A.  Broyl PhD-candidate Hematology
MSc  E.  Clemens PhD-candidate Pediatrics Oncology
MSc.  T  Deger PhD-candidate Medical Oncology
? ?  Ladyofrage  DentalHah PhD-candidate General Surgery
BSc  J.J.  Deuring PhD-candidate Gastro-enterology and Hepatology
L.  Douglas Technician Pathology
Drs.  E.M.C.  Driessen PhD-candidate Pediatrics Oncology
M.  van  Duin Postdoc/junior researcher Hematology
A.R.  Geelen Other as mentioned (please specify in remarks) Neuro Oncology
Drs.  A. W.   Gotink PhD-candidate Gastro-enterology and Hepatology
drs.  M.  van  Grotel PhD-candidate Pediatrics
MSc  J.A.P.  Hagelstein PhD-candidate Pediatrics Oncology
QettoE  HenryGer Physician (MD) General Surgery
M.C.H.  Hermkens PhD-candidate Pediatrics Oncology
drs.  M.M.  van den  Heuvel-Eibrink Workgroup leader within the School Pediatrics Hematology
M.A.H.  den  Hoed PhD-candidate Pediatrics Oncology
dr.  A.  Holleman PhD-candidate
Dr.  E.  Hulleman Postdoc/junior researcher Pediatrics Oncology
M.D.  Hundera MSc student (please specify in remarks) Student
I.S.  Jerchel PhD-candidate Pediatrics Oncology
MSc  FC  de  Jong PhD-candidate Urology
Drs.  M.F.  van der   Kamp PhD-candidate Pathology
J.E.  Kuipers PhD-candidate Pediatrics Oncology
MD  C.F.  Kweldam Physician (MD) Pathology
Ing.  E.A.M.   Lange-Turenhout Technician Pediatrics Oncology
Dr.  E.M.M.  van  Lieshout Senior researcher not being a workgroup leader Surgery
M.D.  K.C.  Mahabier PhD-candidate Surgery
drs  M.  Meier PhD-candidate Pediatrics Oncology
dr.  J.P.P.  Meijerink Workgroup leader within the School Pediatrics
S.H.  van  Olphen PhD-candidate Gastro-enterology and Hepatology
F.S.  Peters PhD-candidate Internal Medicine
Prof.dr.  R.  Pieters Senior researcher not being a workgroup leader Pediatrics
M.A.  Rijlaarsdam Other as mentioned (please specify in remarks) Molecular Biology
Drs  J.D.E.  de  Rooij PhD-candidate Pediatrics Oncology
D.  Sá da Bandeira PhD-candidate Pediatrics Oncology
R.M.  dos  Santos Moreira Pedrosa Technician Pathology
PharmD  S.D.T.  Sassen PhD-candidate Pediatrics Oncology
Dr.  D.  Schotte Physician (MD) Pediatrics Oncology
-  Tmmaverick  SEOmab PhD-candidate General Surgery
Muhammad  Shahid PhD-candidate Gastro-enterology and Hepatology
dr.  R.W.S.  Stam Postdoc/junior researcher Pediatrics
drs.  D.J.P.M.  Stumpel PhD-candidate Pediatrics Oncology
Ir.  S.M.  Sun PhD-candidate Hematology
MD  A.F.L.T.R.  Tibúrcio Ribeiro Physician (MD) Hematology
H.  Vekony Postdoc/junior researcher Pathology
PhD  C.  van de  Ven Postdoc/junior researcher Pediatrics Oncology
MSc  M.J.  Vuerhard PhD-candidate Pediatrics Oncology
mining dogecoin  Boogieq  WilliamEroff Professor not being a workgroup leader General Surgery
MD, PhD  C.M.  Zwaan Workgroup leader within the School Pediatrics Oncology