The impact of UBTF Tandem Duplications in Pediatric Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is a type of blood cancer caused by genetic mutations in the stem cells residing in the bone marrow. AML in children is a serious disease, with only approximately half surviving. Recently, scientists have discovered a new genetic change in a gene named UBTF mainly found in children and young adults. This genetic change (UBTF-TD) is associated with a more aggressive form of AML with a worse survival rate (approximately 40% in 3 years). UBTF-TD often occurs with other common genetic mutations (especially the genes FLT3 and WT1). To better understand how UBTF-TD leads to AML combined with or without WT1/FLT3 mutations, we studied bone marrow samples from patients with specific genetic changes. We used state-of-the-art sequencing techniques to decipher the gene expression, genetic changes, and DNA accessibility to the protein regulators within each cell. Our initial findings suggest that UBTF-TD and its mutational combinations affect the way that the immune system interacts with the cancer cells and also modify critical upstream regulatory pathways. We believe that the UBTF-TD mutation may start the cancer process, and the FLT3/WT1 mutations worsen it by changing the behavior of the affected cells. Our research aims to uncover precisely how these genetic changes work together to drive pediatric AML. It also proposes and tests new drug treatments that can target this oncogenic event (UBTF-TD), hopefully leading to novel curative drug combinations.

Therapeutic targeting of CDK12/13 and PARP in H3G34RN mutant pediatric glioma

One common feature of pediatric brain tumors is the excessive activation of genes that promote the survival and proliferation of the brain cancer cells. A major goal of this research is to identify therapeutic targets to dampen this excessive activation of genes as a strategy for reducing pediatric brain tumor growth. We are pursuing two candidate therapeutic targets called CDK12 and 13 in H3G34R/V mutant pediatric high grade glioma, which are a particularly deadly form of childhood glioma. Currently, there are no drugs are specifically designed to treat children diagnosed with these H3G34R/V mutant glioma. Our preliminary studies indicate that H3G34R/V mutant pediatric brain tumor cells are more sensitive to drugs inhibiting two regulators of cancer gene activation called CDK12 and 13 than normal cells. This finding suggests that these drugs might be used as a treatment for brain tumors. We will determine how CDK12 and 13 regulate pediatric brain tumor growth and unravel roles for CDK12/13 in activating brain tumor-promoting genes. Our further studies will focus on using mouse models of glioma to develop multi-drug combinations of CDK12/13 inhibitors and other drugs under investigation for cancer treatment. This research may lead to new approaches for treating childhood brain tumors with no cure by increasing our understanding of how CDK12/13 might promote brain tumor growth on a mechanistic level, and through the development of CDK12 and 13-targeting combination therapies.

HBEGF as a novel therapeutic target for Diffuse Midline Glioma

Diffuse Midline Glioma or DMG is a type of brain cancer that arises in children and is still mostly incurable. While we have made progress by identifying the genetic drivers that give rise to DMG, this knowledge has so far failed to prolong the lives of most children with DMG. The Becher lab has been developing models for DMG since 2010 and thus far our modeling has required the use of a particular growth factor called PDGFA/B to activate a cell surface receptor called PDGFRA which signals internally within the cell to promote cell division, and tumor growth. Recently we have discovered that another growth factor, HBEGF, -which activates a different cell surface receptor called EGFR- can substitute for PDGF and may be an important ligand, or receptor, to model DMGs with H3.1K27M and EZHIP as co-drivers. We believe this is an important advance in the field and we will share this new model with the research community. Here we propose to characterize this new model of DMG that is driven by HBEGF and EZHIP, to study which genes EZHIP regulates in the context of HBEGF, as well as identify mechanisms of resistance to EGFR inhibition in a mouse model of DMG driven by HBEGF. In summary, this work will provide an important new research tool for the DMG community and may provide new insight as to why several inhibitors of EGFR have failed in the clinic to prolong the lives of children with DMG.

Liquid Biopsy

Liquid biopsy technologies have been employed for numerous purposes that have improved the care of patients with solid tumors including early detection, pre-treatment prognostication, precision medicine, measures of treatment response, detection of minimal residual disease and relapse, and the study of tumor evolution. In pediatrics, numerous studies (primarily from the Crompton lab) now show that circulating tumor DNA (ctDNA) levels and detection of specific variants at diagnosis are prognostic. 

In addition, changes in ctDNA levels early on-treatment also appear to be associated with outcome, a preliminary finding which is similarly being validated. Furthermore, in our published studies, ongoing work, and the published work of others, ctDNA clearly demonstrates the ability to detect prognostic and targetable lesions not identified in sequencing from single-site tumor biopsies, particularly in neuroblastoma, Wilms tumor, and Ewing sarcoma. These results are particularly relevant to sequencing to identify clinical trial and treatment opportunities at relapse. Finally, opportunities also exist for cell-free DNA studies on CSF in brain tumors.

Targeted Nanoparticles for Genetic Engineering of Human Natural Killer Cells for Neuroblastoma Therapy

Neuroblastoma (NB) is the most common solid tumor in children below 5 years of age. It is a devastating illness for children with advanced disease. Survival rates are only around 40%. Those children that do survive suffer life-long medical problems. Immunotherapy, which trains the body’s own immune cells to specifically recognize and kill tumors, offers potential for a less toxic and long-lasting cure for children with NB. However, current methods to train immune cells require removing the cells from the body, modifying them in the lab, and then administering the trained immune cells back to the patient. This process takes a long time, costs a lot, and sometimes results in immune cells that don’t work very well, making it hard for many children to benefit from immunotherapy. To address these big problems, we have developed a nanotechnology called lipid nanoparticles (LNPs) that can deliver gene editing payload for training immune cells directly within the body. In this work, we will test the ability of LNPs to effectively train immune cells to kill NB. We will first optimize the way LNPs are made to maximize training of immune cells. Then, we will test how well the LNP-trained immune cells kill tumors in mice with NB tumors. Our LNP technology has the potential to improve how well immune cells kill NB. Also, LNPs would train immune cells more efficiently. This would reduce costs and improve patient experience, increasing access to life saving immunotherapy for many children.

Cooperative mutagenesis driving pediatric pre-B cell leukemia

Acute lymphoblastic leukemia (ALL) is the most common childhood cancer. ALL, like most cancers, develops because of damage to DNA, the blueprints of every human cell. Many causes of DNA damage in adult-onset cancers are known. For example, tobacco, alcohol, and sunlight cause DNA damage that can lead to cancer. Children are not exposed to these DNA-damaging toxins in the same doses or durations as adults, therefore how DNA mistakes are acquired in pediatric cancers is unknown. We are interested in the causes of DNA damage that spur development of pediatric cancer, especially leukemia. We suspect that damage to DNA in children occurs when normal functions of cells become unregulated.

ALL arises from abnormal immune cells, specifically B cells. In normal B cell development, purposeful DNA damage occurs to optimize cell function. We hypothesize that these sources of purposeful DNA damage can go awry. We have generated a unique system to study sources of purposeful DNA damage in B cells in different contexts. We will determine 1) what causes developing B cells to be vulnerable to DNA damage, and 2) how sources of purposeful DNA damage go awry to cause leukemia to develop. We have generated unique systems to study purposeful sources of DNA damage, therefore mimicking the actual genetic events that drive healthy B cells to become cancer. Our long-term goal is to develop strategies to prevent or harness abnormal DNA damage to limit leukemia development and devise new treatments.

Therapeutic downregulation of core circuitry genes in mutant MYOD1 rhabdomyosarcoma

Rhabdomyosarcoma (RMS) is a highly aggressive childhood cancer with poor outcomes, especially in cases with a specific MYOD1 mutation (MYOD1-L122R). This mutation transforms the MYOD1 protein into a powerful cancer driver, leading to a particularly lethal form of RMS. Current therapies have been ineffective, highlighting the urgent need for new approaches. Our research aims to address this need by focusing on two main objectives. First, we will create detailed maps of how genes are regulated in RMS cells with the MYOD1-L122R mutation. By identifying the key genes and pathways controlled by the mutated MYOD1, we hope to uncover the mechanisms that drive cancer progression in this aggressive RMS subtype. Second, we have discovered that drugs targeting the PI3K signaling pathway can significantly reduce MYOD1-L122R activity and slow RMS cell growth in laboratory models. We will investigate how PI3K inhibition impacts the genetic networks controlled by MYOD1-L122R, with the goal of determining whether these inhibitors could serve as an effective treatment option for MYOD1-mutant RMS. This research has the potential to identify new therapeutic targets and explore the effectiveness of PI3K inhibitors in treating MYOD1-mutant RMS, ultimately aiming to improve survival rates for children with this devastating cancer.

Factsheet

Radiation therapy, alongside surgery and chemotherapy, is crucial for around 30% of children with kidney tumors. Radiation effectively destroys cancer cells, but it can also damage healthy tissue near the treatment area, especially in young children where growing tissues are most vulnerable. While initial side effects might seem mild, they often worsen over time as the tissue fails to grow properly. Children who undergo radiation may face physical problems later in life, such as fatigue, skin issues, pain, or even a heightened risk of developing secondary cancers.

To minimize the harmful effects of radiation, precise marking of the target area is essential. Currently, this marking is done manually by specialists, which is time-consuming and subject to variation between experts. This project aims to explore whether artificial intelligence (AI), specifically deep learning, can make this process faster and more consistent. The goal is to increase efficiency, reduce the risk of side effects, and promote broader acceptance and standardization of this treatment worldwide.

Comprehensive molecular profiling of pediatric adrenal tumors

Pheochromocytomas (PCCs) and paragangliomas (PGLs, together denoted as PPGLs) are rare endocrine tumors originating from chromaffin cells in the adrenal medulla (PCC) and extra-adrenal paraganglia (PGL). While all PPGL are considered malignant, their clinical behavior is variable. Most non-syndromic patients will not suffer from recurrent disease, but the rate of hereditary disease is high, up to 40% of patients, and these are prone to the development of a second primary tumor, that needs to be distinguished from a local or metastatic recurrence. This leads to patient anxiety and high cost of follow up investigations.

Adrenal cortical adenomas and carcinomas are rare endocrine tumors arising from the adrenal cortex. Adenomas are benign and do not need follow up after their removal, whereas carcinomas, mainly in older children frequently lead to death of the patient. Distinction of adenomas and carcinomas can be extremely difficult on the basis of the current multifactorial histological and immunohistochemical classification systems. Thus, there is a need for more accurate classification in relation to clinical behavior for PPGL as well as for adrenal cortical tumors (ACT) in children, where data are scarce, but seem to indicate that classification systems used for adults cannot be applied.

We propose to retrospectively collect a series of approximately 50 PPGL and 50 ACT, both groups enriched for patients with unfavorable clinical behavior. From all patients relevant metadata include age, sex, biochemical status, pathology report, Wieneke classification (for ACT), Ki67 labeling index, and genetic status. Frozen or paraffin- embedded tumor tissue will be used for DNA and RNA extraction according to standard methods and subjected to whole exome or whole genome sequencing, as well as whole transcriptome sequencing in the laboratory for pediatric oncology at Princess Maxima Center, according to established procedures and using automated bioinformatical analysis with manual procurement where needed. Genetic variants will be called in any of the cancer predisposition genes for which the current pipeline has been validated (n=600). Furthermore, gene expression profiles will be used for cluster analysis of tumors in relation to the same or different tumor types for which the classifier has been trained. Specifically, differential clustering within the group of PPGL or of ACT will be analyzed and gene expression profiles will be related to outcome. In depth analysis of most differentially expressed genes between groups will be done, including validation on an independent group of patients or by protein-based techniques such as immunohistochemistry.

Targeting oncogenic transcriptional programs in Juvenile Myelomonocytic Leukemia

Juvenile Myelomonocytic Leukemia (JMML) is a deadly form of childhood cancer. Most children with JMML are diagnosed before the age of three. Children with JMML receive intensive chemotherapy and often receive a bone marrow transplant—where the child’s own bone marrow is destroyed and replaced with that from a donor. Even with these harsh therapies, many children with JMML will have their leukemias return. Indeed, only half of children with JMML will survive for the first five years after their diagnosis. We need better treatments for this aggressive blood cancer. One key to developing new treatments may lie in a gene called SETBP1. Mutations in this gene make the leukemia much more likely to return. If we could target this bad-acting gene, we might be able to stop this leukemia from coming back. We have recently discovered that drugs developed for a different type of childhood leukemia might also work against SETBP1. The goal of our studies is to understand how mutations in SETBP1 cause hard-to-treat disease and to determine whether we can use these drugs to kill SETBP1-mutant leukemia cells.

Development of a novel radiopharmaceutical agent for non-invasive immunotherapy target detection and CAR T combination therapy in neuroblastoma

Neuroblastoma (NB) is a solid tumor in children. Children with disease that has spread to other organs still do poorly. Our lab has developed a CAR T therapy that uses the patient’s immune cells to combat NB. Our CAR T therapy detects a target called glypican 2 (GPC2) that is found in NB cells but not in most healthy tissues. Upon sensing and binding to GPC2, the CAR T-cells can kill NB. However, one challenge in CAR T therapy is that it is often unknown whether the patient’s tumor has the target at the time they receive CAR T therapy. To obtain this information, it is necessary to perform a biopsy which is invasive and can be challenging. No existing test can tell us how much of the target is found in the tumor. So, we propose to develop a radiotracer to do “immunoPET”. We will use an antibody called hCT3 which can bind to GPC2. hCT3 will be bound to zirconium (89Zr), which allows us to measure GPC2 levels in NB tumors and eventually identify patients who benefit the most from GPC2 CAR T therapy. We will also develop a modified version of this. Instead of zirconium (89Zr), we will attach hCT3 to actinium (225Ac). 225Ac-hCT3 can deposit radiation in the tumor, shrink it, and make it more sensitive to CAR T therapy as well. This may make CAR T therapy work better. To summarize, we will develop 89Zr-hCT3 to image immunotherapy targets without the need for invasive procedures and 225Ac-hCT3 to improve CAR T therapy. Our studies are designed to improve patient care and survival.

Pulmonary complications in children with solid tumors

Since 2015, all children in the Netherlands with a tumor in the abdomen or chest cavity are treated in the Máxima. As a result, we notice that there are children with severe pulmonary complications, as a side effect of the treatment. The treatment of these children with cancer is highly specialized and this is taken care of by the specialists of the Máxima, together with radiation doctors from the UMCU, and in the case of these serious pulmonary complications there is intensive cooperation with the pediatric pulmonologists from the WKZ. This cooperation has already led to the creation of protocols that indicate how to treat these patients well; patients with, for example, high blood pressure in the lungs (pulmonary hypertension) and inflammatory reactions after radiation, (radiation pneumonitis and toxic pneumonitis).

The aim is to perform a pilot project to gain a better understanding of the factors that play a role in the development of these serious pulmonary complications and death due to these conditions, in order to recognize them earlier, and to be able to prevent serious complications; through literature research and a retrospective analysis of the first 10-year cohort at the Maxima. This is expected to involve 80-100 children.

Data provided by this project will serve in the future (follow-up research) as the basis for a prospective KIKA application regarding pulmonary complications in children with a solid tumor.

Uncovering the role of chromosome 21 in Down syndrome-associated leukemias

Down syndrome is a genetic condition caused by the presence of an extra copy of chromosome 21. Children with Down syndrome are 150 times more likely to develop leukemia compared to other children. However, we don't fully understand why having an extra chromosome leads to cancer. Our research aims to uncover how this extra copy of chromosome 21 affects leukemia growth in children with Down syndrome.

My laboratory has developed a new technique that allows us to remove specific chromosomes from human cells. Using this method, we plan to create matched sets of leukemia cells- some with an extra chromosome 21 and some without. This will give us a powerful tool to directly compare how this extra chromosome impacts cancer. We will use this technique to discover how gaining a copy of chromosome 21 affects gene expression, cancer cell growth, and responses to chemotherapy drugs.

Through this work, we hope to gain an improved understanding of why children with Down syndrome are at a higher risk for leukemia. Additionally, by identifying key cellular processes affected by the extra chromosome, we hope to uncover new targets to treat leukemias in these children.

Clinical [89Zr]Dinutuximab PET imaging to determine GD2 expression in neuroblastoma patients with PET

We will develop a [89Zr]Dinutuximab for PET/CT to visualize and semiquantify GD2 expression in neuroblastoma for individual patients. Thereby providing insight in treatment effects and changes in GD2 expression. To achieve this we will:
1. Develop [89Zr]dinutuximab under GMP for clinical neuroblastoma imaging.
2. Dosimetry study for dose optimization in children, minimizing the radiation burden.
3. Perform a first-in-children [89Zr]dinutuximab imaging study, before and after therapy and in a relapse setting.

Androgen activity in the normal male embryonic hindbrain drives lethal PFA ependymoma

PFA ependymoma is a rare and aggressive childhood brain tumor that primarily affects infants. PFA ependymoma has a much higher incidence and worse prognosis in males than in females, but the reasons behind this difference are not well understood. Our research aims to uncover why males are more affected by PFA ependymoma and why they have worse outcomes.

Our preliminary study of a single cell RNA sequencing cohort has shown that the cellular hierarchy of male PFA is less differentiated than female PFA. Using a special mouse model, four core genotype (FCG), we were able to separate the effects of sex hormones from genetic differences in sex chromosomes. Our preliminary data has shown that increasing androgen levels stimulate the growth of PFA ependymoma; while blocking androgens reduces the growth of PFA ependymoma. This suggests that androgen signaling in both the normal developing hindbrain, and PFA ependymoma are both necessary and sufficient to promote growth and delay differentiation.

These findings point to a new potential treatment strategy: targeting androgen receptor activity could be an effective way to slow down or stop the growth of PFA ependymoma. Developing anti androgen therapies could provide a much-needed treatment option for this currently untreatable childhood cancer, potentially improving outcomes for affected children, especially males.