B-cell acute lymphoblastic leukemia (B-ALL) is the commonest childhood cancer and the prognosis of children with relapsed or therapy refractory disease remains a challenge. Treatment with chimeric ...antigen receptor-modified T cells targeting the CD19 antigen (CART-19 therapy) has been presented as a promising approach toward improving the outcome of relapsed or refractory disease. However, 10%-20% of the patients suffer another relapse. Epitope-loss under therapy pressure has been suggested as a mechanism of tumor cells to escape the recognition from CART-19 therapy. In this work, we analyzed the expression of CD19 isoforms in a cohort of 14 children with CD19 B-ALL and 6 nonleukemia donors. We showed that an alternatively spliced CD19 mRNA isoform lacking exon 2, and therefore the CART-19 epitope, but not isoforms lacking the transmembrane and cytosolic domains are expressed in leukemic blasts at diagnosis in children and in the bone marrow of nonleukemia donors. Furthermore, we clarified the sequence of a further isoform lacking the epitope recognized by CART-19 therapy and disclosed the presence of new isoforms. In comparison with the children, we showed that alternatively spliced CD19 mRNA isoforms affecting exon 2 are also expressed in 6 adult patients with CD19 B-ALL. On top of that, one of the adults expressed an isoform lacking the CD19 transmembrane and cytosolic domains. In conclusion, we proved that some of the CD19 isoforms contributing to CART-19 escape already preexist at diagnosis and could evolve as a dominant clone during CART-19 therapy suggesting the application of combined treatment approaches.
H3K27M mutant (mut) diffuse midline glioma (DMG) is a lethal cancer with no effective cure. The glycosphingolipids (GSL) metabolism is altered in these tumors and could be exploited to develop new ...therapies. We tested the effect of the glucosylceramide synthase inhibitors (GSI) miglustat and eliglustat on cell proliferation, alone or in combination with temozolomide or ionizing radiation. Miglustat was included in the therapy protocol of two pediatric patients. The effect of H3.3K27 trimethylation on GSL composition was analyzed in ependymoma. GSI reduced the expression of the ganglioside GD2 in a concentration and time-dependent manner and increased the expression of ceramide, ceramide 1-phosphate, sphingosine, and sphingomyelin but not of sphingosine 1-phosphate. Miglustat significantly increased the efficacy of irradiation. Treatment with miglustat according to dose recommendations for patients with Niemann-Pick disease was well tolerated with manageable toxicities. One patient showed a mixed response. In ependymoma, a high concentration of GD2 was found only in the presence of the loss of H3.3K27 trimethylation. In conclusion, treatment with miglustat and, in general, targeting GSL metabolism may offer a new therapeutic opportunity and can be administered in close proximity to radiation therapy. Alterations in H3K27 could be useful to identify patients with a deregulated GSL metabolism.
Despite massive improvements in the treatment of B-ALL through CART-19 immunotherapy, a large number of patients suffer a relapse due to loss of the targeted epitope. Mutations in the CD19 locus and ...aberrant splicing events are known to account for the absence of surface antigen. However, early molecular determinants suggesting therapy resistance as well as the time point when first signs of epitope loss appear to be detectable are not enlightened so far. By deep sequencing of the CD19 locus, we identified a blast-specific 2-nucleotide deletion in intron 2 that exists in 35% of B-ALL samples at initial diagnosis. This deletion overlaps with the binding site of RNA binding proteins (RBPs) including PTBP1 and might thereby affect CD19 splicing. Moreover, we could identify a number of other RBPs that are predicted to bind to the CD19 locus being deregulated in leukemic blasts, including NONO. Their expression is highly heterogeneous across B-ALL molecular subtypes as shown by analyzing 706 B-ALL samples accessed via the St. Jude Cloud. Mechanistically, we show that downregulation of PTBP1, but not of NONO, in 697 cells reduces CD19 total protein by increasing intron 2 retention. Isoform analysis in patient samples revealed that blasts, at diagnosis, express increased amounts of CD19 intron 2 retention compared to normal B cells. Our data suggest that loss of RBP functionality by mutations altering their binding motifs or by deregulated expression might harbor the potential for the disease-associated accumulation of therapy-resistant CD19 isoforms.
Background The CVSS (Cardiac and Vascular Late Sequelae in Long-Term Survivors of Childhood Cancer) study aimed to investigate the prevalence of different stages of heart failure (HF) in childhood ...cancer survivors (CCSs) compared with the general population. Methods and Results A total of 1002 CCSs (age range, 23-48 years) diagnosed with neoplasia before an age of 15 years underwent a comprehensive cardiovascular screening. An age- and sex-matched sample from the population-based GHS (Gutenberg Health Study) served as a comparison group. Although prevalence of HF was significantly higher in CCSs, prevalence of different HF stages varied strongly by specific tumor history. Compared with the population, the prevalence ratio was 2.6 (95% CI, 2.4-2.8) for HF stage A and 4.6 (95% CI, 4.1-5.1) for the composite of HF stage B to D in an age- and sex-adjusted Poisson regression model. Multivariable linear regression, adjusting for tumor entities, age, sex, and cardiovascular risk factors, revealed a lower left ventricular ejection fraction in patients with history of bone tumors (
-4.30 95% CI, -5.70 to -2.80), soft tissue sarcoma (
-1.60 95% CI, -2.90 to -0.30), and renal tumors (
-1.60 95% CI, -2.80 to -0.29) compared with the population. The same model for the diastolic marker, ratio of the peak early diastolic filling velocity/lateral mitral annular early diastolic velocity, showed an association only with cardiovascular risk factors but not with tumor entities. Conclusions The prevalence of HF stage A to D was significantly higher among long-term CCSs compared with the population and varied strongly by tumor entity. Systolic dysfunction was primarily associated with tumor entities, whereas diastolic dysfunction was associated with a higher burden of cardiovascular risk factors in CCSs.
Papillary renal cell carcinoma (PRCC) is a rare entity in children with no established therapy protocols for advanced diseases. Immunotherapy is emerging as an important therapeutic tool for ...childhood cancer. Tumor cells can be recognized and killed by conventional and unconventional T cells. Unconventional T cells are able to recognize lipid antigens presented via CD1 molecules independently from major histocompatibility complex, which offers new alternatives for cancer immunotherapies. The nature of those lipids is largely unknown and α-galactosylceramide is currently used as a synthetic model antigen. In this work, we analyzed infiltrating lymphocytes of two pediatric PRCCs using flow cytometry, immunohistochemistry and qRT-PCR. Moreover, we analyzed the CD1d expression within both tumors. Tumor lipids of PRCC samples and three normal kidney samples were fractionated and the recognition of tumor own lipid fractions by unconventional T cells was analyzed in an
assay. We identified infiltrating lymphocytes including γδ T cells and iNKT cells, as well as CD1d expression in both samples. One lipid fraction, containing ceramides and monoacylglycerides amongst others, was able to induce the proliferation of iNKT cells isolated from peripheral blood mononuclear cells (PBMCs) of healthy donors and of one matched PRCC patient. Furthermore, CD1d tetramer stainings revealed that a subset of iNKT cells is able to bind lipids being present in fraction 2 via CD1d. We conclude that PRCCs are infiltrated by conventional and unconventional T cells and express CD1d. Moreover, certain lipids, present in pediatric PRCC, are able to stimulate unconventional T cells. Manipulating these lipids and T cells may open new strategies for therapy of pediatric PRCCs.
The ganglioside GD2 is an important target in childhood cancer. Nevertheless, the only therapy targeting GD2 that is approved to date is the monoclonal antibody dinutuximab, which is used in the ...therapy of neuroblastoma. The relevance of GD2 as a target in other tumor entities remains to be elucidated. Here, we analyzed the expression of GD2 in different pediatric tumor entities by flow cytometry and tested two approaches for targeting GD2. H3K27M-mutant diffuse midline glioma (H3K27M-mutant DMG) samples showed the highest expression of GD2 with all cells strongly positive for the antigen. Ewing's sarcoma (ES) samples also showed high expression, but displayed intra- and intertumor heterogeneity. Osteosarcoma had low to intermediate expression with a high percentage of GD2-negative cells. Dinutuximab beta in combination with irinotecan and temozolomide was used to treat a five-year-old girl with refractory ES. Disease control lasted over 12 months until a single partially GD2-negative intracranial metastasis was detected. In order to target GD2 in H3K27M-mutant DMG, we blocked ganglioside synthesis via eliglustat, since dinutuximab cannot cross the blood-brain barrier. Eliglustat is an inhibitor of glucosylceramide synthase, and it is used for treating children with Gaucher's disease. Eliglustat completely inhibited the proliferation of primary H3K27M-mutant DMG cells in vitro. In summary, our data provide evidence that dinutuximab might be effective in tumors with high GD2 expression. Moreover, disrupting the ganglioside metabolism in H3K27M-mutant DMG could open up a new therapeutic option for this highly fatal cancer.
Neuroblastoma (NBL) and medulloblastoma (MB) are aggressive pediatric cancers which can benefit from therapies targeting gangliosides. Therefore, we compared the ganglioside profile of 9 MB and 14 ...NBL samples by thin layer chromatography and mass spectrometry. NBL had the highest expression of GD2 (median 0.54 nmol GD2/mg protein), and also expressed complex gangliosides. GD2-low samples expressed GD1a and were more differentiated. MB mainly expressed GD2 (median 0.032 nmol GD2/mg protein) or GM3. Four sonic hedgehog-activated (SHH) as well as one group 4 and one group 3 MBs were GD2-positive. Two group 3 MB samples were GD2-negative but GM3-positive. N-glycolyl neuraminic acid-containing GM3 was neither detected in NBL nor MB by mass spectrometry. Furthermore, a GD2-phenotype predicting two-gene signature (
and
) was applied to RNA-Seq datasets, including 86 MBs and validated by qRT-PCR. The signature values were decreased in group 3 and wingless-activated (WNT) compared to SHH and group 4 MBs. These results suggest that while NBL is GD2-positive, only some MB patients can benefit from a GD2-directed therapy. The expression of genes involved in the ganglioside synthesis may allow the identification of GD2-positive MBs. Finally, the ganglioside profile may reflect the differentiation status in NBL and could help to define MB subtypes.
Objective:
In cancer patients, the impairment in muscle function is a frequently observed phenomenon. However, comprehensive evaluation of the effect of exercise training on muscle function in ...childhood cancer patients (CCPs) is sparse and therefore investigated in the MUCKI trial.
Study Design:
In the randomized controlled MUCKI trial, CCPs during intensive cancer treatment and aged 4–18 years were recruited. Eligible patients were enrolled soon after diagnosis as long as they were physically and mentally able to participate in exercise testing and training. Patients of the exercise group (
n
= 16) participated in average 2.7 ± 1.2 times per week in a combined resistance and endurance training with moderate exercise intensity, for a time period of 8.0 ± 2.1 weeks, while patients of the control group (
n
= 17) received usual care. Leg strength was evaluated as the primary endpoint. Secondary endpoints were 6-min walk performance, arm strength, body composition, fatigue, and health-related quality of life.
Results:
Comparisons of pre- and post-intervention results were evaluated by baseline and stratification criteria adjusted analysis and showed positive effects for the exercise group regarding leg strength
F
(1, 20)
= 5.733;
p
= 0.027
*
;
η
p
2
= 0.223, walking performance
F
(1, 25)
= 4.270;
p
= 0.049
*
;
η
p
2
= 0.146, fatigue
F
(1, 13)
= 8.353;
p
= 0.013
*
;
η
p
2
= 0.391, self-esteem
F
(1, 6)
= 6.823;
p
= 0.040
*
;
η
p
2
= 0.532, and self-reported strength and endurance capacity
F
(1, 6)
= 6.273;
p
= 0.046
*
;
η
p
2
= 0.511. No significant differences were found for the other parameters.
Conclusion:
Within one of the first randomized controlled trials, the present study provides evidence for a positive effect of combined training in CCPs during intensive cancer treatment. Further research is needed to confirm these results and to evaluate their clinical impact.
Clinical Trial Registration Number:
NCT02612025.
The insulin-like growth factor (IGF) pathway plays an important role in several brain tumor entities. However, the lack of inhibitors crossing the blood-brain barrier remains a significant obstacle ...for clinical translation. Here, we targeted the IGF pathway using ceritinib, an off-target inhibitor of the IGF1 receptor (IGF1R) and insulin receptor (INSR), in a pediatric patient with an unclassified brain tumor and a
(
) germline mutation. Pathway analysis of the tumor revealed activation of the sonic hedgehog (SHH), the wingless and integrated-1 (WNT), the IGF, and the Notch pathway. The proliferation of the patient tumor cells (225ZL) was inhibited by arsenic trioxide (ATO), which is an inhibitor of the SHH pathway, by linsitinib, which is an inhibitor of IGF1R and INSR, and by ceritinib. 225ZL expressed INSR but not IGF1R at the protein level, and ceritinib blocked the phosphorylation of INSR. Our first personalized treatment included ATO, but because of side effects, we switched to ceritinib. After 46 days, we achieved a concentration of 1.70 µM of ceritinib in the plasma, and after 58 days, MRI confirmed that there was a response to the treatment. Ceritinib accumulated in the tumor at a concentration of 2.72 µM. Our data suggest ceritinib as a promising drug for the treatment of IGF-driven brain tumors.
Background
Increasing survival rates after childhood cancer have raised the issue of long‐term mental health consequences in adulthood. This study determines mental health distress among long‐term ...survivors of pediatric cancer and compares it to control groups.
Methods
Childhood cancer survivors (CCS; N = 951, aged 24‐49 years) were compared to three age‐matched control groups from the general population collected at three time points. The study compared the prevalence of clinically relevant symptoms of a wide range of common mental disorders (depression, somatic distress, suicidal ideation, generalized anxiety, panic, social anxiety, and sleep disturbances) using identical, validated questionnaires. CCS were identified by the German Childhood Cancer Registry. Controls were approached by a demographic consultation company (USUMA) which assured that the three samples were nationally representative.
Results
Childhood cancer survivors reported higher education than controls and were less often married. All forms of common mental distress were increased among survivors. Twenty‐four percent of male (N = 526) and 41% of female survivors (N = 425) reported some form of clinically relevant mental health symptoms. Somatic distress as the leading complaint was highly frequent among CCS (OR: 10.98, CI 95%: 7.24‐16.64). Complaints by generalized anxiety (OR: 5.04, CI 95%: 2.61‐9.70), panic (OR: 3.28, CI 95%: 1.60‐6.70), depression (OR: 3.36, CI 95%: 2.22‐5.09), and suicidality (OR = 2.22; CI 95%: 1.38‐3.57) were also strongly increased. Female sex, low education, low income, and unemployment were associated with increased distress.
Conclusions
Findings indicate a need to integrate psycho‐oncological screening and care into long‐term aftercare. Somatic distress, as cause and indicator of psychological distress, should receive stronger attention, especially tiredness, low energy, and pain.
The article compares self‐reported distress by common mental disorders (depression, somatic symptoms, suicidal ideation, generalized anxiety, panic, social anxiety, and sleep disturbances) among adult survivors of childhood cancer (N = 951) with the general population. Survivors suffer more frequently from symptoms of common mental disorders than controls.
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