Personalized dendritic cell vaccine in multimodal individualized combination therapy improves survival in high-risk pediatric cancer patients.

A lot of hope for high-risk cancers is being pinned on immunotherapy but the evidence in children is lacking due to the rarity and limited efficacy of single-agent approaches. Here, we aim to assess the effectiveness of multimodal therapy comprising a personalized dendritic cell (DC) vaccine in children with relapsed and/or high-risk solid tumors using the N-of-1 approach in real-world scenario. A total of 160 evaluable events occurred in 48 patients during the 4-year follow-up. Overall survival of the cohort was 7.03 years. Disease control after vaccination was achieved in 53.8% patients. Comparative survival analysis showed the beneficial effect of DC vaccine beyond 2 years from initial diagnosis (HR = 0.53, P = .048) or in patients with disease control (HR = 0.16, P = .00053). A trend for synergistic effect with metronomic cyclophosphamide and/or vinblastine was indicated (HR = 0.60 P = .225). A strong synergistic effect was found for immune check-point inhibitors (ICIs) after priming with the DC vaccine (HR = 0.40, P = .0047). In conclusion, the personalized DC vaccine was an effective component in the multimodal individualized treatment. Personalized DC vaccine was effective in less burdened or more indolent diseases with a favorable safety profile and synergized with metronomic and/or immunomodulating agents.

Precision immuno-oncology approach for four malignant tumors in siblings with constitutional mismatch repair deficiency syndrome.

Constitutional mismatch repair deficiency (CMMRD) is a rare syndrome characterized by an increased incidence of cancer. It is caused by biallelic germline mutations in one of the four mismatch repair genes (MMR) genes: MLH1, MSH2, MSH6, or PMS2. Accurate diagnosis accompanied by a proper molecular genetic examination plays a crucial role in cancer management and also has implications for other family members. In this report, we share the impact of the diagnosis and challenges during the clinical management of two brothers with CMMRD from a non-consanguineous family harbouring compound heterozygous variants in the PMS2 gene. Both brothers presented with different phenotypic manifestations and cancer spectrum. Treatment involving immune checkpoint inhibitors significantly contributed to prolonged survival in both patients affected by lethal gliomas. The uniform hypermutation also allowed immune-directed treatment using nivolumab for the B-cell lymphoma, thereby limiting the intensive chemotherapy exposure in this young patient who remains at risk for subsequent malignancies.

Tazemetostat in the therapy of pediatric INI1-negative malignant rhabdoid tumors.

Rhabdoid tumors are aggressive tumors that may arise in the kidney, soft tissue, central nervous system, or other organs. They are defined by SMARCB1 (INI1) or SMARCA4 alterations. Often, very young children are affected, and the prognosis is dismal. Four patients with primary atypical teratoid rhabdoid tumor (AT/RT, a rhabdoid tumor in the central nervous system) were treated by resection and high dose chemotherapy. Tazemetostat was introduced after completion of chemotherapy. Three patients have achieved an event free survival of 32, 34, and 30 months respectively. One progressed and died. His overall survival was 20 months. One patient was treated for a relapsed atypical teratoid rhabdoid tumor. The treatment combined metronomic therapy, radiotherapy, tazemetostat and immunotherapy. This patient died of disease progression, with an overall survival of 37 months. One patient was treated for a rhabdoid tumor of the ovary. Tazemetostat was given as maintenance after resection, chemotherapy, and radiotherapy, concomitantly with immunotherapy. Her event free survival is 44 months. Only approximately 40% of patients with rhabdoid tumors achieve long-term survival. Nearly all relapses occur within two years from diagnosis. The event free survival of four of the six patients in our cohort has exceeded this timepoint. Tazemetostat has been mostly tested as a single agent in the relapsed setting. We present promising results when applied as maintenance or add on in the first line treatment.

Skeletal dysplasias of the fetus and infant: comprehensive review and our experience over a 10-year period.

We present a comprehensive review dealing with rare genetic skeletal disorders. More than 400 entities are included in the latest classification. The most severe or lethal phenotypes are identifiable in the prenatal period and the pregnancy can be terminated. Perinatal autopsy and posmortem X-rays are crucial in providing a definitive diagnosis. The number of cases confirmed by genetic testing is increasing. We report our own experience with genetic skeletal disorders based on 41 illustrative fetal and neonatal cases which we encountered over a 10-year period. Thanatophoric dysplasia and osteogenesis imperfecta represent approximately half of the cases coming to autopsy. Achondrogenesis type 2 and hypochondrogenesis, short-rib dysplasia, chondrodysplasia punctata, campomelic dysplasia and achondroplasia are less common. Skeletal dysplasias with autosomal recessive inheritance are the least frequent, e.g. perinatally lethal hypophophatasia, achondrogenesis type 1A, diastrophic dysplasia/atelosteogenesis type 2 or mucolipidosis type 2 (I cell disease).

Two sisters with cardiac-urogenital syndrome secondary to pathogenic splicing variant in the MYRF gene with unaffected parents: A case of gonadal mosaicism?

BACKGROUND: Cardiac-urogenital syndrome [MIM # 618280] is a newly described very rare syndrome associated with pathogenic variants in the myelin regulatory factor (MYRF) gene that leads to loss of protein function. MYRF is a transcription factor previously associated only with the control of myelin-related gene expression. However, it is also highly expressed in other tissues and associated with various organ anomalies. The clinical picture is primarily dominated by complex congenital cardiac developmental defects, pulmonary hypoplasia, congenital diaphragmatic hernia, and urogenital malformations. CASE PRESENTATION: We present case reports of two siblings of unrelated parents in whom whole-exome sequencing was indicated due to familial occurrence of extensive developmental defects. A new, previously undescribed splicing pathogenic variant c.1388+2T>G in the MYRF gene has been identified in both patients. Both parents are unaffected, tested negative, and have another healthy daughter. The identical de novo event in siblings suggests gonadal mosaicism, which can mimic recessive inheritance. CONCLUSIONS: To our knowledge, this is the first published case of familial cardiac-urogenital syndrome indicating gonadal mosaicism.

Improved osteosarcoma survival with addition of mifamurtide to conventional chemotherapy - Observational prospective single institution analysis.

PURPOSE: Conventional osteosarcoma is an orphan disease. Current treatment approaches include combining a three drug chemotherapy schedule and surgery. The 3- and 5-year event-free survival (EFS) in localized disease is roughly 65 and 60%, respectively. The registration study of mifamurtide reported survival benefit, but some methodological controversies have been insufficient for FDA market authorization in contrast to EMA. METHODS: prospective single centre survival analysis of a mifamurtide addition to conventional therapy in 23 patients over a 5.5 year enrolment period is reported and compared to a historical control of 26 patient with localized disease. Bias arising from observational methodology was addressed using Landmark analysis and time-dependent Cox models. Blood count dynamics were analysed during the treatment. RESULTS: The adverse event profile was as expected with no dose limiting toxicities. There were no local relapses observed, one patient died in the first complete remission due to doxorubicin cardiotoxicity, one patient had pulmonary metastatic relapse. The observed 3- and 5-year EFS was 87.4% (CI 72.4-100%) and 87.4% (CI 72.4-100%), progression free survival (PFS) was 92.9% (CI 80.3-100%) and 92.9% (CI 80.3-100%), overall survival was 94.1% (CI 83.6-100) and 80.7% (CI 58.3-100), respectively. Comparison to the historical control showed statistically significant better PFS for mifamurtide patients (Landmark analysis; p = 0.044). Risk of progression was 5-times lower for the mifamurtide group (Cox model; HR 0.21, p = 0.136). Only subtle differences in lymphocyte counts were observed across treatment. CONCLUSION: the PFS benefit of mifamurtide is reported herein. The addition of mifamurtide could be considered as a best treatment option for localized osteosarcoma.

Spindle Cell Hemangioma and Atypically Localized Juxtaglomerular Cell Tumor in a Patient with Hereditary BRIP1 Mutation: A Case Report.

Spindle cell hemangioma is a benign vascular tumor typically occurring in the dermis or subcutis of distal extremities as red-brown lesions that can grow in both size and number over time. They can be very painful and potentially disabling. A family history of cancer or previous history may be relevant and must be taken into consideration. Juxtaglomerular cell tumor (reninoma) is an extremely rare cause of secondary hypertension diagnosed mostly among adolescents and young adults. Excessive renin secretion results in secondary hyperaldosteronism. Subsequent hypokalemia and metabolic alkalosis, together with high blood pressure, are clues for clinical diagnosis. Histological examination of the excised tumor leads to a definitive diagnosis. Reninoma is found in subcapsular localization, in most cases as a solitary mass, in imaging studies of kidneys. Exceptionally, it can be located in another part of a kidney. Both spindle cell hemangioma and reninoma are extremely rare tumors in children and adolescents. Herein, the authors present a case report of a patient with hereditary BRCA1 interacting protein C-terminal helicase 1 (BRIP1) mutation, spindle cell hemangioma, and secondary hypertension caused by atypically localized reninoma.

SDHC Methylation Pattern in Patients With Carney Triad.

Carney triad is a multitumor syndrome affecting almost exclusively young women in a nonfamilial setting, which manifests by multifocal gastric gastrointestinal stromal tumors, paragangliomas, and pulmonary chondroma. The Carney triad-associated tumors are characterized by a deficiency of the mitochondrial succinate dehydrogenase enzymatic complex. Recently, it has been observed that the deficiency results from epigenetic silencing of the SDHC gene by its promoter hypermethylation. To elucidate anatomic distribution of SDHC promoter methylation in Carney triad patients and thus to shed some light on the possible natural development of this epigenetic change, both neoplastic and available non-neoplastic tissues of 3 patients with Carney triad were tested for hypermethylation at the SDHC promoter site. SDHC promoter hypermethylation was proven in all tumors studied. Lack of SDHC epigenetic silencing in the non-neoplastic lymphoid and duodenal tissue (ie, tissues not involved in the development of Carney triad-associated tumors) together with the finding of SDHC promoter hypermethylation in the non-neoplastic gastric wall favors the hypothesis of postzygotic somatic mosaicism as the biological background of Carney triad; it also offers an explanation of the multifocality of gastrointestinal stromal tumors of the stomach occurring in this scenario as well. However, the precise mechanism responsible for the peculiar organ-specific distribution of Carney triad-associated tumors is still unknown.

Salivary Gland Choristoma of the Middle Ear in a Child: A Case Report.

Salivary gland choristoma is an extremely rare middle ear pathology. We present the case of a 10-year-old girl with unilateral conductive hearing loss. Tympanotomy showed a nonspecific middle ear mass, absence of stapes, anomaly of incus, and displaced facial nerve. It was not possible to remove the mass completely. Histology confirmed salivary gland choristoma. The hearing in this case can be improved with a bone-anchored hearing aid.

Histiocytoses and neoplasms of the macrophage-dendritic cell lineages. Comparison of recent WHO classification published 2017 and classification of Histiocyte Society published 2016.

The histiocytoses are rare disorders characterized by the accumulation of cells thought to be derived from dendritic cells or macrophages. Their clinical behaviour ranges from mild to disseminated and, sometimes, life-threatening forms. The incidence of this diseases is much smaller, then the incidence of diseases derived from lymphocytic or myeloid lineage. Langerhans cell histiocytosis is most frequent disease from this group. The last version of WHO classification from 2017 and last version of classification published by Histiocyte Society is summarised in this paper.

A Newly Observed Mutation of the ABCA3 Gene Causing Lethal Respiratory Failure of a Full-Term Newborn: A Case Report.

Respiratory distress syndrome caused by a secondary surfactant deficiency is one of the most common diagnoses requiring admission to the Neonatal Intensive Care Unit. We illustrate the case of a term female newborn without prenatal and peripartal risks. There had been significant signs of respiratory distress 4 h after delivery. The condition gradually worsened to the point of needing oscillatory ventilation. The most common infectious and non-infectious causes were excluded. Considering the course of illness, a congenital surfactant deficiency was suspected. There nevertheless was no significant improvement after administration of surfactant. Following a short period of palliative care, the child died at 34 days of age due to respiratory failure. DNA diagnostics revealed compound heterozygosity of ABCA3 functional mutations leading to the p.Pro147Leu and p.Pro246Leu exchanges. The second identified mutation of ABCA3 c.737C>T had not to date been described in connection with primary surfactant deficiency.

Assessment of Tumor Mutational Burden in Pediatric Tumors by Real-Life Whole-Exome Sequencing and In Silico Simulation of Targeted Gene Panels: How the Choice of Method Could Affect the Clinical Decision?

BACKGROUND: Tumor mutational burden (TMB) is an emerging genomic biomarker in cancer that has been associated with improved response to immune checkpoint inhibitors (ICIs) in adult cancers. It was described that variability in TMB assessment is introduced by different laboratory techniques and various settings of bioinformatic pipelines. In pediatric oncology, no study has been published describing this variability so far. METHODS: In our study, we performed whole exome sequencing (WES, both germline and somatic) and calculated TMB in 106 patients with high-risk/recurrent pediatric solid tumors of 28 distinct cancer types. Subsequently, we used WES data for TMB calculation using an in silico approach simulating two The Food and Drug Administration (FDA)-approved/authorized comprehensive genomic panels for cancer. RESULTS: We describe a strong correlation between WES-based and panel-based TMBs; however, we show that this high correlation is significantly affected by inclusion of only a few hypermutated cases. In the series of nine cases, we determined TMB in two sequentially collected tumor tissue specimens and observed an increase in TMB along with tumor progression. Furthermore, we evaluated the extent to which potential ICI indication could be affected by variability in techniques and bioinformatic pipelines used for TMB assessment. We confirmed that this technological variability could significantly affect ICI indication in pediatric cancer patients; however, this significance decreases with the increasing cut-off values. CONCLUSIONS: For the first time in pediatric oncology, we assessed the reliability of TMB estimation across multiple pediatric cancer types using real-life WES and in silico analysis of two major targeted gene panels and confirmed a significant technological variability to be introduced by different laboratory techniques and various settings of bioinformatic pipelines.

Comparative Analysis of Putative Prognostic and Predictive Markers in Neuroblastomas: High Expression of PBX1 Is Associated With a Poor Response to Induction Therapy.

The survival rate for patients with high-risk neuroblastomas remains poor despite new improvements in available therapeutic modalities. A detailed understanding of the mechanisms underlying clinical responses to multimodal treatment is one of the important aspects that may provide precision in the prediction of a patient's clinical outcome. Our study was designed as a detailed comparative analysis of five selected proteins (DDX39A, HMGA1, HOXC9, NF1, and PBX1) in one cohort of patients using the same methodical approaches. These proteins were already reported separately as related to the resistance or sensitivity to retinoids and as useful prognostic markers of survival probability. In the cohort of 19 patients suffering from high-risk neuroblastomas, we analyzed initial immunohistochemistry samples obtained by diagnostic biopsy and post-induction samples taken after the end of induction therapy. The expression of DDX39A, HMGA1, HOXC9, and NF1 showed varied patterns with almost no differences between responders and non-responders. Nevertheless, we found very interesting results for PBX1: non-responders had significantly higher expression levels of this protein in the initial tumor samples when compared with responders; this expression pattern changed inversely in the post-induction samples, and this change was also statistically significant. Moreover, our results from survival analyses reveal the prognostic value of PBX1, NF1, and HOXC9 expression in neuroblastoma tissue. In addition to the prognostic importance of PBX1, NF1, and HOXC9 proteins, our results demonstrated that PBX1 could be used for the prediction of the clinical response to induction chemotherapy in patients suffering from high-risk neuroblastoma.

Association of rare non-coding SNVs in the lung-specific FOXF1 enhancer with a mitigation of the lethal ACDMPV phenotype.

Haploinsufficiency of FOXF1 causes alveolar capillary dysplasia with misalignment of pulmonary veins (ACDMPV), a lethal neonatal lung developmental disorder. We describe two similar heterozygous CNV deletions involving the FOXF1 enhancer and re-analyze FOXF1 missense mutation, all associated with an unexpectedly mitigated disease phenotype. In one case, the deletion of the maternal allele of the FOXF1 enhancer caused pulmonary hypertension and histopathologically diagnosed MPV without the typical ACD features. In the second case, the deletion of the paternal enhancer resulted in ACDMPV rather than the expected neonatal lethality. In both cases, FOXF1 expression in lung tissue was higher than usually seen or expected in patients with similar deletions, suggesting an increased activity of the remaining allele of the enhancer. Sequencing of these alleles revealed two rare SNVs, rs150502618-A and rs79301423-T, mapping to the partially overlapping binding sites for TFAP2s and CTCF in the core region of the enhancer. Moreover, in a family with three histopathologically-diagnosed ACDMPV siblings whose missense FOXF1 mutation was inherited from the healthy non-mosaic carrier mother, we have identified a rare SNV rs28571077-A within 2-kb of the above-mentioned non-coding SNVs in the FOXF1 enhancer in the mother, that was absent in the affected newborns and 13 unrelated ACDMPV patients with CNV deletions of this genomic region. Based on the low population frequencies of these three variants, their absence in ACDMPV patients, the results of reporter assay, RNAi and EMSA experiments, and in silico predictions, we propose that the described SNVs might have acted on FOXF1 enhancer as hypermorphs.

Individualization of Treatment Improves the Survival of Children With High-Risk Solid Tumors: Comparative Patient Series Analysis in a Real-Life Scenario.

Introduction: The individualization of treatment is attractive, especially in children with high-risk cancer. In such a rare and very heterogeneous group of diseases, large population-based clinical randomized trials are not feasible without international collaboration. We therefore propose comparative patient series analysis in a real-life scenario. Methods: Open cohort observational study, comparative analysis. Seventy patients with high-risk solid tumors diagnosed between 2003 and 2015 and in whom the treatment was individualized either empirically or based on biomarkers were analyzed. The heterogeneity of the cohort and repeated measurements were advantageously utilized to increase effective sample size using appropriate statistical tools. Results: We demonstrated a beneficial effect of empirically given low-dose metronomic chemotherapy (HR 0.46 for relapses, p = 0.017) as well as various repurposed or targeted agents (HR 0.15 for deaths, p = 0.004) in a real-life scenario. However, targeted agents given on the basis of limited biological information were not beneficial. Conclusions: Comparative patient series analysis provides institutional-level evidence for treatment individualization in high-risk pediatric malignancies. Our findings emphasize the need for a comprehensive, multi omics assessment of the tumor and the host as well whenever molecularly driven targeted therapies are being considered. Low-dose metronomic chemotherapy or local control of the disease may be a more rational option in situations where targeted treatment cannot be justified by robust evidence and comprehensive biological information. "Targeted drugs" may be given empirically with a realistic benefit expectation when based on robust rationale.

Prediction of neuroblastoma cell response to treatment with natural or synthetic retinoids using selected protein biomarkers.

Although the administration of retinoids represents an important part of treatment for children suffering from high-risk neuroblastomas, approximately 50% of these patients do not respond to this therapy or develop resistance to retinoids during treatment. Our study focused on the comparative analysis of the expression of five genes and corresponding proteins (DDX39A, HMGA1, HMGA2, HOXC9 and PBX1) that have recently been discussed as possible predictive biomarkers of clinical response to retinoid differentiation therapy. Expression of these five candidate biomarkers was evaluated at both the mRNA and protein level in the same subset of 8 neuroblastoma cell lines after treatment with natural or synthetic retinoids. We found that the cell lines that were HMGA2-positive and/or HOXC9-negative have a reduced sensitivity to retinoids. Furthermore, the experiments revealed that the retinoid-sensitive cell lines showed a uniform pattern of change after treatment with both natural and sensitive retinoids: increased DDX39A and decreased PBX1 protein levels. Our results showed that in NBL cells, these putative protein biomarkers are associated with sensitivity or resistance to retinoids, and their endogenous or induced expression can distinguish between these two phenotypes.

Noonan syndrome from a fetopathologist perspective.

We present our experience with four cases of fetal autopsies with abnormal prenatal ultrasound findings and suspicion of Noonan syndrome. These were fetuses from the 17th to the 24th age of gestation (GA). In all cases, prenatal ultrasound examination recorded increased nuchal translucency (NT) and presence of lymphatic neck sacs. Some fetuses showed signs of fetal hydrops and polyhydramnion was found. Similar signs and congenital developmental defects were confirmed in the autopsy examination. These were primarily signs of developing fetal hydrops with increased nuchal edema, in some cases up to the character of cystic hygroma, pleural and abdominal effusions, congenital heart and kidney defects, skeletal defects and facial dysmorphism. A karyotype was examined in all cases without chromosome aneuploidy. The diagnosis of NS was confimed by subsequent genetic analysis of causal gene mutations (mainly PTPN11, KRAS, RAF 1,). Our cases demonstrate a wide range of signs of prenatal presentation of this syndrome. Because of wide differential diagnosis, summarizing prenatal ultrasound findings, autopsy examination and molecular genetic testing is essential.

Personalized Treatment of H3K27M-Mutant Pediatric Diffuse Gliomas Provides Improved Therapeutic Opportunities.

Diffuse gliomas with K27M histone mutations (H3K27M glioma) are generally characterized by a fatal prognosis, particularly affecting the pediatric population. Based on the molecular heterogeneity observed in this tumor type, personalized treatment is considered to substantially improve therapeutic options. Therefore, clinical evidence for therapy, guided by comprehensive molecular profiling, is urgently required. In this study, we analyzed feasibility and clinical outcomes in a cohort of 12 H3K27M glioma cases treated at two centers. Patients were subjected to personalized treatment either at primary diagnosis or disease progression and received backbone therapy including focal irradiation. Molecular analyses included whole-exome sequencing of tumor and germline DNA, RNA-sequencing, and transcriptomic profiling. Patients were monitored with regular clinical as well as radiological follow-up. In one case, liquid biopsy of cerebrospinal fluid (CSF) was used. Analyses could be completed in 83% (10/12) and subsequent personalized treatment for one or more additional pharmacological therapies could be recommended in 90% (9/10). Personalized treatment included inhibition of the PI3K/AKT/mTOR pathway (3/9), MAPK signaling (2/9), immunotherapy (2/9), receptor tyrosine kinase inhibition (2/9), and retinoic receptor agonist (1/9). The overall response rate within the cohort was 78% (7/9) including one complete remission, three partial responses, and three stable diseases. Sustained responses lasting for 28 to 150 weeks were observed for cases with PIK3CA mutations treated with either miltefosine or everolimus and additional treatment with trametinib/dabrafenib in a case with BRAFV600E mutation. Immune checkpoint inhibitor treatment of a case with increased tumor mutational burden (TMB) resulted in complete remission lasting 40 weeks. Median time to progression was 29 weeks. Median overall survival (OS) in the personalized treatment cohort was 16.5 months. Last, we compared OS to a control cohort (n = 9) showing a median OS of 17.5 months. No significant difference between the cohorts could be detected, but long-term survivors (>2 years) were only present in the personalized treatment cohort. Taken together, we present the first evidence of clinical efficacy and an improved patient outcome through a personalized approach at least in selected cases of H3K27M glioma.

Comprehensive Molecular Profiling for Relapsed/Refractory Pediatric Burkitt Lymphomas-Retrospective Analysis of Three Real-Life Clinical Cases-Addressing Issues on Randomization and Customization at the Bedside.

In order to identify reasons for treatment failures when using targeted therapies, we have analyzed the comprehensive molecular profiles of three relapsed, poor-prognosis Burkitt lymphoma cases. All three cases had resembling clinical presentation and histology and all three patients relapsed, but their outcomes differed significantly. The samples of their tumor tissue were analyzed using whole-exome sequencing, gene expression profiling, phosphoproteomic assays, and single-cell phosphoflow cytometry. These results explain different treatment responses of the three histologically identical but molecularly different tumors. Our findings support a personalized approach for patient with high risk, refractory, and rare diseases and may contribute to personalized and customized treatment efforts for patients with limited treatment options like relapsed/refractory Burkitt lymphoma. SUMMARY: The main aim of this study is to analyze three relapsed Burkitt lymphoma patients using a comprehensive molecular profiling, in order to explain their different outcomes and to propose a biomarker-based targeted treatment. In cases 1 and 3, the tumor tissue and the host were analyzed prospectively and appropriate target for the treatment was successfully implemented; however, in case 2, analyses become available only retrospectively and his empirically based rescue treatment did not hit the right target of his disease.