To evaluate plasma cell‐free DNA (cfDNA) as a promising biomarker for neuroblastoma (NB) tumor burden. Seventy‐nine eligible patients with newly diagnosed NB were recruited from Beijing Children's Hospital between April 2016 and April 2017. Additionally, from September 2011 to June 2017, 79 patients with stable NB were evaluated with a median follow‐up time of 21 months. Approximately 2 mL of peripheral blood was drawn upon enrollment, and plasma cfDNA levels were measured via quantitative polymerase chain reaction (qPCR). Total cfDNA analysis was performed using the long interspersed nuclear element 1 (LINE‐1) 79 bp fragment, and DNA integrity was calculated by the ratio of the LINE‐1 300 bp fragment to the LINE‐1 79 bp fragment. A total of 79 NB patients with a median age of 36 months comprised the group of newly diagnosed NB patients. The main primary tumor site was the retroperitoneal and adrenal region (81%). Three or more metastatic sites were found in 17.7% of patients. Stable NB patients older than 18 months comprised 98.7% of the stable NB patients. Neuron‐specific enolase (NSE), lactate dehydrogenase (LDH), and cfDNA levels were dramatically increased in the newly diagnosed NB patients and significantly different from those in the stable NB patients. Moreover, the concentration of cfDNA was much higher in patients with larger tumors. By analyzing the area under the receiver operator characteristic (ROC) curve (AUC), the areas of total cfDNA, NSE, and LDH levels were 0.953, 0.929, and 0.906, respectively. The sensitivity and specificity data clarified that the level of circulating cfDNA in plasma can be considered as a reliable biomarker for describing tumor load in NB. The plasma cfDNA concentration was as good as the levels of LDH and NSE to discriminate the tumor burden in children with NB.

Importance: Retinoblastoma (Rb) is the most common primary malignant intraocular cancer in children. Systemic chemotherapy combined with local therapy is safe and effective for intraocular Rb. Objective: To summarize the short-term outcomes of patients with Rb to provide evidence for optimizing treatment protocols and improving therapeutic safety and efficacy. Methods: The clinical data of 356 patients (486 eyes) with intraocular Rb admitted to our center from December 2009 to April 2017 were retrospectively analyzed. The measures included drug toxicity, eyepreservation rate, and survival rate, with an emphasis on safety and shortterm efficacy. The date of last follow-up was 30 November, 2017. results: The patients comprised 226 unilateral Rb and 130 bilateral Rb. Enucleation before chemotherapy was performed in 72 patients. Among the 174 patients with unilateral Rb, enucleation after chemotherapy was performed in 80 patients (46.0%), and the eye was not enucleated in 89 (51.1%); 68 eyes were preserved (68/114, 59.6%) in Group D and 20 eyes (20/59, 33.8%) in Group E. Among the 220 eyes in patients with bilateral Rb, enucleation after chemotherapy was performed for 35 eyes; the eyepreservation rate was 91.7% in Group C, 79.1% in Group D, and 52.1% in Group E. All patients developed grade II to IV myelosuppression after chemotherapy, among whom 18 patients (5%) requiring transfusion. Fourteen patients (3.9%) died of intracranial metastasis following selfelected discontinuation of treatment (n = 7). Patients were followed up for a median of 47 (range, 1-96) months. The expected 5-year overall survival rate was 95.3% (96.7% for unilateral Rb and 92.9% for bilateral Rb, P = 0.074). Interpretation: The VEC (vincristine, etoposide, and carboplatin) regimen with local treatment was safe for intraocular Rb. Intracranial metastasis remains the most common cause of Rb-related death. How to cite this article: Jin M, Zhao J, Zhang D, et al. Analysis of the short-term curative effect of 356 cases of intraocular retinoblastoma in children. Pediatr Invest. 2018;2:236-241.

Background To improve cure rates for neuroblastoma (NB), it is important and necessary to evaluate therapy response. Our investigation focuses on using plasma cell free DNA (cfDNA) as a biomarker to determine tumor burden and minimal residual disease (MRD) of NB patients during chemotherapy. Methods Total 58 NB patients were recruited from July 2016 to December 2017. Therapy regime and risk classification were based on COG standard and BCH‐NB‐2007 protocol. RECIST study was used to judge response to therapy at the end of fourth cycle of chemotherapy (CC4) and maintenance stage (MS) respectively. Serial quantifications of cfDNA, NSE, and LDH were examined at four stages, including newly diagnosed, second and CC4, and maintenance. Results During early chemotherapy, 65.5% of NB kids responded well. Consistently, cfDNA, NSE, and LDH levels were down‐regulated in NB patients with partial remission (PR) compared to those with stable disease (SD). In both training and predicting sets, the levels of cfDNA were significantly comparable between PR and SD only at CC4 stage. To predict the insufficient response to early chemotherapy, the optimal AUC value of cfDNA was 0.732 and 0.747 in training and predicting sets respectively, with a sensitivity of 63.2% and 80% specificity at 11.59 ng/ml and a sensitivity of 68.4% and 90% specificity at 10.35 ng/ml. At MS, responded NB patients were slightly increased up to 70%. This evaluation was confirmed by further decrease in cfDNA and NSE levels during intermediate chemotherapy in comparison with early stage. Conclusion The dynamic change of cfDNA was considered as a surrogate biomarker to evaluate tumor burden and MRD of NB during early and intermediate therapy periods.

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Detection of amplification of the MYCN gene is essential for determining optimal treatment and estimating prognosis of patients with neuroblastoma (NB). DNA FISH with neuroblastoma tissues or patient-derived bone marrow cells is the standard clinical practice for the detection of MYCN amplification. As tumor cells may often be unavailable, we developed a method to detect MYCN amplification in the plasma of patients with neuroblastoma. Taking single-copy NAGK DNA as reference, we used real-time quantitative PCR (qPCR) to determine the MYCN/NAGK ratio in the plasma of 115 patients diagnosed with NB. An increased MYCN/NAGK ratio in the plasma was consistent with MYCN amplification as assessed by DNA FISH. The AUC for a MYCN/NAGK ratio equal to 6.965 was 0.943, with 86% sensitivity and 100% specificity. Beyond the threshold of 6.965, the MYCN/NAGK ratio correlated with a heavier tumor burden. Event-free and overall survival of two years were significantly shortened in stage 4 patients with a MYCN/NAGK ratio higher than 6.965. Plasma MYCN/NAGK ratios increased in patients with progressive disease and relapse. Thus, we conclude that the determination of the plasma MYCN/ NAGK ratio by qPCR is a noninvasive and reproducible method to measure MYCN amplification in patients with NB.

Our aim is to examine the impact of DICER1 mutations on the pathogenesis of pleuropulmonary blastoma (PPB) by evaluating the mutation frequency and investigating the family history of Chinese patients with PPB. The family histories of 12 children with PPB recruited consecutively were surveyed. Blood samples from patients and their first-degree relatives were tested for DICER1 mutations. Whole-genome sequencing of blood samples and formalin-fixed and paraffin-embedded (FFPE) tumor tissue was performed in one family with twins. Twelve patients with PPB included six type II and six type III cases. Seven of the 12 patients harbored DICER1 mutations, six of which were frameshift or nonsense mutations. Another case carried a germline DICER1 mutation affecting the splice site. FFPE sample had a nonsense mutation in TDG and missense mutations in DICER1. In addition, two cases with DICER1 mutations were found to have lung cysts preceding the diagnosis of PPB. Furthermore, one patient had a family history remarkable for thyroid diseases. Our results indicate that the germline mutation frequency in Chinese patients with PPB is similar to the ones reported for patients from USA, UK, and Japan. Moreover, our study strongly suggests that investigating the family history and detecting germline DICER1 mutations might be of benefit to increasing awareness and improving the accuracy of the differential diagnosis of PPB from non-malignant lung cysts.

BackgroundPrevious studies have shown that γδ TFH cells are capable of modulating antibody production in immunized and infected mouse model. In recent studies, human γδ TFH cells are shown to contribute to the activation of humoral immunity and promote the maturation of B cells. However, little information is available on their involvement in neuroblastoma (NB) pathogenesis.ResultsIn the present study, the frequency of γδ TFH cells in 74 NB patients was significantly higher compared with that in 60 healthy controls. Moreover, most γδ TFH cells in NB patients had a naive phenotype with up-regulation of CD25, CD69, HLA-DR and CD40L and down-regulation of ICOS. Importantly, γδ TFH cells in NB patients produced more IL-4 and IL-10 than those in healthy controls. Furthermore, serum total IgG level was significantly increased in NB patients compared with healthy controls. The expression of CD23 on B cells was up-regulated while CD80 expression was significantly down-regulated in NB patients. Further analysis of B cell compartment showed that the frequency of CD19+CD27hi plasma cells was enhanced in NB patients. Spearman’s correlation analysis revealed that the frequency of γδ TFH cells was positively correlated to serum total IgG level and CD19+CD27hi plasma cells in NB patients, but negatively correlated to CD19+ B cells.ConclusionsWe concluded that γδ TFH cells might promote B cell maturation and antibody production in NB patients.

Pleuropulmonary blastoma (PPB) is the most common primary malignant neoplasm of the lung in children that is associated with a germline mutation in DICER1. In this report, we share an interesting case of a pair of monozygotic twins: one of them developed PPB when she was 4-year old, while the other developed acute transient hepatitis when she was 5-year old. Next-Gen sequencing for DICER1 mutations of their family revealed that both twins and their mother had c.C3675A mutation. The mother also had a history of multinodular goiter. Identification of DICER1 mutation carriers and close surveillance of individuals at risk for DICER1 syndrome may allow early detection and hence better outcome.