Neuroblastoma is a childhood cancer that resembles developmental stages of the neural crest. It is not established what developmental processes neuroblastoma cancer cells represent. Here, we sought to reveal the phenotype of neuroblastoma cancer cells by comparing cancer (n = 19,723) with normal fetal adrenal single-cell transcriptomes (n = 57,972). Our principal finding was that the neuroblastoma cancer cell resembled fetal sympathoblasts, but no other fetal adrenal cell type. The sympathoblastic state was a universal feature of neuroblastoma cells, transcending cell cluster diversity, individual patients, and clinical phenotypes. We substantiated our findings in 650 neuroblastoma bulk transcriptomes and by integrating canonical features of the neuroblastoma genome with transcriptional signals. Overall, our observations indicate that a pan-neuroblastoma cancer cell state exists, which may be attractive for novel immunotherapeutic and targeted avenues.

Background-Intestinal neuronal dysplasia (IND) of the colonic submucous plexus is considered to be a congenital malformation of the enteric nervous system causing symptoms resembling those of Hirschsprung's disease. In contrast with the established diagnosis of aganglionosis using enzyme histochemistry, controversy exists over the diagnostic criteria of IND on rectal biopsies previously defined by a consensus report and the causal relation between morphological findings and clinical symptoms. Aims-The interobserver variability was prospectively investigated with respect to final diagnoses and several histological features in rectal biopsy specimens from children suspected of having colonic motility disturbances. Methods-377 biopsy specimens from 108 children aged 4 days to 15 years were independently coded without knowledge of clinical symptoms by three experienced pathologists for 20 histological features, and a final diagnosis was given for every case. Interobserver variation for the different items and the final diagnosis were analysed using Cohen's statistic. Clinical data at biopsy and outcome after 12 months were related to morphological findings. Results-The three pathologists agreed completely with respect to the diagnosis Hirschsprung's disease ( = 1), but in only 14% of the children without aganglionosis. In 15 (17%) of the 87 children without aganglionosis, at least one pathologist judged the case as normal, while another diagnosed IND. values were close to the zero value expected by chance for the diagnoses normal and IND. Young age was related to the presence of several morphological features-for example, acetylcholine esterase staining and presence of giant ganglia. Children with chronic constipation diagnosed as having IND, given no other specific diagnosis by any of the pathologists, were significantly younger (median 8.8 months) and had a higher cure rate after one year (60%) than constipated patients considered by all observers to have no histological abnormalities (median 6.1 years, cure rate 23%).Conclusions-In contrast with Hirschsprung's disease, there is a high interobserver variation with regard to the diVerent morphological features and final diagnosis of IND, based on the criteria and conditions of the previous consensus report. The high frequency of histological "abnormalities" in young infants suggests that some of the features may represent a normal variant of postnatal development rather than a pathological process. Investigations using more refined and morphometric methods in rectal specimens from infants and children without bowel disease are needed to define the normal range of morphological appearance at diVerent ages. These preliminary data indicate that, with current knowledge, rectal biopsy for diagnostic purposes should only be performed in constipated children for diagnosis of Hirschsprung's disease. (Gut 1999;44:853-861)

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(Gut 2001;48:671-675)

High-risk neuroblastoma, a pediatric tumor originating from the sympathetic nervous system, has a low mutation load but highly recurrent somatic DNA copy number variants. Previously, segmental gains and/or amplifications allowed identification of drivers for neuroblastoma development. Using this approach, combined with gene dosage impact on expression and survival, we identified ribonucleotide reductase subunit M2 (RRM2) as a candidate dependency factor further supported by growth inhibition upon in vitro knockdown and accelerated tumor formation in a neuroblastoma zebrafish model coexpressing human RRM2 with MYCN. Forced RRM2 induction alleviates excessive replicative stress induced by CHK1 inhibition, while high RRM2 expression in human neuroblastomas correlates with high CHK1 activity. MYCN-driven zebrafish tumors with RRM2 co-overexpression exhibit differentially expressed DNA repair genes in keeping with enhanced ATR-CHK1 signaling activity. In vitro, RRM2 inhibition enhances intrinsic replication stress checkpoint addiction. Last, combinatorial RRM2-CHK1 inhibition acts synergistic in high-risk neuroblastoma cell lines and patient-derived xenograft models, illustrating the therapeutic potential.

Benign recurrent intrahepatic cholestasis (BRIC) is an autosomal recessive liver disease characterized by multiple episodes of cholestasis without progression to chronic liver disease. The gene was previously assigned to chromosome 18q21, using a shared segment analysis in three families from the Netherlands. In the present study we report the linkage analysis of an expanded sample of 14 BRIC families, using 15 microsatellite markers from the 18q21 region. Obligate recombinants in two families place the gene in a 7-cM interval, between markers D18S69 and D18S64. All intervening markers had significant LOD scores in two-point linkage analysis. Moreover, we identified one family in which the BRIC gene seems to be unlinked to the 18q21 region, or that represents incomplete penetrance of the BRIC genotype. IntroductionBenign recurrent intrahepatic cholerstasis (BRIC) is a rare autosomal recessive liver disease (De Koning et al. 1995) characterized by intermittent attacks of cholestasis without extrahepatic bile duct obstruction (Summerskil and Walshe 1959;Brenard et al. 1989). Each cholestatic attack starts with rising serum bile acid levels, usually followed by hyperbilirubinemia and overt cholestasis (Bijleveld et al. 1989;Brenard et al. 1989). The attacks vary in duration (weeks to months) and resolve spontaneously. No biochemical abnormalities are found in patients between attacks. The initial episode can occur at any age from infancy to adulthood, although most patients present before the age of 25 years. The frequency of attacks varies from several times a year to less than once per decade (Brenard et al. 1989). Progression to chronic liver disease has not been observed, but cholestatic episodes result in substantial morbidity, including pronounced weight loss and deficiency of fat-soluble vitamins. No established treatment is available, although cholestyramine may alleviate symptoms in some patients. Development of an effective therapy is hampered because the basic defect in BRIC is unknown.The gene for BRIC was originally mapped to chromosome 18q21 through the identification of a shared chromosomal segment in three distantly related patients from an isolated community in the Netherlands (Houwen et al. 1994). The region shared in these patients extended from D18S363 to D18S55, a genetic distance of more than 20 cM. To facilitate future molecular genetic studies we subsequently started to delimit the region of interest by studying additional BRIC families with a more densly spaced set of markers. We now present an analysis of 14 BRIC families from different ethnic sources with microsatellites selected primarily from the Généthon map, with the addi- Benign recurrent intrahepatic cholestasis (BRIC): evidence of genetic heterogeneity and delimitation of the BRIC locus to a 7-cM interval between D18S69 and D18S64

BackgroundThe addition of monoclonal antibody therapy against GD2 to the treatment of high-risk neuroblastoma led to improved responses in patients. Nevertheless, administration of GD2 antibodies against neuroblastoma is associated with therapy-limiting neuropathic pain. This severe pain is evoked at least partially through complement activation on GD2-expressing sensory neurons.MethodsTo reduce pain while maintaining antitumor activity, we have reformatted the approved GD2 antibody ch14.18 into the IgA1 isotype. This novel reformatted IgA is unable to activate the complement system but efficiently activates leukocytes through the FcαRI (CD89).ResultsIgA GD2 did not activate the complement system in vitro nor induced pain in mice. Importantly, neutrophil-mediated killing of neuroblastoma cells is enhanced with IgA in comparison to IgG, resulting in efficient tumoricidal capacity of the antibody in vitro and in vivo.ConclusionsOur results indicate that employing IgA GD2 as a novel isotype has two major benefits: it halts antibody-induced excruciating pain and improves neutrophil-mediated lysis of neuroblastoma. Thus, we postulate that patients with high-risk neuroblastoma would strongly benefit from IgA GD2 therapy.

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