Polygenic risk scores, radiation treatment exposures and subsequent cancer risk in childhood cancer survivors.

Survivors of childhood cancer are at increased risk for subsequent cancers attributable to the late effects of radiotherapy and other treatment exposures; thus, further understanding of the impact of genetic predisposition on risk is needed. Combining genotype data for 11,220 5-year survivors from the Childhood Cancer Survivor Study and the St Jude Lifetime Cohort, we found that cancer-specific polygenic risk scores (PRSs) derived from general population, genome-wide association study, cancer loci identified survivors of European ancestry at increased risk of subsequent basal cell carcinoma (odds ratio per s.d. of the PRS: OR = 1.37, 95% confidence interval (CI) = 1.29-1.46), female breast cancer (OR = 1.42, 95% CI = 1.27-1.58), thyroid cancer (OR = 1.48, 95% CI = 1.31-1.67), squamous cell carcinoma (OR = 1.20, 95% CI = 1.00-1.44) and melanoma (OR = 1.60, 95% CI = 1.31-1.96); however, the association for colorectal cancer was not significant (OR = 1.19, 95% CI = 0.94-1.52). An investigation of joint associations between PRSs and radiotherapy found more than additive increased risks of basal cell carcinoma, and breast and thyroid cancers. For survivors with radiotherapy exposure, the cumulative incidence of subsequent cancer by age 50 years was increased for those with high versus low PRS. These findings suggest a degree of shared genetic etiology for these malignancy types in the general population and survivors, which remains evident in the context of strong radiotherapy-related risk.

Development of a novel definitive scoring system for an enteral feed-only model of necrotizing enterocolitis in piglets.

Introduction: Necrotizing enterocolitis (NEC) is a complex inflammatory disorder of the human intestine that most often occurs in premature newborns. Animal models of NEC typically use mice or rats; however, pigs have emerged as a viable alternative given their similar size, intestinal development, and physiology compared to humans. While most piglet NEC models initially administer total parenteral nutrition prior to enteral feeds, here we describe an enteral-feed only piglet model of NEC that recapitulates the microbiome abnormalities present in neonates that develop NEC and introduce a novel multifactorial definitive NEC (D-NEC) scoring system to assess disease severity. Methods: Premature piglets were delivered via Caesarean section. Piglets in the colostrum-fed group received bovine colostrum feeds only throughout the experiment. Piglets in the formula-fed group received colostrum for the first 24 h of life, followed by Neocate Junior to induce intestinal injury. The presence of at least 3 of the following 4 criteria were required to diagnose D-NEC: (1) gross injury score ≥4 of 6; (2) histologic injury score ≥3 of 5; (3) a newly developed clinical sickness score ≥5 of 8 within the last 12 h of life; and (4) bacterial translocation to ≥2 internal organs. Quantitative reverse transcription polymerase chain reaction was performed to confirm intestinal inflammation in the small intestine and colon. 16S rRNA sequencing was performed to evaluate the intestinal microbiome. Results: Compared to the colostrum-fed group, the formula-fed group had lower survival, higher clinical sickness scores, and more severe gross and histologic intestinal injury. There was significantly increased bacterial translocation, D-NEC, and expression of IL-1α and IL-10 in the colon of formula-fed compared to colostrum-fed piglets. Intestinal microbiome analysis of piglets with D-NEC demonstrated lower microbial diversity and increased Gammaproteobacteria and Enterobacteriaceae. Conclusions: We have developed a clinical sickness score and a new multifactorial D-NEC scoring system to accurately evaluate an enteral feed-only piglet model of NEC. Piglets with D-NEC had microbiome changes consistent with those seen in preterm infants with NEC. This model can be used to test future novel therapies to treat and prevent this devastating disease.

EGFR internal tandem duplications in fusion-negative congenital and neonatal spindle cell tumors.

Next-generation sequencing (NGS) assays can sensitively detect somatic variation, and increasingly can enable the identification of complex structural rearrangements. A subset of infantile spindle cell sarcomas, particularly congenital mesoblastic nephromas with classic or mixed histology, have structural rearrangement in the form of internal tandem duplications (ITD) involving EGFR. We performed prospective analysis to identify EGFR ITD through clinical or research studies, as well as retrospective analysis to quantify the frequency of EGFR ITD in pediatric sarcomas. Within our institution, three tumors with EGFR ITD were prospectively identified, all occurring in patients less than 1 year of age at diagnosis, including two renal tumors and one mediastinal soft tissue tumor. These three cases exhibited both cellular and mixed cellular and classic histology. All patients had no evidence of disease progression off therapy, despite incomplete resection. To extend our analysis and quantify the frequency of EGFR ITD in pediatric sarcomas, we retrospectively analyzed a cohort of tumors (n = 90) that were previously negative for clinical RT-PCR-based fusion testing. We identified EGFR ITD in three analyzed cases, all in patients less than 1 year of age (n = 18; 3/18, 17%). Here we expand the spectrum of tumors with EGFR ITD to congenital soft tissue tumors and report an unusual example of an EGFR ITD in a tumor with cellular congenital mesoblastic nephroma histology. We also highlight the importance of appropriate test selection and bioinformatic analysis for identification of this genomic alteration that is unexpectedly common in congenital and infantile spindle cell tumors.

Calretinin Staining in Anorectal Line Biopsies Accurately Distinguished Hirschsprung Disease in a Retrospective Study.

INTRODUCTION: The absence of submucosal ganglion cells does not reliably distinguish Hirschsprung disease from non Hirschsprung disease in anorectal line biopsies. Calretinin staining might be helpful in these biopsies. To determine its value, we analyzed calretinin positive mucosal neurites in anorectal line biopsies. METHODS: Two pediatric pathologists, without access to patient data, evaluated calretinin positive mucosal neurites in anorectal line junctional mucosa in archival rectal biopsies contributed by 17 institutions. A separate investigator compiled patient information and sent data for statistical analysis. RESULTS: Biopsies with anorectal junctional mucosa from 115 patients were evaluated for calretinin positive mucosal neurites. 20/20 Hirschsprung disease biopsies were negative. 87/88 non Hirschsprung disease biopsies and 7/7 post pullthrough Hirschsprung disease neorectal biopsies were positive. Statistical analysis of the 108 non pullthrough biopsies yielded an accuracy of 99.1% (sensitivity 100%, specificity 98.9%). Age range was preterm to 16 years. Biopsy size was less than 1 mm to over 1 cm. CONCLUSIONS: Absence of calretinin positive mucosal neurites at the anorectal line was highly accurate in distinguishing Hirschsprung disease from non Hirschsprung disease cases in this blinded retrospective study. Calretinin staining is useful for interpreting biopsies from the physiologic hypoganglionic zone up to the anorectal line.

Expanding the clinical phenotype of FGFR1 internal tandem duplication.

Closed spinal dysraphism (SD) is a type of neural tube defect originating during early embryonic development whereby the neural tissue of the spinal defect remains covered by skin, often coinciding with markers of cutaneous stigmata. It is hypothesized that these events are caused by multifactorial processes, including genetic and environmental causes. We present an infant with a unique congenital midline lesion associated with a closed SD. Through comprehensive molecular profiling of the intraspinal lesion and contiguous skin lesion, an internal tandem duplication (ITD) of the kinase domain of the fibroblast growth factor receptor 1 (FGFR1) gene was found. This ITD variant is somatic mosaic in nature as supported by a diminished variant allele frequency in the lesional tissue and by its absence in peripheral blood. FGFR1 ITD results in constitutive activation of the receptor tyrosine kinase to promote cell growth, differentiation, and survival through RAS/MAPK signaling. Identification of FGFR1 ITD outside of central nervous system tumors is exceedingly rare, and this report broadens the phenotypic spectrum of somatic mosaic FGFR1-related disease.

Subsequent malignant neoplasms in the Childhood Cancer Survivor Study: Occurrence of cancer types in which human papillomavirus is an established etiologic risk factor.

BACKGROUND: Human papillomavirus (HPV)-associated subsequent malignant neoplasms (SMNHPV ) in childhood cancer survivors are poorly understood. METHODS: The cumulative risk of SMNHPV was assessed among 24,363 Childhood Cancer Survivor Study participants. Standardized incidence ratios (SIRs) and absolute excess risk were calculated using age-matched, sex-matched, and calendar year rates from the Surveillance, Epidemiology, and End Results program. Poisson regression models identified SMNHPV risk factors, evaluating relative SIRs (rSIR) and 95% confidence intervals (95% CIs). RESULTS: In total, 46 survivors developed an SMNHPV (median age, 31 years [range, 10-56 years]; median time from primary cancer, 21 years [range, 9-35 years]). SMNHPV sites included oropharynx (N = 44), anorectum (N = 6), uterine cervix (N = 2), and vulva (N = 2). The 33-year cumulative incidence was 0.3% (95% CI, 0.2%-0.4%), and the SIR was nearly 3-fold that of the general population (SIR, 2.86; 95% CI, 2.05-4.00). Female survivors were not at increased risk of cervical or vulvar cancers compared with the general population. All survivors had an elevated risk of oropharyngeal SMNHPV (males: SIR, 4.06; 95% CI, 2.37-6.97; females: SIR, 8.44; 95% CI 4.88-14.61) and anorectal SMNHPV (males: SIR, 13.56; 95% CI, 5.09-36.13; females: SIR, 9.15; 95% CI, 2.29-36.61). Males (vs females: rSIR, 1.99; 95% CI, 1.00-3.94); head, neck, and pelvic radiotherapy doses >3000 centigray (vs none: rSIR, 2.35; 95% CI, 1.11-4.97); and cisplatin-equivalent doses >400 mg/m2 (vs none: rSIR, 4.51; 95% CI, 1.78-11.43) were associated with increased SMNHPV SIRs in multivariable analysis. CONCLUSIONS: Childhood cancer survivors are at increased risk for SMN in sites susceptible to HPV-associated malignancies. Further research examining HPV in the etiology of SMN and the promotion of HPV vaccination and surveillance guidelines for SMNHPV in cancer survivors is warranted.

Submucosal Nerve Diameter in the Rectum Increases With Age: An Important Consideration for the Diagnosis of Hirschsprung Disease.

INTRODUCTION: Hypertrophic submucosal nerves, defined as ≥40 µm in diameter, are considered supportive of a diagnosis of HSCR, but the effect of age on nerve diameter has not been well-studied. We sought to determine the distribution of the largest nerve diameter in ganglionic rectal biopsies and the significance of hypertrophic submucosal nerves in the diagnosis of Hirschsprung disease (HSCR) based on age. METHODS: Rectal biopsies performed in the evaluation of HSCR were retrospectively reviewed from 179 patients (151 ganglionic biopsies, 28 aganglionic biopsies), and the diameter of the largest submucosal nerve was measured. RESULTS: In non-Hirschsprung disease (non-HSCR) biopsies, submucosal nerve diameter increased with age. In patients <1 year, the average diameter was 34.1 ± 11.6 µm but increased to 50.8 ± 17.3 µm after 1 year of age. Submucosal nerves ≥40 µm in diameter were significantly associated with HSCR across all ages [HSCR = 25/28 (89.3%) vs non-HSCR = 59/151 (39.1%), p < 0.0001] and remained significant in patients <1 year of age [HSCR = 22/24 (91.7%) vs non-HSCR = 19/91 (20.9%), p < 0.0001]. CONCLUSIONS: The diameter of submucosal nerves increases with age, and ≥40 µm nerves are common after 1 year of age.

Novel morphologic findings in PLAG1-rearranged soft tissue tumors.

Oncogenesis in PLAG1-rearranged tumors often results from PLAG1 transcription factor overexpression driven by promoter-swapping between constitutively expressed fusion partners. PLAG1-rearranged tumors demonstrate diverse morphologies. This study adds to this morphologic heterogeneity by introducing two tumors with PLAG1 rearrangements that display distinct histologic features. The first arose in the inguinal region of a 3-year-old, appeared well-circumscribed with a multinodular pattern, and harbored two fusions: ZFHX4-PLAG1 and CHCHD7-PLAG1. The second arose in the pelvic cavity of a 15-year-old girl, was extensively infiltrative and vascularized with an adipocytic component, and demonstrated a COL3A1-PLAG1 fusion. Both showed low-grade cytomorphology, scarce mitoses, no necrosis, and expression of CD34 and desmin. The ZFHX4-/CHCHD7-PLAG1-rearranged tumor showed no evidence of recurrence after 5 months. By contrast, the COL3A1-PLAG1-rearranged tumor quickly recurred following primary excision with positive margins; subsequent re-excision with adjuvant chemotherapy resulted in no evidence of recurrence after 2 years. While both tumors show overlap with benign and malignant fibroblastic and fibrovascular neoplasms, they also display divergent features. These cases highlight the importance of appropriate characterization in soft tissue tumors with unusual clinical and histologic characteristics.

Neuropathology of Mowat-Wilson Syndrome.

Mowat-Wilson syndrome (MWS) is a syndromic form of Hirschsprung disease that is characterized by variable degrees of intellectual disability, characteristic facial dysmorphism, and a diverse set of other congenital malformations due to haploinsufficiency of ZEB2. A variety of brain malformations have been described in neuroimaging studies of MWS patients, and the role of ZEB2 in the brain has been studied in a multitude of genetically engineered mouse models that are now available. However, a paucity of autopsy information limits our ability to correlate data from neuroimaging studies and animal models with actual MWS patient tissues. Here, we report the autopsy neuropathology of a 19-year-old male patient with MWS. Autopsy neuropathology findings correlated well with the reported MWS neuroimaging data and are in keeping with data from genetically engineered MWS mouse models. This autopsy enhances our understanding of ZEB2 function in human brain development and demonstrates the reliability of MWS murine models.

Cytoplasmic Fibrillar Aggregates in Gallbladder Epithelium Are a Frequent Mimic of Cystoisospora in Pediatric Cholecystectomy Specimens.

CONTEXT.­: Cystoisospora belli is an intracellular parasite associated with gastrointestinal disease in immunocompromised hosts. Although infection has been classically associated with intestinal disease, studies have identified Cystoisospora in the gallbladder of immunocompetent patients based on hematoxylin-eosin morphology. Recently, the identity of this histologic finding as Cystoisospora has been questioned based on negative results of nucleic acid studies. OBJECTIVE.­: To determine the prevalence of this histologic feature in pediatric patients, we retrospectively reviewed all cholecystectomy specimens from a pediatric hospital during a 24-month period. DESIGN.­: In 180 cholecystectomy specimens, we identified 11 cases (6.1%) with classical histologic features previously described to represent Cystoisospora organisms. To further investigate these structures, we retrieved tissue from paraffin-embedded blocks and performed electron microscopy. RESULTS.­: Ultrastructural examination identified ovoid perinuclear cytoplasmic structures composed of dense fibrillar aggregates rather than organisms. Patients with positive cases were similar in age to controls (positive cases: mean patient age 13.4 years [range, 2-23 years]; negative cases: mean patient age 14.7 years [range, 12 weeks-31 years]; P = .35). There was no significant association of this finding with cholelithiasis (54.5% versus 65.1%, P = .52), cholesterolosis (0% versus 22.5%, P = .12), acute cholecystitis (9.1% versus 10.1%, P > .99), or chronic cholecystitis (45.5% versus 66.3%, P = .20). CONCLUSIONS.­: To our knowledge, this is the first positive identification of these structures as cytoplasmic fibrillar aggregates rather than parasitic inclusions by ultrastructural examination, and the first study of this histologic finding in pediatric cholecystectomies.

Infra-anastomotic Innervation of Residual Aganglionic Distal Rectum After Pull-through Surgery for Hirschsprung Disease.

BACKGROUND: Descending neurons are important for intestinal reflex activities, including the recto-anal inhibitory reflex involved in normal defecation. Pull-through surgery for Hirschsprung disease results in the anastomosis of ganglionic bowel to native aganglionic rectum just superior to the internal anal sphincter, which potentially could allow for physiologically significant infra-anastomotic innervation. METHODS: The density and distribution of intramuscular neuronal nitric oxide synthase (nNOS)- and mucosal calretinin-immunoreactive nerves were evaluated proximal and distal to the anastomosis in redo resection specimens after pull-through surgery for Hirschsprung disease. The findings were compared with data collected from cadaveric controls with no history of dysmotility and the distal aganglionic segments of primary rectal resections from patients with Hirschsprung disease. RESULTS: Native aganglionic rectum of Hirschsprung patients lacks the normal lush intramuscular nNOS- and mucosal calretinin-immunoreactive nerves present in normal bowel. In post-pull-through resection specimens obtained more than 7 months after pull-through surgery, nNOS- and calretinin-positive innervation is at least partially restored for variable distances up to 10 to 12 mm inferior to the anastomosis, respectively. CONCLUSIONS: Innervation of infra-anastomotic muscularis propria and mucosa in the aganglionic distal rectum occurs to a variable degree after pull-through surgery for Hirschsprung disease and may contribute to individual differences in postoperative obstructive symptoms. Strategies to enhance infra-anastomotic innervation may improve clinical outcome.

Remodeling of Rectal Innervation After Pullthrough Surgery for Hirschsprung Disease: Relevance to Criteria for the Determination of Retained Transition Zone.

BACKGROUND: After pullthrough surgery for Hirschsprung disease (HSCR), Glut1-positive submucosal nerve hypertrophy is used to diagnose retained transition zone in the neorectum. We hypothesized that pelvic nerves, severed during pullthrough surgery, sprout into the neorectum to mimic transition zone. METHODS: The density (nerves/100x field) and maximum diameter of Glut1-positive submucosal nerves were measured in biopsies and redo resections from 20 patients with post-pullthrough obstructive symptoms. Their original and/or redo resections excluded unequivocal features of transition zone (myenteric hypoganglionosis or partial circumferential aganglionosis) in 17. Postoperative values were compared with control data from 28 cadaveric and 6 surgical non-HSCR specimens, and 14 primary HSCR resections. When possible, nerves were tracked from attached native pelvic soft tissue or aganglionic rectal cuff into the pulled-through colon. RESULTS: Glut1-positive submucosal nerves were not present in the 11 colons of non-HSCR infants less than 1 year of age, except sparsely in the rectum. In 17 older non-HSCR controls, occasional Glut1-positive nerves were observed in prerectal colon and were larger and more numerous in the rectum. In redo resections, Glut1-positive submucosal innervation in post-pullthrough specimens did not differ significantly from age-appropriate non-HSCR rectal controls and pelvic Glut1-positive nerves were never observed to penetrate the pulled-through colon. However, the density and caliber of Glut1-positive nerves in the neorectums were significantly greater than expected based on the prerectal location from which the pulled-through bowel originated. CONCLUSIONS: Submucosal innervation in post-pullthrough specimens does not support the hypothesis that native pelvic nerves innervate the neorectum, but suggests remodeling occurs to establish the age-appropriate density and caliber of rectal Glut1-positive innervation. The latter should not be interpreted as transition zone pullthrough in a rectal biopsy from a previously done pullthrough.

Adenovirus and "Culture-Negative Sepsis" in a Preterm Neonate.

Background Respiratory viral infections remain an underrecognized cause of morbidity and mortality among preterm infants in the neonatal intensive care unit (NICU). Case Report An eight day old, 650 gram birth weight, 23 weeks' gestational age female developed "culture-negative" sepsis manifested by respiratory deterioration, hypoxia, leukocytosis, and thrombocytopenia. She was diagnosed with pneumonia and hepatitis due to adenovirus HAdV-D (H29F9) by polymerase chain reaction (PCR) testing, but died at the age of 18 days despite treatment with cidofovir and immune globulin intravenous. Conclusion As the ability to diagnosis respiratory viral infections in the NICU has improved greatly with the use of PCR testing, the impact and contribution of these viruses to neonatal disease is now being recognized and the notion of "culture-negative" sepsis needs reassessment. The diagnosis of these infections in high risk infants is important not only for etiologic and epidemiologic reasons but ultimately for informing antimicrobial stewardship efforts.

A Strategy for Helicobacter Immunohistochemistry Utilization in Pediatric Practice: Insights From Morphologic and Cost-Benefit Analyses.

OBJECTIVES: Recent studies in adults have examined the utility of immunohistochemistry (IHC) in detecting Helicobacter in gastric biopsy specimens and reached differing conclusions. Dedicated cost-benefit analysis of Helicobacter IHC in pediatric gastric biopsy specimens has not been performed. METHODS: From 1,955 pediatric gastric biopsies in a 1-year period, we identified 63 Helicobacter -positive and 120 Helicobacter -negative biopsy specimens. All cases were scored according to the Updated Sydney System for the severity of inflammation. RESULTS: We observed that pediatric Helicobacter infection was significantly associated with germinal center formation, active inflammation, oxyntic mucosa with moderate to severe chronic inflammation, and antral mucosa with any chronic inflammation, exclusive of mild and superficial chronic inflammation. At least one associated pattern was seen in each Helicobacter -positive biopsy specimen. In comparison with adults, pediatric Helicobacter -positive biopsy specimens are more likely to lack acute inflammation and more likely to show moderate to marked chronic inflammation. CONCLUSIONS: We recommend performing Helicobacter IHC on pediatric gastric biopsy specimens with any of the above inflammatory patterns. This approach can sensitively identify pediatric patients with Helicobacter gastritis, limit IHC staining to approximately 30% of all gastric biopsy specimens, and reduce costs by up to $55,306.90 per 1,000 biopsy specimens.

Urethral caruncle: a lesion related to IgG4-associated sclerosing disease?

AIMS: Urethral caruncle is a benign, polypoid urethral mass that occurs almost exclusively in postmenopausal women. Despite that these lesions are routinely managed with topical medications or excision, their pathogenesis is not well understood. We investigated the possibilities of autoimmune, viral and inflammatory myofibroblastic proliferations as possible aetiologies. METHODS: In 38 patients with urethral caruncle, we utilised immunohistochemistry for immunoglobulin G (IgG) and IgG4 to assess for a potential autoimmune aetiology. Immunohistochemistry was performed in nine patients for Epstein-Barr virus, BK virus, human herpesvirus 8, human papillomavirus, adenovirus and anaplastic lymphoma kinase. RESULTS: Four patients (11%) showed infiltrates of ≥50 IgG4-positive plasma cells per high power field, of which all showed an IgG4 to IgG ratio greater than 40%. A statistically significant difference (p<0.01) was detected in the mean number of IgG4-positive cells (14.73 per high power field) compared with control benign urethral specimens (mean, 1.19). One patient with increased counts below this threshold had rheumatoid arthritis; none had documented autoimmune pancreatitis or other known manifestations of systemic IgG4-related sclerosing disease. All lesions showed negative reactions for the viral and inflammatory myofibroblastic markers. CONCLUSIONS: Urethral caruncle is a benign inflammatory and fibrous polypoid urethral mass of unclear aetiology. It appears unrelated to viral infection and lacks the abnormal expression of anaplastic lymphoma kinase protein, as seen in inflammatory myofibroblastic tumours. Increased numbers of IgG4-positive plasma cells in a subset of lesions raise the possibility that some cases may be related to the autoimmune phenomena of IgG4-associated disease.

Urethral caruncle: clinicopathologic features of 41 cases.

Urethral caruncle is a benign polypoid mass of the urethral meatus in primarily postmenopausal women. Although a conclusive association with malignancy, urologic disorder, or systemic disease has not been established, often the lesion carries a challenging clinical differential diagnosis that includes malignancy. Conversely, unexpected malignancy is identified in some cases resembling caruncle clinically. We examined clinical and histopathologic characteristics in 41 patients. Medical records were assessed for presentation, clinical diagnosis, associated urothelial carcinoma, radiation treatment, tobacco use, immunologic/urologic disorder, and treatment strategy/outcome. Average patient age was 68 years (range, 28-87 years). Presenting symptoms were pain (37%), hematuria (27%), and dysuria (20%), in contrast to asymptomatic (32%). Clinical diagnosis favored malignancy in 10% of cases. Concurrent or subsequent urothelial carcinoma was present for 5 patients (12%), although none developed urethral carcinoma. Histologic features included mixed hyperplastic urothelial and squamous lining, overlying a variably fibrotic, edematous, inflamed, and vascular stroma. Invaginations of urothelium extending into the stroma were common (68%), showing rounded nests with cystic or glandular luminal spaces, similar to urethritis cystica/glandularis, without intestinal metaplasia. Two lesions included an organizing thrombus, 1 with intravascular papillary endothelial hyperplasia. Twenty patients were treated with topical medications without resolution. Three lesions recurred (7%) after excision. A subset of patients had history of smoking or previous pelvic irradiation. Urethral caruncle is an uncommon lesion that may clinically mimic benign and malignant conditions. Awareness of the spectrum of clinical and histologic differential diagnoses is important in dealing with this unusual disease.