Antisense Oligonucleotide Therapy for Calmodulinopathy.

BACKGROUND: Calmodulinopathies are rare inherited arrhythmia syndromes caused by dominant heterozygous variants in CALM1, CALM2, or CALM3, which each encode the identical CaM (calmodulin) protein. We hypothesized that antisense oligonucleotide (ASO)-mediated depletion of an affected calmodulin gene would ameliorate disease manifestations, whereas the other 2 calmodulin genes would preserve CaM level and function. METHODS: We tested this hypothesis using human induced pluripotent stem cell-derived cardiomyocyte and mouse models of CALM1 pathogenic variants. RESULTS: Human CALM1F142L/+ induced pluripotent stem cell-derived cardiomyocytes exhibited prolonged action potentials, modeling congenital long QT syndrome. CALM1 knockout or CALM1-depleting ASOs did not alter CaM protein level and normalized repolarization duration of CALM1F142L/+ induced pluripotent stem cell-derived cardiomyocytes. Similarly, an ASO targeting murine Calm1 depleted Calm1 transcript without affecting CaM protein level. This ASO alleviated drug-induced bidirectional ventricular tachycardia in CalmN98S/+ mice without a deleterious effect on cardiac electrical or contractile function. CONCLUSIONS: These results provide proof of concept that ASOs targeting individual calmodulin genes are potentially effective and safe therapies for calmodulinopathies.

Professional Master of Health Science in Laboratory Medicine: Training Clinical Laboratory Scientists in an Integrated Program at a Research-Intensive University.

A 2-year professional master of health science program at the University of Toronto provides a unique integrated educational program to train allied health science personnel to practice as physician extenders and health care professionals in two high-demand clinical laboratory disciplines, Pathologists' Assistant (PA) and Clinical Embryologist (CE). This report describes an integrated graduate program developed and delivered in a research-intensive laboratory medicine department. The core courses in fundamental biomedical science and in general medical laboratory function and operations formed the foundation on which the requisite clinical skills required to practice as a PA or CE were subsequently delivered as comprehensive CE and PA specialty courses and practicums. Students acquired research skills through courses that teach research methods, critical analysis of research articles, and biostatistics for clinical research scientists. A capstone research project provided students the opportunity to design a research project relevant to the CE or PA fields, perform and analyze the findings, and present the project as an oral abstract and a written scientific article. Students learn to face the clinical challenges by focusing on critical analysis of evidence-based professional practice. The PA field received a 5-year accreditation. CE and PA students presented their clinical research at national and international meetings, with some receiving awards, and published scientific articles. All graduates found meaningful employment in their respective fields, and initial employer response has been favorable.

Clinicopathologic Features of Gastrointestinal Tract Langerhans Cell Histiocytosis.

Gastrointestinal (GI) tract involvement by Langerhans cell histiocytosis (LCH) is rare and its clinicopathologic characteristics have only been described in case reports and small series. We reviewed hematoxylin and eosin and CD1a, S100, and Langerin immunohistochemical-stained slides from 47 patients with well-documented demographic and clinical findings. Our cases included 8 children and 39 adults, with a mean follow-up of 63 months. All pediatric patients had concurrent multisystem LCH, presented with GI symptoms, and showed nonpolypoid lesions. Seven (88%) showed multifocal GI disease, including 5 with multiple GI organ involvement. All sampled lesions from children exhibited infiltrative growth. More than half had died of the disease or manifested persistent LCH at last follow-up. Twenty-five of 39 (64%) adults had LCH involving only the GI tract (single system), with the remaining 14 (36%) exhibiting multisystem disease. Adult single-system GI LCH was typically encountered incidentally on screening/surveillance endoscopy (72%). Most exhibited isolated colorectal involvement (88%) as a solitary polyp (92%), with a well-demarcated/noninfiltrative growth pattern (70%), and excellent prognosis (100%). In comparison, adult patients with multisystem LCH more frequently presented with GI symptoms (92%, P < .001), noncolorectal GI site involvement (50%, P = .02), multifocal GI lesions (43%, P = .005), nonpolypoid lesions (71%, P < .001), infiltrative histologic growth pattern (78%, P = .04), and persistent disease (57%, P < .001). Adult patients with multisystem LCH appear to exhibit similar clinicopathologic features to those of pediatric patients. These results demonstrated that adults with single-system LCH involving the GI tract have an excellent prognosis, whereas multisystem LCH occurring at any age carries an unfavorable prognosis. High-risk features of GI LCH include pediatric age, GI symptomatology, noncolorectal GI involvement, multifocal GI disease, nonpolypoid lesions, and infiltrative growth pattern.

Diagnostic approach to hepatic vascular lesions: a paediatric perspective.

The pathological evaluation of hepatic vascular lesions in children requires special consideration. Inconsistent terminology, rarity of pathology specimens and overlapping pathological features between various lesions may pose a serious diagnostic challenge. In this review, we highlight the importance of using the International Society for the Study of Vascular Anomalies (ISSVA) classification scheme to characterise these lesions. Selected entities are discussed, including hepatic vascular tumours exclusively seen in the paediatric age group, hepatic infantile haemangioma and hepatic congenital haemangioma. Vascular malformations, with emphasis on their syndromic associations (venous malformation in blue rubber bleb naevus syndrome) and complications (hepatocellular nodules in Abernethy malformation) are also covered.

Management of undifferentiated embryonal sarcoma of the liver: A Pediatric Surgical Oncology Research Collaborative study.

BACKGROUND: Undifferentiated embryonal sarcoma of the liver (UESL) is a rare tumor for which there are few evidence-based guidelines. The aim of this study was to define current management strategies and outcomes for these patients using a multi-institutional dataset curated by the Pediatric Surgical Oncology Research Collaborative. METHODS: Data were collected retrospectively for patients with UESL treated across 17 children's hospitals in North America from 1989 to 2019. Factors analyzed included patient and tumor characteristics, PRETEXT group, operative details, and neoadjuvant/adjuvant regimens. Event-free and overall survival (EFS, OS) were the primary and secondary outcomes, respectively. RESULTS: Seventy-eight patients were identified with a median age of 9.9 years [interquartile range [IQR): 7-12]. Twenty-seven patients underwent resection at diagnosis, and 47 patients underwent delayed resection, including eight liver transplants. Neoadjuvant chemotherapy led to a median change in maximum tumor diameter of 1.6 cm [IQR: 0.0-4.4] and greater than 90% tumor necrosis in 79% of the patients undergoing delayed resection. R0 resections were accomplished in 63 patients (81%). Univariate analysis found that metastatic disease impacted OS, and completeness of resection impacted both EFS and OS, while multivariate analysis revealed that R0 resection was associated with decreased expected hazards of experiencing an event [hazard ratio (HR): 0.14, 95% confidence interval (CI): 0.04-0.6]. At a median follow-up of 4 years [IQR: 2-8], the EFS was 70.0% [95% CI: 60%-82%] and OS was 83% [95% CI: 75%-93%]. CONCLUSION: Complete resection is associated with improved survival for patients with UESL. Neoadjuvant chemotherapy causes minimal radiographic response, but significant tumor necrosis.

Survey Data and Experience from a Pediatric Pathology Workshop in Indonesia: Understanding Practice Needs and Utility of Outreach Teachings.

Objective: We aimed to share the post-workshop survey results of a pediatric pathology course held in Jakarta, Indonesia. Methods: Questionnaires were distributed to participants; responses from practicing pathologists and pathologists-in-training were analyzed. Results: The respondents (107 pathologists of 143 attendees) were predominantly female (83.2%) and 31-60 years of age (77.5%). Over half (71.7%) signed out pediatric and perinatal specimens but only a third (34.3%) were comfortable handling such cases. Most (70.0%) felt that their exposure to pediatric and perinatal cases during their training was inadequate. All respondents thought that the workshop was helpful, and would highly recommend it to their colleagues. Post-workshop, the respondents claimed expansion of differential diagnoses (49.5%) and better understanding of what to include in pathology reports (41.1%). Conclusions: Our experience affirms the need for subspecialty courses to address training gaps in developing countries. Post-workshop surveys are helpful in determining actionable deficiencies and effectiveness of outreach teachings.

Using Deep Learning to Automate Eosinophil Counting in Pediatric Ulcerative Colitis Histopathological Images.

Background: Accurate identification of inflammatory cells from mucosal histopathology images is important in diagnosing ulcerative colitis. The identification of eosinophils in the colonic mucosa has been associated with disease course. Cell counting is not only time-consuming but can also be subjective to human biases. In this study we developed an automatic eosinophilic cell counting tool from mucosal histopathology images, using deep learning. Method: Four pediatric IBD pathologists from two North American pediatric hospitals annotated 530 crops from 143 standard-of-care hematoxylin and eosin (H & E) rectal mucosal biopsies. A 305/75 split was used for training/validation to develop and optimize a U-Net based deep learning model, and 150 crops were used as a test set. The U-Net model was then compared to SAU-Net, a state-of-the-art U-Net variant. We undertook post-processing steps, namely, (1) the pixel-level probability threshold, (2) the minimum number of clustered pixels to designate a cell, and (3) the connectivity. Experiments were run to optimize model parameters using AUROC and cross-entropy loss as the performance metrics. Results: The F1-score was 0.86 (95%CI:0.79-0.91) (Precision: 0.77 (95%CI:0.70-0.83), Recall: 0.96 (95%CI:0.93-0.99)) to identify eosinophils as compared to an F1-score of 0.2 (95%CI:0.13-0.26) for SAU-Net (Precision: 0.38 (95%CI:0.31-0.46), Recall: 0.13 (95%CI:0.08-0.19)). The inter-rater reliability was 0.96 (95%CI:0.93-0.97). The correlation between two pathologists and the algorithm was 0.89 (95%CI:0.82-0.94) and 0.88 (95%CI:0.80-0.94) respectively. Conclusion: Our results indicate that deep learning-based automated eosinophilic cell counting can obtain a robust level of accuracy with a high degree of concordance with manual expert annotations.

Metastatic Lymphoepithelioma-like Hepatocellular Carcinoma: a Potential Diagnostic Pitfall and Demonstration of Pd-l1 Expression

Abstract: Lymphoepithelioma-like hepatocellular carcinoma (LEL-HCC) is a rare primary hepatic neoplasm with female predominance and relatively good prognosis. We report a 73-year-old female with chronic hepatitis B who developed metastatic lesions 5 years after underwent resection for LEL-HCC. The metastatic lesions showed a spectrum of morphologic findings, which could be mistaken for other entities such as lymphoma, particularly in lesions with single-cell infiltrative pattern and abundant tumor-infiltrating lymphocytes. Immunohistochemical study to confirm the origin of the neoplastic cells is important to make the diagnosis. We also highlighted the clinicopathologic correlation and potential therapeutic implication of programmed death ligand-1 expression in LEL-HCC.