ObjectivesWe present an adolescent in whom olfactory neuroblastoma (ONB) was detected on follow-up magnetic resonance imaging (MRI) 2.5 years after SIADH diagnosis. Our case contrasts prior pediatric reports in which ONB and SIADH were diagnosed concurrently.Case presentationA previously healthy 13-year-old girl was found to have SIADH during evaluation for restrictive eating. Work-up ruled out adrenal, thyroid and paraneoplastic causes, diuretic use, and vasopressin receptor and aquaporin channel mutations. Brain MRI was normal except for paranasal sinus (PNS) inflammatory changes to the left fronto-maxillary sinuses and frontoethmoidal recess. The sodium levels normalized with fluid restriction (800-900 ml/m2/day). Multiple repeated attempts to liberalize fluid intake resulted in recurrent hyponatremia. Follow-up brain MRIs 4 and 11 months after the initial presentation showed persistent PNS inflammatory changes. A subsequent brain MRI 31 months after initial presentation demonstrated a lesion in the left frontoethmoidal recess extending into the left nasal cavity and biopsy showed low grade ONB. The patient underwent surgery with normalization of serum sodium on liberalized fluid intake. Seven days after surgery, she had recurrence of SIADH, and brain MRI showed remnant of the ONB at the fovea ethmoidalis. She completed adjuvant radiotherapy though her SIADH persisted.ConclusionsOur case highlights the importance of considering ONB in the evaluation of children with SIADH. Idiopathic SIADH is rare in children and if no cause is identified, computed tomography of sinuses and nasal endoscopy should be considered earlier in the work-up of these patients, particularly in the absence of sinus symptoms.

Due to the rarity and genomic disparity of soft tissue and bone sarcomas, actionable therapeutic targets have been elusive. However, inhibition of GSK-3β has emerged as a potentially promising therapy that could be of great benefit to pediatric and adult sarcoma patients. Saos-2 osteosarcoma and 93T449 liposarcoma cell lines were pretreated at IC5072 with small molecule GSK-3β inhibitor 9-ING-41 (elraglusib). The cells were harvested for western blot analysis after 48 hours of treatment. Additionally, both cancer cell lines as well as TALL-104 T-cells and NK-92 NK cells were treated with 0.5 μM 9-ING-41 for 24 hours and harvested for Luminex cytokine profiling. The western blots demonstrated an increase in cPARP, an apoptotic marker, in both cancer cell lines after 9-ING-41 treatment. Additionally, an increase in PD-L1 expression was observed. The cytokine analysis revealed stimulation of immune cell activity in response to 9-ING-41. Treated T-cells had an increase in CXCL11, which is associated with T-cell recruitment, as well as an increased level of IL-18, which is shown to induce increased IFN-γ in Th1 cells. Additionally, NK-92 cells demonstrated an increase in IL-8 chemokine and an increase in soluble TRAIL (TRAIL/TNFSF10). The cancer cell lines showed a homogenous increase in growth factor TGF-ɑ, however, only the Saos-2 osteosarcoma cell line demonstrated an increase of IL-6. We are further validating the data with follow-up cytokine profiling. The increase in immunostimulatory cytokines as well as the increased expression of PD-L1 suggest a rationale for combining 9-ING-41 with immune checkpoint blockade therapy. The potential synergistic effect of these two therapies is currently under investigation with co-culture experiments of sarcoma and immune cells. The treatment cohorts for these experiments include 9-ING-41 combined with either anti-PD-L1, anti-PD-1, or anti-CTLA-4 immune checkpoint inhibitors. Our results suggest a promising combination therapeutic strategy for patients with soft tissue and bone sarcomas and future work will strive to better elucidate the mechanisms of efficacy. Citation Format: William J. MacDonald, Samuel Lew, Kelsey E. Huntington, Rishi R. Lulla, Bradley D. DeNardo, Wafik S. El-Deiry. The immunostimulatory effect of 9-ING-41, a small molecule GSK-3 inhibitor, in sarcomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3251.

Our Laboratory was established in 1994 at Univ. of Pennsylvania. Lab members demonstrated initial competencies by performing cell culture, western blots, immunofluorescence, and flow cytometry showing induction of p53/p21(WAF1) in cells treated with chemotherapy. Years later, our Laboratory of Translational Oncology & Experimental Cancer Therapeutics moved to Penn State Univ., Fox Chase Cancer Center/Temple Univ. and then Brown Univ. By 2020, with desire for inclusiveness (everyone succeeds), scientific rigor/reproducibility mandated by NIH, and as a training and mentoring activity (lab scientists/trainees/students mentoring others at High School level and beyond), we established a process for onboarding and training new cancer researchers. By Fall of 2022, there were 17 current Brown University undergraduate students (10 receiving research credit and 7 not receiving credit), HS students, 7 graduate students (PhD, masters, MD/PhD), and 6 medical students working with collaborating faculty at our laboratory at Brown’s Legorreta Cancer Center. After completion of biosafety training, and required trainings such as by IACUC, new lab members complete basic competencies in cell culture, cell viability, and western blot analysis that include technical, presentation quality output, and quantitative/statistical rigor to satisfy current standards for journal publication. For cell culture this includes pathogen free conditions, authentication, attention to details of routine procedures, documentation of morphology, freezing, thawing, passaging, seeding density, and managing cell populations to not run out of cells. Cell viability assessment includes attention to culture conditions, synergy analysis, data robustness, and presentation, and for western blots attention to quality of blots, protein quantification, loading, labeling, antibody specificity and sensitivity controls, presentation at 2022 standards, conventions for splicing, and issues with reproducibility including biological replicates, and generalizability. Additional and advanced competencies include RT-PCR, long-term colony assays, 3-D cultures (spheroids, organoids), transfection (overexpression, knockdown, CRISPR), co-culture and triculture with immune cells and fibroblasts, cytokine profiling, in vivo studies, in vivo imaging, immunohistochemistry, flow cytometric analysis, single cell techniques, viral infection, circulating tumor cell isolation, blood immune and cytokine analysis, and work with transgenic organoids and inducible cancer predisposing alleles. Modeling the tumor microenvironment, relevance to human cancer and translational directions are emphasized. Shared online lab resources, protocols, practices, videos, and manuscripts are available for lab members. The framework herein may be of interest to others involved in similar training programs. Citation Format: Wafik S. El-Deiry, Andrew George, Francesca Di Cristofano, Praveen Srinivasan, Lindsey Carlsen, Kelsey E. Huntington, Arielle De La Cruz, Leiqing Zhang, Marina Hahn, Shuai Zhao, Attila Seyhan, Bradley D. DeNardo, Aaron W. Maxwell, Dae Hee Kim, Alex Raufi, Hina Khan, Stephanie L. Graff, Don S. Dizon, Christopher Azzoli, Abbas E. Abbas, Roxanne Wood, Rishi R. Lulla, Howard P. Safran, Benedito A. Carneiro, Arunasalam Navaraj, Xiaobing Tian, Shengliang Zhang, Lanlan Zhou. Inclusive basic and advanced translational laboratory research competencies for research in cancer biology and therapeutics. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4185.

Vincristine, a key agent in the treatment of many pediatric malignancies, causes sensory, motor and autonomic neuropathy. We report the clinical courses of 5 patients who required cessation of vincristine after developing severe neurotoxicity during treatment for acute lymphoblastic leukemia. All 5 patients lost the ability to ambulate and 3 had additional severe neurotoxic side effects including vision loss and vocal cord dysfunction. Although prior literature reports poor outcomes for children in whom vincristine was discontinued during acute lymphoblastic leukemia therapy, all 5 patients described here achieved and have maintained complete continuous remission.

• Extrapulmonary lymphangioleiomyomatosis is rare and can be associated with tuberous sclerosis. • Recognition of lymphangioleiomyomatosis is important for early disease screening and genetic testing. • Lymphangioleiomyomatosis in lower uterine segment is very rare and can be overlooked.

Alveolar soft part sarcoma (ASPS) is a rare soft tissue sarcoma that presents most commonly in the extremities of adolescents and young adults. Involvement of the head and neck is more common in younger patients. A 2-year-old found to have localized lingual ASPS and was treated successfully with partial glossectomy. We describe this rare form of sarcoma, discuss management options, review updated literature and present our patient's outcome after 5 years of close follow-up.

Targeted next-generation sequencing (NGS) identified a novel loss of function mutation in GARS , a gene linked to Charcot–Marie–Tooth disease (CMT), in a paediatric acute lymphoblastic leukaemia patient with severe chemotherapy-induced peripheral neuropathy (CIPN) due to vincristine. The patient was clinically asymptomatic, and lacked a family history of neuropathy. The effect of the mutation was modelled in a zebrafish knockdown system that recapitulated the symptoms of the patient both prior to and after treatment with vincristine. Confocal microscopy of pre- and post-synaptic markers revealed that the GARS knockdown results in changes to peripheral motor neurons, acetylcholine receptors and their co-localisation in neuromuscular junctions (NMJs), whereas a sensitive and reproducible stimulus–response assay demonstrated that the changes correlating with the GARS mutation in themselves fail to produce peripheral neuropathy symptoms. However, with vincristine treatment the GARS knockdown exacerbates decreased stimulus response and NMJ lesions. We propose that there is substantial benefit in the use of a targeted NGS screen of cancer patients who are to be treated with microtubule targeting agents for deleterious mutations in CMT linked genes, and for the screening in zebrafish of reagents that might inhibit CIPN.

Neuroblastoma is an embryonal tumor of childhood with a heterogenous clinical presentation that reflects differences in activation of complex biological signaling pathways. Protein phosphorylation is a key component of cellular signal transduction and plays a critical role in processes that control cancer cell growth and survival. We used shotgun LC/MS to compare phosphorylation between a human MYCN amplified neuroblastoma cell line (NB10), modeling a resistant tumor, and a human neural precursor cell line (NPC), modeling a normal baseline neural crest cell. 2181 unique phosphorylation sites representing 1171 proteins and 2598 phosphopeptides were found. Protein kinases accounted for 6% of the proteome, with a predominance of tyrosine kinases, supporting their prominent role in oncogenic signaling pathways. Highly abundant receptor tyrosine kinase (RTK) phosphopeptides in the NB10 cell line relative to the NPC cell line included RET, insulin-like growth factor 1 receptor/insulin receptor (IGF-1R/IR), and fibroblast growth factor receptor 1 (FGFR1). Multiple phosphorylated peptides from downstream mediators of the PI3K/AKT/mTOR and RAS pathways were also highly abundant in NB10 relative to NPC. Our analysis highlights the importance of RET, IGF-1R/IR and FGFR1 as RTKs in neuroblastoma and suggests a methodology that can be used to identify potential novel biological therapeutic targets. Furthermore, application of this previously unexploited technology in the clinic opens the possibility of providing a new wide-scale molecular signature to assess disease progression and prognosis.