Norma M. Quintanilla

Diagnostic Yield of Clinical Tumor and Germline Whole-Exome Sequencing for Children With Solid Tumors.

Importance Whole-exome sequencing (WES) has the potential to reveal tumor and germline mutations of clinical relevance, but the diagnostic yield for pediatric patients with solid tumors is unknown. Objective To characterize the diagnostic yield of combined tumor and germline WES for children with solid tumors. Design Unselected children with newly diagnosed and previously untreated central nervous system (CNS) and non-CNS solid tumors were prospectively enrolled in the BASIC3 study at a large academic childrens hospital during a 23-month period from August 2012 through June 2014. Blood and tumor samples underwent WES in a certified clinical laboratory with genetic results categorized on the basis of perceived clinical relevance and entered in the electronic health record. Main Outcomes and Measures Clinical categorization of somatic mutations; frequencies of deleterious germline mutations related to patient phenotype and incidental medically-actionable mutations. Results Of the first 150 participants (80 boys and 70 girls, mean age, 7.4 years), tumor samples adequate for WES were available from 121 patients (81%). Somatic mutations of established clinical utility (category I) were reported in 4 (3%) of 121 patients, with mutations of potential utility (category II) detected in an additional 29 (24%) of 121 patients. CTNNB1 was the gene most frequently mutated, with recurrent mutations in KIT, TSC2, and MAPK pathway genes (BRAF, KRAS, and NRAS) also identified. Mutations in consensus cancer genes (category III) were found in an additional 24 (20%) of 121 tumors. Fewer than half of somatic mutations identified were in genes known to be recurrently mutated in the tumor type tested. Diagnostic germline findings related to patient phenotype were discovered in 15 (10%) of 150 cases: 13 pathogenic or likely pathogenic dominant mutations in adult and pediatric cancer susceptibility genes (including 2 each in TP53, VHL, and BRCA1), 1 recessive liver disorder with hepatocellular carcinoma (TJP2), and 1 renal diagnosis (CLCN5). Incidental findings were reported in 8 (5%) of 150 patients. Most patients harbored germline uncertain variants in cancer genes (98%), pharmacogenetic variants (89%), and recessive carrier mutations (85%). Conclusions and Relevance Tumor and germline WES revealed mutations in a broad spectrum of genes previously implicated in both adult and pediatric cancers. Combined reporting of tumor and germline WES identified diagnostic and/or potentially actionable findings in nearly 40% of newly diagnosed pediatric patients with solid tumors.

Integrating Molecular Testing in the Diagnosis and Management of Children with Thyroid Lesions

Thyroid nodules occur in 1–2% of children, and identifying which nodules are malignant is often challenging. Cytologic evaluation facilitates the diagnosis of thyroid lesions (TLs), but in 10–40% of cases the interpretation is indeterminate. Patients with indeterminate diagnoses are often treated with hemithyroidectomy followed by completion thyroidectomy, if cancer is found in the initial specimen. Exposing patients to multiple surgeries increases costs and morbidity. The American Thyroid Association states that a combination of molecular markers is likely to optimize the management of patients with indeterminate cytology. However, few studies have addressed the molecular alterations present in pediatric TL. Twenty-seven thyroid carcinomas from patients 10 to 19 years of age were tested for alterations common in adult TL, including BRAF V600E mutation, RET fusions, and TERT promoter mutations. Mutation-negative cases were subsequently analyzed with a next-generation sequencing (NGS) mutation panel to search for additional targets. Histologic diagnoses included 12 classic papillary thyroid carcinomas (PTCs), 13 follicular variant PTCs, 1 medullary thyroid carcinoma, and 1 follicular carcinoma. Fourteen cases showed lymph node involvement, and 13 cases demonstrated lymphovascular invasion. The BRAF V600E mutation was detected in 10/27 cases, and RET fusions were detected in 6/27 cases. No TERT promoter mutations were identified in any of the cases. The NGS panel revealed additional RET and CTNNB1 pathogenic missense mutations. Our results demonstrate that molecular abnormalities are common in pediatric TLs and suggest that incorporation of molecular testing will be helpful in optimizing patient management.

Atypical fibroxanthoma and squamous cell carcinoma of the conjunctiva in xeroderma pigmentosum.

Patients with xeroderma pigmentosum (XP) have defective DNA repair and a high predisposition to developing abnormalities and neoplasia in the sun-exposed areas of the skin and mucous membranes. The most common tumors reported in patients with XP are squamous cell carcinomas, basal cell carcinomas, and melanomas. Atypical fibroxanthoma (AFX) is a pleomorphic tumor that arises predominantly in the sun-damaged skin of the head and neck regions of the elderly. We describe a unique case of a 6-year-old African American boy with XP who developed an atypical fibroxanthoma and 2 squamous cell carcinomas in the conjunctiva. The clinical and histopathologic findings of AFX are discussed.

Mucoepidermoid Carcinoma in Children: A Single Institutional Experience

To determine the clinicopathologic and molecular features and outcome of children with mucoepidermoid carcinoma (MEC).

Mucoepidermoid Carcinoma in Children

To determine the clinicopathologic and molecular features and outcome of children with mucoepidermoid carcinoma (MEC).

Lysosomal Acid Lipase Deficiency Unmasked in Two Children With Nonalcoholic Fatty Liver Disease

Lysosomal acid lipase deficiency (LAL-D) is a classic lysosomal storage disorder characterized by accumulation of cholesteryl ester and triglyceride. Although it is associated with progressive liver injury, fibrosis, and end-stage liver disease in children and adolescents, LAL-D frequently presents with nonspecific signs that overlap substantially with other, more common, chronic conditions like nonalcoholic fatty liver disease (NAFLD), metabolic syndrome, and certain inherited dyslipidemias. We present 2 children with NAFLD who achieved clinically significant weight reduction through healthy eating and exercise, but who failed to have the anticipated improvements in aminotransferases and γ-glutamyl transferase. Liver biopsies performed for these “treatment failures” demonstrated significant microvesicular steatosis, prompting consideration of coexisting metabolic diseases. In both patients, lysosomal acid lipase activity was low and LIPA gene testing confirmed LAL-D. We propose that LAL-D should be considered in the differential diagnosis when liver indices in patients with NAFLD fail to improve in the face of appropriate body weight reduction.

Response to Lenvatinib in Children with Papillary Thyroid Carcinoma

BACKGROUND Papillary thyroid carcinoma (PTC) is the most common thyroid malignancy in children and adolescents. Infrequently, children with PTC may present with or develop disease not amenable to surgery or radioactive iodine (RAI), and systemic therapy may be an option. Lenvatinib is an oral tyrosine kinase inhibitor that is approved by the Food and Drug Administration for the treatment of adults with locally recurrent or metastatic, progressive, RAI-refractory well-differentiated thyroid carcinoma. The effect of lenvatinib in children with PTC has not been reported. PATIENT FINDINGS Three children with metastatic PTC not amenable or refractory to RAI who responded to lenvatinib are reported. All of them developed respiratory distress requiring oxygen caused by extensive bilateral metastatic pulmonary disease. The first patient is a 14-year-old female who was initially treated with sorafenib for extensive PTC not amenable to upfront surgery or RAI. She had progressive pulmonary disease after five months, and was subsequently treated with oral lenvatinib (14 mg/m2/day). She was weaned to room air after eight weeks. The second patient is a 15-year-old male who was treated with lenvatinib (14 mg/m2/day) for iodine non-avid diffuse pulmonary disease after initial total thyroidectomy and cervical lymph node dissection. He was weaned off oxygen in six weeks. The third patient is a five-year-old male who was treated with lenvatinib (14 mg/m2/day) for pulmonary disease progression 24 months after treatment with total thyroidectomy, cervical lymph node dissection, and RAI treatment. He was weaned off oxygen one day after starting lenvatinib. Two of the patients required dose adjustments secondary to proteinuria. Otherwise, all patients tolerated lenvatinib well. The first two patients remained clinically stable on lenvatinib 23 months and 11 months after initiation of therapy, respectively, and the third patient transitioned to a tumor-specific targeted therapy after one month. SUMMARY Three pediatric patients are reported with metastatic PTC not amenable or refractory to RAI who achieved a response on lenvatinib. CONCLUSION Lenvatinib therapy is well tolerated and demonstrated clinical activity in children with advanced PTC. Lenvatinib should be considered in children with PTC that is refractory or not amenable to conventional management.

Therapeutic response of metastatic angiomatoid fibrous histiocytoma carrying EWSR1-CREB1 fusion to the interleukin-6 receptor antibody tocilizumab

Angiomatoid fibrous histiocytoma (AFH) is a rare soft tissue tumor that has been associated with EWSR1‐CREB1 gene fusion. Outcome in patients with unresectable distant metastases is generally fatal. Interleukin‐6 (IL‐6) secretion has been described in tumors with EWSR1‐CREB1 fusion, and may promote tumor growth due to autocrine stimulation. Tocilizumab is an IL‐6 receptor antibody that has potential benefit as a targeted therapy in refractory neoplasms with IL‐6 secretion. We describe a child with metastatic AFH with EWSR1‐CREB1 fusion and elevated IL‐6 whose disease progressed during treatment with traditional chemotherapeutic agents, but improved after targeted therapy with tocilizumab.

Abstract B24: Patient-derived tumor xenograft to study cancer stem cells is pediatric sarcomas

Tumor heterogeneity is recognized as an important hallmark in cancer. This characteristic allows cancer cells to evolve in time, to survive radiation and cyotoxic therapies, and to thrive in different organ microenvironments. Therefore, the presence of distinct tumor subpopulations, as mainstay feature of tumor heterogeneity, may play a role in tumor progression and treatment resistance. Patient-derived xenografts (PDXs) have been used to discover the molecular signaling pathways involved in tumorigenesis, metastasis formation and radiation/chemotherapy resistance in different epithelial-adult cancers. Our research interest in pediatric sarcomas is to understand the processes that drive tumor heterogeneity, to isolate and characterize cancer stem cells (CSCs), and to elucidate the mechanisms of chemoresistance using a PDX model. Pediatric patients with suspected or established diagnosis of sarcoma evaluated at Texas Children9s Hospital signed an informed consent to provide tumor samples for subcutaneous implantation of tumor explants in the flank of nonobese diabetic severe combined immunodeficient gamma (NSG) mice. Tumor explants were implanted on the same day of collection when was possible. First and subsequent PDX generations were harvested for tumor propagation, tumor histology, tumor cell karyotyping , RNA/DNA extraction for genetic studies and fluorescent activated cell sorting (FACS) analysis for ALDH, CD133, EphA2 and CD47. Forty-six tumor samples have been collected since June 2013. Half of tumor samples (n=21) corresponded to osteosarcoma patients and 59% of all tumor specimens were obtained at the time of the initial biopsy. Approximately 66% of biopsies have been performed by Interventional Radiology in our institution. All PDXs maintained the same histological characteristics that original tumors, and also presented specific genetic translocations associated with the different sarcoma types. Chemotherapy resistant and very aggressive tumors (metastatic) had short tumor latency and generate tumors in all transplanted mice. We did not observe the formation of metastases (lungs or liver) in any of the transplanted PDX mice. PDXs significantly increased the amount of tumor material necessary to perform cell culture under stem-like cell conditions and to analyze the expression of CSC markers by FACS. These studies would be very difficult to perform directly from patient tumor biopsies. All tumors harvested (n=15) contained 2-9% of ALDH Hi or stem-like cells. None of the PDXs analyzed expressed CD133 but synovial sarcoma. We found that >90% of sarcoma cells expressed CD47 and EphA2 at different intensity levels. Whole genome sequencing of different tumor cell subpopulations and limiting dilution experiments to validate the presence of CSCs are ongoing. Core needle biopsies would represent the most common source to obtain tumor specimens before treatment in pediatric sarcomas. The development of PDXs allows the propagation of tumor cell subpopulations, including CSCs, without the biological changes impose by cell culture. PDXs resemble the original tumor and constitute a powerful tool to study tumor heterogeneity. We plan to perform orthotopic transplantation and tail vein injection of PDX cells in an attempt to develop a PDX model of metastatic disease in pediatric sarcomas. Citation Format: Nino Rainusso, Jose Hernandez, Rex Marco, Sanjeev Vasudevan, Norma Quintanilla, John Hicks, Wendy Allen-Rhoades, Matteo Trucco, Jason Yustein. Patient-derived tumor xenograft to study cancer stem cells is pediatric sarcomas. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr B24.

Perianal Papillomatous Crohn's Disease in a Male Adolescent

16-year-old male patient presented with hemaAtochezia. He had a history of a perianal abscess treated with antibiotics 2 years prior. He had lost 15 pounds in the previous 3 months and had 1 month of daily epigastric pain and loose stools. On physical examination, perianal skin tags and multiple nontender papillomatous plaques were noted (Figure A). Laboratory analysis revealed hemoglobin 11.5 g/dL, platelets 371,000/mL, erythrocyte sedimentation rate 52 mm/h, and C-reactive protein 2.5 mg/dL (normal, <1.0). Endoscopic evaluation showed aphthous ulceration of the esophagus and duodenum, gastric edema, mucopurulent debris in the terminal ileum, and diffuse, scattered aphthous ulceration, erythema, and edema of the entire colon, most prevalent in the rectum. Although mucosal edema, erythema, and ulceration were noted on endoscopic retroflexed view of the rectal vault, extension into the rectum of the external papillomatous lesions was noticeably absent (Figure B). The dermatology service was consulted, and punch biopsy of the lesion revealed dermal noncaseating granulomatous inflammation consistent with Crohn’s disease (CD) (Figure C). Endoscopic biopsies confirmed the diagnosis, demonstrating lymphoplasmacytic gastritis as well as chronic and acute duodenitis, ileitis, and colitis. A granuloma was noted on a rectosigmoid colonic biopsy. The patient had marked improvement of his perianal lesions and constitutional symptoms 2 months into infliximab therapy (Figure D). Cutaneous CD can manifest in multiple ways. Papillomatous lesions are one of the least common symptoms. Such presentation is even rarer in children, with a handful of case reports in the literature, all in female patients. This case reports CD complicated by perianal papillomatous vegetation in a male adolescent. These lesions can be mistaken for condylomata acuminata or pseudoverrucous papules and nodules. In addition, secondary skin lesions from occluding undergarments, stool seepage, and infections can manifest as oval reddish-purple granulomatous nodules known as granuloma gluteale infantum/adultorum or granuloma intertriginosum infantum. Clinical and histologic description allows for accurate diagnosis. Treatment with an anti–tumor necrosis factor-a antibody may significantly improve such perianal CD.