Jinhua Wu

PURPOSE: NTRK gene fusions are oncogenic drivers for a variety of adult and pediatric tumors, making them a target for tumor-agnostic precision medicine. Tropomyosin receptor kinase (TRK) inhibitors are approved by the US Food and Drug Administration for cancers driven by TRK fusions. However, NTRK genes can fuse with many different partner genes, leading to diverse TRK fusion proteins, highlighting the importance of identifying the specific fusion partner with optimal pan-cancer diagnostics. This analysis aims to provide an updated descriptive compendium of NTRK gene fusions. METHODS: NTRK gene fusions were identified via literature searches (PubMed), a search of a clinical trials database (larotrectinib), and searches of two genomic databases (Memorial Sloan Kettering and Children's Hospital of Philadelphia). RESULTS: In total, 358 distinct NTRK gene fusion-tumor pairings were identified across 25 tumor types. Primary CNS tumors were observed to harbor 86 distinct NTRK gene fusions, followed by sarcomas (n = 73). Overall, 229 different fusion partners were identified across tumor types (regardless of NTRK gene). Twenty-three fusion partners were found to fuse with >1 NTRK gene across tumor types, while 183 fusion partners were associated with only a single NTRK gene in one tumor type. ETV6 :: NTRK3 was found in the highest number of different tumor types. CONCLUSION: This analysis illustrates the diversity of NTRK gene fusion partners across various tumor types and highlights the importance of selecting a pan-tumor fusion-partner agnostic test that can identify both known and novel fusion partners to identify patients who may benefit from treatment with TRK inhibitors.

TP53 alterations are common in certain pediatric cancers, making identification of putative germline variants through tumor genomic profiling crucial for patient management. We analyzed TP53 alterations in 3123 tumors from 2788 pediatric patients sequenced using tumor-only or tumor-normal paired panels. Germline confirmatory testing was performed when indicated. Somatic and germline variants were classified following published guidelines. In 248 tumors from 222 patients, 284 Tier 1/2 TP53 sequence and small copy number variants were detected. Following germline classification, 73.9% of 142 unique variants were pathogenic/likely pathogenic (P/LP). Confirmatory testing on 118 patients revealed germline TP53 variants in 28 patients (23 P/LP and 5 uncertain significance), suggesting a minimum Li-Fraumeni syndrome (LFS) incidence of 0.8% (23/2788) in this cohort, 10.4% (23/222) in patients with TP53 variant-carrying tumors, and 19.5% (23/118) with available normal samples. About 25% (7/28) of patients with germline TP53 variants did not meet LFS diagnostic/testing criteria while 20.9% (28/134) with confirmed or inferred somatic origins did. TP53 biallelic inactivation occurred in 75% of germline carrier tumors and was also prevalent in other groups, causing an elevated tumor-observed variant allelic fraction (VAF). However, somatic evidence including low VAF correctly identified only 27.8% (25/90) of patients with confirmed somatic TP53 variants. The high incidence and variable phenotype of LFS in this cohort highlights the importance of assessing germline status of TP53 variants identified in all pediatric tumors. Without clear somatic evidence, distinguishing somatic from germline origins is challenging. Classifying germline and somatic variants should follow appropriate guidelines.

Fibroblast growth factor receptors (FGFRs) are a family of receptor tyrosine kinases expressed on the cell membrane that play crucial roles in cellular lineage commitment, differentiation, proliferation, and apoptosis. Deregulated FGFR signaling is observed in a subset of tumors across various histologies, making FGFRs ideal therapeutic targets. We sought to determine the genetic landscape of FGFR-family variations in a cohort of pediatric and young adult patients with solid tumors. The CHOP Comprehensive Solid Tumor Panel was performed on 1,420 patients. The panel covers 238 cancer genes and screens for single nucleotide variants (SNVs), indels, copy number alterations, and 117 fusion gene partners interrogating over 700 exons for known and novel fusions. Identified variants were categorized and reported according to the AMP/ASCO/CAP guidelines. Fifty-six patients (4.1%), including 47 children and 9 young adults, were found to carry at least one FGFR alteration in their tumors. CNS tumors accounted for most of the cases (51 total, 87.9%), with pilomyxoid astrocytoma/pilocytic astrocytoma and dysembryoplastic neuroepithelial tumor the most common (13 and 12 patients, respectively). Non-CNS solid tumors included rhabdomyosarcoma (4 patients), neuroblastoma/ganglioneuroblastoma (2), and follicular thyroid carcinoma (1). FGFR somatic alterations were found in 56 tumors including 41 SNVs and small indels, 6 internal tandem duplications (ITDs), and 15 fusions genes. The most common SNVs observed were hotspot mutations p.K656E and p.N546K of FGFR1. Sequence alterations in FGFR1 contained 35 SNVs and small indels, mostly gain of function mutations located in the kinase domain, and 6 kinase domain ITDs. One SNV was identified in FGFR2 in the immunoglobulin domain. Two SNVs were reported in FGFR3, both of which were in the fibroblast growth factor receptor family domain, and 3 SNVs were identified in FGFR4, all occurring at the p.V550 codon located on the kinase domain. Companion mutations in non-FGFR genes were detected in 27 tumors, predominantly involving RAS signaling pathway genes including NF1 (14 variants), PIK3CA (8), PTPN11 (6) and PIK3R1 (4). Among fusion variants, FGFR1-TACC1 fusions were found in 5 patients, mostly in pediatric patients. One FGFR3-TACC3 fusion was identified in one young adult patient. Seven pediatric patients tested positive for FGFR2 fusions; all with different 3’ partners. The detection of an FGFR alteration defined or changed the histologic diagnosis for 22 patients. Our results reveal that FGFR alterations account for 4.1% (56/1420) of the patients with solid tumors tested in our laboratory. The majority of the FGFR-positive tumors are low-grade CNS tumors. Further, the identification of FGFR alterations can significantly improve the tumor diagnosis and provide genomic evidence for potential targeted treatment with FGFR inhibitors. Citation Format: Jinhua Wu, Jeffrey Schubert, Feng Xu, Ariel Long, Maha Patel, Netta Golenberg, Weixuan Fu, Kajia Cao, Jiani Chen, Elizabeth H. Denenberg, Elizabeth A. Fanning, Rochelle Bagatell, Theodore W. Laetsch, Adam Resnick, Mariarita Santi, Phillip Jay B. Storm, Minjie Luo, Lea F. Surrey, Yiming Zhong, Marilyn M. Li. The spectrum of FGFR mutations in pediatric and young adult solid tumor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5268.

Yiming Zhong, Ph.D, Feng Xu, Ph.D, Jinhua Wu, Ph.D, Jeffrey Schubert, Ph.D, and Marilyn M. Li, M.D. Ann Lab Med 2021;41:25-43. https://doi.org/10.3343/alm.2021.41.1.25

PALB2 (partner and localizer of BRCA2) gene encodes a protein that colocalizes with BRCA2 in nuclear foci and likely permits the stable intranuclear localization and accumulation of BRCA2. PALB2 plays a critical role in maintaining genome integrity through its role in the Fanconi anemia and homologous recombination DNA repair pathways. It has a known loss-of-function disease mechanism. Biallelic PALB2 pathogenic variants have been described in autosomal recessive Fanconi anemia. Heterozygous pathogenic variants in PALB2 are associated with increased risk for female and male breast cancer and pancreatic cancer (Science 324: 217; Cancer Res 71: 2222–2229; N Engl J Med 371: 497–506). Heterozygous germline PALB2 mutations have also been observed in patients with medulloblastoma (Lancet Oncol 19: 785–798). However, PALB2-related cancer predisposition to high-grade gliomas has not been reported. Here we report a germline PALB2 pathogenic variant (c.509_510delGA, p.Arg170Ilefs*14, NM_024675.3) found in a pediatric patient with high-grade glioma. This variant was first identified by tumor sequencing using the Children's Hospital of Philadelphia (CHOP) Comprehensive Solid Tumor Panel and then confirmed to be a germline change using the CHOP Comprehensive Hereditary Cancer Panel on DNA from a blood sample of this patient. Parental studies showed that this variant was paternally inherited. Further studies are needed to illustrate if pathogenic variants in PALB2 convey increased risk to developing brain tumor. This case also highlights the potential of identifying germline mutation through tumor sequencing.

Inhibition of ATM kinase upregulates levels of cell death induced by cannabidiol and -irradiation in human glioblastoma cells Inhibition of ATM kinase upregulates levels of cell death induced by cannabidiol and γ-irradiation in human glioblastoma cells

Even though the first ultraviolet microbeam was described by S. Tschachotin back in 1912, the development of sophisticated micro-irradiation facilities only began to flourish in the late 1980s. In this article, we highlight significant microbeam experiments, describe the latest microbeam irradiator configurations and critical discoveries made by using the microbeam apparatus. Modern radiological microbeams facilities are capable of producing a beam size of a few micrometers, or even tens of nanometers in size, and can deposit radiation with high precision within a cellular target. In the past three decades, a variety of microbeams has been developed to deliver a range of radiations including charged particles, X-rays, and electrons. Despite the original intention for their development to measure the effects of a single radiation track, the ability to target radiation with microbeams at sub-cellular targets has been extensively used to investigate radiation-induced biological responses within cells. Studies conducted using microbeams to target specific cells in a tissue have elucidated bystander responses, and further studies have shown reactive oxygen species (ROS) and reactive nitrogen species (RNS) play critical roles in the process. The radiation-induced abscopal effect, which has a profound impact on cancer radiotherapy, further reaffirmed the importance of bystander effects. Finally, by targeting sub-cellular compartments with a microbeam, we have reported cytoplasmic-specific biological responses. Despite the common dogma that nuclear DNA is the primary target for radiation-induced cell death and carcinogenesis, studies conducted using microbeam suggested that targeted cytoplasmic irradiation induces mitochondrial dysfunction, cellular stress, and genomic instability. A more recent development in microbeam technology includes application of mouse models to visualize in vivo DNA double-strand breaks. Microbeams are making important contributions towards our understanding of radiation responses in cells and tissue models.