Partner Genes Research Articles

LncRNA-mRNA integrated profiling analysis in response to white spot syndrome virus in hepatopancreas in Penaeus japonicus

Penaeus japonicas is an important shrimp species, which is exposed to stressors including a variety of epidemic diseases. To date, little is known about the mechanisms involved in the response to white spot syndrome virus (WSSV) mediated by long non-coding RNAs (lncRNAs). A total of 6544 putative lncRNAs were identified in the hepatopancreas in P. japonicas, which provides a useful lncRNA reference resource for use in future studies. In addition, a total of 444 differentially expressed mRNAs and 457 differentially expressed lncRNAs were identified at 6, 12, and 24 h after WSSV infection in the hepatopancreas of P. japonicas. Functional enrichment analysis showed that the differentially expressed mRNAs were enriched in terms related to immune response and viral infectivity such as defense response, aminopeptidase activity, whereas the differentially expressed lncRNA partner genes were enriched in ubiquitin-dependent protein catabolic process, lipoprotein metabolic process, and antigen processing and presentation. Moreover, several lncRNAs were induced by WSSV infection, indicating these lncRNAs might participate in regulating many immune processes referring to their partner genes. Co-expression analysis of the lncRNAs and their partner genes identified some high lncRNA-mRNA correlations. These results suggest that WSSV stimulates the immune response in the hepatopancreas potentially through an important coding and non-coding gene network, thereby providing valuable information regarding non-coding responses to WSSV in Penaeus species.

PAX5 fusion genes are frequent in poor risk childhood acute lymphoblastic leukaemia and can be targeted with BIBF1120.

SummaryBackgroundDespite intensive risk-based treatment protocols, 15% of paediatric patients with B-Cell Precursor Acute Lymphoblastic Leukaemia (BCP-ALL) experience relapse. There is urgent need of novel strategies to target poor prognosis subgroups, like PAX5 translocated.MethodsWe considered 289 childhood BCP-ALL cases consecutively enrolled in Italy in the AIEOP-BFM ALL2000/R2006 protocols and we performed extensive molecular profiling, integrating gene expression, copy number analyses and fusion genes discovery by target-capture NGS. We developed preclinical strategies to target PAX5 fusion genes.FindingsWe identified 135 cases without recurrent genetic rearrangements. Among them, 59 patients (43·7%) had a Ph-like signature; the remaining cases were identified as ERG-related (26%), High-Hyperdiploid-like (17%), ETV6::RUNX1-like (8·9%), MEF2D-rearranged (2·2%) or KMT2A-like (1·5%). A poor prognosis was associated with the Ph-like signature, independently from other high-risk features.Interestingly, PAX5 was altered in 54·4% of Ph-like compared to 16·2% of non-Ph-like cases, with 7 patients carrying PAX5 fusions (PAX5t), involving either novel (ALDH18A1, IKZF1, CDH13) or known (FBRSL1, AUTS2, DACH2) partner genes. PAX5t cases have a specific driver activity signature, extending to multiple pathways including LCK hyperactivation. Among FDA-approved drugs and inhibitors, we selected Dasatinib, Bosutinib and Foretinib, in addition to Nintedanib, known to be LCK ligands. We demonstrated the efficacy of the LCK-inhibitor BIBF1120/Nintedanib, as single agent or in combination with conventional chemotherapy, both ex vivo and in patient-derived xenograft model, showing a synergistic effect with dexamethasone.InterpretationThis study provides new insights in high-risk Ph-like leukaemia and identifies a potential therapy for targeting PAX5-fusion poor risk group.FundingRicerca Finalizzata-Giovani Ricercatori (Italian Ministry of Health), AIRC, Transcall, Fondazione Cariparo.

GLI1 -Rearranged Enteric Tumor : Expanding the Spectrum of Gastrointestinal Neoplasms With GLI1 Gene Fusions.

GLI1 encodes a transcription factor that targets cell cycle regulators affecting stem cell proliferation. GLI1 gene fusions were initially described in pericytomas with a t[7;12] translocation and more recently in gastric plexiform fibromyxomas and gastroblastomas. This study describes the clinicopathologic, immunohistochemical, and molecular features of three intestinal-based neoplasms harboring GLI1 gene fusions. We studied three unique mesenchymal small bowel tumors. Paraffin embedded tumor tissues from these cases and 62 additional tumor samples that included a plexiform fibromyxoma were sequenced using a targeted RNAseq method to detect fusion events. The study patients included two women and one man who were 52, 80, and 22 years of age at the time of diagnosis. The tumors involved the submucosa and muscularis propria of the duodenum, jejunum, and ileum. All 3 tumors contained a proliferation of monotonous oval or spindle cells with scattered, somewhat dilated vessels. Two cases showed epithelioid structures such as glands, tubules, or nests. Immunohistochemical analysis revealed cytokeratin expression in the epithelioid components of both tumors displaying these features, and variable numbers of mesenchymal cells. Diffuse CD56 positivity was seen in the mesenchymal component of 2 tumors and desmin and smooth muscle actin staining in the other tumor. Immunostains for S-100 protein, DOG-1, and CD117 were negative in all cases. GLI1 fusions with different partner genes were detected in all tumors, and in the plexiform fibromyxoma, used as a control. Validation by fluorescence in situ hybridization was performed. None of the tumors have recurred or metastasize after surgery. We describe novel GLI1 fusions in 3 mesenchymal neoplasms of the small intestine, including 2 with biphenotypic features. Thus far, all cases have pursued indolent clinical courses. We propose the term " GLI1 -rearranged enteric tumor" to encompass this group of unique neoplasms of the small intestine that harbor GLI1 gene fusions and expand the spectrum of gastrointestinal neoplasms with these alterations.

Single-cell diploid Hi-C reveals the role of spatial aggregations in complex rearrangements and KMT2A fusions in leukemia

BackgroundSimple translocations and complex rearrangements are formed through illegitimate ligations of double-strand breaks of fusion partners and lead to generation of oncogenic fusion genes that affect cellular function. The contact first hypothesis states that fusion partners tend to colocalize prior to fusion in normal cells. Here we test this hypothesis at the single-cell level and explore the underlying mechanism.ResultsBy analyzing published single-cell diploid Hi-C datasets, we find partner genes fused in leukemia exhibit smaller spatial distances than those fused in solid tumor and control gene pairs. Intriguingly, multiple partners tend to colocalize with KMT2A in the same cell. 3D genome architecture has little association with lineage decision of KMT2A fusion types in leukemia. Besides simple translocations, complex rearrangement-related KMT2A fusion genes (CRGs) also show closer proximity and belong to a genome-wide mutual proximity network. We find CRGs are co-expressed, co-localized, and enriched in the targets of the transcriptional factor RUNX1, suggesting they may be involved in RUNX1-mediated transcription factories. Knockdown of RUNX1 leads to significantly fewer contacts among CRGs. We also find CRGs are enriched in active transcriptional regions and loop anchors, and exhibit high levels of TOP2-mediated DNA breakages. Inhibition of transcription leads to reduced DNA breakages of CRGs.ConclusionsOur results demonstrate KMT2A partners and CRGs may form dynamic and multipartite spatial clusters in individual cells that may be involved in RUNX1-mediated transcription factories, wherein massive DNA damages and illegitimate ligations of genes may occur, leading to complex rearrangements and KMT2A fusions in leukemia.

Rapid and highly sensitive approach for multiplexed somatic fusion detection

AbstractSomatic gene translocations are key to making an accurate diagnosis in many cancers including many pediatric sarcomas. Currently available molecular diagnostic approaches to identifying somatic pathognomonic translocations have limitations such as minimal multiplexing, high cost, complex computational requirements, or slow turnaround times. We sought to develop a new fusion-detection assay optimized to mitigate these challenges. To accomplish this goal, we developed a highly sensitive multiplexed digital PCR-based approach that can identify the gene partners of multiple somatic fusion transcripts. This assay was validated for specificity with cell lines and synthetized DNA fragments. Assay sensitivity was optimized using a tiered amplification approach for fusion detection from low input and/or degraded RNA. The assay was then tested for the potential application of fusion detection from FFPE tissue and liquid biopsy samples. We found that this multiplexed PCR approach was able to accurately identify the presence of seven different targeted fusion transcripts with a turnaround time of 1 to 2 days. The addition of a tiered amplification step allowed the detection of targeted fusions from as little as 1 pg of RNA input. We also identified fusions from as little as two unstained slides of FFPE tumor biopsy tissue, from circulating tumor cells collected from tumor-bearing mice, and from liquid biopsy samples from patients with known fusion-positive cancers. We also demonstrated that the assay could be easily adapted for additional fusion targets. In summary, this novel assay detects multiple somatic fusion partners in biologic samples with low tumor content and low-quality RNA in less than two days. The assay is inexpensive and could be applied to surgical and liquid biopsies, particularly in places with inadequate resources for more expensive and expertise-dependent assays such as next-generation sequencing.

NUT Carcinoma-An Underdiagnosed Malignancy.

NUT carcinoma (NC) is a rare and highly aggressive malignancy with a dismal prognosis and a median survival of 6–9 months only. Although very few cases of NC are reported each year, the true prevalence is estimated to be much higher, with NC potentially widely underdiagnosed due to the lack of awareness. NC primarily occurs in midline structures including thorax, head, and neck; however, other sites such as pancreas and kidney are also affected, albeit at lower frequencies. NC is characterized by a single translocation involving the NUTM1 (NUT midline carcinoma family member 1) gene and different partner genes. The resulting fusion proteins initiate tumorigenesis through a mechanism involving BET (bromo-domain and extra-terminal motif) proteins such as Bromodomain-containing protein 4 (BRD4) and inordinate acetylation of chromatin, leading to the dysregulation of growth and differentiation genes. While no clinical characteristics are specific for NC, some histologic features can be indicative; therefore, patients with these tumor characteristics should be routinely tested for NUTM1. The diagnosis of NC using immunohistochemistry with a highly specific antibody is straightforward. There are currently no standard-of-care treatment options for patients with NC. However, novel therapies specifically addressing the unique tumorigenic mechanism are under investigation, including BET inhibitors. This review aims to raise awareness of this underdiagnosed cancer entity and provide all patients the opportunity to be properly diagnosed and referred to a clinical study.

074 ALK-positive desmoplastic Spitz naevus in a patient with corresponding ALK-positive anaplastic large cell lymphoma

Aim:Spitz naevus is a benign melanocytic tumour with distinct histopathological features of spindle-shaped melanocytes. An understanding of genetic aberrations can help to characterise the different subsets of Spitzoid neoplasms. Anaplastic lymphoma kinase (ALK) is a tyrosine kinase receptor that was first described in the pathogenesis of anaplastic large cell lymphoma (ALCL). ALK mutations have been reported in up to 10% of Spitz tumours.This study describes the rare occurrence of an ALK mutation in two different diseases in the same patient – ALCL and Spitz naevus. Method:Presentation, management, and clinical outcomes were retrospectively reviewed for a patient with ALK-positive ALCL and ALK-positive Spitz naevus at a tertiary paediatric hospital. Result:The patient was a 14-year-old female with a 12-month history of a red-brown plaque on the right shoulder.This was on a background stage III ALCL treated with chemotherapy and stem cell transplantation. She underwent surgical excision of the skin lesion. Histopathological examination showed an ALK-positive desmoplastic Spitz naevus. A bone marrow aspirate and trephine (BMAT) of the right axillary lymph node at the time of ALCL diagnosis also revealed an ALK mutation. She is currently in remission 2 years post stem cell transplantation. Conclusion:Our case reports on a rare coincidence of a patient having the same mutation for two distinct disease processes. There has been one similarly reported case in the literature in a patient with ALK-positive cutaneous ALCL and Spitz naevi with the same fusion partner gene (TPM3). Whilst this occurrence is not fully understood, further studies may identify whether this is an independent coincidence in two different cell lineages, or a fragile site similarity which may predispose to the same translocation event.

Approaches in Gene Coexpression Analysis in Eukaryotes.

Simple SummaryGenes whose expression levels rise and fall similarly in a large set of samples, may be considered coexpressed. Gene coexpression analysis refers to the en masse discovery of coexpressed genes from a large variety of transcriptomic experiments. The type of biological networks that studies gene coexpression, known as Gene Coexpression Networks, consist of an undirected graph depicting genes and their coexpression relationships. Coexpressed genes are clustered in smaller subnetworks, the predominant biological roles of which can be determined through enrichment analysis. By studying well-annotated gene partners, the attribution of new roles to genes of unknown function or assumption for participation in common metabolic pathways can be achieved, through a guilt-by-association approach. In this review, we present key issues in gene coexpression analysis, as well as the most popular tools that perform it.Gene coexpression analysis constitutes a widely used practice for gene partner identification and gene function prediction, consisting of many intricate procedures. The analysis begins with the collection of primary transcriptomic data and their preprocessing, continues with the calculation of the similarity between genes based on their expression values in the selected sample dataset and results in the construction and visualisation of a gene coexpression network (GCN) and its evaluation using biological term enrichment analysis. As gene coexpression analysis has been studied extensively, we present most parts of the methodology in a clear manner and the reasoning behind the selection of some of the techniques. In this review, we offer a comprehensive and comprehensible account of the steps required for performing a complete gene coexpression analysis in eukaryotic organisms. We comment on the use of RNA-Seq vs. microarrays, as well as the best practices for GCN construction. Furthermore, we recount the most popular webtools and standalone applications performing gene coexpression analysis, with details on their methods, features and outputs.

The lncRNA PRINS-miRNA-mRNA Axis Gene Expression Profile as a Circulating Biomarker Panel in Psoriasis.

BackgroundThe interaction between genes and the environment in psoriasis is firmly coupled by epigenetic modification. Epigenetic modifications are inherited variations in gene expression devoid of DNA sequence alterations. Non-coding RNAs are regarded as one of the epigenetic modifications that lead eventually to enduring heritable variations in gene expression. In the present study, we chose the lncRNA, Psoriasis-susceptibility-Related RNA Gene Induced by Stress (PRINS) known to have a regulatory role in psoriasis and deduced its axis of lncRNA-miRNA-mRNA through an in silico data analysis. We aimed to assess the expression levels of this lncRNA-miRNA-mRNA in patients with psoriasis to elucidate their possible roles in psoriasis management.MethodsWe investigated the lncRNA-PRINS and its target microRNAs (miRNA124-3p, miRNA203a-5p, miRNA129-5p, miRNA146a-5p, miRNA9-5p) and partner genes (NPM, G1P3) expression levels in the plasma of 120 patients with psoriasis compared to 120 healthy volunteers using quantitative real-time polymerase chain reaction and correlated the results with the patients’ clinicopathological data. Finally, we performed a function, disease, and pathway enrichment analysis for the LncRNA-miRNA-mRNA axis under study.ResultsThe lncRNA PRINS, G1P3, and NPM genes showed significantly under-expressed levels while all miRNAs included in the study showed significant over-expression in patients with psoriasis relative to controls. The lncRNA PRINS, G1P3, and NPM genes showed a significant direct correlation with each other and inverse significant correlations with all miRNAs under study. All the study biomarkers showed significant results for discriminating between patients with psoriasis and controls using a receiver operating curve analysis with sensitivity over 90% except for PRINS, which was 74.2%. The G1P3 gene showed a direct significant correlation with body mass index in patients with psoriasis (p = 0.009) and an inverse significant correlation with age (p = 0.034). The NPM gene showed a significant correlation with body mass index in patients with psoriasis (p = 0.002).ConclusionsBased on our results, we suggest that restoring the altered PRINS-miRNA-mRNA axis gene expression levels might represent a tool to prevent psoriasis worsening, along with standard therapy. Thus, on the clinical practice level, the PRINS-miRNA-mRNA axis expression profile can be utilized in designing specific targeted therapy aimed at applying a personalized medicine approach among patients with psoriasis.

PB1799: KMT2A-REARRANGED PEDIATRIC ACUTE MYELOID LEUKEMIA

Background: Histone-lysine N-methyltransferase 2A gene rearrangements (KMT2A-r) are common genetic events in acute leukemia. They demonstrate great molecular heterogeneity with more than 90 partner genes and multiple KMT2A breakpoints involved. These characteristics challenge both the initial molecular diagnostics and MRD monitoring in ALs with KMT2A-r. On the other hand, the profile of accompanying mutations is relatively quiet in this type of leukemia due to strong oncogenic potential of KMT2A rearrangement itself. Aims: Our recent study aims to characterize KMT2A-rearranged acute myeloid leukemia in children in Russian Federation both in terms of partner genes involved and in accompanying mutations study. Methods: A cohort of 313 KMT2A-positive AMLs in children aged 4 days to 18 y.o. (median 2 y.o.; 162 boys and 151 girls) was analyzed using G-banded karyotyping and FISH together with real-time and nested RT-PCR. Transcripts were Sanger sequenced. Patients with rare or unknown translocations were subjected to long-distance inverse PCR (LDI-PCR), anchored multiplex PCR (ArcherDX, CO, USA) or custom targeted KMT2A enrichment panel (Illumina, CA, USA; Meyer et al., 2019). Additional events were characterized by fragment analysis, Sanger sequencing and NGS (Qiagen Myeloid Neoplasms panel, Hilden, Germany) in 113 cases included in Russian AML registration study and AML-MRD-2018 local treatment protocol. Results: The incidence of KMT2A-rs was the highest in children under 2 y.o. inclusive (n=120, 38%) and goes down with age. A total of 26 different KMT2A-rs were found in 296 patients (94.6%). The prevailing translocations were as follows: t(9;11)(p21;q23.3)/KMT2A-MLLT3 (n=129, 41.21%), t(10;11)(p12;q23.3)/KMT2A-MLL10 (n=71, 22.68%), t(11;19)(q23.3;p13.1)/KMT2A-ELL (n=24, 7.67%), t(6;11)(q27;q23.3)/KMT2A-AFDN (n=15, 4.79%), t(11;19)(q23.3;p13.3)/KMT2A-MLLT1 (n=13, 4.15%). Other translocation partners were much rarer, for 15 partner genes only single cases were found. Three novel KMT2A-rs were identified, namely t(11;16)(q23;q23.3)/KMT2A-USP10, t(X;11)(q22.1;q23.3)/KMT2A-BTK and t(X;11)(q26.3;q23.3)/KMT2A-CT45A. KMT2A breakpoint location was analyzed in 274 patients (87,54%). It was found in introns 7 to 12, which is narrower than in ALL, and was most often situated in introns 9 (n=131, 47.81%) and 10 (89 cases, 32.48%) as numbered by Meyer et al. [Meyer et al., 2006]. Intron 9 was also predominant in rearrangements with all but MLLT1 frequent translocation partners and most rare translocation partners. KMT2A break outside the main cluster was found in intron 7 on 4 cases of KMT2A-MLL10. The achieved data on KMT2A translocation partners and breakpoint locations was used for subsequent patient-specific MRD monitoring. The molecular profile of myeloid neoplasms-associated gene was studied in 113 samples. One to 9 additional pathogenic somatic events (median 4) with variant allele frequency above 1% were found in the studied cohort with most of them being subclonal (VAF<10%). Only 62 samples contained additional mutations with VAF greater above 10%. Among them NRAS (n=11 of 62, 17.74%), KRAS (n= 8, 12,90%) and PTEN11 (4, 6,45%) were prevalent. Accompanying mutations did not exhibit translocation partner preference. Summary/Conclusion: Molecular profiling of KMT2A-rearranged AML in Russian Federation reveals the highest incidence of KMT2A-MLLT3 fusion gene and KMT2A intron 9 involvement. Our data also confirms the relatively quiet landscape of accompanying mutations in KMT2A-rearranged AML with RAS pathway most frequently involved.

P343: GENOMIC CHARACTERISATION OF B-OTHER ALL IN UKALL2003 PATIENTS BY NEXT GENERATION SEQUENCING

Background: Incorporating genetics into risk stratification for the treatment of childhood acute lymphoblastic leukaemia (ALL) has contributed to increased survival rates. About 30% of patients (B-other-ALL) harbour none of the known major chromosomal changes. Recently, we estimated that up to two thirds of B-other-ALL can be classified into novel subtypes using FISH and MLPA alone. We showed that ABL-class fusions and ERG deletions were linked to prognosis (Schwab et al, BJHaem, 2022). Aims: To characterise B-other-ALL in the UKALL2003 trial using next generation sequencing (NGS), to evaluate the added benefits of these techniques and validate the prognostic significance of B-other-ALL subtypes. Methods: B-other-ALL patients were tested using whole genome sequencing (WGS) (n=158) and bespoke targeted NGS (t-NGS) for the detection of abnormalities in 64 genes commonly mutated or rearranged in B-ALL (n=180), 42 patients had both WGS and t-NGS. Data were integrated with results from FISH, MLPA and cytogenetics. Results: A representative cohort of 347 B-other-ALL was classified into subgroups, including six with ≥20 cases (Figure). Among patients tested by WGS, 92% (n=146/158) were classified compared to 75% (n=104/138) tested by t-NGS. NGS-based approaches allowed for the detection of PAX5 P80R, IKZF1 N169Y and ZEB2 H1038R and fusion partner genes. Identification of DUX4-r was not possible using t-NGS. PAX5alt was the most frequently observed (n=90), including dic(9;20) (n=27), dic(9;12) (n=11), PAX5 fusions (n=21), PAX5 mutation (n=11) and PAX5-ITD (n=12). Additionally, 11 patients were observed with PAX5 P80R mutations, classified as a separate subgroup due to the associated gene expression signature. PAX5alt had an outcome similar to B-other-ALL overall (overall survival (OS) at 10yrs 82% vs 86%). There was no significant difference in outcome between PAX5 abnormalities, although an association with age was observed: dic(9;20) was more commonly observed in children aged 1-4 (p< 0.001), while both dic(9;12) and PAX5 P80R were seen in older children aged 10-15 years (p=0.005). One subgroup with DUX4 rearrangements and/or ERG deletions (DUX4-r/ERG-d) (n=78) had DUX4 involvement confirmed by WGS in 57 patients. In the remaining 21 patients ERG-d were identified by t-NGS and/or MLPA. ERG-d is exclusive to DUX4-r patients, thus can be used as a surrogate marker. In line with other studies, patients with DUX4-r had a lower relapse rate (RR) (5%) and improved OS (96%) compared to other subgroups. Patients classified as ETV6-RUNX1-like (n=21) were characterised by multiples abnormalities of ETV6, including rearrangements (n=17) and/or deletion (n=12) as well as rearrangements (n=7) and/or deletions (n=6) of IKZF1. An ETV6-RUNX1-like gene expression signature was confirmed by RNA-Seq (n=6). No relapses or deaths were reported within this subgroup. As previously reported, ABL-class fusions (n=25) were associated with an inferior outcome compared to other subgroups (RR=63% and OS=50%). Patients with rearrangements of CRLF2 (n=52) and ZNF384 (n=37) had similar outcomes to the cohort overall. Image:Summary/Conclusion: A combination of techniques has classified a representative cohort of B-other-ALL patients from UKALL2003 into clinically relevant subgroups. The identification of DUX4-r, subgroup defining mutations and fusion partner genes demonstrates the value of NGS-based approaches. We have confirmed the good and poor prognostic associations of DUX4-r and ABL-class fusions, respectively, and identified ETV6-RUNX1-like subgroup to have a good prognosis.

Abstract 2014: Mosaic BRAF fusions are a recurrent cause of multiple congenital melanocytic naevi

Abstract Congenital melanocytic naevi (CMN) are moles present at birth, and when multiple or very extensive can involve other organ systems as well as predisposing to melanoma. Some CMN patients develop a highly proliferative multinodular phenotype leading to chronic intense pruritus, and the causes for that specific phenotype progression are poorly understood. Genotypically CMN are mosaic single gene disorders. Thus far only two recurrent variants have been described as clearly causative: NRAS missense variants affecting codon 61 accounting for 68% of cases, and BRAF missense variants affecting codon 600 accounting for 7% of cases. Small numbers of cases of CMN have been reported to carry gene-fusions, and in 2 cases in the world literature (one BRAF-fusion and one RAF1-fusion) these have been shown to be clonal and therefore likely causal. We sought to address the issue of causation of the remaining 25%. From an initial large cohort study, skin biopsies from 19 patients were shown to be double wildtype for NRAS/BRAF and had sufficient tissue for further study after deep whole exome sequencing. These 19 samples then underwent transcriptome-wide paired-end RNA sequencing with bioinformatics analysis (STAR-Fusion v1.6 and Fusion Inspector v2.3) for gene fusion transcripts. 11/19 patients were found to have BRAFgene fusions, of which 7 had the multinodular proliferative phenotype. Fusions were confirmed on Sanger sequencing of the cDNA across the fusion junction, specifically demonstrated in 8 children from more than one separate skin lesion, confirming clonality. In the fusions identified, BRAF was fused to 11 different partner genes (GOLGA4, QKI, STRN3, AGAP3, MKRN2, PHIP, LCA5, EEA1, AKAP9, SEC31A, MIER3). This resulted in loss of the 5’ regulatory domain of BRAF but preservation of the kinase domain, such that expression was driven by the 5’ fusion partner. This structure follows the pattern of somatic BRAF-fusions reported previously in solid tumors including melanoma. Potential dimerization domains in the partner genes were identified in 9 cases. We identify here mosaic BRAF fusions as a recurrent cause of multiple CMN allowing genetic diagnosis in a further 15% of cases and linking this genotype to a highly proliferative pruritic phenotype. In vitro data from melanoma cell lines have suggested that higher expression levels of the fusion protein as well as dimerization domains in the partner gene correlate with resistance and/or paradoxical MAP-kinase activation after treatment with RAF and MEK inhibitors. This may have implications for the use of targeted therapies for attempted reduction of the nodular phenotype, or where melanoma arises in CMN patients caused by mosaic BRAF-fusions. Citation Format: Sara Barberan Martin, Satyamaanasa Polubothu, Alicia Bruzos, Neil Bulstrode, Gavin Kelly, Veronica Kinsler. Mosaic BRAF fusions are a recurrent cause of multiple congenital melanocytic naevi [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 2014.

Abstract CT032: Activity and safety of alectinib for ALK-altered solid tumors from MyPathway

Abstract Purpose: Alectinib is FDA-approved for ALK-positive, metastatic non-small cell lung cancer (NSCLC). We analyzed alectinib treatment in a pan-tumor population with ALK alterations from MyPathway (NCT02091141), a multi-basket study assessing approved therapies in non-indicated advanced solid tumors with relevant alterations. Methods: Enrolled patients (pts) were ≥18 yrs old and had metastatic tumors with ALK gene rearrangements, putative activating non-synonymous ALK mutations, and/or ALK gene amplification. Pts received alectinib 600 mg PO BID. The primary endpoint was investigator-assessed objective response rate (ORR; complete response [CR] + partial response [PR]). Other endpoints included duration of response (DOR), disease control rate (DCR; CR + PR + stable disease [SD] &amp;gt;4 mos), progression-free survival (PFS), and safety. Results: By the 11-18-2021 data cutoff, 21 pts with various tumor types had been enrolled and treated (ALK mutations or amplification, n=11 [52.4%]; ALK rearrangements +/- other ALK alterations, n=10 [47.6%]). Pts had a median of 2 (range, 1-5) prior lines of therapy. In the 10 pts with ALK rearrangements, there were 3 PRs (30.0%) with a median DOR of 6.8 mos (Table). An additional 3 pts in this group had SD &amp;gt;4 mos; DCR was 60.0% (6/10). In contrast, there were no responses among the 11 pts with ALK mutations or amplification. Confirmed ORR for the entire group was 14.3% (3/21), and DCR was 42.9% (9/21). Median PFS was 8.2 mos in pts with ALK rearrangements vs 1.8 mos for those with other ALK alterations. Alectinib-related adverse events (AEs) were observed in 85.7% of pts, with 3 (14.3%) experiencing grade 3 AEs (anemia; hypokalemia; and changes in AST, ALT, and/or blood creatinine levels). AEs were consistent with the known alectinib safety profile. Conclusions: Although the number of pts is small, alectinib appears active in those with non-NSCLC advanced solid tumors with ALK rearrangements. As in NSCLC, cancers with ALK mutations or amplification were not responsive to ALK inhibition. Table. Clinical outcomes Clinical Response Rearrangement (n=10) Mutation (n=7) Amplification (n=4) Median PFS, mos (all alterations) n Indications n Indications n Indications PR 3a Melanoma, Papillary urothelial carcinoma, Colon adenocarcinoma 0 NA 0 NA 9.3 SD &amp;gt;4 mos 3b,c Colon adenocarcinoma, Uterine leiomyosarcoma, Pancreatic adenocarcinoma 2 Squamous cell carcinoma, Soft tissue sarcoma 1 Uterine body clear cell carcinoma 5.7 SD ≤4 mos 0 NA 1 Fallopian tube serous carcinoma 0 NA PD 3d,e Colon adenocarcinoma, Esophageal adenocarcinoma, Uterine serous carcinoma 4 Anaplastic thyroid carcinoma, Esophageal adenocarcinoma, Colon adenocarcinoma (2) 3 Ovarian serous carcinoma, Gastric squamous cell carcinoma, Peritoneal non-small cell carcinoma 1.7 Non-evaluable 1f Uterine inflammatory myofibroblastic tumor 0 NA 0 NA NA aFusion gene partners: EMILIN1, DCTN1, and DIAPH2. bOne pt with ALK rearrangement and SD &amp;gt;4 mos also had ALK amplification. cFusion gene partners: STRN, IGFBP5, and EML4. dPts with ALK rearrangement and PD also had an ALK mutation (n=1) or ALK amplification (n=1). eFusion gene partner: STRN; fusion genes unknown for 2 pts. fPt withdrew prior to clinical assessment and was censored for PFS at 0.03 mos. NA, not applicable; PD, progressive disease; PFS, progression-free survival; PR, partial response; SD, stable disease. Citation Format: Charles Swanton, Claire F. Friedman, Christopher J. Sweeney, Funda Meric-Bernstam, David Spigel, Ron Bose, Howard Burris, Walter C. Darbonne, Julia Malato, Jonathan Levy, Yong Wang, Tania Szado, Katja Schulze, John Hainsworth, Razelle Kurzrock. Activity and safety of alectinib for ALK-altered solid tumors from MyPathway [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 CT032.

Abstract 751: Transcriptome profiling reveals novel gene fusions in early-onset sporadic rectal cancer

Abstract Implementation of regular screening and adoption of a healthier lifestyle has resulted in a steady decline of colorectal cancer (CRC) incidence in the USA over the past few decades. On the contrary, early-onset sporadic rectal cancer (EOSRC) has exhibited an opposite trend in the more recent past. In addition, EOSRC appears to be the predominant CRC subtype in developing nations where CRC incidence has risen significantly during the past 2 decades. Our previous work revealed the absence of canonical CRC tumorigenesis pathways including Wnt (Wingless, Int-1) activation and mismatch repair inactivation in a significant proportion of EOSRC cases. More importantly, we revealed the enrichment of Ca2+/NFAT signaling leading to chromosomal instability in samples devoid of Wnt activation besides identifying ARID2 as a novel tumor suppressor for EOSRC. In order to improve our understanding of EOSRC driver events, we endeavored to characterize the landscape of gene fusions (GFs); given that the latter provide an attractive druggable target. To this end, we performed paired-end RNA Sequencing on 37 carefully selected and well annotated microsatellite stable EOSRC samples. We employed in silico prediction algorithms viz. FusionCatcher, SOAPFuse and EricScript to identify GFs after removing those predicted (GTEx and ConjoinG databases) to occur in normal tissues. The GFs were further filtered based on read support and identification through at least 2 of the 3 algorithms and validated using reverse transcription PCR and Sanger sequencing. Overall, we identified several known (EIF3E-RSPO2, NCEH1-SPATA16) and novel GFs that arose as inter- or intra-chromosomal events. Computational analyses revealed selective enrichment of Gene Ontologies (GOs) in the 5’- (as compared to 3’-) gene partners which differed from GOs enriched in GFs identified from the TCGA CRC data. More importantly, we analyzed the potential role of chromatin architecture in determining the fusion breakpoints. Analysis of CRC Hi-C data revealed that a majority of EOSRC GFs arose from genes located in open chromatin regions. Interestingly, fusions between 5’ ‘open’ and 3’ ‘closed’ partners often resulted in transcriptional activation of the latter revealing a novel mode of oncogenic activation in cancer. Further analyses to correlate the predicted fusion breakpoints with topologically associating domain (TAD) boundaries as well as with common fragile sites (CFSs) that associate with transcription-replication collision are currently ongoing. Validation of potential oncogenic role of few selected GFs using CRC cell lines and nude mice xenografts is also underway. The study is expected to provide significant insights into the patterns of genomic rearrangements in EOSRC besides highlighting the critical role GFs might play in propagating tumorigenesis in CRC. Citation Format: Asmita Gupta, Vasanth Kumar Mandla, Pratyusha Bala, Murali Dharan Bashyam. Transcriptome profiling reveals novel gene fusions in early-onset sporadic rectal cancer [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 751.

Abstract 5725: HOXA9 demonstrates allelic imbalance in KMT2A-rearranged infant acute lymphoblastic leukemia

Abstract Background: KMT2A-rearrangement is a characteristic finding in approximately 70% of infant acute lymphoblastic leukemia (ALL) cases. We previously utilized a computational screen to find that KMT2A-r infant ALL demonstrated somatic allele specific expression for 431 genes when compared to non-KMT2A-r samples (Abstract Number 3649, AACR 2019). The gene that exhibited the most differential allele specific expression (ASE) pattern was HOXA9. Here, we follow up that analysis with manual examination of additional infant ALL samples. Methods: We performed whole genome sequencing (WGS) and RNA sequencing (RNAseq) on peripheral blood or bone marrow samples from 48 infants with ALL (33 KMT2A-r cases and 15 non-KMT2A-r cases) treated on Children’s Oncology Group protocol AALL0631, at diagnosis (DX), at remission (MD), and at relapse (RL), if applicable. WGS data at MD was phased using a 1000 Genomes reference panel. HOXA9 expression was quantified in transcripts per million (TPM). Biallelic expression was manually calculated on the phased genome for samples with at least 1 germline single nucleotide polymorphism (SNP) within HOXA9 and at least 5 aligned RNAseq reads over the HOXA9 transcript. ASE was defined as minor allele fraction (MAF; allele fraction for the less prevalent allele) of ≤ 0.35. Whole genome bisulfite sequencing data and sequencing-derived copy number data were also analyzed to investigate possible explanations for HOXA9 ASE. Results: We identified diagnostic samples from 21 patients, 9 KMT2A-AFF1, 9 KMT2A-MLLT1, and 3 non-KMT2A-r, that passed manual filtering. Data included 5 patients with paired DX and RL samples. Of the 15 KMT2A-r samples unable to be analyzed for ASE, 13 lacked a SNP for phasing and 2 had &amp;lt; 5 reads. For the 12 non-KMT2A-r samples unanalyzable for ASE, 6 lacked a SNP and 6 had &amp;lt; 5 reads. Average HOXA9 expression was 15.17 TPM (range 0.01 to 44.34) in KMT2A-r samples and 2.91 TPM (range 0 to 29.29) in non-KMT2A-r samples. ASE of HOXA9 (MAF ≤ 0.35) was observed in 14 samples, 72% of KMT2A-r (13 of 18 samples) and 33% of non-KMT2A-r (1 of 3 samples). RL samples were found to have HOXA9 ASE in all cases with ASE of HOXA9 in the DX sample. Of the 18 KMT2A-r samples, 6 of 9 with KMT2A-AFF1 and 7 of 9 with KMT2A-MLLT1 demonstrated HOXA9 ASE. Frequency of HOXA9 ASE at diagnosis was similar between patients known to later relapse versus those known to remain in remission. Allele specific methylation, mono-allelic deletion, or copy-neutral loss of heterozygosity (cn-LOH) for HOXA9 were not observed in KMT2A-r nor non-KMT2A-r samples. Conclusion: Our expanded cohort supports the ASE of HOXA9 in KMT2A-r infant ALL. This phenomenon did not appear to segregate by KMT2A-r partner gene, nor could it be explained by allele specific methylation or copy number variation (e.g., deletion of one HOXA9 allele or cn-LOH). Overall, further investigation into a causal mechanism is needed. Citation Format: Sarah E. Mc Dermott, Byunggil Yoo, Warren A. Cheung, Emily Farrow, Bing Ge, Margaret Gibson, Patrick A. Brown, Erin Guest, Tomi Pastinen, Midhat S. Farooqi. HOXA9 demonstrates allelic imbalance in KMT2A-rearranged infant acute lymphoblastic leukemia [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 5725.

Abstract 2503: Identification of clinically relevant gene fusions in archived pediatric solid and liquid tumor samples using Arima-HiC sequencing

Abstract Gene fusions are known to have oncogenic effects in various cancers and serve as disease biomarkers. Obtaining gene-level resolution of these large structural variants is not possible with lower resolution karyotyping approaches and can be challenging using RNA sequencing approaches due to low transcript abundance or low-quality RNA. Furthermore, the ability of fluorescence in situ hybridization (FISH) techniques to detect fusions is limited by the targeted nature of these assays and subject to which probes were run. Here, we used a novel method, Arima-HiC sequencing, on 6 archived pediatric cancer samples to determine the technique’s effectiveness in detecting gene fusions. We first adapted the Arima Hi-C method for use with formalin-fixed paraffin-embedded (FFPE) tissue. We then selected four archived pediatric alveolar rhabdomyosarcoma (ARMS) tumors (FFPE archival period range: 8-12 years)—known to be fusion-positive via prior clinical testing—for Arima-HiC sequencing. Briefly, FFPE tissue scrolls were dewaxed, and the tissue rehydrated; it then underwent chromatin digestion, end-labeling, and proximity ligation prior to DNA purification per the Arima-HiC FFPE protocol. Purified DNA was next prepared as a short-read sequencing library and sequenced on a HiSeq 2500. The raw reads were aligned and deduplicated, and structural variants were called using the HiC-Breakfinder software. We additionally selected 2 non-FFPE cases, both pediatric leukemia cases with cryopreserved blasts (archival period range: 1-3 years), for Arima-HiC sequencing (as above). These cases had undergone standard of care cytogenetic (karyotyping, FISH) and molecular (targeted cancer NGS sequencing panel) testing clinically and a genetic driver/known gene fusion had not been identified. Using Arima-HiC sequencing, we identified either a PAX3-FOXO1 (n=2) or PAX7-FOXO1 (n=2) gene fusion in each of our 4 ARMS cases, consistent with the original diagnostic cytogenetic finding. HiC data was additionally able to provide partner gene information (PAX3 or PAX7) in 2 of the 4 cases where this was unknown. We also test two cases of leukemia, one, a case of precursor B-cell acute lymphoblastic leukemia (B-ALL) in a 3-year-old male, and the other, a case of acute myeloid leukemia (AML) in a 2-year-old female. In the first case, we detected an EP300-ZNF384 gene fusion, which is a known, but rare, gene fusion seen in B-ALL. This finding is clinically relevant as ZNF384 fusions are associated with a more favorable prognosis. In the second case, we detected a SCN4B-MLLT10 gene fusion. This finding is also clinically relevant as MLLT10 fusions are rare in AML but indicate an unfavorable prognosis. In summary, this study demonstrates how Arima-HiC sequencing provides molecular diagnostic value in archived pediatric solid and liquid tumor specimens via the identification of clinically relevant gene fusions. Citation Format: Midhat S. Farooqi, Kristin Sikkink, Derek Reid, Tomi Pastinen, Anthony Schmitt, Atif Ahmed. Identification of clinically relevant gene fusions in archived pediatric solid and liquid tumor samples using Arima-HiC sequencing [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 2503.

Abstract CT545: Variability in NTRK gene fusions does not appear to impact response to larotrectinib

Abstract Introduction: Neurotrophic tyrosine receptor kinase (NTRK) gene fusions are oncogenic drivers in various tumor types. Gene fusions arise from inter- and intrachromosomal rearrangements involving the 3’ region of the NTRK gene and the 5’ end of a fusion partner gene, leading to the expression of a constitutively active tropomyosin receptor kinase (TRK) fusion protein. Larotrectinib is a highly selective and central nervous system (CNS)-active TRK inhibitor approved for the treatment of adult and pediatric patients with TRK fusion cancer. Larotrectinib has previously demonstrated an objective response rate (ORR) of 75% in 206 patients with non-primary CNS solid tumors (Hong et al, ASCO 2021). Here, we report on the TRK fusion exon junction organization in patients with TRK fusion cancer receiving larotrectinib. Methods: Patients with non-primary CNS TRK fusion cancer treated with larotrectinib were identified from three clinical trials (NCT02122913, NCT02576431, and NCT02637687). NTRK gene fusion data were determined by local molecular testing. The testing method, NTRK gene fusions, and exon structure were extracted from the molecular pathology reports from 212 patients. The data cut-off was July 20, 2020. Results: TRK fusion exon junction data were available for 80 patients from 14 different tumor types. Median age was 47 years (range 0.5-84 years). The ORR for these patients was 82.5% (95% confidence interval [CI] 72.4-90.1). Patients had gene fusions involving NTRK1 (40%), NTRK2 (4%), or NTRK3 (56%), a pattern similar to that seen in the larger dataset. We detected 20 different NTRK gene fusions; the most common were ETV6-NTRK3, TPM3-NTRK1, and LMNA-NTRK1 detected in 39 (49%), ten (13%), and seven (9%) patients, respectively. There were 12, three, and five different fusion partners associated with NTRK1, NTRK2, and NTRK3, respectively. The majority (12/20; 60%) of fusions were intrachromosomal rearrangement events: 8/12 (67%) related to NTRK1, 2/3 (67%) related to NTRK2, and 2/5 (40%) related to NTRK3. There were 35 different 5’/3’ gene breakpoint combinations: 3’ fusion points were located at NTRK1 exon 8 (n = 2), NTRK1 exon 9 (n = 5), NTRK1 exon 10 (n = 15), NTRK1 exon 11 (n = 6), NTRK1 exon 12 (n = 3), NTRK1 exon 13 (n = 1), NTRK2 exon 15 (n = 2), NTRK2 exon 16 (n = 1), NTRK3 exon 14 (n = 21), and NTRK3 exon 15 (n = 24). For ETV6-NTRK3 fusions, the NTRK3 fusion point was at exon 14 in ten of 11 thyroid carcinomas, but in only three of 13 salivary gland tumors. Conclusions: Among 80 patients with TRK fusion cancer, we detected 20 different NTRK gene fusions, of which 55% occurred in only one patient each. There were 36 different fusion variants. Despite the variability of the fusion structure, larotrectinib was equally efficacious. The large number of potential different fusion partners and involvement of various breakpoints highlight the need for validated and appropriate testing methodologies that are agnostic of 5’ partners and breakpoints. Citation Format: Marc Ladanyi, Sinchita Roy-Chowdhuri, Lauren Ritterhouse, Felix Sahm, Ricarda Norenberg, Kui Shen, Chi Chen, Marc Fellous, Nicoletta Brega, Cornelis M. van Tilburg, Jessica J. Lin, Marcia S. Brose, Ulrik N. Lassen, Ray McDermott, Theodore W. Laetsch, David S. Hong, Alexander Drilon, Erin R. Rudzinski. Variability in NTRK gene fusions does not appear to impact response to larotrectinib [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 CT545.

Abstract 6390: Chromosome arm aneuploidies as genetic vulnerabilities of triple-negative breast cancer

Abstract Triple-negative breast cancer (TNBC) is a subtype of breast cancer which lacks the expression of key biomarkers, namely Estrogen and Progesterone receptors and HER2. Therefore, it lacks targeted therapies, resulting in the worst prognosis compared to other breast cancer subtypes. Chromosome 4p (chr4p) loss is recurrent in TNBC, correlates with poor prognosis, is an early event in tumor evolution, and confers a proliferative advantage onto cells. Here, we propose that chr4p loss can be leveraged as a genetic vulnerability of TNBC by identifying its specific synthetic lethal (SL) genetic interactions. An SL interaction occurs when the inactivation of individual genes is tolerated, however, the combined inactivation of the corresponding genes leads to a loss of cellular viability. SL interactions offer promising avenues for precision oncology therapeutic strategies of TNBC. We leveraged the publicly available CRISPR-mediated genome-wide gene inactivation screens (DepMap project) dataset to computationally predict pairwise SL interactions that are specific to individual genes within chr4p, as well as complex SL genetic interactions involving multiple genes spanning large segments of chr4p and the entire chromosome arm. We developed regression models to compare the inactivation effect of a given SL partner gene, between cell lines that harbor chr4p loss and those characterized by a copy-neutral status of chr4p, while effectively accounting for potential confounding effects. We integrated the putative SL interactions identified from these regression models with those identified using other methods, drugZ and MAGECK, therefore, resulting in a robust set of putative SL interactions. We have prioritized the SL interactions for their subsequent experimental validation in TNBC PDX-derived 2D and 3D cell models, which we will carry out using CRISPR-based gene editing. The prediction model and collective set of SL candidates identified in this study is a unique resource for the development of SL-based therapeutic strategies for TNBC and other cancers harboring chr4p loss. Citation Format: Rohan Dandage, Michael Schwartz, Lynn Karam, Traver Hart, Elena Kuzmin. Chromosome arm aneuploidies as genetic vulnerabilities of triple-negative breast cancer [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 6390.

Abstract 881: Functional characterization and clinical prevalence of ESR1 fusions in advanced endocrine resistant breast cancer

Abstract Breast cancer is the most frequently diagnosed women’s cancer with one in eight women being diagnosed during her lifetime. Even within the era of novel therapeutics and improved diagnostic screening, mortality rates remain stable, with an estimated 43,600 deaths occurring in 2021 alone. Malignant transformation of breast cancer to advanced disease is the mainstay driver of mortality and is largely attributed to treatment resistance. Fortunately, identification of therapeutic resistance mechanisms has rapidly transformed our understanding of cancer evasion and is enabling personalized treatment guidance and regimens. Notably, mutations and amplifications of the estrogen receptor (ER, ESR1) remain a prominent source of endocrine therapy resistance. Two-thirds of all breast cancer are positive for ER, making it an attractive druggable target that has revolutionized patient care. Unfortunately, however, 30% of tumors exposed to endocrine therapy subsequently develop resistance. Recently identified ER fusion proteins, discovered in metastatic endocrine resistant disease, result in substitution of the C-terminal ER ligand binding domain with a variety of fused in-frame gene partners due to genomic translocations occurring at ESR1 intron 6. Our lab has demonstrated that ER fusion proteins drive robust ER hyperactivation and metastatic potential independent of ER ligand estradiol and in the presence of endocrine therapies such as Fulvestrant. Overexpression of ER fusion proteins, particularly ESR1-SOX9 and ESR1-YAP1, in invasive ductal and invasive lobular carcinoma models with endogenous ER knockdown, resulted in enhanced 3D growth, increased migration by wound scratch assay, enhanced colony formation as a proxy of survival and ER pathway hyperactivation. Targeted endogenous ER knockdown utilizing short hairpin RNA enhances fusion driven phenotypes that are unique to fusion partner proteins and cellular context. ER fusion prevalence in advanced breast cancer remains a challenge due to the promiscuous fusion partners disrupting traditional sequencing techniques. Our group has developed a novel method of ESR1 fusion detection, EnRich, which exploits the ESR1 fusion breakpoints. Importantly, we have assessed EnRich fusion discovery with the use of circulating tumor DNA (ctDNA) from non-invasive liquid biopsies. Our pilot study of 15 blood ctDNA samples detected three different ESR1 fusion gene products which have been further validated by PCR. EnRich will help us identify and monitor treatment resistance of known and novel ESR1 fusions in patient samples irrespective of treatment regime as well as in longitudinal samples collected from patients undergoing endocrine therapy. The in vitro resistant properties of fusion proteins emphasize the necessity to better detect these mutations in patients with advanced disease and determine more appropriate treatment measures. Citation Format: Megan E. Yates, Tiantong Liu, Jagmohan Hooda, Jennifer M. Atkinson, Adrian V. Lee, Steffi Oesterreich. Functional characterization and clinical prevalence of ESR1 fusions in advanced endocrine resistant breast cancer [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 881.

Abstract 5268: The spectrum of FGFR mutations in pediatric and young adult solid tumor

Abstract 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.