Identification of Novel Fusion Genes in Pediatric B-ALL patients Using Whole Transcriptome Sequencing

Identification of Novel Fusion Genes in Pediatric B-ALL patients Using Whole Transcriptome Sequencing | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Identification of Novel Fusion Genes in Pediatric B-ALL patients Using Whole Transcriptome Sequencing Osheen Sajjad, Ali Amar, Abu Bakar, Muhammad Asif Naveed, Ayman Ali Mohammed Alameen, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7808597/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background: Fusion genes (FGs), serving as diagnostic, prognostic, and therapeutic markers, are key molecular aberrations in acute leukemia. They are also essential for risk stratification and measurable residual disease (MRD) monitoring. This study aimed to characterize the distribution of FGs using whole transcriptome sequencing (WTS). Materials and Methods: A total of 12 newly diagnosed treatment naive pediatric BALL cases from a local tertiary care hospital were enrolled in this study. Following the nucleic acids isolation procedures, the RNA sequencing was done for 12 BALL patients to find the fusion genes. Results: In the present cohort of 12 pediatric BALL patients, 19 high-confidence in-frame gene fusion events were identified involving 29 unique partner genes. The commonly reported sub-type defining rearrangements in BALL , including ETV6-RUNX1, TCF3-PBX1 and BCR-ABL1, were found in 8.3 % of the patients whereas the rearrangements in commonly prevalent genes in BALL like PAX5, ABL1 and ATXN3 were also found in 8.3 % of the patients with different unreported partner genes but reported earlier in various studies i.e. PAX-ETV6 (8.3%), ABL1-SNX2 (8.3%) and CMC2-ATXN3. Conclusion: The present work expands the scope of fusions in pediatric BALL by revealing unreported and domain-retaining fusions, as well as co-occurring rearranged fusions with possible combinatorial outcomes. By introducing WTS into clinical workflows, the genetic classification can be done more accurately and new pathogenic drivers missed by traditional methods can be identified, highlighting the increased significance of transcriptomic profiling in the diagnosis, prognosis, and personalized therapy of leukemia. Pediatric B-cell acute lymphoblastic leukemia Fusion genes Whole transcriptome sequencing RNA sequencing Kinase fusions Transcription factor fusions Domain retention Precision oncology Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Full Text Additional Declarations No competing interests reported. Table 1 is available in the Supplementary Files section. Supplementary Files Table.docx Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7808597","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":532329311,"identity":"cff25464-01de-440e-9a44-f5c35f3ca374","order_by":0,"name":"Osheen Sajjad","email":"","orcid":"","institution":"University of Health Sciences Lahore","correspondingAuthor":false,"prefix":"","firstName":"Osheen","middleName":"","lastName":"Sajjad","suffix":""},{"id":532329312,"identity":"eac431c5-8787-427b-8a7c-a7bdba238830","order_by":1,"name":"Ali Amar","email":"","orcid":"","institution":"Pakistan Kidney and Liver Institute and Research Center 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13:05:45","extension":"json","order_by":3,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":8813,"visible":true,"origin":"","legend":"","description":"","filename":"09800d118c0d4cbf97699485b444f422.json","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/b791a9927b970fe923172d5f.json"},{"id":94361879,"identity":"e6c57dce-9b9b-46d8-bed5-173a953a764f","added_by":"auto","created_at":"2025-10-27 13:04:29","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":68947,"visible":true,"origin":"","legend":"\u003cp\u003eDistribution of Fusion Gene (FG) Positivity and Multiplicity in Pediatric B-ALL Patients. The left pie chart shows the proportion of patients with detectable fusion genes (FGs) (n=9, red) versus FG-negative patients (n=3, green). The right exploded pie chart highlights the number of distinct fusion genes identified per FG-positive patient: 4 patients harbored a single FG (orange), 3 had two FGs (gray), while 1 patient each showed three (yellow) and four (blue) FGs. This illustrates both the frequency and heterogeneity of FG events in the cohort.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/03abff41e221af338a48ded2.png"},{"id":94362910,"identity":"0d9e4886-f8b3-4165-a7eb-65714881fbd0","added_by":"auto","created_at":"2025-10-27 13:05:42","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":500003,"visible":true,"origin":"","legend":"\u003cp\u003eFusion gene interaction network in pediatric B-ALL\u003cstrong\u003e. \u003c/strong\u003eSTRING-based protein–protein interaction network of fusion gene partners identified in pediatric B-ALL. Nodes represent gene products, and edges indicate predicted functional associations.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/4b399ba6900192846adf192b.png"},{"id":94362518,"identity":"821b97ff-4e25-444e-a2a2-3562878fefd0","added_by":"auto","created_at":"2025-10-27 13:05:11","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":166209,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eFusion transcript arising from intra-chromosomal inversion of the BCAR1 gene.\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/5081070379ec9604e67cda40.png"},{"id":94362579,"identity":"56fb41b7-112b-4480-a66a-2e31aa6b7a5e","added_by":"auto","created_at":"2025-10-27 13:05:15","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":220845,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eStructural representation and domain retention of the not reported NPR1::GALNT8 fusion gene\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/027b273fa0f5f9e18753ec09.png"},{"id":94362887,"identity":"e745d1b7-1768-4613-8bad-ded237fd4e3c","added_by":"auto","created_at":"2025-10-27 13:05:36","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":198641,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eStructural depiction and domain retention of the TAF3::TAF3 tandem duplication fusion in B-ALL\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/e0699a0ee8769ebb5a7f727a.png"},{"id":94362771,"identity":"df3dbc35-f423-4cf7-9d97-ba416a955c6f","added_by":"auto","created_at":"2025-10-27 13:05:25","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":186684,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eInter -chromosomal fusion of PCBP1::ARPC3 showing domain retention in an out-of-frame transcript\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"6.png","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/4ec2284ebf2215eb16c5e669.png"},{"id":94362760,"identity":"32c59f8a-5760-45f2-b64e-bfe04be6d2c7","added_by":"auto","created_at":"2025-10-27 13:05:25","extension":"png","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":145626,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eInter -chromosomal CMC2::ATXN3 fusion involving retained ubiquitin-interaction domains in an unclear reading frame.\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"7.png","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/bdf230707b2d06cab27deab9.png"},{"id":94362886,"identity":"2e667421-c6c0-4f34-9687-28e08342091f","added_by":"auto","created_at":"2025-10-27 13:05:35","extension":"png","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":384332,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCircos plot showing chromosomal locations of fusion genes identified in pediatric B-ALL patients\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"8.png","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1/83cfbc2b03dc7a40b64b3afb.png"},{"id":95018009,"identity":"12e7a596-44b9-4e8f-b37d-5dbf541c474c","added_by":"auto","created_at":"2025-11-03 11:39:07","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1092189,"visible":true,"origin":"","legend":"","description":"","filename":"FusionGenesinBALL.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7808597/v1_covered_6658b22c-bc7d-46f0-a13a-6f6c8204a821.pdf"},{"id":94362928,"identity":"054e3837-a4ab-469f-848d-8a839504e1e4","added_by":"auto","created_at":"2025-10-27 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Sequencing","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Pediatric B-cell acute lymphoblastic leukemia, Fusion genes, Whole transcriptome sequencing, RNA sequencing, Kinase fusions, Transcription factor fusions, Domain retention, Precision oncology","lastPublishedDoi":"10.21203/rs.3.rs-7808597/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7808597/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eBackground: Fusion genes (FGs), serving as diagnostic, prognostic, and therapeutic markers, are key molecular aberrations in acute leukemia. They are also essential for risk stratification and measurable residual disease (MRD) monitoring. This study aimed to characterize the distribution of FGs using whole transcriptome sequencing (WTS).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eMaterials and Methods: A total of 12 newly diagnosed treatment naive pediatric BALL cases from a local tertiary care hospital were enrolled in this study. Following the nucleic acids isolation procedures, the RNA sequencing was done for 12 BALL patients to find the fusion genes.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eResults: In the present cohort of 12 pediatric BALL patients, 19 high-confidence in-frame gene fusion events were identified involving 29 unique partner genes. The commonly reported sub-type defining rearrangements in BALL , including ETV6-RUNX1, TCF3-PBX1 and BCR-ABL1, were found in 8.3 % of the patients whereas the rearrangements in commonly prevalent genes in BALL like PAX5, ABL1 and ATXN3 were also found in 8.3 % of the patients with different unreported partner genes but reported earlier in various studies i.e. PAX-ETV6 (8.3%), ABL1-SNX2 (8.3%) and CMC2-ATXN3.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eConclusion: The present work expands the scope of fusions in pediatric BALL by revealing unreported and domain-retaining fusions, as well as co-occurring rearranged fusions with possible combinatorial outcomes. By introducing WTS into clinical workflows, the genetic classification can be done more accurately and new pathogenic drivers missed by traditional methods can be identified, highlighting the increased significance of transcriptomic profiling in the diagnosis, prognosis, and personalized therapy of leukemia.\u003c/p\u003e","manuscriptTitle":"Identification of Novel Fusion Genes in Pediatric B-ALL patients Using Whole Transcriptome Sequencing","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-24 18:13:13","doi":"10.21203/rs.3.rs-7808597/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"edec4248-c13d-4053-98a2-d749df5e8ed1","owner":[],"postedDate":"October 24th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-11-03T11:38:32+00:00","versionOfRecord":[],"versionCreatedAt":"2025-10-24 18:13:13","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7808597","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7808597","identity":"rs-7808597","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}