{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,22]],"date-time":"2026-06-22T10:53:46Z","timestamp":1782125626107,"version":"3.54.5"},"reference-count":51,"publisher":"MDPI AG","issue":"12","license":[{"start":{"date-parts":[[2026,6,20]],"date-time":"2026-06-20T00:00:00Z","timestamp":1781913600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"UCSF Department of Pathology"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Diagnostics"],"abstract":"<jats:p>Background\/Objectives: There remains a need for additional prognostic markers in classic follicular lymphoma (cFL) to identify aggressive disease. Immunohistochemical stains such as Ki-67, MYC, and p53 have shown variable associations with histologic grade and adverse outcomes. In this study, we aimed to assess intrafollicular Ki-67, MYC, and p53 expression in cFL via immunohistochemistry, quantified by both manual and digital methods, and evaluate their relation to histologic grade and clinical outcomes. Methods: We evaluated 37 cases of cFL from 2000 to 2019 and performed immunohistochemistry for Ki-67, MYC, and p53 on tumor microarray tissue. Stains were assessed within follicles by digital pathology means on QuPath software and via manual low-power estimates. Results:MYC expression was greater in FL3A compared to FL1\u20132 across all digital and manual scoring methods (all p &lt; 0.05). Ki-67 and p53 expression did not differ by histologic grade group. No biomarker showed a significant association with adverse clinicopathologic features or outcomes, including FLIPI risk group, bulky disease, clinical stage, event-free survival, or overall survival. Manual and digital scores demonstrated strong correlations for all markers (\u03c1 = 0.71\u20130.89, all p &lt; 0.001). Conclusions: In our cohort, MYC expression was increased in FL3A compared to FL1\u20132, while no intrafollicular biomarker measurement was associated with adverse clinicopathologic features or clinical outcomes in exploratory analyses. These findings should be interpreted with caution in light of our limited cohort size. Strong concordance between manual and digital scoring supports the feasibility of digital IHC quantification in cFL.<\/jats:p>","DOI":"10.3390\/diagnostics16121917","type":"journal-article","created":{"date-parts":[[2026,6,22]],"date-time":"2026-06-22T09:49:55Z","timestamp":1782121795000},"page":"1917","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Digital and Manual Assessment of Intrafollicular Ki-67, MYC, and p53 in Classic Follicular Lymphoma"],"prefix":"10.3390","volume":"16","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-7353-0641","authenticated-orcid":false,"given":"George C.","family":"de Castro","sequence":"first","affiliation":[{"name":"Department of Pathology, University of California San Francisco, San Francisco, CA 94143, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4941-7643","authenticated-orcid":false,"given":"Morgan L.","family":"Shannon","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine, Stanford Health Care, Palo Alto, CA 94305, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ruth","family":"Zhang","sequence":"additional","affiliation":[{"name":"Peninsula Pathologists Medical Group, South San Francisco, CA 94063, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9024-5751","authenticated-orcid":false,"given":"Kunwar","family":"Singh","sequence":"additional","affiliation":[{"name":"Guardant Health, Redwood City, CA 94063, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robert S.","family":"Ohgami","sequence":"additional","affiliation":[{"name":"Department of Pathology, University of Utah and ARUP Laboratories, Salt Lake City, UT 84108, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8251-902X","authenticated-orcid":false,"given":"Kwun Wah","family":"Wen","sequence":"additional","affiliation":[{"name":"Department of Pathology, University of California San Francisco, San Francisco, CA 94143, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,20]]},"reference":[{"key":"ref_1","unstructured":"WHO Classification of Tumours Editorial Board (2024). Follicular Lymphoma. Hematolymphoid Tumors, International Agency for Research on Cancer. WHO Classification of Tumours Series."},{"key":"ref_2","first-page":"443","article-title":"2016 US Lymphoid Malignancy Statistics by World Health Organization Subtypes","volume":"66","author":"Teras","year":"2016","journal-title":"CA Cancer J. Clin."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"316","DOI":"10.1002\/ajh.25696","article-title":"Follicular Lymphoma: 2020 Update on Diagnosis and Management","volume":"95","author":"Freedman","year":"2020","journal-title":"Am. J. Hematol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"e151","DOI":"10.3324\/haematol.2017.175059","article-title":"Impact of Histological Grading on Survival in the SWOG S0016 Follicular Lymphoma Cohort","volume":"103","author":"Rimsza","year":"2018","journal-title":"Haematologica"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"696","DOI":"10.1111\/bjh.18404","article-title":"The Impact of Histological Grade on Outcomes in Follicular Lymphoma: An Analysis of Patients in the SEER Database in the Context of Evolving Disease Classification and Treatment","volume":"199","author":"Naik","year":"2022","journal-title":"Br. J. Haematol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"16","DOI":"10.1186\/s40364-023-00462-z","article-title":"Differences in Clinical Characteristics and Outcomes between Patients with Grade 3a and Grades 1\u20132 Follicular Lymphoma: A Real-World Multicenter Study","volume":"11","author":"Zha","year":"2023","journal-title":"Biomark. Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"209","DOI":"10.1007\/s00277-017-3154-z","article-title":"Current Prognostic and Predictive Factors in Follicular Lymphoma","volume":"97","author":"Sorigue","year":"2018","journal-title":"Ann. Hematol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"539","DOI":"10.1093\/ajcp\/aqz008","article-title":"Stratification of Follicular Lymphoma: Time for a Paradigm Shift?","volume":"151","author":"Kroft","year":"2019","journal-title":"Am. J. Clin. Pathol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1229","DOI":"10.1182\/blood.2022015851","article-title":"The International Consensus Classification of Mature Lymphoid Neoplasms: A Report from the Clinical Advisory Committee","volume":"140","author":"Campo","year":"2022","journal-title":"Blood"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"225","DOI":"10.1111\/j.1365-2141.2011.08942.x","article-title":"Clinical Significance of the WHO Grades of Follicular Lymphoma in a Population-Based Cohort of 505 Patients with Long Follow-up Times","volume":"156","author":"Wahlin","year":"2012","journal-title":"Br. J. Haematol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"31","DOI":"10.1038\/s41408-020-0299-0","article-title":"Patterns of Change in Treatment, Response, and Outcome in Patients with Follicular Lymphoma over the Last Four Decades: A Single-Center Experience","volume":"10","author":"Mozas","year":"2020","journal-title":"Blood Cancer J."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"2516","DOI":"10.1200\/JCO.2014.59.7534","article-title":"Early Relapse of Follicular Lymphoma After Rituximab Plus Cyclophosphamide, Doxorubicin, Vincristine, and Prednisone Defines Patients at High Risk for Death: An Analysis From the National LymphoCare Study","volume":"33","author":"Casulo","year":"2015","journal-title":"J. Clin. Oncol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1258","DOI":"10.1182\/blood-2003-12-4434","article-title":"Follicular Lymphoma International Prognostic Index","volume":"104","author":"Roy","year":"2004","journal-title":"Blood"},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"He, X., Chen, Z., Fu, T., Jin, X., Yu, T., Liang, Y., Zhao, X., and Huang, L. (2014). Ki-67 Is a Valuable Prognostic Predictor of Lymphoma but Its Utility Varies in Lymphoma Subtypes: Evidence from a Systematic Meta-Analysis. BMC Cancer, 14.","DOI":"10.1186\/1471-2407-14-153"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"3307","DOI":"10.1182\/bloodadvances.2022009521","article-title":"Molecular Profiling Identifies at Least 3 Distinct Types of Posttransplant Lymphoproliferative Disorder Involving the CNS","volume":"7","author":"Guney","year":"2023","journal-title":"Blood Adv."},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Cui, Y., and Guo, G. (2016). Immunomodulatory Function of the Tumor Suppressor P53 in Host Immune Response and the Tumor Microenvironment. Int. J. Mol. Sci., 17.","DOI":"10.3390\/ijms17111942"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1554","DOI":"10.3324\/haematol.2013.086173","article-title":"MYC Protein Expression and Genetic Alterations Have Prognostic Impact in Patients with Diffuse Large B-Cell Lymphoma Treated with Immunochemotherapy","volume":"98","author":"Valera","year":"2013","journal-title":"Haematologica"},{"key":"ref_18","first-page":"139","article-title":"Cross-Talk between Myc and P53 in B-Cell Lymphomas","volume":"5","author":"Yu","year":"2019","journal-title":"Chronic Dis. Transl. Med."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"165","DOI":"10.1038\/nrc2072","article-title":"The P53 Family in Differentiation and Tumorigenesis","volume":"7","author":"Stiewe","year":"2007","journal-title":"Nat. Rev. Cancer"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"68","DOI":"10.1016\/j.cell.2012.08.033","article-title":"C-Myc Is a Universal Amplifier of Expressed Genes in Lymphocytes and Embryonic Stem Cells","volume":"151","author":"Nie","year":"2012","journal-title":"Cell"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Mao, X., Li, Y., Liu, S., He, C., Yi, S., Kuang, D., Xiao, M., Zhu, L., and Wang, C. (2023). Multicolor Flow Cytometric Assessment of Ki67 Expression and Its Diagnostic Value in Mature B-Cell Neoplasms. Front. Oncol., 13.","DOI":"10.3389\/fonc.2023.1108837"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"646","DOI":"10.1111\/bjh.12667","article-title":"Association of Quantitative Assessment of the Intrafollicular Proliferation Index with Outcome in Follicular Lymphoma","volume":"164","author":"Kedmi","year":"2014","journal-title":"Br. J. Haematol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"380","DOI":"10.1093\/ajcp\/aqad161","article-title":"Digital and Manual Interfollicular Ki-67 Are Associated with a Progression-Free Survival in Patients with Low-Grade Follicular Lymphoma","volume":"161","author":"Nasir","year":"2024","journal-title":"Am. J. Clin. Pathol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"2085","DOI":"10.1016\/j.humpath.2014.06.019","article-title":"Identification of MUM1 as a Prognostic Immunohistochemical Marker in Follicular Lymphoma Using Computerized Image Analysis","volume":"45","author":"Xerri","year":"2014","journal-title":"Hum. Pathol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"3330","DOI":"10.1200\/JCO.2006.10.5833","article-title":"Tumor Sclerosis but Not Cell Proliferation or Malignancy Grade Is a Prognostic Marker in Advanced-Stage Follicular Lymphoma: The German Low Grade Lymphoma Study Group","volume":"25","author":"Klapper","year":"2007","journal-title":"J. Clin. Oncol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"384","DOI":"10.1097\/PAS.0000000000001609","article-title":"Biomarkers for Risk Stratification in Patients With Previously Untreated Follicular Lymphoma Receiving Anti\u2013CD20-Based Biological Therapy","volume":"45","author":"Sohani","year":"2021","journal-title":"Am. J. Surg. Pathol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"665","DOI":"10.1002\/hon.2778","article-title":"Prognostic Significance of Histologic Grade and Ki-67 Proliferation Index in Follicular Lymphoma","volume":"38","author":"Xue","year":"2020","journal-title":"Hematol. Oncol."},{"key":"ref_28","first-page":"1508","article-title":"Prognostic Value of P53 and Myc Protein Expression in Follicular Lymphomas in the PRIMA Trial in the Lysa Group","volume":"130","author":"Canioni","year":"2017","journal-title":"Blood"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"97","DOI":"10.1097\/00129039-200406000-00001","article-title":"Immunophenotypic and Genotypic Characterization of Progression in Follicular Lymphomas","volume":"12","author":"Natkunam","year":"2004","journal-title":"Appl. Immunohistochem. Mol. Morphol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1385\/MO:18:1:15","article-title":"Prognostic Significance of Ki-67 Nuclear Proliferative Antigen, Bcl-2 Protein, and P53 Expression in Follicular and Diffuse Large B-Cell Lymphoma","volume":"18","author":"Llanos","year":"2001","journal-title":"Med. Oncol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1994","DOI":"10.1182\/blood.V82.7.1994.1994","article-title":"P53 Mutation Is Associated With Progression in Follicular Lymphomas","volume":"82","author":"Sander","year":"1993","journal-title":"Blood"},{"key":"ref_32","first-page":"7559","article-title":"Utility of Immunohistochemistry with an Antibody against MYC at the Initial Diagnosis of Follicular Lymphoma, Grade 3A, for Predicting a More Aggressive Clinical Course: A Case Report and Review of the Literature","volume":"8","author":"Ichikawa","year":"2015","journal-title":"Int. J. Clin. Exp. Pathol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"960","DOI":"10.1111\/his.13316","article-title":"MYC Expression and Translocation Analyses in Low-Grade and Transformed Follicular Lymphoma","volume":"71","author":"Aukema","year":"2017","journal-title":"Histopathology"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"16878","DOI":"10.1038\/s41598-017-17204-5","article-title":"QuPath: Open Source Software for Digital Pathology Image Analysis","volume":"7","author":"Bankhead","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_35","unstructured":"R Core Team (R: A Language and Environment for Statistical Computing, 2023). R: A Language and Environment for Statistical Computing, version 4.5.2."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"100958","DOI":"10.1016\/j.modpat.2026.100958","article-title":"Biomarkers Informed by Single-Cell and Spatial Transcriptomics\u2014Biomarkers for Grade 3 Follicular Lymphoma","volume":"39","author":"Kanzaria","year":"2026","journal-title":"Mod. Pathol."},{"key":"ref_37","first-page":"3366","article-title":"Ki-67 Expression of Immunohistochemistry Using Computerized Image Analysis Is a Useful Prognostic Marker in Follicular Lymphomas","volume":"11","author":"Kawaguchi","year":"2018","journal-title":"Int. J. Clin. Exp. Pathol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"1084","DOI":"10.5858\/2007-131-1084-GOFLCO","article-title":"Grading of Follicular Lymphoma: Comparison of Routine Histology with Immunohistochemistry","volume":"131","author":"Martinez","year":"2007","journal-title":"Arch. Pathol. Lab. Med."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"184","DOI":"10.3324\/haematol.10384","article-title":"van The Prognostic Significance of the Intra-Follicular Tumor Cell Proliferative Rate in Follicular Lymphoma","volume":"92","author":"Koster","year":"2007","journal-title":"Haematologica"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"1490","DOI":"10.1097\/01.pas.0000172191.87176.3b","article-title":"Low Histologic Grade Follicular Lymphoma with High Proliferation Index: Morphologic and Clinical Features","volume":"29","author":"Wang","year":"2005","journal-title":"Am. J. Surg. Pathol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"313","DOI":"10.1038\/modpathol.2017.134","article-title":"Clinicopathological and Genomic Analysis of Double-Hit Follicular Lymphoma: Comparison with High-Grade B-Cell Lymphoma with MYC and BCL2 and\/or BCL6 Rearrangements","volume":"31","author":"Miyaoka","year":"2018","journal-title":"Mod. Pathol."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"307","DOI":"10.1002\/(SICI)1096-9896(199707)182:3<307::AID-PATH873>3.0.CO;2-6","article-title":"Bcl-2 and P53 Protein Expression in Follicular Lymphoma","volume":"182","author":"Cooper","year":"1997","journal-title":"J. Pathol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"538","DOI":"10.1093\/ajcp\/105.5.538","article-title":"Immunohistochemical Detection of P53, Bcl-2, and Retinoblastoma Proteins in Follicular Lymphoma","volume":"105","author":"Nguyen","year":"1996","journal-title":"Am. J. Clin. Pathol."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"3806","DOI":"10.1182\/blood.V99.10.3806","article-title":"Cytomorphologic, Immunohistochemical, and Cytogenetic Profiles of Follicular Lymphoma: 2 Types of Follicular Lymphoma Grade 3","volume":"99","author":"Ott","year":"2002","journal-title":"Blood"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"6685","DOI":"10.21873\/anticanres.14692","article-title":"EZH2 Expression in Follicular Lymphoma Is Variable and Independent from the Progression of Disease Within 24 Months of First Treatment","volume":"40","author":"Poleszczuk","year":"2020","journal-title":"Anticancer Res."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"19","DOI":"10.1016\/j.humpath.2021.04.014","article-title":"Relative Frequency and Clinicopathologic Characteristics of MYC-Rearranged Follicular Lymphoma","volume":"114","author":"Chaudhary","year":"2021","journal-title":"Hum. Pathol."},{"key":"ref_47","doi-asserted-by":"crossref","unstructured":"Narita, H., Kuroiwa, K., Kawaguchi, Y., Murai, S., Sasaki, Y., Homma, M., Kawamata, N., Hayashi, H., Nagao, K., and Okamura, R. (2024). High Ki-67 Expression Predicting a Risk Factor for the Progression of Disease within 24 Months and Microenvironment in Follicular Lymphoma. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms252011057"},{"key":"ref_48","first-page":"38","article-title":"Prognostic Value of the Ki67 Proliferation Index, CD10 and MUM1 in Follicular Lymphoma","volume":"5","author":"Vu","year":"2022","journal-title":"Int. J. Oncol. Res."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"972","DOI":"10.1016\/j.humpath.2010.10.011","article-title":"Differential Expression of Cell-Cycle Regulatory Proteins Defines Distinct Classes of Follicular Lymphoma","volume":"42","author":"Alhejaily","year":"2011","journal-title":"Hum. Pathol."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"667","DOI":"10.1111\/j.1440-1827.2004.01678.x","article-title":"Efficacy of Rituximab plus Chemotherapy in Follicular Lymphoma Depends on Ki-67 Expression","volume":"54","author":"Saito","year":"2004","journal-title":"Pathol. Int."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"1459","DOI":"10.1038\/sj.leu.2403802","article-title":"A Limited Role for TP53 Mutation in the Transformation of Follicular Lymphoma to Diffuse Large B-Cell Lymphoma","volume":"19","author":"Davies","year":"2005","journal-title":"Leukemia"}],"container-title":["Diagnostics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2075-4418\/16\/12\/1917\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,22]],"date-time":"2026-06-22T10:31:53Z","timestamp":1782124313000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2075-4418\/16\/12\/1917"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,6,20]]},"references-count":51,"journal-issue":{"issue":"12","published-online":{"date-parts":[[2026,6]]}},"alternative-id":["diagnostics16121917"],"URL":"https:\/\/doi.org\/10.3390\/diagnostics16121917","relation":{},"ISSN":["2075-4418"],"issn-type":[{"value":"2075-4418","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,6,20]]}}}