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Diagnostic value of MDM2 RNA in situ hybridization for low-grade osteosarcoma: Consistency comparison of RNA in situ hybridization, fluorescence in situ hybridization, and immunohistochemistry

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Abstract

Detection of MDM2 gene amplification via fluorescence in situ hybridization (FISH) and MDM2 overexpression by immunohistochemistry (IHC) have been utilized for the diagnosis of low-grade osteosarcoma (LGOS). The aim of this study was to evaluate the diagnostic value of MDM2 RNA in situ hybridization (RNA-ISH) and compare this assay with MDM2 FISH and IHC in distinguishing LGOS from its histologic mimics. MDM2 RNA-ISH, FISH and IHC were performed on nondecalcified samples of 23 LGOSs and 52 control cases. Twenty (20/21, 95.2%) LGOSs were MDM2-amplified, and two cases failed in FISH. All control cases were MDM2-nonamplified. All 20 MDM2-amplified LGOSs and one MDM2-nonamplified LGOS harboring TP53 mutation and RB1 deletion showed positivity for RNA-ISH. Fifty of the 52 (96.2%) control cases were negative for RNA-ISH. The diagnostic sensitivity and specificity of MDM2 RNA-ISH were 100.0% and 96.2%, respectively. Nineteen of the 23 LGOSs were evaluated by MDM2 RNA-ISH and FISH in decalcified samples simultaneously. All decalcified LGOSs failed in FISH and most samples (18/19) were no staining in RNA-ISH. Fifteen (15/20, 75%) MDM2-amplified LGOSs were positive for IHC and 96.2% (50/52) of control cases were negative. The sensitivity of RNA-ISH (100%) was higher than that of IHC (75%). In conclusion, MDM2 RNA-ISH has great value for the diagnosis of LGOS, with excellent consistency with FISH and better sensitivity than IHC. Acid decalcification still has an adverse impact on RNA. Some MDM2-nonamplified tumors may show positivity for MDM2 RNA-ISH, which needs to be analyzed comprehensively in combination with clinicopathological features.

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Data availability

The datasets used during the current study are available from the corresponding author upon reasonable request.

References

  1. Hang JF, Chen PC (2014) Parosteal osteosarcoma. Arch Pathol Lab Med 138:694–699. https://doi.org/10.5858/arpa.2013-0030-RS

    Article  PubMed  Google Scholar 

  2. Schwab JH, Antonescu CR, Athanasian EA, Boland PJ, Healey JH, Morris CD (2008) A comparison of intramedullary and juxtacortical low-grade osteogenic sarcoma. Clin Orthop Relat Res 466:1318–1322. https://doi.org/10.1007/s11999-008-0251-2

    Article  PubMed  PubMed Central  Google Scholar 

  3. Antonescu CR, Blay J-Y, Bovee JVMG, Bridge JA, Cunha IW, Dei Tos AP, Flanagan AM, Fletcher CDM, Folpe A, Hornick JL, Mertens F, Miettinen M, Nielsen GP, Oda Y, Yoshida A (2020) World Health Organization classification of tumours of soft tissue and bone, 5th edn. IARC Press, Lyon

    Google Scholar 

  4. Kurt AM, Unni KK, McLeod RA, Pritchard DJ (1990) Low-grade intraosseous osteosarcoma. Cancer 65:1418–1428. https://doi.org/10.1002/1097-0142(19900315)65:6%3c1418::aid-cncr2820650629%3e3.0.co;2-q

    Article  CAS  PubMed  Google Scholar 

  5. Duhamel LA, Ye H, Halai D, Idowu BD, Presneau N, Tirabosco R, Flanagan AM (2012) Frequency of Mouse Double Minute 2 (MDM2) and Mouse Double Minute 4 (MDM4) amplification in parosteal and conventional osteosarcoma subtypes. Histopathology 60:357–359. https://doi.org/10.1111/j.1365-2559.2011.04023.x

    Article  PubMed  Google Scholar 

  6. Dujardin F, Binh MB, Bouvier C, Gomez-Brouchet A, Larousserie F, Muret A, Louis-Brennetot C, Aurias A, Coindre JM, Guillou L, Pedeutour F, Duval H, Collin C, de Pinieux G (2011) MDM2 and CDK4 immunohistochemistry is a valuable tool in the differential diagnosis of low-grade osteosarcomas and other primary fibro-osseous lesions of the bone. Mod Pathol 24:624–637. https://doi.org/10.1038/modpathol.2010.229

    Article  CAS  PubMed  Google Scholar 

  7. Mejia-Guerrero S, Quejada M, Gokgoz N, Gill M, Parkes RK, Wunder JS, Andrulis IL (2010) Characterization of the 12q15 MDM2 and 12q13–14 CDK4 amplicons and clinical correlations in osteosarcoma. Genes Chromosomes Cancer 49:518–525 https://doi.org/10.1002/gcc.20761

  8. He X, Pang Z, Zhang X, Lan T, Chen H, Chen M, Yang H, Huang J, Chen Y, Zhang Z, Jing W, Peng R, Zhang H (2018) Consistent Amplification of FRS2 and MDM2 in Low-grade Osteosarcoma: A Genetic Study of 22 Cases With Clinicopathologic Analysis. Am J Surg Pathol 42:1143–1155. https://doi.org/10.1097/pas.0000000000001125

    Article  PubMed  Google Scholar 

  9. Miquelestorena-Standley E, Jourdan ML, Collin C, Bouvier C, Larousserie F, Aubert S, Gomez-Brouchet A, Guinebretière JM, Tallegas M, Brulin B, Le Nail LR, Tallet A, Le Loarer F, Massiere J, Galant C, de Pinieux G (2020) Effect of decalcification protocols on immunohistochemistry and molecular analyses of bone samples. Mod Pathol 33:1505–1517. https://doi.org/10.1038/s41379-020-0503-6

    Article  CAS  PubMed  Google Scholar 

  10. Mueller C, Gambarotti M, Benini S, Picci P, Righi A, Stevanin M, Hombach-Klonisch S, Henderson D, Liotta L, Espina V (2019) Unlocking bone for proteomic analysis and FISH. Lab Invest 99:708–721. https://doi.org/10.1038/s41374-018-0168-7

    Article  CAS  PubMed  Google Scholar 

  11. Limbach AL, Lingen MW, McElherne J, Mashek H, Fitzpatrick C, Hyjek E, Mostofi R, Cipriani NA (2020) The Utility of MDM2 and CDK4 Immunohistochemistry and MDM2 FISH in Craniofacial Osteosarcoma. Head Neck Pathol 14:889–898. https://doi.org/10.1007/s12105-020-01139-x

    Article  PubMed  PubMed Central  Google Scholar 

  12. Yoshida A, Ushiku T, Motoi T, Shibata T, Beppu Y, Fukayama M, Tsuda H (2010) Immunohistochemical analysis of MDM2 and CDK4 distinguishes low-grade osteosarcoma from benign mimics. Mod Pathol 23:1279–1288. https://doi.org/10.1038/modpathol.2010.124

    Article  CAS  PubMed  Google Scholar 

  13. Wang F, Flanagan J, Su N, Wang LC, Bui S, Nielson A, Wu X, Vo HT, Ma XJ, Luo Y (2012) RNAscope: a novel in situ RNA analysis platform for formalin-fixed, paraffin-embedded tissues. J Mol Diagn 14:22–29. https://doi.org/10.1016/j.jmoldx.2011.08.002

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Kulkarni AS, Wojcik JB, Chougule A, Arora K, Chittampalli Y, Kurzawa P, Mullen JT, Chebib I, Nielsen GP, Rivera MN, Ting DT, Deshpande V (2019) MDM2 RNA In Situ Hybridization for the Diagnosis of Atypical Lipomatous Tumor: A Study Evaluating DNA, RNA, and Protein Expression. Am J Surg Pathol 43:446–454. https://doi.org/10.1097/pas.0000000000001199

    Article  PubMed  Google Scholar 

  15. Hung YP, Michal M, Dubuc AM, Rosenberg AE, Nielsen GP (2020) Dysplastic lipoma: potential diagnostic pitfall of using MDM2 RNA in situ hybridization to distinguish between lipoma and atypical lipomatous tumor. Hum Pathol 101:53–57. https://doi.org/10.1016/j.humpath.2020.05.004

    Article  CAS  PubMed  Google Scholar 

  16. Zhang H, Bu H, Chen H, Wei B, Liu W, Guo J, Li F, Liao D, Tang Y, Zhang Z (2008) Comparison of immunohistochemical markers in the differential diagnosis of adrenocortical tumors: immunohistochemical analysis of adrenocortical tumors. Appl Immunohistochem Mol Morphol 16:32–39. https://doi.org/10.1097/PAI.0b013e318032cf56

    Article  CAS  PubMed  Google Scholar 

  17. Zhang H, Erickson-Johnson M, Wang X, Oliveira JL, Nascimento AG, Sim FH, Wenger DE, Zamolyi RQ, Pannain VL, Oliveira AM (2010) Molecular testing for lipomatous tumors: critical analysis and test recommendations based on the analysis of 405 extremity-based tumors. Am J Surg Pathol 34:1304–1311. https://doi.org/10.1097/PAS.0b013e3181e92d0b

    Article  PubMed  Google Scholar 

  18. Weaver J, Downs-Kelly E, Goldblum JR, Turner S, Kulkarni S, Tubbs RR, Rubin BP, Skacel M (2008) Fluorescence in situ hybridization for MDM2 gene amplification as a diagnostic tool in lipomatous neoplasms. Mod Pathol 21:943–949. https://doi.org/10.1038/modpathol.2008.84

    Article  CAS  PubMed  Google Scholar 

  19. Touqan N, Diggle CP, Verghese ET, Perry S, Horgan K, Merchant W, Anwar R, Markham AF, Carr IM, Achuthan R (2013) An observational study on the expression levels of MDM2 and MDMX proteins, and associated effects on P53 in a series of human liposarcomas. BMC Clin Pathol 13:32. https://doi.org/10.1186/1472-6890-13-32

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Chen CY, Zhang HZ, Jiang ZM, Zhou J, Chen J, Liu L (2016) Value of MDM2, CDK4 and SATB2 immunohistochemistry in histologic diagnosis of low-grade osteosarcoma. Zhonghua Bing Li Xue Za Zhi 45:387–392. https://doi.org/10.3760/cma.j.issn.0529-5807.2016.06.007

    Article  CAS  PubMed  Google Scholar 

  21. Yoshida A, Ushiku T, Motoi T, Beppu Y, Fukayama M, Tsuda H, Shibata T (2012) MDM2 and CDK4 immunohistochemical coexpression in high-grade osteosarcoma: correlation with a dedifferentiated subtype. Am J Surg Pathol 36:423–431. https://doi.org/10.1097/PAS.0b013e31824230d0

    Article  PubMed  Google Scholar 

  22. Anderson CM, Zhang B, Miller M, Butko E, Wu X, Laver T, Kernag C, Kim J, Luo Y, Lamparski H, Park E, Su N, Ma XJ (2016) Fully Automated RNAscope In Situ Hybridization Assays for Formalin-Fixed Paraffin-Embedded Cells and Tissues. J Cell Biochem 117:2201–2208. https://doi.org/10.1002/jcb.25606

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Choi SE, Hong SW, Yoon SO (2015) Proposal of an appropriate decalcification method of bone marrow biopsy specimens in the era of expanding genetic molecular study. J Pathol Transl Med 49:236–242. https://doi.org/10.4132/jptm.2015.03.16

    Article  PubMed  PubMed Central  Google Scholar 

  24. Flørenes VA, Maelandsmo GM, Forus A, Andreassen A, Myklebost O, Fodstad O (1994) MDM2 gene amplification and transcript levels in human sarcomas: relationship to TP53 gene status. J Natl Cancer Inst 86:1297–1302. https://doi.org/10.1093/jnci/86.17.1297

    Article  PubMed  Google Scholar 

  25. Jiao YF, Nakamura S, Sugai T, Habano W, Uesugi N, Oikawa M, Sato T (2002) p53 gene mutation and MDM2 overexpression in a case of primary malignant fibrous histiocytoma of the jejunum. APMIS 110:165–171. https://doi.org/10.1034/j.1600-0463.2002.100207.x

    Article  CAS  PubMed  Google Scholar 

  26. Meltzer PS (1994) MDM2 and p53: a question of balance. J Natl Cancer Inst 86:1265–1266. https://doi.org/10.1093/jnci/86.17.1265

    Article  CAS  PubMed  Google Scholar 

  27. Peng Y, Chen L, Li C, Lu W, Agrawal S, Chen J (2001) Stabilization of the MDM2 oncoprotein by mutant p53. J Biol Chem 276:6874–6878. https://doi.org/10.1074/jbc.C000781200

    Article  CAS  PubMed  Google Scholar 

  28. Lianes P, Orlow I, Zhang ZF, Oliva MR, Sarkis AS, Reuter VE, Cordon-Cardo C (1994) Altered patterns of MDM2 and TP53 expression in human bladder cancer. J Natl Cancer Inst 86:1325–1330. https://doi.org/10.1093/jnci/86.17.1325

    Article  CAS  PubMed  Google Scholar 

  29. Bouska A, Lushnikova T, Plaza S, Eischen CM (2008) MDM2 promotes genetic instability and transformation independent of p53. Mol Cell Biol 28:4862–4874. https://doi.org/10.1128/mcb.01584-07

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Pan Y, Haines DS (2000) Identification of a tumor-derived p53 mutant with novel transactivating selectivity. Oncogene 19:3095–3100. https://doi.org/10.1038/sj.onc.1203663

    Article  CAS  PubMed  Google Scholar 

  31. Simons A, Schepens M, Jeuken J, Sprenger S, van de Zande G, Bjerkehagen B, Forus A, Weibolt V, Molenaar I, van den Berg E, Myklebost O, Bridge J, van Kessel AG, Suijkerbuijk R (2000) Frequent loss of 9p21 (p16(INK4A)) and other genomic imbalances in human malignant fibrous histiocytoma. Cancer Genet Cytogenet 118:89–98. https://doi.org/10.1016/s0165-4608(99)00178-8

    Article  CAS  PubMed  Google Scholar 

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Funding

This work was supported by the National Natural Science Foundation of China (No. 81972520) and the 135 Project for Disciplines of Excellence–Clinical Research Incubation Project, West China Hospital, Sichuan University (No. 2018HXFH011).

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  1. Chen Chen and Xin He these authors contributed equally to this work.

Authors and Affiliations

  1. Department of Pathology, West China Hospital, Sichuan University, GuoXueXiang 37, Chengdu, 610041, Sichuan, China

    Chen Chen, Xin He, Min Chen, Tianhai Du, Weiji Qin, Wenyi Jing & Hongying Zhang

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Chen Chen and Xin He contributed equally to this work. Chen Chen collected the clinicopathological data, performed the histopathological examinations, molecular detection and prepared the manuscript. Xin He performed the histopathological examinations, analyzed the molecular data and prepared the manuscript. Min Chen, Tianhai Du and Weiji Qin helped molecular experiments. Wenyi Jing helped data review. Hongying Zhang was responsible for the diagnosis, study design and the manuscript revision.

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Imaging findings of LGOSs

a. X-ray of POS (case 7) showed multiple mixed high-density nodules in the posterior portion of the left lower segment of femur. The adjacent bone cortex was thickened, and periosteal reaction can be seen. b. MRI of POS (case 7) showed mixed signal mass in the posterior muscle space of the lower segment of the left thigh. The cortex of adjacent femur bone was thickened and coarse. c. CT of LGCOS (case 2) demonstrated an expansive change, with thinning of bone cortex and clear boundary. d. MRI of dedifferentiated POS (case 17) showed a mass with mixed signals in the marrow cavity of right middle upper arm. The adjacent bone cortex was thickened and patchy signal in the soft tissues can be seen. (PNG 339 kb)

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Chen, C., He, X., Chen, M. et al. Diagnostic value of MDM2 RNA in situ hybridization for low-grade osteosarcoma: Consistency comparison of RNA in situ hybridization, fluorescence in situ hybridization, and immunohistochemistry. Virchows Arch 482, 999–1010 (2023). https://doi.org/10.1007/s00428-023-03530-9

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