Ewing Sarkoma ile Primitif Nöroektodermal Tümör Hücre Hatlarında Agresiflik Paterninin Karşılaştırılması

GİRİŞ VE AMAÇ: Ewing Sarkoma (EWS), nöral orijinli, andiferansiye özellikte primer malign kemik tümörüdür. Primitif Nöroektodermal Tümör’ler (PNET), kötü diferansiye ve prognozları kötü seyreden malign lezyonlardır. PNET’ler morfolojik ve histolojik olarak kemiğe ait gelişen EWS’den ayırt edilememektedir. Metastaz özellik gösteren ve farklı tedavi gerektiren PNET/EWS’nin birbirinden ayrılması tanıda büyük önem arz etmektedir. Çalışmamızda; EWS (TC71, TC106, CHLA99) ve PNET (CHLA32, CHLA10, CHLA9) hücre hatlarında agresiflik paternlerini karşılaştırmayı ve hücrelerin metastaz durumunu belirlemek için epitelyalmezenkimal geçiş (EMT) belirteçlerindeki değişimleri incelemeyi amaçladık. YÖNTEM VE GEREÇLER: Children’s Oncology Group’tan temin edilen EWS ve PNET hücre hatlarının ikilenme süreleri belirlendi. Hücrelerde fonksiyonel çalışmalar kapsamında migrasyon kapasitelerinin tespiti için scratch assay yapıldı. EWS ve PNET hücrelerinde EMT belirteçleri Real-Time PZR ile belirlendi. BULGULAR: Hücre hatlarında ikilenme süresinin sırasıyla; EWS’de TC71, TC106, CHLA99, PNET’te CHLA32, CHLA9 ve CHLA10 olduğu belirlendi. Hücrelerin migrasyon kapasitelerinde ikilenme süreleri ile benzerlik saptandı. Metastatik belirteçlerden; SNAIL1, SNAIL 2, ZEB1, ZEB2 ve Vimentin ifade düzeylerinin TC-71’de TC-106 ve CHLA-99’a göre daha yüksek olduğu, NCad ifadesinin ise CHLA99’da en yüksek olduğu saptandı. PNET hücre hatlarından CHLA32’de ZEB1, ZEB2, SNAIL1 SNAIL 2, NCad ve Fibronektin ifade düzeylerinin CHLA-9 ve CHLA-10’a göre daha yüksek olduğu, ECad’ın en fazla CHLA-10'da, Vimentin için sırasıyla CHLA-9 CHLA-32 ve CHLA-10 olduğu bulundu. TARTIŞMA VE SONUÇ: Çalışma sonucunda, EWS’de agresiflik düzeyleri sırayla azalarak, TC71, TC106 CHLA99, PNET’te CHLA32, CHLA10, CHLA9 hücreleri olduğu saptandı. Hücrelerin agresiflik düzeylerinin aydınlatılması EWS ve PNET’in metastaz tanısında rol alabilecek biyobelirteçlerin belirlenmesi, erken tanı ve yeni tedavi yöntemlerinin geliştirilmesinde önem arz etmektedir.

Comparison of Aggressive Pattern in Ewing Sarcoma and Primitive Neuroectodermal Tumor Cell Lines

INTRODUCTION: Ewing Sarcoma (EWS) is a primary malignant bone tumor of neural origin with undifferentiated characteristics. Primitive Neuroectodermal Tumors (PNET) are malignant lesions with poorly differentiated and poor prognosis. PNETs cannot be morphologically and histologically distinguishable from EWS of the bone.In this study,we aimed to compare the patterns of aggression in EWS and PNET cell lines and to investigate the changes in epithelial-mesenchymal transition (EMT) markers to determine the metastasis status of the cells. METHODS: The doubling times of EWS and PNET cell lines obtained from Children's Oncology Group were determined. Scratch assay was performed to determine the migration capacity of cells within the scope of functional studies.EMT markers in EWS and PNET cells were determined by RealTime PCR. RESULTS: The doubling time in cell lines was TC71, TC106, CHLA99 in EWS, CHLA32, CHLA9 and CHLA10 in PNET.The migration capacity of the cells was similar to the doubling time. SNAIL1, SNAIL 2, ZEB1, ZEB2 and Vimentin expression levels were higher in TC71 than TC106 and CHLA99, and NCad expression was highest in CHLA99.The expression levels of ZEB1, ZEB2, SNAIL1, SNAIL 2, NCad and Fibronectin were higher in CHLA32 compared to CHLA9 and CHLA10 in PNET.ECad was found to be at most CHLA10 and CHLA9 for Vimentin. DISCUSSION AND CONCLUSION: The most aggressive and least aggressive cells were TC71, TC106 CHLA99 in EWS, CHLA32, CHLA10, CHLA9 in PNET, respectively. In the diagnosis of EWS and PNET metastasis, early diagnosis, and the development of new treatment modalities to clarify the level of aggression of the cells are important.

PDF

___

1. Whelan J, McTiernan A, Cooper N, Wong YK, Francis M, Vernon S, et al. Incidence and survival of malignant bone sarcomas in England 1979-2007. Int J Cancer. 2012;131(4):E508-17.
2. Delattre O, Zucman J, Melot T, Garau XS, Zucker JM, Lenoir GM, et al. The Ewing family of tumors--a subgroup of small-round-cell tumors defined by specific chimeric transcripts. N Engl J Med. 1994;331(5):294-9.
3. Prasad AN. Supratentorial PNET in a young child. Indian J Pediatr. 2011;78(5):613-5.
4. Smoll NR. Relative survival of childhood and adult medulloblastomas and primitive neuroectodermal tumors (PNETs). Cancer. 2012;118(5):1313-22.
5. Shi L, Guo Z, Wu X. Primary pulmonary primitive neuroectodermal tumor metastasis to the pancreas: a rare case with seven-year follow-up. Diagn Pathol. 2013;8:51.
6. Picarsic J, Reyes-Múgica M. Phenotype and immunophenotype of the most common pediatric tumors. Appl Immunohistochem Mol Morphol. 2015;23(5):313-26.
7. Pappo AS, Douglass EC, Meyer WH, Marina N, Parham DM. Use of HBA 71 and anti-beta 2- microglobulin to distinguish peripheral neuroepithelioma from neuroblastoma. Hum Pathol. 1993;24(8):880-5.
8. Vargo MM. Brain Tumors and Metastases. Phys Med Rehabil Clin N Am. 2017;28(1):115-41.
9. Thiery JP. Epithelial-mesenchymal transitions in tumour progression. Nat Rev Cancer. 2002;2(6):442- 54.
10. Poste G, Fidler IJ. The pathogenesis of cancer metastasis. Nature. 1980;283(5743):139-46.
11. Pouysségur J, Dayan F, Mazure NM. Hypoxia signalling in cancer and approaches to enforce tumour regression. Nature. 2006;441(7092):437-43.
12. Teicher BA, Bagley RG, Rouleau C, Kruger A, Ren Y, Kurtzberg L. Characteristics of human Ewing/PNET sarcoma models. Ann Saudi Med. 2011;31(2):174-82.
13. Suer I, Karatas OF, Yuceturk B, Yilmaz M, Guven G, Buge O, et al. Characterization of stem-like cells directly isolated from freshly resected laryngeal squamous cell carcinoma specimens. Curr Stem Cell Res Ther. 2014;9(4):347-53.
14. Casas E, Kim J, Bendesky A, Ohno-Machado L, Wolfe CJ, Yang J. Snail2 is an essential mediator of Twist1-induced epithelial mesenchymal transition and metastasis. Cancer Res. 2011;71(1):245-54.
15. Thompson LD. Small round blue cell tumors of the sinonasal tract: a differential diagnosis approach. Mod Pathol. 2017;30(s1):S1-S26.
16. Travis WD, Asamura H, Bankier AA, Beasley MB, Detterbeck F, Flieder DB, et al. The IASLC Lung Cancer Staging Project: Proposals for Coding T Categories for Subsolid Nodules and Assessment of Tumor Size in Part-Solid Tumors in the Forthcoming Eighth Edition of the TNM Classification of Lung Cancer. J Thorac Oncol. 2016;11(8):1204-23.
17. Giovannini M, Biegel JA, Serra M, Wang JY, Wei YH, Nycum L, et al. EWS-erg and EWS-Fli1 fusion transcripts in Ewing's sarcoma and primitive neuroectodermal tumors with variant translocations. J Clin Invest. 1994;94(2):489-96.
18. https://www.cccells.org/cellreqs-eft.php [
19. Thiery JP, Acloque H, Huang RY, Nieto MA. Epithelial-mesenchymal transitions in development and disease. Cell. 2009;139(5):871-90.
20. Sarrió D, Rodriguez-Pinilla SM, Hardisson D, Cano A, Moreno-Bueno G, Palacios J. Epithelialmesenchymal transition in breast cancer relates to the basal-like phenotype. Cancer Res. 2008;68(4):989-97.
21. Kumarswamy R, Mudduluru G, Ceppi P, Muppala S, Kozlowski M, Niklinski J, et al. MicroRNA-30a inhibits epithelial-to-mesenchymal transition by targeting Snai1 and is downregulated in non-small cell lung cancer. Int J Cancer. 2012;130(9):2044-53.
22. Sannino G, Marchetto A, Kirchner T, Grünewald TGP. Epithelial-to-Mesenchymal and Mesenchymal-to-Epithelial Transition in Mesenchymal Tumors: A Paradox in Sarcomas? Cancer Res. 2017;77(17):4556-61.
23. Kahlert UD, Joseph JV, Kruyt FAE. EMT- and MET-related processes in nonepithelial tumors: importance for disease progression, prognosis, and therapeutic opportunities. Mol Oncol. 2017;11(7):860- 77.
24. Wiles ET, Bell R, Thomas D, Beckerle M, Lessnick SL. ZEB2 Represses the Epithelial Phenotype and Facilitates Metastasis in Ewing Sarcoma. Genes Cancer. 2013;4(11-12):486-500.
25. Schuetz AN, Rubin BP, Goldblum JR, Shehata B, Weiss SW, Liu W, et al. Intercellular junctions in Ewing sarcoma/primitive neuroectodermal tumor: additional evidence of epithelial differentiation. Mod Pathol. 2005;18(11):1403-10.