Interaction between p53 and MDM2 in Human Lung Cancer Cells

Czech version

Authors: S. Rybárová ¹; I. Hodorová ¹; J. Vecanová ¹; J. Muri ²; J. Mihalik ¹
Authors‘ workplace: Ústav anatómie, LF UPJŠ, Košice, Slovenská republika ¹; Národný ústav tuberkulózy, pľúcnych chorôb a hrudníkovej chirurgie, Vyšné Hágy, Slovenská Republika ²
Published in: Klin Onkol 2014; 27(1): 33-37
Category: Original Articles

Overview

Background:
The oncoprotein p53 protein induces cell growth arrest (apoptosis) in response to endo‑ or exogenous stimuli. Mutation of TP53 (gene encoding the p53 protein) is common in human malignancies and alters the conformation of p53. The result is a more stable protein which accumulates in nuclei of tumor cells with loss of function. Mutant p53 is stabilized, and it is possible to detect this form very clearly by immunohistochemistry (IHC). Expression of the MDM2 protein is used as a potential marker of p53 function. P53 levels in normal cells are highly determined by the MDM2 protein (murine double minute‑ 2) – mediated degradation of p53. MDM2 overexpression represents at least one mechanism by which p53 function can be abrogated during tumorigenesis.

Material and Methods:
Lung carcinoma samples were obtained from patients, who underwent radical resection (lobectomy or pulmonectomy and lymphadectomy). Pathological diagnosis was based on the WHO criteria. In our study, we investigated the expression of p53 and MDM2 protein that might improve IHC as a marker for p53 status. Proteins were IHC detected in 136 samples of primary lung carcinoma. Immunostaining results of p53 positive samples were compared to IHC expression of MDM2 positive and MDM2 negative samples.

Results:
Strong brown nuclear staining was visible in p53 and MDM2 positive cells. The most p53 positive cases were samples of squamocellular carcinoma (55%), then samples of large cell carcinoma (53%) and 26% adenocarcinoma samples showed the p53 immunoreactivity. No one sample of different types was p53 positive. When we compared the p53 expression and grade of tumor, we found that the p53 expression increased with the grade of tumor. For statistical evaluation, the chi‑ square test was used. The relationship between p53 expression and type of tumor, also the p53 expression and grade of tumor was statistically significant (p = 0.000425; p = 0.00157). Regarding p53 and MDM2 expression, only nine samples (7%) were simultaneously p53 and MDM2 positive. In 46 (34%) cases, elevation of p53 was combined with MDM2 negative expression. Other tumor samples were either negative for both proteins (71/ 52%), or p53 negative and MDM2 positive in 10 (7%) tumor samples.

Conclusion:
Absence of p53 staining in most studies indicates absence of p53 mutation, and on the contrary, positive expression of p53 is a sign of p53 mutations with loss of function. In our study, 34% of cases with extensively high level of p53 without increased level of MDM2 were identified. We suppose that these are tumors with inactivating mutations that stabilize p53. On the other hand, tumors with high level of stabilized wild‑type p53 protein and simultaneously with increased MDM2 staining (9 samples/7%) represent group with functional p53. In this group of patients, we could expect better prognosis with regard to function of p53 protein.

Key words:
oncoprotein p53 – MDM2 protein – lung cancer – immunohistochemistry

This work was supported by the Grant Agency of the Ministry of Education of the Slovak Republic VEGA:
1/0224/12; 1/0925/11; 1/0928/11.

The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study.

The Editorial Board declares that the manuscript met the ICMJE “uniform requirements” for biomedical papers.

Submitted:
13. 6. 2013

Accepted:
23. 7. 2013

Sources

1. Bates S, Vousden KH. p53 in signaling checkpoint arrest or apoptosis. Curr Opin Genet Dev 1996; 6(1): 12– 18.

2. Li FP, Fraumeni JF Jr. Soft‑ tissue sarcomas, breast cancer, and other neoplasms: a familial syndrome? Ann Intern Med 1969; 71(4): 747– 752.

3. Foretová L, Štěrba J, Opletal P et al. Li‑ Fraumeni syndrom – návrh komplexní preventivní péče o nosiče TP53 mutace s použitím celotělové magnetické rezonance. Klin Onkol 2012; 25 (Suppl 1): 49– 54.

4. Mao L, Hruban RH, Boyle JO et al. Detection of oncogene mutations in sputum precedes diagnosis of lung cancer. Cancer Res 1994; 54(7): 1634– 1637.

5. Han H, Landreneau RJ, Santucci TS et al. Prognostic value of immunohistochemical expressions of p53, HER‑ 2/ neu, and bcl‑ 2 in stage I non‑small‑cell lung cancer. Hum Pathol 2002; 33(1): 105– 110.

6. Bouska A, Eischen CM. Mdm2 affects genome stability independent of p53. Cancer Res 2009; 69(5): 1697– 1701.

7. Mantovani A. Molecular pathways linking inflammation and cancer. Curr Mol Med 2010; 10(4): 369– 373.

8. Eischen CM, Lozano G. p53 and MDM2: antagonists or partners in crime? Cancer Cell 2009; 15(3): 161– 162.

9. Marine JC, Lozano G. Mdm2– mediated ubiquitylation: p53 and beyond. Cell Death Differ 2010; 17(1): 93– 102.

10. Clegg HV, Itahana K, Zhang Y. Unlocking the Mdm2– p53 loop: ubiquitin is the key. Cell Cycle 2008; 7(3): 287– 292.

11. Beasley MB, Brambilla E, Travis WD. The 2004 World Health Organization classification of lung tumors. Semin Roentgenol 2005; 40(2): 90– 97.

12. IASLC Staging Manual in Thoracic Oncology. Orange Park, FL, USA: Editorial Rx Press 2009: 129– 140.

13. Bergqvist M, Brattstrom D, Gullbo J et al. p53 Status and its in vitro relationship to radiosensitivity and chemosensitivity in lung cancer. Anticancer Res 2003; 23(2B): 1207– 1212.

14. Wang T, Xu J, Zhong NS. Relationship between the acquired multi‑drug resistance of human large cell lung cancer cell line NCI‑ H460 by cisplatin selection and p53 mutation. Zhonghua Jie He He Hu Xi Za Zhi 2005; 28(2): 102– 107.

15. Brattström D, Bergqvist M, Lamberg K et al. Complete sequence of p53 gene in 20 patients with lung cancer: comparison with chemosensitivity and immunohistochemistry. Med Oncol 1998; 15(4): 255– 261.

16. Blandino G, Levine AJ, Oren M. Mutant p53 gain of function: differential effects of different p53 mutants on resistance of cultured cells to chemotherapy. Oncogene 1999; 18(2): 477– 485.

17. Lai SL, Perng RP, Hwang J. p53 Gene status modulates the chemosensitivity of non‑small cell lung cancer cells. J Biomed Sci 2000; 7(1): 64– 70.

18. Inoue A, Narumi K, Matsubara N et al. Administration of wild‑type p53 adenoviral vector synergistically enhances the cytotoxicity of anti‑cancer drugs in human lung cancer cells irrespective of the status of p53 gene. Cancer Lett 2000; 157(1): 105– 112.

19. Wang Y, Blandino G, Oren M et al. Induced p53 expression in lung cancer cell line promotes cell senescence and differentially modifies the cytotoxicity of anti‑cancer drugs. Oncogene 1998; 17(15): 1923– 1930.

20. Ling YH, Zou Y, Perez‑ Soler R. Induction of senescence‑like phenotype and loss of paclitaxel sensitivity after wild‑type p53 gene transfection of p53– null human non‑small cell lung cancer H358 cells. Anticancer Res 2000; 20(2A): 693– 702.

21. He Y, Fan SZ, Jiang YG et al. Effect of p73 gene on chemosensitivity of human lung adenocarcinoma cells H1299. Ai Zheng 2004; 23(6): 645– 649.

22. Mori T, Okamoto H, Takahashi N et al. Aberrant overexpression of 53BP2 mRNA in lung cancer cell lines. FEBS Lett 2000; 465(2– 3): 124– 128.

23. Ji D, Deeds SL, Weinstein EJ. A screen of shRNAs targeting tumor suppressor genes to identify factors involved in A549 paclitaxel sensitivity. Oncol Rep 2007; 18(6): 1499– 1505.

24. Vogt U, Zaczek A, Klinke F et al. p53 Gene status in relation to ex vivo chemosensitivity of non‑small cell lung cancer. J Cancer Res Clin Oncol 2002; 128(3): 141– 147.

25. Kandioler‑ Eckersberger D, Kappel S, Mittlbock M et al. The TP53 genotype but not immunohistochemical result is predictive of response to cisplatin‑based neoadjuvant therapy in stage III non‑small cell lung cancer. J Thorac Cardiovasc Surg 1999; 117(4): 744– 750.

26. Kandioler D, Stamatis G, Eberhardt W et al. Growing clinical evidence for the interaction of the p53 genotype and response to induction chemotherapy in advanced non‑small cell lung cancer. J Thorac Cardiovasc Surg 2008; 135(5): 1036– 1041.

27. Ludovini V, Gregorc V, Pistola L et al. Vascular endothelial growth factor, p53, Rb, Bcl‑ 2 expression and response to chemotherapy in advanced non‑small cell lung cancer. Lung Cancer 2004; 46(1): 77– 85.

28. Miyatake K, Gemba K, Ueoka H et al. Prognostic significance of mutant p53 protein, P‑ glycoprotein and glutathione S‑ transferase‑ pi in patients with unresectable non‑small cell lung cancer. Anticancer Res 2003; 23(3C): 2829– 2836.

29. Brooks KR, To K, Joshi MB et al. Measurement of chemoresistance markers in patients with stage III non‑small cell lung cancer: a novel approach for patient selection. Ann Thorac Surg 2003; 76(1): 187– 193.

30. Azuma K, Komohara Y, Sasada T et al. Excision repair cross‑ complementation group 1 predicts progression‑free and overall survival in non‑small cell lung cancer patients treated with platinum‑based chemotherapy. Cancer Sci 2007; 98(9): 1336– 1343.

31. Gajra A, Tatum AH, Newman N et al. The predictive value of neuroendocrine markers and p53 for response to chemotherapy and survival in patients with advanced non‑small cell lung cancer. Lung Cancer 2002; 36(2): 159– 165.

32. Berrieman HK, Cawkwell L, O’Kane SL et al. Hsp27 may allow prediction of the response to single‑agent vinorelbine chemotherapy in non‑small cell lung cancer. Oncol Rep 2006; 15(1): 283– 286.

33. Gregorc V, Darwish S, Ludovini V et al. The clinical relevance of Bcl‑ 2, Rb and p53 expression in advanced non‑small cell lung cancer. Lung Cancer 2003; 42(3): 275– 281.

34. van de Vaart PJ, Belderbos J, de Jong D et al. DNA‑ adduct levels as a predictor of outcome for NSCLC patients receiving daily cisplatin and radiotherapy. Int J Cancer 2000; 89(2): 160– 166.