P53 is the best known tumor suppressor gene. If p53 is mutated, the ability of the cell to sense and repair DNA defects is lost. Failure of this mechanism increases the risk of malignant transformation and tumorigenesis. P53 overexpression is implicated in many carcinomas. P53 alterations appear to be frequent in bone and soft tissue sarcoma and have a strong negative impact on survival in various subtypes of sarcoma like Ewings sarcoma, synovial sarcoma, and myxoid liposarcoma. There is also evidence in the literature that p53 may be implicated in bone giant cell tumor behavior.
The goal of this pilot retrospective study was to detect p53 mutation in giant cell tumor of bone and correlate it with clinical outcome. We analyzed the presence of p53 mutation in 39 patients with giant cell tumor of bone by means of immunohistochemical staining; 8 tumors expressed mutated p53 protein. Seven of them recurred locally (P<.001) and 2 metastasized to the lung (P<.05). In multivariate analysis/subgroup analysis, local recurrence was still strongly correlated, while metastasis had a weaker correlation. Our findings suggest that p53 mutation in giant cell tumor of bone can be useful in predicting tumor behavior, especially in regard to local recurrence.
Limitations of this study include the retrospective data collection, the limited number of patients, and the multifactorial nature of the disease; tumor grade, surgical margins, use of adjuvant therapy, and thoroughness of excision may influence the therapeutic outcome. Despite these limitations, this correlation should be further investigated with larger clinical studies. P53 may be used as a marker for the biologic behavior of giant cell tumor of bone.
P53 is the best known tumor suppressor gene. It is considered the guardian of the genome and is the key factor in the pathway of programmed cell death. If a mistake occurs during the replication of DNA during cell division, the cell cycle is halted until the mistake is repaired. If the damage is too extensive to repair, the cell self-destructs. If p53 is mutated, the ability of the cell to sense and repair DNA defects is lost. Failure of this mechanism increases the risk of malignant transformation and tumorigenesis.1,2 P53 alterations have been found in a variety of tumors, with approximately 33% prevalence in sarcomas.3,4
Giant cell tumors of bone are rare, usually benign connective tissue neoplasms characterized by localized bone destruction.5 They comprise osteoclast-like giant cells, stromal cells, and CD68-positive monocytes.6,7 The stromal cells are thought to be the neoplastic component of the tumor, as they can be propagated in culture and stain positive for the proliferation marker, Ki67.7,8
Giant cell tumors of bone have an unpredictable biologic behavior. Although regarded as a benign neoplasm, they can be locally aggressive, and 1% to 3% give metastases to the lungs, while 3% to 5% appear as giant cell sarcomas. Giant cell tumor usually grows slowly but sometimes grows rapidly, destroying the cortex and invading surrounding tissues.9,10
Bibliographic data suggest that p53 is implicated in bone giant cell tumor behavior.11-14 Based on our clinical observation that p53 mutation was related with worse clinical outcome, we designed a retrospective pilot study looking at p53 mutation in giant cell tumors of bone and correlated these finding with the biologic course and prognosis of the tumor. More specifically, our hypothesis was that P53 mutation is related with more aggressive behavior (higher incidence of local recurrence and/or pulmonary metastasis).
Materials and Methods
Thirty-nine cases of giant cell tumor of bone were surgically treated over the past 11 years (minimum follow-up, 2 years) at 2 orthopedic oncology departments. Twenty-two men and 17 women had a mean age of 28.8 years (range, 16-58 years).
Preoperative evaluation included precision imaging techniques (plain radiographs, computed tomography [CT], magnetic resonance imaging [MRI], CT of the chest, full Tc99m body scan) plus biopsy (fine needle, incisional, or intraoperative-frozen section biopsy). According to the radiographic grading system of Campanacci et al,5 5 tumors were classified as grade I, 19 as grade II and 15 as grade III (Table 1).
The operation type according to Ennekings15 criteria was intralesional excision in 25 cases, marginal excision in 3, and wide excision in 11. We aimed for wide surgical margins for all grade III tumors, except in a few cases where this would lead to severe morbidity, while we performed thorough curettage plus cement as an adjuvant in the majority of grade I and II cases.
Diagnosis was confirmed in all cases by the same pathologist (I.I.); tissue specimens from all cases were fixed in 10% neutral-buffered formalin and embedded in paraffin. P53 overexpression was detected by immunohistochemical staining. This is an indirect test for mutation and is based on the fact that mutant p53 proteins have a longer half-life than normal p53 proteins4,13,16,17; therefore, p53 mutation leads to cellular accumulation and overexpression in immunochemistry. For this examination, each section was deparaffinized with xylene for 30 minutes, washed in phosphate-buffered saline 0.85%, and treated with 0.3% hydrogen peroxide for 30 minutes.
To investigate for any association between categorical variables, 2×2 contingency tables were constructed and Fishers exact test was calculated (univariate analysis). Multivariate analysis (logistic regression) was performed to eliminate the effect of the confounding variables (grade, excision type). Finally, subgroup analysis was performed in regard to the operation type: (1) intralesional/marginal excision vs (2) wide excision (chi-square test). All reported P values are based on 2-sided tests and compared to a significance level of 5%. Data were analyzed using SPSS version 17.0 (SPSS Inc, Chicago, Illinois).
Four grade II and 4 grade III tumors recurred locally, while 2 grade III tumors metastasized to the lung. The mean time from initial operation to lung metastasis was 6 months.
P53 was overexpressed by stromal cells and/or giant cells in 8 of the 39 cases (Figures 1, 2). From these 8 cases with p53 overexpression, 2 metastasized to the lung and 7 recurred locally, while in 31 cases with normal p53 status, only 1 recurred locally and no lung metastasis occurred (Table 2).
| || |
Figure 1: Giant cell tumor nuclear positivity for p53 mainly in stromal cells (hematoxylin-eosin ×200). Figure 2: Nuclear positivity for p53 in both stromal and giant cells (hematoxylin-eosin ×400).
Based on the results of the univariate analysis, significant correlation was found between p53 mutation local recurrence (Fishers exact test, P<.001) and metastasis (Fishers exact test, P=.038). Correlation with grade and excision was weak (Fishers exact test, P=.221 and .188, respectively). Multivariate analysis (binary logistic regression analysis, stepwise method including all the variables) showed that local recurrence again was strongly correlated with the presence of p53 mutation (Wald 13.138 Sig P=.000 Exp[B] 210.000). However, the resulting model was weak (Hosmer and Lemeshow test, P=1.000). We used the logistic regression enter method individually for recurrence and the result was reproduced (Wald Sig P=.000). Metastasis was found to correlate less strongly with p53 in the binary logistic regression analysis stepwise method model, but it was important if local recurrence was removed from the variables (metastasis, P=.002; excision, P=.074; grade, P=.703). When we used logistic regression enter method individually for metastasis, it was found to be of low significance (Wald Sig P=.0999). The local recurrence and metastasis model was also useless, with a low Hosmer and Lemeshow test significance (P=1.000), and both the variables in the equation were found to be of low value to the model (local recurrence Wald Sig. P=.997; metastasis P=.0999). Therefore, the presence of P53 mutation correlated strongly with the possibility of local recurrence of the tumor. A weaker correlation existed with the presence of metastases. Unfortunately, a model that combined all of the variables in 1 prediction model could not be created in our series.
For subgroup analysis, we divided our sample into 2 groups depending on the surgical operation (non-wide vs wide excision), since excision type is the most significant variable affecting the outcome; grade is not considered by some authorities as significant factor for the surgical outcome; however, this division also correlated with tumor grade, since in group 1, 96.4% of cases were grade I to II, and all cases in group 2 were grade III. P53 mutation seemed to correlate strongly with the possibility of local recurrence both in Ennekings excision type I or II (P=.000) and in type III (wide excision) (P=.003). The correlation between p53 mutation and metastasis in cases of grade III and wide excision was weak but still existed. In contrast, cases with grade I and II and intralesional excision seemed to have no risk of metastasis (0% in our series).
P53 is the best known tumor suppressor gene. If p53 is mutated, the ability of the cell to sense and repair DNA defects is lost. Failure of this mechanism increases the risk of malignant transformation and tumorigenesis.1,2 P53 overexpression is implicated in many carcinomas, such as breast cancer,18 lung cancer,19 colorectal cancer,20 and brain tumors.21 P53 alterations appear to be frequent in bone and soft tissue sarcoma3,4,22 and have a strong negative impact on survival in various subtypes of sarcoma like Ewings sarcoma,23 synovial sarcoma,24 and myxoid liposarcoma.25
There is also evidence in the literature that p53 is implicated in bone giant cell tumor behavior. Loss of heterozygosity of the 17p (in proximity to the p53 locus) and 9p chromosomes occurred in bone giant cell tumor with pulmonary metastases.12 Another study reports that p53 inactivation by MDM2 expression may be involved in the pathogenesis of giant cell lesions of the jaw and long bones.14 In another report of 2 cases, p53 mutation was revealed by immunohistochemistry.13 Finally, Masui et al11 reported a correlation between tumor behavior and p53 expression.
Limitations of our study include the retrospective data collection, the limited number of patients, and the multifactorial nature of the disease; tumor grade, surgical margins, use of adjuvant therapy, and thoroughness of excision may influence the therapeutic outcome.
It is not clear whether tumor stage correlates with the therapeutic outcome; Campanacci et al5 found no correlation between radiographic grading and recurrence rate, while Rock26 found an increased risk for recurrence in grade III tumors. In a recent series of 384 tumors, a reduced recurrence rate was found for grade III vs stage II tumors, and this paradox was attributed to increased surgical thoroughness and wide excision.27
The type of surgery affects the outcome, since intralesional excision is associated with higher recurrence rate5,11,27; the same applies for the surgical technique and thoroughness of excision. Capanna et al28 reported a much higher recurrence rate for intralesional excision using a small cortical window compared to a thorough intralesional procedure through a window larger than half of the tumor size. Meticulous excision is important, and this may explain the nonsignificant difference in recurrence rate between primary and recurrent tumors encountered in some series.5,27
The use of adjuvant therapy, especially bone cement, has been widely accepted after intralesional treatment.10,27,29,30 We tried to overcome these confounding variables with multivariate analysis. Unfortunately, a model combining all of the variables in 1 prediction model could not be created in our series. We additionally divided our sample into 2 populations depending on the surgical operation (wide vs non-wide excision). From both statistical methods we may conclude that P53 mutation correlates strongly with the possibility of local recurrence of the tumor, while a weaker correlation exists with the presence of metastases in grade III tumors treated with wide excision.
Another limitation is that immunochemistry does not detect the wild type of p53, which has a short half-life, and nonsense mutations that lead to no expression of p53 protein; however, p53 missense mutations represent about 85% of the p53 mutations observed. Thus, these types of mutations represent the majority of cases detectable by immunohistochemistry.22,31
Our pilot study demonstrates that p53 mutation correlates with tumor aggressiveness, especially with local recurrence. Despite the limitations, this correlation should be further investigated with larger clinical studies. P53 may be used as a marker for the biologic behavior of giant cell tumor of bone.
- Wadayama B, Toguchida J, Yamaguchi T, Sasaki MS, Yamamuro T. p53 expression and its relationship to DNA alterations in bone and soft tissue sarcomas. Br J Cancer. 1993; 68(6):1134-1139.
- Levine AJ, Perry ME, Chang A, et al. The 1993 Walter Hubert Lecture: the role of the p53 tumour-suppressor gene in tumorigenesis. Br J Cancer. 1994; 69(3):409-416.
- Toguchida J, Yamaguchi T, Ritchie B, et al. Mutation spectrum of the p53 gene in bone and soft tissue sarcomas. Cancer Res. 1992; 52(22):6194-6199.
- Porter PL, Gown AM, Kramp SG, Coltrera MD. Widespread p53 overexpression in human malignant tumors. An immunohistochemical study using methacarn-fixed, embedded tissue. Am J Pathol. 1992; 140(1):145-153.
- Campanacci M, Baldini N, Boriani S, Sudanese A. Giant-cell tumor of bone. J Bone Joint Surg Am. 1987; 69(1):106-114.
- Goldring SR, Roelke MS, Petrison KK, Bhan AK. Human giant cell tumors of bone identification and characterization of cell types. J Clin Invest. 1987; 79(2):483-491.
- Wülling M, Delling G, Kaiser E. The origin of the neoplastic stromal cell in giant cell tumor of bone. Hum Pathol. 2003; 34(10):983-993.
- Ghert M, Simunovic N, Cowan RW, Colterjohn N, Singh G. Properties of the stromal cell in giant cell tumor of bone. Clin Orthop Relat Res. 2007; (459):8-13.
- Szendröi M, Kiss J, Antal I. Surgical treatment and prognostic factors in giant-cell tumor of bone. Acta Chir Orthop Traumatol Cech. 2003; 70(3):142-150.
- ODonnell RJ, Springfield DS, Motwani HK, Ready JE, Gebhardt MC, Mankin HJ. Recurrence of giant-cell tumors of the long bones after curettage and packing with cement. J Bone Joint Surg Am. 1994; 76(12):1827-1833.
- Masui F, Ushigome S, Fujii K. Giant cell tumor of bone: a clinicopathologic study of prognostic factors. Pathol Int. 1998; 48(9):723-729.
- Rao UN, Goodman M, Chung WW, Swalski P, Pal R, Finkelstein S. Molecular analysis of primary and recurrent giant cell tumors of bone. Cancer Genet Cytogenet. 2005; 158(2):126-136.
- Oda Y, Sakamoto A, Saito T, et al. Secondary malignant giant-cell tumour of bone: molecular abnormalities of p53 and H-ras gene correlated with malignant transformation. Histopathology. 2001; 39(6):629-637.
- de Souza PE, Paim JF, Carvalhais JN, Gomez RS. Immunohistochemical expression of p53, MDM2, Ki-67 and PCNA in central giant cell granuloma and giant cell tumor. J Oral Pathol Med. 1999; 28(2):54-58.
- Enneking WF. A system of staging musculoskeletal neoplasms. Clin Orthop Relat Res. 1986; (204):9-24.
- Soini Y, Vähäkangas K, Nuorva K, Kamel D, Lane DP, Pääkkö P. p53 immunohistochemistry in malignant fibrous histiocytomas and other mesenchymal tumours. J Pathol. 1992; 168(1):29-33.
- Andreassen A, Oyjord T, Hovig E, et al. p53 abnormalities in different subtypes of human sarcomas. Cancer Res. 1993; 53(3):468-471.
- Thor AD, Moore DH II, Edgerton SM, et al. Accumulation of p53 tumor suppressor gene protein: an independent marker of prognosis in breast cancers. J Natl Cancer Inst. 1992; 84(11):845-855.
- Quinlan DC, Davidson AG, Summers CL, Warden HE, Doshi HM. Accumulation of p53 protein correlates with a poor prognosis in human lung cancer. Cancer Res. 1992; 52(17):4828-4831.
- Starzynska T, Bromley M, Ghosh A, Stern PL. Prognostic significance of p53 overexpression in gastric and colorectal carcinoma. Br J Cancer. 1992; 66(3):558-562.
- Jaros E, Perry RH, Adam L, et al. Prognostic implications of p53 protein, epidermal growth factor receptor, and Ki-67 labelling in brain tumours. Br J Cancer. 1992; 66(2):373-385.
- Cordon-Cardo C, Latres E, Drobnjak M, et al. Molecular abnormalities of mdm2 and p53 genes in adult soft tissue sarcomas. Cancer Res. 1994; 54(3):794-799.
- de Alava E, Antonescu CR, Panizo A, et al. Prognostic impact of P53 status in Ewing sarcoma. Cancer. 2000; 89(4):783-792.
- Antonescu CR, Leung DH, Dudas M, et al. Alterations of cell cycle regulators in localized synovial sarcoma: A multifactorial study with prognostic implications. Am J Pathol. 2000; 156(3):977-983.
- Antonescu CR, Tschernyavsky SJ, Decuseara R, et al. Prognostic impact of P53 status, TLS-CHOP fusion transcript structure, and histological grade in myxoid liposarcoma: a molecular and clinicopathologic study of 82 cases. Clin Cancer Res. 2001; 7(12):3977-3987.
- Rock M. Curettage of giant cell tumor of bone. Factors influencing local recurrences and metastasis. Chir Organi Mov. 1990; 75(1 Suppl):204-205.
- Arbeitsgemeinschaft Knochentumoren, Becker WT, Dohle J, et al. Local recurrence of giant cell tumor of bone after intralesional treatment with and without adjuvant therapy. J Bone Joint Surg Am. 2008; 90(5):1060-1067.
- Capanna R, Fabbri N, Bettelli G. Curettage of giant cell tumor of bone. The effect of surgical technique and adjuvants on local recurrence rate. Chir Organi Mov. 1990; 75(1 Suppl):206.
- Persson BM, Wouters HW. Curettage and acrylic cementation in surgery of giant cell tumors of bone. Clin Orthop Relat Res. 1976; (120):125-133.
- Krishnan EC, Nelson C, Neff JR. Thermodynamic considerations of acrylic cement implant at the site of giant cell tumors of the bone. Med Phys. 1986; 13(2):233-239.
- Reich NC, Oren M, Levine AJ. Two distinct mechanisms regulate the levels of a cellular tumor antigen, p53. Mol Cell Biol. 1983; 3(12):2143-2150.
Drs Papanastassiou, Ioannou, Iakovidou, and Demertzis are from Metaxa Anticancer Hospital, Pireaus, Dr Papagelopoulos is from Attikon University General Hospital, Athens University Medical School, Athens, Dr Arealis is from the Orthopedic Department, Edesa General Hospital, Edesa, and Dr Mihas is from Kimi General Hospital, Kimi, Greece.
Drs Papanastassiou, Ioannou, Papagelopoulos, Arealis, Mihas, Iakovidou, and Demertzis have no relevant financial relationships to disclose.
Correspondence should be addressed to: Ioannis Papanastassiou, MD, 8413 Lucuya Way, Apt 104, Tampa, FL 33637 (firstname.lastname@example.org).