P53 gene mutations in surgical margins and primary tumor tissues of patients with squamous cell carcinoma of the head and neck

AIMS AND BACKGROUND: The frequency of p53 mutations in primary tumors, the effect of the mutations on some clinical and pathological features of head and neck squamous cell carcinoma, and the impact of p53 mutations in the surgical margins on local recurrence were determined, MATERIAL AND METHODS: We investigated the presence of p53 mutations in primary tumor samples and in the surgical margins of 34 patients with head and neck cancer using single strand conformational polymorphism and sequencing analysis. RESULTS: The p53 mutations (codons 175addAT, 175delGC, 206G --> A, and 248delC) were found in the primary tumor samples of 15 of 34 patients (44.12%) and in the surgical margins of 5 of the 15 tumors (33.33%) with p53 mutations. CONCLUSIONS: We found no statistically significant association between the presence of p53 mutations in the primary tumor, the clinical and pathological features, or outcome of head and neck squamous cell carcinoma in this study. Furthermore, the presence of p53 mutations in the surgical margins may not increase the risk of local-regional recurrence, but probably increases the risk of developing distant metastases or second primary tumors.     

Mutant p53 protein overexpression in women with ipsilateral breast tumor recurrence following lumpectomy and radiation therapy

BACKGROUND: The p53 tumor suppressor gene encodes a nuclear phosphoprotein that is thought to be important to cell cycle regulation and DNA repair and that also may regulate induction of apoptosis by ionizing radiation. Somatic p53 gene mutations occur in 30-50% of breast carcinomas and are associated with poor prognosis. Mutations in the p53 gene result in prolonged stability of the protein that can be detected by immunohistochemical techniques. In a matched case-control study of breast carcinoma patients with ipsilateral breast tumor recurrence (IBTR) following lumpectomy and radiation therapy, the authors investigated the frequency and prognostic significance of somatic p53 mutations as well as the clinical characteristics of patients with these mutations. METHODS: Between 1973 and 1995, there were 121 breast carcinoma patients with IBTR following lumpectomy and radiation therapy, and the authors identified 47 patients in whom the paraffin embedded tissue blocks from the primary breast tumors were available for further molecular analysis. Forty-seven control breast carcinoma patients from the breast carcinoma data base were individually matched to the index cases who did not have IBTR for age, treatment date, follow-up, histology, margin status, radiation dose, and adjuvant treatment. Immunohistochemistry using a monoclonal antibody to mutant p53 protein was used to determine mutant p53 protein overexpression in breast tumors and appropriately scored. RESULTS: A total of 12 of 47 tumor specimens (26%) from index patients with breast tumor relapses demonstrated mutant p53 protein overexpression, whereas only 4 of 47 specimens from controls (9%) demonstrated high mutant p53 immunoreactivity (P = 0.02). The authors found that 9 of 23 patients (39%) with early breast tumor recurrences (recurrences within 4 years of diagnosis) had overexpression of mutant p53 protein, whereas only 1 of 23 control cases (4%) had high mutant p53 protein immunoreactivity (P = 0.003). In contrast, index cases from patients with late breast tumor relapses (more than 4 years after diagnosis), which are more likely to represent de novo breast tumors, and control cases from the breast carcinoma data base without IBTR had similar levels of mutant p53 protein overexpression (P = not significant). The 10-year distant disease free survival for patients with mutant p53 protein was 48%, compared with 67% for breast carcinoma patients without detection of mutant p53 protein (P = 0. 08). The authors found that 13 of 14 primary breast tumors (93%) with mutant p53 protein overexpression were estrogen receptor negative (P = 0.01) and 11 of 14 (79%) were progesterone receptor negative (P = not significant). CONCLUSIONS: In a matched case-control study, overexpression of mutant p53 protein has prognostic significance with respect to IBTR following lumpectomy and radiation therapy. Breast tumors with p53 mutations are generally estrogen receptor negative and are associated with compromised distant disease free survival.     

Effect of human papillomavirus-16 infection on CD8+ T-cell recognition of a wild-type sequence p53264-272 peptide in patients with squamous cell carcinoma of the head and neck.

PURPOSE: Wild-type sequence (wt) p53 peptides are attractive candidates for broadly applicable cancer vaccines, currently considered primarily for patients whose tumors overexpress p53. Circumstances exist, however, where increased p53 degradation may result in appreciable presentation of p53-derived peptides, despite low p53 expression. Squamous cell carcinoma of the head and neck is associated with oncogenic human papillomavirus (HPV) subtypes, which inactivate p53 through proteasomal degradation. The criterion of p53 overexpression would exclude these individuals from wt p53-based immunotherapy. EXPERIMENTAL DESIGN: We tested the correlation of HPV infection with enhanced antigenicity of the p53 protein and postulated that removal of HPV-16(+) tumors with enhanced p53(264)-(272) peptide presentation might lead to a drop in T cells specific for this peptide in vivo. Circulating frequencies of T cells specific for the HLA A*0201:p53(264)-(272) complex were measured ex vivo using dimeric HLA:peptide complexes in 15 head and neck cancer patients before and 6 months after tumor excision. RESULTS: CD8+ T-cell recognition of HLA A*0201 restricted wt p53(264)-(272) peptide presented by HPV-16(-) squamous cell carcinoma of the head and neck lines was enhanced by HPV-16 E6 expression, sometimes exceeding that of a naturally transformed, HPV-16(+) wt p53 expressing squamous cell carcinoma of the head and neck cell line. In patients with HPV-16(-) tumors, the frequency of wt p53(264-272)-specific T cells remained largely unchanged after tumor removal. However, a significant decline in frequency of anti-p53(264-272) T cells was observed postoperatively in HPV-16(+) patients (P < 0.005). CONCLUSIONS: Recognition of HPV-associated squamous cell carcinoma of the head and neck appears associated with levels of wt p53-specific T cells and inversely with p53 expression. p53 peptides may be useful tumor antigens for squamous cell carcinoma of the head and neck immunotherapy in addition to viral gene products.     

RhoC GTPase expression as a potential marker of lymph node metastasis in squamous cell carcinomas of the head and neck.

PURPOSE: Survival rates for squamous cell carcinoma of the head and neck (SCCHN) have remained unchanged for several decades due to local tumor recurrences as well as regional and distant metastases. Recent evidence has shown that RhoC GTPase is overexpressed in stages III and IV regionally metastatic SCCHN compared with stages I and II localized disease. This study evaluated the expression of RhoC in head and neck carcinoma and investigated the prognostic use of this marker on a large cohort of previously untreated patients with SCCHN. EXPERIMENTAL DESIGN: Standard Western blot techniques were used to evaluate RhoC protein expression in nine established head and neck cancer cell lines and in normal oral epithelium. In vivo expression of RhoC in metastatic and nonmetastatic SCCHN was investigated using immunohistochemical analysis on a tissue microarray composed of 113 independent tumor samples. RhoC expression was analyzed as it related to clinical and pathologic variables of interest. RESULTS: Levels of RhoC protein were increased in the SCCHN cell lines compared with normal oral epithelium. The in vivo expression of RhoC correlated with advanced clinical stage and lymph node metastases for the entire patient cohort as well as in small primary tumors (T(1) and T(2)). CONCLUSIONS: This study is the first to examine the expression of RhoC GTPase protein in SCCHN and normal squamous epithelium. It is clear from the results that RhoC is a specific marker of lymph node metastases in patients with this challenging form of carcinoma. RhoC levels seem to identify a subset of patients with early tumor stage primary tumors and high metastatic potential that might benefit from more aggressive therapy. Through continued investigation, blockade of RhoC activity may be a potential target in the development of novel strategies for treating metastases of head and neck cancer.     

p53 protein overexpression in infiltrating ductal carcinoma of female breast and its association with lymph node metastasis

Traditionally, prognosis in carcinoma of the breast is evaluated based on size and differentiation of the tumor and status of lymph node metastasis. In addition to these established markers, in this molecular age other parameters such as overexpression of p53, c-erbB-2 and c-myc proteins are increasingly used to assess the prognosis. At present, the prognostic value of the molecular markers, at best, is controversial and conflicting. In this study, we examined 67 infiltrating ductal. carcinomas of female breast with and without lymph node metastasis for p53 protein overexpression by immunohistochemistry on formalin-fixed, paraffin-embedded tissue sections to ascertain if p53 positive tumors have greater metastatic potential than p53 negative tumors. In addition, p53 overexpression was also correlated with tumor size, grade, expression of estrogen and progesterone receptors, and age of the patient to clarify the issue of relevance of p53 overexpression in prognostication, p53 overexpression was observed in 39% of tumors and showed strong correlation only with the histological grade of the tumor. The incidence of p53 overexpression in grade 1, grade 2 and grade 3 tumors was 0%, 33% and 58% respectively. Lymph node metastasis was less frequent in tumors that overexpressed p53 protein. Twenty-seven percent of primary tumors with lymph node metastasis showed p53 protein overexpression, in contrast to 44% of tumors without metastasis. No correlation was observed between p53 overexpression and all the other parameters evaluated except progesterone receptor negative status. These results suggest that p53 overexpression is not an independent prognostic indicator and should not be used to predict lymph node metastasis or aggressive behavior of the tumor.     

p53 alteration in regional lymph node metastases from prostate carcinoma: a marker for progression?

BACKGROUND: Alterations of the p53 tumor suppressor gene are associated with advanced stage prostate carcinoma. The biologic significance of p53 nuclear accumulation in prostate cancer patients with regional lymph node metastases is uncertain. METHODS: The authors investigated p53 alterations by immunohistochemistry in 220 lymph node positive patients who were treated with radical prostatectomy, bilateral pelvic lymphadenectomy, and androgen deprivation therapy between 1987-1992 at the Mayo Clinic. The mean follow-up was 6.3 years. Tumor volume of lymph node metastases was measured using the grid method. RESULTS: p53 immunoreactivity was detected in 109 of 211 primary tumors (52%) and 83 of 144 matched regional lymph node metastases (58%); this expression was strongly concordant (correlation coefficient 0.53; P = 0.0001). Overexpression of p53 protein in lymph node metastases was associated with distant metastasis free survival by univariate analysis (P = 0.03), but did not reach statistical significance by multivariate analysis (P = 0.07). Regional lymph node cancer volume was the single most important predictor of distant metastases after adjusting for Gleason score, DNA ploidy, and p53 expression. CONCLUSIONS: The findings of the current study suggest that assessment of biologic changes (including p53 alterations in regional lymph node metastases) could be of value in the assessment of the biologic aggressiveness of prostate carcinoma, whereas p53 expression in the primary tumor does not appear to influence patient outcome.     

Detection of minimal residual cancer to investigate why oral tumors recur despite seemingly adequate treatment.

Improvements in surgery and radiotherapy techniques have led to only a modest increase in the 5-year survival rate for patients with head and neck cancer. This is because the pattern of clinical disease is changing, such that locoregional recurrence now accounts for fewer treatment failures, but more patients develop a second primary cancer or distant metastatic disease. In this study, we have used the p53 phage plaque assay, immunocytochemistry, and mutational analysis to assess the contribution of minimal residual cancer and genetic aberrations in clinically normal upper aerodigestive tract mucosa to treatment failure. Eighteen consecutive patients with oral tumors, with conventional clear margins, have been followed for a minimum of 36 months. Molecular assessment identified tumor-positive surgical margins for 6 of 11 assessable patients and additional tumor-positive lymph nodes for three cases. Disseminated malignant cells were detected in the hematopoietic cell compartment for six cases, and one patient had molecular evidence of field cancerization. Locoregional recurrence developed in five patients with tumors harboring a p53 gene mutation; four of these were associated with tumor-positive surgical margins, and one was associated with molecular evidence of field cancerization. Radiotherapy to the primary site did not prevent development of local recurrence when the residual tumor harbored a p53 gene mutation. Three of six cases with a tumor-positive bone marrow aspirate developed distant metastases. These findings reveal that molecular and immunocytochemical detection of minimal residual cancer and field cancerization can help identify patients who may develop locoregional or distant recurrence and justify further studies to evaluate the contribution of these remaining malignant cells to treatment failure.     

mdm2 expression as a prognostic indicator in clear cell renal cell carcinoma: comparison with p53 overexpression and clinicopathological parameters.

The present study was designed to analyze the expression of p53 and mdm2 in clear cell renal cell carcinoma with special emphasis on their association with tumor grade and clinical outcome. In particular, the value of individual protein overexpression as well as combined p53/mdm2 positivity was evaluated because both proteins are functionally connected, and their expression is controlled by an autoregulatory feedback loop. A cohort of 97 clear cell renal cell carcinomas was analyzed. The overexpression of mdm2 and p53 proteins was investigated on paraffin-embedded material by using monoclonal antibodies. Eighteen tumors showed mdm2 positivity, whereas 35 of the tumors overexpressed p53. Whereas p53 and mdm2 positivity correlated significantly (P = 0.00004), no correlation could be found between mdm2 protein overexpression and tumor stage, lymph node involvement, and presence of distant metastases. mdm2 positivity was found significantly more frequently in tumors of higher grade. In univariate analysis, there was a statistically significant correlation between p53 and mdm2 overexpression in the same tumor and poor survival (P = 0.00179). Multivariate analysis revealed that coincident mdm2/p53 overexpression, the presence of distant metastases, and tumor grade were independent predictors for tumor progression. Our results indicate that mdm2/p53 co-overexpression, nuclear grade, and preoperative presence of distant metastasis are independent predictors for poor survival.     

Her-2/neu overexpression in muscle-invasive urothelial carcinoma of the bladder: prognostic significance and comparative analysis in primary and metastatic tumors.

PURPOSE: The prognostic significance of Her-2/neu overexpression in muscle-invasive urothelial carcinoma of the bladder is largely unknown. Accurate determination of Her-2/neu overexpression may have therapeutic importance. EXPERIMENTAL DESIGN: Eighty consecutive cases of muscle-invasive urothelial carcinoma of the bladder treated by radical cystectomy with available follow-up were analyzed. In each case, one representative section was stained with anti-Her-2/neu. Staining was graded as 1 = faint/equivocal, 2 = moderate, and 3 = strong and was considered positive if > or =2. In those cases with a metastasis, the stain was also performed in the metastatic tumor. Results were correlated with survival. RESULTS: Twenty-two (28%) cases were considered Her-2/neu-positive in the primary tumor, and 17 of 32 (53%) were considered Her-2/neu-positive in the lymph node metastasis. Median survival for Her-2/neu-positive primary tumors was 33 months, compared with 50 months for Her-2/neu-negative cases (P = 0.46). Similarly, Her-2/neu overexpression in the lymph node metastasis did not predict survival. Sixty metastatic urothelial carcinomas were further studied by comparing Her-2/neu expression in the primary tumor with that of the lymph node and/or distant metastasis. Forty-five percent of Her-2/neu-negative primary tumors had a Her-2/neu-positive lymph node metastasis, whereas only one case (8%) of Her-2/neu-positive primary tumors was Her-2/neu-negative in the lymph node metastasis (P = 0.009). Similarly, 67% of Her-2/neu-negative primary tumors had a Her-2/neu-positive distant metastasis, whereas no Her-2/neu-positive primary tumor was negative in the metastasis (P = 0.429). CONCLUSIONS: Her-2/neu overexpression in primary or metastatic tumor did not predict survival in this cohort of muscle-invasive tumors. Overexpression in the primary tumors consistently predicts overexpression in a distant or regional metastasis. However, some Her-2/neu-negative primary tumors may show overexpression in their corresponding metastasis. Her-2/neu analysis in a metastasis may be necessary to accurately determine Her-2/neu status in metastatic bladder urothelial carcinoma.     

P53 expression in squamous cell carcinomas of the supraglottic larynx and its lymph node metastases: new results for an old question

BACKGROUND: Although p53 overexpression is frequent in head and neck squamous cell carcinomas (HNSCCs), controversy remains regarding the prognostic significance of that overexpression. The objective of this study was to investigate the expression pattern and prognostic significance of p53 expression in HNSCC of the same location, treated in the same way, and with long-term follow-up. METHODS: P53 expression was determined by immunohistochemistry in paraffin-embedded tissue specimens from 107 consecutive patients (107 primary squamous cell carcinomas of the supraglottic larynx and 46 matched lymph node metastases). All patients underwent surgical resection and bilateral neck dissection. RESULTS: A strong correlation was observed between p53 expression in the primary tumor and in the matched lymph node metastases (P=.0001). P53 overexpression in the lymph nodes was an independent predictor of regional recurrence (P=.027). Likewise, expression of p53 in the lymph nodes correlated significantly with disease-specific survival (P=.018). Five years after treatment, 70% of patients with p53-negative, metastatic lymph nodes remained alive, whereas only 30% of patients with p53-positive lymph nodes remained alive. In multivariate analysis, lymph node status and p53 expression in the lymph nodes remained associated with survival. CONCLUSIONS: The current data suggested that, although p53 overexpression is common in supraglottic carcinomas, its expression in the primary tumor is of limited clinical significance. However, the results supported the role of p53 in the lymph node metastases as an independent predictor of regional failure and a poor prognosis in patients with HNSCC. A prospective trial is indicated to validate these findings.     

p53 mutation in plasma DNA and its prognostic value in breast cancer patients.

PURPOSE: Tumor-specific DNA has recently been detected in the plasma of lung, head and neck, breast, and colon cancer patients. Detection of tumor-specific genetic materials in cancer patients at sites distant from the tumor, such as in the blood, may provide a unique and valuable tumor marker for diagnosis and prognosis. EXPERIMENTAL DESIGN: The present investigation was aimed at determining the presence of p53 mutations in the peripheral blood of breast cancer patients and its prognostic value in these patients. RESULTS: In this study, we found that the mean concentration of plasma DNA in healthy women was 21 ng/ml, whereas in patients with breast cancer the mean concentration was 211 ng/ml (P < 0.01). p53 mutations were detected in the primary tumors of 46 of 126 (36.5%) breast cancer patients. Of these 46 patients, 30 (65.1%) were found to have p53 mutations in their plasma DNA. p53 mutations in plasma DNA were strongly correlated with clinical stage, tumor size, lymph node (LN) metastasis, and estrogen receptor status (P < 0.05). After a median follow-up of 29 months, univariate and multivariate analysis revealed that both primary tumor and plasma DNA p53 mutations were significant prognostic factors for both relapse-free and overall survival. Furthermore, we demonstrated that patients with both primary tumor and plasma p53 mutations have the worst survival. This outcome occurs in both LN-positive and LN-negative groups. Thirteen of the 22 (59%) patients with recurrence and/or metastasis later had detectable p53 mutations in their plasma DNA. CONCLUSIONS: Detection of p53 mutations in plasma DNA may be used as a prognostic factor and an early marker to indicate recurrence or distant metastasis.     

p53 is overexpressed in fifty percent of pre-invasive lesions of head and neck epithelium

To identify the stage in head and neck carcinogenesis at whichp53 abnormalities become important, we evaluated 512 squamousepithelial tissue samples (201 pre-invasive and 209 invasivelesions, as well as 102 normal epithelia) in specimens fromtwo institutions. Of 311 patients evaluated, 128 did not havean invasive carcinoma. The frequency of p53 overexpression inthe pre-invasive lesions was not influenced by an adjacent invasivetumor. There was an increasing frequency of p53 overexpressionin specimens from histologically normal epithelium (5%, withonly scattered basal layers positive) to mild (28%) and moderatedysplasia (47%). p53 overexpression was also significantly morecommon in lesions with severe dysplasia (54%) and carcinomain situ (CIS; 50%) than in those with mild dysplasia. To furtherevaluate the timing of p53 alterations in the development ofhead and neck cancer, paired epithelial samples (one pre-invasiveand one invasive) from the same patient were evaluated. A discordancein the p53 staining pattern was seen in approximately one-thirdof the cases. When a discordance was present, the frequencywith which the invasive lesion was positive but the noninvasivenegative decreased from 97% (invasive versus normal epithelium)to 50% (invasive versus severe dysplasia or CIS). When p53 expressionwas evaluated by site, buccal lesions appeared to overexpressp53 more frequently than tongue lesions. In conclusion, p53protein overexpression is an early event in head and neck carcinogenesisand may represent a biomarker for patients with pre-invasivelesions.     

Interaction between potential doubling time and TP53 mutation: predicting radiotherapy outcome in squamous cell carcinoma of the head and neck

PURPOSE: To investigate the correlation between tumor potential doubling time, Tpot, and mutations in the p53 gene, TP53, and the potential of these parameters to predict outcome of head and neck cancer patients treated with radiotherapy. METHODS AND MATERIALS: Data from two independent studies on Tpot and TP53 mutations were combined, including 58 patients with squamous cell carcinoma of the head and neck. Tpot was estimated on biopsies obtained 6-9 h after infusion of iododeoxyuridine by combined flow cytometry and immunohistology. TP53 mutations were detected using DGGE and sequenced. All patients received primary radiotherapy alone. RESULTS: The predictive value of Tpot alone was of borderline significance. However, in TP53 wild-type tumors, Tpot was a strong predictor of outcome, whereas Tpot in TP53 mutant tumors failed to provide any information. Tpot and TP53 were not associated with nodal control; however, there was a strong relationship with control in the T-position, disease-specific survival, and overall survival. CONCLUSION: Tpot can to be a relevant parameter for predicting outcome of radiotherapy in head and neck cancer but only in the subset of patients without mutations in the p53 gene.     

Immunohistochemical analysis of p53 in colorectal cancer regarding clinicopathological correlation and prognostic significance

The overexpression of the tumor suppressor gene p53 was investigated immunohistochemically in 144 cases of primary colorectal cancer and in 8 cases with cancer in the corresponding metastatic lymph nodes. Abnormalities in p53 expression were found in 36 cases (25%) of the 144 primary cancer cases. In addition, p53-positive tumors were found to metastasize frequently to the lymph nodes, as compared to p53-negative tumors (61.1% vs. 41.7%, P=0.0428). p53 staining was identical in 7 of 8 (87.5%) cases in primary and metastatic lesions. When the DNA content of the tumor was determined by flow cytometry, the DNA index (mean ± SD) was significantly higher in p53-positive tumors than in p53-negative tumors (1.57 ± 0.38 vs. 1.39 ± 0.37, P=0.012). Therefore, the immunohistochemical data of p53 in colorectal cancer may help in potentially predicting metastatic spread to the lymph nodes. © 1995 Wiley-Liss, Inc.     

Apoptin induces apoptosis in an oral cancer mouse model

Apoptin, a chicken anemia virus-derived protein, induces apoptosis in various tumor cell lines and xenografted tumors. Its apoptotic activity is not hampered by tumor-suppressor p53 mutations or overexpression of anti-apoptosis proteins Bcl-2 or Bcl-x(L). We report for the first time the effects of apoptin expression in primary oral tumors, induced by the carcinogen 4-Nitroquinoline- 1-oxide in immunocompetent mice. In vivo a significant amount of primary oral tumor cells expressing apoptin cells underwent apoptosis, whereas synthesis of the LacZ control product did not. Ectopical expression of apoptin in passage 1 cell cultures derived from these oral tumors also resulted in apoptin-induced. Both in-vivo and in-vitro treated cells underwent apoptosis via the activation of caspase-3. The fact that apoptin induces apoptosis in primary squamous cell carcinoma cells indicates that apoptin is a potential therapeutic agent for treatment of head and neck squamous cell carcinoma.     

Cyclin D1 and p16INK4A expression predict reduced survival in carcinoma of the anterior tongue.

Cyclin D1 and p16INK4A are molecules with pivotal roles in cell cycle control and the development of diverse human cancers, and overexpression of cyclin D1 and loss of p16INK4A expression are common genetic events in head and neck squamous cell carcinoma. The prognostic significance of these molecular events at different sites within the head and neck, however, remains controversial. Thus, we sought to determine the relationship between cyclin D1 and/or p16INK4A expression and disease outcome in squamous cell carcinoma of the anterior tongue. Immunohistochemical detection of nuclear proteins cyclin D1, p53, and p16INK4A, and the Ki-67 labeling index was undertaken in tissue sections from 148 tongue cancers treated by surgical resection. Nuclear antigen status was analyzed in relation to pathological variables, tumor recurrence, and patient survival. Statistical significance was assessed using chi2 analysis for pathological variables and the Kaplan-Meier method, log rank test, and the Cox proportional hazards model for survival parameters. Overexpression of cyclin D1 occurred in 68% of tumors (100 of 147) and was associated with increased lymph node stage (P = 0.014), increased tumor grade (P = 0.003), and reduced disease-free (P = 0.006) and overall (P = 0.01) survival. Loss of p16INK4A expression was demonstrated in 55% of tumors (78 of 143) and was associated with reduced disease-free (P = 0.007) and overall (P = 0.014) survival. Multivariate analysis confirmed that in addition to pathological stage and regional lymph node status, cyclin D1 overexpression and loss of p16INK4A expression are independent predictors of death from tongue cancer. Loss of p16INK4A in the presence of cyclin D1 overexpression conferred a significantly worse disease-free (P = 0.011) and overall (P = 0.002) survival at 5 years. p53 nuclear accumulation and the Ki-67 labeling index were not prognostic. These data indicate that cyclin D1 overexpression and loss of p16INK4A expression predict early relapse and reduced survival in squamous cell carcinoma of the anterior tongue. Simultaneous assessment of cyclin D1 and p16INK4A protein levels define subgroups of patients at increased risk of relapse and may be of clinical utility in optimizing therapy.     

P53和P21过表达与胃癌淋巴结转移呈正相关

应用Ｓ－Ｐ免疫组化方法研究Ｐ５３和Ｐ２１表达变化与胃癌肿瘤组织分型、细胞增殖和浸润深度及淋巴结转移的关系。发现Ｐ５３阳性染色率为４８．９％，Ｐ５３阳性染色与胃癌肿瘤浸润深度及癌细胞增殖活性呈正相关（Ｐｅａｒｓｏｎ列联系数分别为Ｐ＝０．３２和Ｐ＝０．３５，Ｐ＜０．０５）。同时发现Ｐ２１阳性染色率为７０．０％，Ｐ２１阳性染色与胃癌肿瘤组织分型及癌细胞增殖活性呈正相关（Ｐｅａｒｓｏｎ列联系数分别为Ｐ＝０．３３和Ｐ＝０．５２，Ｐ＜０．０５）。Ｐ５３和Ｐ２１阳性肿瘤的淋巴结转移率（９３％和８１％）分别均明显高于其阴性表达的肿瘤（６０％和５５．５％，Ｐ＜０．０５）。Ｐ５３和Ｐ２１过表达对胃癌淋巴结转移率的贡献为以Ｐ５３过表达为主的独立的联合作用。提示Ｐ５３和Ｐ２１过表达在胃癌淋巴结转移和肿瘤增殖中起重要作用。Ｐ５３过表达与肿瘤浸润有关，而Ｐ２１过表达与肿瘤组织分型有关。     

Head and neck tumor sites differ in prevalence and spectrum of p53 alterations but these have limited prognostic value

The tumor site is a strong clinical factor in head and neck squamous cell carcinoma (HNSCC). To clarify the biologic and clinical role of p53 alterations in HNSCC, we have examined the prevalence and the nature of p53 alterations in a large cohort of tumors from the different sites. For immunohistochemical analysis of p53 protein expression, we introduced tyramide signal amplification immunohistochemistry (TSA-IHC) on a tissue microarray. This allowed the discrimination between normal low-level expression and reduced or lost expression. Two hundred fifty-three tumors were subjected to mutational analysis by genomic DNA sequencing, employing also the p53 GeneChip from Affymetrix. The prevalence of all p53 alterations, i.e., mutations, overexpression and loss of expression, was significantly higher in hypopharyngeal tumors than in the other sites (p = 0.001). Laryngeal tumors showed the lowest rate of p53 alterations, but revealed a distinct mutation spectrum: most mutations affected exon 5 (p = 0.013) and the S2' domain (p = 0.002), and most hot-spot 248 mutations occurred in the larynx (p < 0.001). Sequencing by p53GeneChip technology was shown to be only insignificantly more sensitive than dideoxy sequencing. In agreement with p53 mutations occurring prior to invasiveness, their prevalence did not increase with tumor stage, and all mutation classes lacked prognostic significance. The large patient cohort of this study showed that p53 is differentially affected in the different tumor sites of the head and neck, but its mode of inactivation does not play a major role in tumor progression.     

Molecular diagnosis of surgical margins and local recurrence in head and neck cancer patients: a prospective study.

PURPOSE: Approximately 10-30% of surgically treated head and neck cancer patients develop local recurrences while the resection margins are histologically tumor free. These recurrences may arise from cancer cells left behind but not detected by the pathologist, or they may develop from precursor lesions adjacent to the tumor that were not completely resected. We have investigated whether TP53-mutated DNA in the surgical margins is suitable to identify patients with head and neck squamous cell carcinoma at risk for local and locoregional recurrence. EXPERIMENTAL DESIGN: In a prospective cohort study of 76 patients with histologically tumor-free margins, the presence of TP53-mutated DNA was determined in the surgical margins using the phage plaque assay and correlated to clinical outcome. Immunostaining of the molecular-positive margins for mutated p53 protein was used to identify whether unresected precursor lesions or residual tumor cells were left behind. RESULTS: The absence of TP53-mutated DNA in surgical margins was significantly associated with remaining free of local and locoregional recurrence (P = 0.027 and P = 0.028, respectively). Moreover, the presence of TP53-mutated DNA in the surgical margins was an independent prognosticator for locoregional recurrence (relative risk = 7.1; P = 0.021; 95% confidence interval, 0.9-56). In 20% of the cases, the presence of TP53-mutated DNA in the surgical margins was found to be related to the presence of tumor-related precursor lesions. CONCLUSIONS: This study shows the value of TP53-mutated DNA as a molecular marker to predict locally recurrent head and neck squamous cell carcinoma. The observation that all patients who were negative for TP53-mutated DNA in the surgical margins remained free of local recurrence raises hope that molecular analysis of histologically tumor-free surgical margins can be exploited to decide on postoperative radiotherapy. Furthermore, our data provide evidence that local recurrences originate mainly from tumor cells left behind but also originate, in part, from unresected precursor lesions.     

Maintenance of head and neck tumor gene expression profiles upon lymph node metastasis

Spread of cancer and development of solid metastases at distant sites is the main cause of cancer-related deaths. To understand and treat metastases, it is important to determine at which stages the most pivotal steps for development of metastases occur. In head and neck squamous cell carcinoma (HNSCC), metastasis nearly always occurs first in local lymph nodes before development of distant metastasis. Here, we have investigated gene expression patterns in HNSCC lymph node metastases using DNA microarrays. Several types of analyses show that the gene expression patterns in lymph node metastases are most similar to the corresponding primary tumors from which they arose, as long as samples contain sufficient proportions of tumor cells. Strikingly, gene expression patterns of metastatic primary HNSCC are largely maintained upon spread to the lymph node. Only a single gene, metastasis-associated gene 1 (MTA1), was found to show consistently changed expression between a large number of matched primary tumor-lymph node metastasis pairs. The maintained expression pattern includes the predictive signature for HNSCC lymph node metastasis. These results underscore the importance of the primary tumor gene expression profile for development and treatment of metastasis. The findings also agree with the concept that disseminated cancer cells alter the surrounding tissue into a metastatic environment that resembles the primary tumor microenvironment.