Fluorescence in situ hybridization for detecting genomic alterations of cyclin D1 and p16 in oral squamous cell carcinomas

BACKGROUND: Cyclin D1 (CCND1) and p16 alterations have been detected in oral squamous cell carcinomas (SCCs), suggesting that abnormalities of these genes may play an important role in the genesis or progression of oral SCCs and serve as independent prognostic indicators. The detection of CCND1 and p16 aberrations using a simple and sensitive method would be valuable for the development of effective treatment modalities for oral cancer. The objective of the current study was to determine whether CCND1 numerical aberrations and p16 deletions in oral SCCs detected by fluorescence in situ hybridization (FISH) have any impact on clinical outcome. METHODS: Using genomic DNA probes for CCND1 and p16, FISH was performed on specimens that were obtained by fine-needle aspiration (FNA) from 57 primary oral SCCs. RESULTS: The CCND1 numerical aberration was observed in 28 of 57 patients (49%) with oral SCCs and was associated significantly with reduced disease-free survival (P = .0004) and overall survival (P = .0179). Conversely, p16 deletion was detected in 22 of 57 patients (39%). The disease-free and overall survival rates for patients with p16 deletion were lower than those among patients without the p16 deletion, although the difference just failed to reach statistical significance (P = .0516 and P = .1878, respectively). The p16 deletion in the presence of the CCND1 numerical aberration conferred significantly worse disease-free survival (P = .0002) and overall survival (P = .0153). CONCLUSIONS: Although the CCND1 numerical aberration was a good predictor of aggressive tumors, recurrence, and poor prognosis in patients with oral SCCs, the authors were able to identify subgroups of patients that had early disease recurrence and a poor prognosis more efficiently by assessment of p16 deletion in addition to CCND1 genetic status using FISH on FNA biopsy samples compared with the analysis of either alteration alone.     

Cyclin D1 gene numerical aberration is a predictive marker for occult cervical lymph node metastasis in TNM Stage I and II squamous cell carcinoma of the oral cavity

BACKGROUND: The management of occult cervical lymph node metastases originating from oral squamous cell carcinomas (OSCCs) remains controversial. The purpose of this study was to evaluate the value of cyclin D1 gene (CCND1) numerical aberrations in predicting the risk of late lymph node metastases. METHODS: Fluorescence in situ hybridization (FISH), using a BAC clone specific for CCND1, was performed on OSCC specimens obtained by fine-needle aspiration (FNA) biopsy from 45 patients with previously untreated TNM Stage I and II (T1-2N0M0) disease who had not undergone elective cervical lymph node dissection. RESULTS: CCND1 numerical aberrations were observed in 15 (33.3%) of the 45 patients and were significantly associated with the mode of invasion of the primary tumor (P = 0.01) and the presence of occult lymph node metastases (P < 0.001). Twelve of these 15 patients (80%) developed late cervical lymph node metastases within 2 years of surgery for primary OSCCs. All patients with cluster-type amplification of CCND1 developed late lymph node metastases. Multivariate analysis showed that only CCND1 numerical aberrations (risk ratio, 8.685%, 95% confidence interval = 2.232-33.802, P = 0.002) independently predicted late cervical lymph node metastasis. CONCLUSIONS: Aberrations in CCND1 numbers appear to be valuable in identifying patients at high risk of late lymph node metastasis in Stage I and II OSCCs. Analysis of CCND1 numerical aberrations using FISH on FNA biopsy specimens may be useful in selecting patients for elective cervical lymph node dissection.     

Prognostic significance of cyclin D1 amplification and overexpression in oral squamous cell carcinomas

Amplification and overexpression of cyclin D1 (CCND1) have been reported as independent prognostic indicators of several tumors. To investigate the association between CCND1 amplification and overexpression in oral squamous cell carcinomas (OSCCs), and to determine which is more reliable as a prognostic indicator, fluorescence in situ hybridization (FISH) on fine-needle aspiration (FNA) biopsies and immunohistochemistry were performed on 41 primary OSCCs (26 males, 15 females; mean age; 58.4 years, range 21-89 years). Thirteen patients were stage I, 13 were stage II, nine were stage III, and six were stage IV. CCND1 amplification and overexpression was detected in 13 (31.7%) and 27 (65.9%) of 41 cases. CCND1 was overexpressed in all cases showing CCND1 amplification. On the other hand, CCND1 overexpression was also detected in 14 of 28 cases (50.0%) lacked such amplification. Statistical analysis showed that the correlation between CCND1 overexpression and decreased survival just failed to reach statistical significance, and CCND1 amplification and nodal status were independent prognostic indicators. In conclusion, it will be necessary to investigate the other pathways that regulate CCND1 expression besides CCND1 amplification. From the present study, CCND1 amplification is a more reliable prognostic indicator than CCND1 overexpression in OSCCs.     

EMS1 gene amplification correlates with poor prognosis in squamous cell carcinomas of the head and neck.

The relationship between CCND1 and/or EMS1 amplification and disease outcome was studied in a prospective series of 104 head and neck squamous cell carcinomas treated by surgical resection. The CCND1 and EMS1 copy number in tumor samples was estimated by differential PCR. The presence or absence of amplification was analyzed in relation to clinicopathological variables, tumor recurrence, and patient survival. CCND1 amplification occurred in 32 cases (31%) and was associated with increased lymph node stage (P = 0.005) and advanced disease stage (P = 0.003). EMS1 amplification was identified in 21 cases (20%) and was related with advanced T stages (P = 0.001), increased lymph node stage (P = 0.02), advanced disease stage (P = 0.041), poor histological differentiation (P = 0.018), recurrent disease (P = 0.0004), and reduced disease-specific survival (P < 0.0001). Coamplification of both genes occurred in 11 cases (11.5%). Multivariate analysis confirmed that in addition to regional lymph node status, EMS1 amplification is an independent predictor of death from the tumor (P = 0.0027). CCND1 amplification was not prognostic. These data indicate that EMS1 amplification, but not CCND1 amplification, predicts early recurrence and reduced survival in squamous cell carcinoma of the head and neck. The prognostic significance previously attributed to CCND1 amplification may be attributable to its frequent coamplification with EMS1.     

Epidermal growth factor receptor gene copy number aberration at the primary tumour is significantly associated with extracapsular spread in oral cancer

Background:

Extracapsular spread (ECS) of lymph node metastasis in head and neck cancers, including oral squamous cell carcinomas (OSCCs), is known to reflect tumour aggressiveness, and is significantly associated with high rates of loco-regional recurrence, distant metastasis, and poor outcome. The purpose of this study was to confirm ECS as an important prognostic indicator and to determine the significant factors associated with ECS in OSCCs.

Methods:

We investigated the incidence of ECS and impact of ECS on survival in 127 OSCC patients. To determine the factors significantly correlated with ECS, we examined many factors, including the clinicopathological features of primary tumours, lymph node metastasis, and copy number aberrations of the cyclin D1 gene (CCND1) and epidermal growth factor receptor gene (EGFR) at primary tumours, and evaluated the value of predicting the risk of ECS of the metastatic lymph node.

Results:

Kaplan–Meier and multivariate disease-free and overall survival analysis clearly demonstrated that ECS is an independent prognostic factor in OSCCs. Moreover, logistic regression analysis showed that the number of pathologically positive nodes and copy number aberrations of EGFR at the primary tumour are independent predictors of ECS.

Conclusions:

The findings suggest that ECS is an independent prognostic factor in OSCCs. Moreover, the number of pathologically positive lymph nodes and EGFR numerical aberrations of the primary tumour were also shown to be excellent predictors of ECS in OSCCs. Preoperative evaluation of EGFR numerical aberrations might therefore be a useful tool for selecting patients at high risk of ECS, who would benefit from targeted aggressive multimodality therapy.     

Cyclin D1 amplification correlates with early recurrence of squamous cell carcinoma of the tongue

Amplification of CCND1 was studied in 23 tongue carcinoma patients by fluorescence in situ hybridization (FISH) using a paraffin embedded specimen. All the patients received complete resection of the primary site with or without neck dissection. CCND1 amplification was positive in 13 (56.5%) out of 23 cases. Correlations between CCND1 amplification and histological grading, T category, N category, and Stages were not significant. The 5-year disease-free survival rate, which was 23.1% for CCND1 amplification positive patients and 80.0% for negative patients, was significantly better for the CCND1 amplification negative patients (P=0.0070). Nine patients were examined by dual-color FISH with the probe for centromere of chromosome 11 and 11q13. In five patients, who had positive amplification for CCND1, cell numbers with a larger number of signals for 11q13 than the centromere of chromosome 11 were significantly higher than those of CCND1 amplification negative patients (P=0.013). This indicates that amplification of 11q13 occurs more frequently than aberration of chromosome 11 in CCND1 amplification positive patients. From these results, the amplification of CCND1 is a key factor in predicting the aggressiveness of tongue cancer. Furthermore, FISH proved to be a useful method for such evaluation.     

Expression and prognostic relevance of MUC1 in stage IB non-small cell lung cancer

The goal of this study was to evaluate the expression of MUC1 in stage IB non-small cell lung cancer (NSCLC) and its prognostic significance. The expression of MUC1 in 178 NSCLC specimens was evaluated via immunohistochemistry. A reproducible semiquantitative method which took both staining percentage and intensity into account was applied for immunohistochemical scoring, and receiver operating characteristic curve analysis was utilized to select the cut-off score for high or low MUC1 expression. Then, the correlations between MUC1 expression and clinicopathological features and its prognostic relevance were determined. In this study, high MUC1 expression was detected more frequently in adenocarcinomas (86.3%) and other NSCLCs (74.1%) than in squamous cell carcinomas (39.1%, P < 0.001). The Kaplan-Meier survival curves showed that up-regulated expression of MUC1 indicated poorer overall survival (OS) and disease-free survival (DFS) (P = 0.011 and P = 0.008, respectively), especially for those with non-squamous cell carcinomas (P = 0.033 and P = 0.011, respectively). Multivariate analysis also confirmed that MUC1 expression was an independent prognostic factor for both OS and DFS in stage IB NSCLC (P = 0.008 and P = 0.004, respectively). MUC1 might be correlated with the histogenesis of lung adenocarcinoma, and its elevated expression may be an adverse prognostic indicator for the patients with stages IB NSCLC, particularly for those with non-squamous cell carcinomas.     

Proliferative activity is the most significant predictor of recurrence in noninvasive papillary urothelial neoplasms of low malignant potential and grade 1 papillary carcinomas of the bladder

BACKGROUND: Recurrence of transitional cell carcinoma of the bladder cannot be predicted accurately by traditional criteria alone. This study examined the value of cell proliferative activity, morphometry, and expression of p53, c-erbB-2, and bcl-2 oncogenes in predicting recurrence of superficial papillary urothelial neoplasms of low malignant potential (LMP) and Grade 1 (G1) papillary carcinomas of the bladder. METHODS: Sixty-two patients (mean age, 62 years) with newly diagnosed superficial pTa bladder tumors (19 LMP, and 43 G1) were analyzed retrospectively. All patients underwent transurethral resection (TUR). Median follow-up was 69 months. Serial sections from formalin-fixed, paraffin-embedded material at initial TUR were stained with monoclonal antibodies (MoAbs) DO7, CB11, and bcl-2-124. Cell proliferation was assessed by MIB-1 MoAb, the quantity of argyrophilic nucleolar organizer region-associated proteins (AgNORs), and mitotic count. RESULTS: Of the 62 patients, 42 (67.7%) had one or more recurrences. Recurrence rates were higher in MIB-1 (P < 0.0001) and p53 immunopositive cases (P = 0.02), when the mitotic count was greater than 5 (P = 0.004), and in G1 carcinomas (P = 0.04). In univariate analysis, the disease-free period was shorter for MIB-1 (P < 0.0001) and p53 immunopositive (P = 0.0001) cases, for cases with high AgNOR quantity (P = 0.04), mitotic count greater than 5 (P = 0.01), and in G1 carcinomas (P = 0.002). In multivariate analysis, only MIB-1 immunoreactivity retained independent prognostic significance. CONCLUSIONS: Despite the small cohort, the results confirm the prognostic value of cell proliferation and p53 expression in patients with bladder neoplasms. The results also indicate that MIB-1 immunopositivity is the most significant predictor of recurrence and disease-free survival in superficial LMP and G1 papillary bladder carcinomas.     

Numerical aberrations of chromosome 17 and the 9p21 locus are independent predictors of tumor recurrence in non-invasive transitional cell carcinoma of the urinary bladder

Since the majority of patients with non-invasive (stage pTa) bladder carcinoma experience tumor recurrence, the identification of prognostic markers for assessing the biologic behavior of tumors is of high clinical interest. In this retrospective study, we investigated the prognostic value of numerical aberrations of chromosomes 3, 7, 17 and of the 9p21 gene locus in 71 pTa bladder carcinomas using the UroVysion test. This recently developed multi-probe fluorescence in situ hybridization (FISH) test, which simultaneously stains the 9p21 locus and the centromer regions of chromosomes 3, 7 and 17, was applied on paraffin sections from formalin-fixed tumor specimens. All tumors were evaluated for twelve different criteria, among which were: polysomy 3, 7 or 17 in > or =10% of tumor cells, pentasomy or higher polysomy 3, 7 or 17 in > or =1 tumor cell, and loss of both 9p21 alleles in > or =10% of tumor cells. The latter parameter was the most frequent chromosomal aberration (detected in 83% of the tumors), while polysomies 3, 7 and 17 were registered in 27, 13 and 12% of the cases, respectively. Most of the parameters were found at higher frequencies in G3 tumors compared to G1 or G2 tumors. None of the parameters correlated with progression-free survival, but polysomy 3 (p=0.029), polysomy of at least one of the chromosomes 3, 7 or 17 (p=0.032), pentasomy/higher polysomy 17 (p=0.007), and loss of 9p21 (p=0.026), correlated significantly with recurrence-free survival. Of these, pentasomy/higher polysomy 17 (p=0.015) and loss of 9p21 (p=0.036) were identified as independent predictors of tumor recurrence in a multivariate Cox regression analysis that also included tumor grade. In conclusion, we suggest that evaluation of numerical aberrations of chromosome 17 and the 9p21 locus may represent a useful tool to assess tumor recurrence of pTa bladder carcinomas more accurately.     

Tissue microarray analysis reveals site-specific prevalence of oncogene amplifications in head and neck squamous cell carcinoma

Fluorescence in situ hybridization was applied on a collection of 609 squamous cell carcinomas of the head and neck (HNSCCs),including 511 primary carcinomas of different clinical stage and anatomical localization and 98 recurrent carcinomas, second primary carcinomas, and regional metastases on a tissue microarray. The overall prevalence of amplifications of five oncogenes analyzed was 34.5% for CCND1, 12.7% for EGFR, 8.8% for MYC, 6.2% for ZNF217, and 3.6% for ERBB2. CCND1 amplifications were associated with the pharyngeal site in primary carcinomas (P < 0.001), whereas amplifications of ZNF217 were less frequent in pharyngeal carcinomas as compared with primary oral and laryngeal carcinomas (P = 0.02). The amplification pattern of these oncogenes suggests that different molecular pathways are involved in HNSCCs of different localizations.     

早期舌鳞状细胞癌的疗效分析

目的 分析不同方法治疗Ⅰ、Ⅱ期舌鳞癌的疗效.方法 回顾性分析103例Ⅰ、Ⅱ期舌鳞癌患者,比较单纯手术、单纯放疗和综合治疗(术前或术后放疗)的疗效及其影响因素.结果Ⅰ、Ⅱ期患者的5年生存率分别为82.4%和80.O%(P=0.361),其中单纯手术组、单纯放疗组和综合治疗组的5年生存率分别为90.3%、68.4%和84.0%(P=0.104),局部复发率分别为2.5%、35.7%和5.7%(P＜0.001).隐匿性淋巴结转移率为23.8%,其中Ⅱ区的转移率(19.0%)最高.局部复发、区域复发是影响患者预后的独立因素,单纯手术即能获得满意的局部控制,而低分化鳞癌易出现区域复发.结论 选用单纯手术治疗早期舌癌,如采用择区性颈淋巴清扫术,清扫范围为Ⅰ～Ⅳ区.     

Cyclin D1 G870A polymorphism and squamous cell carcinoma of the uterine cervix in Korean women

Though many investigators have reported relationships between the CCND1 polymorphism and susceptibility to various carcinomas, to our knowledge, no report has been issued concerning its relationship with uterine cervical cancer. Thus, we undertook this study to investigate the association between CCND1 polymorphisms and susceptibility to cervical cancer in Korean women. This study was carried on 222 patients with squamous cell carcinoma of uterine cervix and on 314 normal controls. CCND1 genotyping was determined by polymerase chain reaction and restriction fragment length polymorphism. The allelic frequencies of the cases (A, 0.53; G, 0.47) were not significantly different from those of the controls (A, 0.49; G, 0.51) (P=0.238). Regression analysis after adjusting for age showed that the CCND1 G870A genotypes are not related to the risk of squamous cell carcinoma of the uterine cervix. Our findings suggest that the CCND1 polymorphism is not associated with an increased risk of squamous cell carcinoma of uterine cervix in Korean women.     

局部晚期宫颈鳞癌流式细胞术分析与放射治疗预后的关系

 目的 研究局部晚期宫颈鳞癌流式细胞术分析结果（DNA倍体、SPF和PI）与放疗预后的关系，探讨它们在预测宫颈癌放疗效果中的价值。方法 68例ⅢB期宫颈鳞癌病人在放疗前钳取宫颈癌组织，制备成单细胞悬液，采用流式细胞术检测癌细胞DNA倍体、S期比例（SPF）及增殖指数（PI），分析他们及其他临床参数与患者根治性放疗后疾病复发和生存的关系。结果 在68例标本中，异倍体检出率为47．1％（32／68），SPF和PI值为分别为（7．49±2．91）和（12．89±3．75）；在随访期间，全组宫颈癌患者复发率为44．1％（30／68）；二倍体组和异倍体组的5年无复发生存率分别为52．7％和35．2％（P〈0．（15），高SPF组和低SPF组5年无复发生存率分别为25．3％和56．6％（P〈0．05），高PI组和低PI组5年无复发生存率分别为37．5％和49．3％（P〈0．05）；经多因素分析显示，肿瘤大小和SPF是影响宫颈癌患者无复发生存的独立预后因素。结论 通过流式细胞术检测宫颈癌组织SPF可以预测局部晚期宫颈鳞癌放射治疗后的预后。     

Bone morphogenetic protein 7 expression associates with bone metastasis in breast carcinomas.

BACKGROUND: We recently showed that bone morphogenetic protein 7 (BMP7) is overexpressed in primary breast tumors. Here we explored the clinical significance of BMP7 expression in breast cancer. MATERIALS AND METHODS: This study included 483 breast cancer patients with complete clinicopathological information and up to 15 years of follow-up. Samples contained 241 lobular carcinomas, 242 ductal carcinomas, and 40 local recurrences. BMP7 protein expression was determined using immunohistochemistry. RESULTS: BMP7 was expressed in 47% of the primary tumor samples and 13% of the local recurrences. The primary tumors expressed BMP7 more often than the corresponding local recurrences (P = 0.004). BMP7 expression was dependent on the tumor subtype; 57% of the lobular carcinomas but only 37% of the ductal carcinomas were BMP7 positive (P = 0.0001). BMP7 expression was associated with accelerated bone metastasis formation (P = 0.040), especially in ductal carcinomas (P = 0.033), and multivariate analysis confirmed that BMP7 is an independent prognostic indicator for early bone metastasis development (P = 0.032). CONCLUSION: BMP7 is clearly associated with bone metastasis formation and thus might have clinical utility in identification of patients with increased risk of bone metastasis. This is the first time that bone inducing factor BMP7 has been linked to the bone metastasis process in breast cancer.     

Cyclin D1 overexpression correlates with poor prognosis in patients with tongue squamous cell carcinoma

Tongue squamous cell carcinoma makes up a large percentage of head and neck cancers, and the incidence among young patients is increasing. The aim of this study was to reveal the correlation between cyclin D1 (CCND1) expression and clinical and histologic features. We performed an immunohistochemical study on the level of CCND1 expression in tumor specimens obtained from 94 patients with tongue squamous cell carcinoma. The relationship between the expression and the following features such as age, sex, smoking and alcohol intake history, T, N, histologic grade, and multiple primary cancer was analyzed. Eighteen patients (19%) showed CCND1 overexpression (tumor cell nuclei positivity >/=50%). The 5-year survival rate of high CCND1 expressors was 39%, which was significantly poor (p=0.04). N classification correlated with CCND1 expression. CCND1 overexpression is associated with poor survival associated with progression of lymph node spread in patients with tongue squamous cell carcinomas. CCND1 expression may be a useful biologic marker for prognosis.     

The gene ratios c-MYC:cyclin-dependent kinase (CDK)N2A and CCND1:CDKN2A correlate with poor prognosis in squamous cell carcinoma of the head and neck.

PURPOSE: Tumor-Node-Metastasis classification does not fully predict outcome of treatment and prognosis in patients with squamous cell carcinoma of the head and neck. Different biomarkers have been suggested to yield additional prognostic information, but no single marker has thus far been introduced in the clinic. The objective of the present study was to analyze the copy number of the frequently amplified oncogenes CCND1 and c-MYC in relation to the commonly deleted tumor suppressor gene cyclin-dependent kinase (CDK)N2A (p16) to enhance the clinical significance. EXPERIMENTAL DESIGN: Extracted DNA from diagnostic biopsies of 78 untreated patients were analyzed by real-time PCR with specific primers for c-MYC, CCND1, and CDKN2A. Gene copy number ratios were calculated by dividing the copy number of c-MYC or CCND1 with CDKN2A. Ratios > 2 were defined as enhanced. These data were related to disease-free interval and disease-specific survival. RESULTS: Enhanced gene ratio of c-MYC:CDKN2A was detected in 35 of 78 (45%) and enhanced ratio of CCND1:CDKN2A in 36 of 78 (46%) of the cases. The c-MYC:CDKN2A and CCND1:CDKN2A ratios correlated with disease-specific survival with respect to death (P = 0.042 and 0.049, respectively; Log-rank test). Furthermore, enhancement of c-MYC:CDKN2A was associated with a shorter disease-free interval as marked by the development of recurrences or metastases (P = 0.014; Log-rank test). CONCLUSIONS: We conclude that CCND1 and/or c-MYC amplification, when combined with CDKN2A deletion, yield additional prognostic information as compared with analysis of single genetic aberrations. These gene ratios, as analyzed by a sensitive method like real-time PCR on diagnostic biopsies, might help clinicians to individualize the treatment of squamous cell carcinoma of the head and neck as they reflect the biological properties of the tumors. This could be used as an adjunct to the Tumor-Node-Metastasis classification system.     

Prognostic significance of p27KIP1 expression in resected non-small cell lung cancers: analysis in combination with expressions of p16INK4A,pRB, and p53

BACKGROUND AND OBJECTIVES: Whether a prognostic role for expression of the tumor suppressor gene (TSG) products exists in resected non-small call lung cancers (NSCLCs) remains controversial. Our study was performed to determine the value of TSGs expressions for patients survival in NSCLCs. METHODS: We examined 108 resected NSCLCs for the expression of TSG products, p27(KIP1), p16(INK4A), pRB, and p53 that govern cell cycle transition by immunohistochemistry and compared them with patient clinical characteristics and prognoses. RESULTS: Abnormal expressions of p27(KIP1), p16(INK4A), pRB, and p53 were found in 61 (57%), 53 (49%), 42 (39%), and 48 (44%), respectively, of the 108 NSCLCs. Univariate analysis showed abnormal expression of p27(KIP1) to be a strong indicator for poor patient survival, not only in the total cohort (P = 0.0024), but also in subgroups with T1-T2 (P = 0.016), N0 (P = 0.047), and squamous cell carcinomas (P = 0.026), but not according to the expression of p16(INK4A), pRB, or p53. In the Cox regression analysis, p27(KIP1) expression was found to be an independent prognostic factor (P = 0.0148) and associated with pathological stage (P = 0.0278). CONCLUSIONS: Our results suggest that abnormal p27(KIP1) expression may be a useful indicator to predict postoperative prognosis, especially in patients with early stage NSCLCs, as compared to other TSG products examined.     

Lack of Elevated HER2/neu Expression in Epithelial Dysplasia and Oral Squamous Cell Carcinoma in Iran

Purpose: The role of the HER family in oral squamous cell carcinomas (OSCCs) is not well-defined. Thisstudy was aimed to assess the frequency of HER2/neu overexpression in oral carcinogenesis. Materials andMethods: Expression of HER2/neu oncoprotein in OSCCs (N=18), oral epithelial dysplasia (N=18) and normaloral mucosa (N= 18) was assessed by immunohistochemistry using a cerbB2 antibody kit. Results: HER2/neuwas almost undetectable in normal oral mucosa and only 1/18 (0/05) of cases was positive. In oral epithelialdysplasia, 2/18 (11.1%) demonstrated staining, as did 3/18 OSCCs. Membrane staining was observed in allcases and there was no significant variation in frequency/intensity between normal oral mucosa / oral epithelialdysplasia and OSCCs (p>0/05). Conclusions: Aberrant expression of HER2/neu apparently does not contributeto carcinogenesis in the oral epithelium. The lack of overexpression in OSCCs indicates that molecular targetingis not feasible for adjuvant treatment.     

Image cytometry of cyclin D1: a prognostic marker for head and neck squamous cell carcinomas.

CCND1 gene amplification and cyclin D1 protein overexpression are indicators for poor prognosis in invasive head and neck carcinomas. Increased CCND1 gene dosage is a more sensitive prognostic factor than protein overexpression as evaluated by conventional immunohistochemical techniques. Qualitative immunohistochemistry cannot distinguish cyclin D1 overexpression accompanied by amplification of the CCND1 gene from overexpression associated with normal CCND1 gene copy number. To improve the sensitivity of cyclin D1 protein determination, we applied quantitative techniques of image analysis to evaluate cyclin D1 in 54 head and neck carcinomas. There was a significantly higher rate of occurrence of adverse events (P = 0.043) among patients with CCND1 gene amplification than among those without gene amplification. There was a strong association between CCND1 gene amplification (as detected by Southern blot analysis) and the highest nuclear score (by image cytometry of the immunostained tumor sections). The predominance of cells in the lowest nuclear score category was significantly associated with normal copy number (P = 0.005). Conversely, the highest nuclear score was a significant predictor of gene dosage (P = 0.02). Similarly, high nuclear score was a good predictor of death as the final outcome of the disease (P = 0.01). Although somewhat less accurate than Southern blotting, image cytometry of immunohistochemical cyclin D1 stain appears to be a promising tool that could be useful for other tumor marker expression studies.