Immunodetection of GLUT1, p63 and phospho-histone H1 in invasive head and neck squamous carcinoma: correlation of immunohistochemical staining patterns with keratinization

Immunodetection of GLUT1, p63 and phospho-histone H1 in invasive head and neck squamous carcinoma: correlation of immunohistochemical staining patterns with keratinizationAims : To examine invasive head and neck squamous carcinomas for expression of GLUT1, a glucose transporter and marker of increased glucose uptake, glycolytic metabolism and response to tissue hypoxia; p63, a p53 homologue that is a marker of the undifferentiated proliferative basaloid phenotype; and phospho-histone H1, a marker of activation of the cell cycle-promoting cyclin-dependent kinases 1 and 2. Methods : Routinely processed slides from 34 invasive squamous carcinomas, including 25 with intraepithelial components, were immunostained with anti-GLUT1 (Chemicon), anti-p63 (4A4, Santa Cruz), and antiphospho-histone H1 (monoclonal 12D11). Results : In keratinizing carcinomas, all three markers were most commonly immunodetected peripherally, with loss of expression in central keratinized zones. In contrast, in non-keratinizing carcinomas, p63 and phospho-histone H1 expression was most commonly observed throughout tumour nests and anti-GLUT1 stained in a pattern suggestive of hypoxia-induced expression ('antistromal' staining), in which cells at the tumour-stromal interface were GLUT1- and cells in central, perinecrotic zones showed progressive induction of GLUT1. Intraepithelial components also displayed basal and 'antibasal' GLUT1 staining patterns, homologous to the pro- and antistromal patterns in invasive carcinoma; basal patterns in intraepithelial lesions appeared to be more predictive of keratinizing invasive carcinoma and antibasal intraepithelial staining more predictive of non-keratinizing poorly differentiated carcinomas. Conclusions : Keratinizing and non-keratinizing squamous carcinomas differ in expression patterns of GLUT1, p63 and phospho-histone H1. In the former, all three markers were typically suppressed in conjunction with keratinization; in the latter, GLUT1 expression was more likely to occur in a hypoxia-inducible pattern and expression of p63 and phospho-histone H1 was unsuppressed. GLUT1 expression patterns in intraepithelial lesions may be predictive of the differentiation status of the associated invasive carcinoma.     

Feasibility of immunocytochemical detection of tumor markers (XIAP, phosphohistone H1 and p63) in FNA cellblock samples from head and neck squamous cell carcinoma

One of the major handicaps in contemporary clinical oncology is the inability to predict the responsiveness of any individual's malignancy to specific therapies. The purpose of this study was to test the feasibility of immunocytochemically detecting markers that may be affected by therapy or are predictive of therapeutic responsiveness, including phosphohistone H1 (anti-p-H1 MoAb 12D11) and X-linked inhibitor of apoptosis (XIAP) in small samples obtained via fine-needle aspiration (FNA) biopsy procedure, thus improving therapeutic monitoring. p63, a squamous stem cell regulatory protein, was also examined. These three markers were studied in FNA cell block samples of head and neck squamous cell carcinoma (HNSCC). Twenty-eight alcohol-fixed formalin-postfixed paraffin-embedded cell-block samples from FNAs of patients with HNSCC were subjected to antigen retrieval and then incubated with anti-XIAP, anti-p-H1, and anti-p63, and developed using EnVision-Plus reagents and diaminobenzidine as chromagen; Granular or heterogeneous cytoplasmic staining for XIAP and nuclear staining for p63 and p-H1 were considered positive. Among the 28 cases studied, the overall positive rates for XIAP, p-H1, and p63 were 60.7%, 96.4%, and 92.8%, respectively. The staining intensity for XIAP: + 70.6%, ++ 23.5%, +++ 0%, and ++++ 5.9%; for p-H1: + 48.1%, ++ 11.1%, +++37.0%, and ++++ 3.7%; and for p63: + 11.5%, ++ 23.1%, +++ 53.9%, and ++++ 11.5%. The expression of p-H1 and p63 appeared to be higher and stronger than that of XIAP in HNSCC. This study demonstrated the feasibility of monitoring expression of three tumor markers using FNA samples. p-H1 and XIAP may be useful for monitoring actions of cyclin-dependent kinase inhibitors, XIAP-lowering, and/or apoptosis-inducing drugs, respectively. Future studies will focus on the impact of therapies upon these staining profiles.     

GLUT1 and p63 expression in endometrial intraepithelial and uterine serous papillary carcinoma

AIMS: To analyse the expression patterns of GLUT1, p63 and p53 and the possible correlation between these markers in uterine serous papillary carcinoma (USPC) and endometrial intraepithelial carcinoma (EIC). METHODS AND RESULTS: Fourteen cases of USPC, 12 of which also showed EIC, were examined for GLUT1, p63 and p53 immunoreactivity. Four-micrometre sections from formalin-fixed paraffin-embedded tissue were immunostained using commercially available primary antibodies and Dako Envision Plus reagents for visualization. Membranous GLUT1 immunoreactivity was observed in all cases, including all 14 invasive tumours (100%) and 11 of 12 associated EICs (92%), and was confined to neoplastic cells. In USPC, staining tended to be strongest in superficial antistromal regions. p63 positivity was found in 8/14 (57%) USPCs and 9/12 (75%) associated EICs. In 11 cases p53 was overexpressed in both invasive USPC (11/14; 79%) and EIC (11/12; 92%). p53+ USPCs tended to be positive for p63, whereas p53- USPCs were also negative for p63. CONCLUSIONS: GLUT1 is expressed in the vast majority of USPC and EIC, suggesting a biological role during the early steps of carcinogenesis in endometrial serous neoplasms. GLUT1 expression may be induced by hypoxia-related as well as other mechanisms.     

Expression of the GLUT1 glucose transporter, p63 and p53 in thyroid carcinomas

Only few studies have evaluated the usefulness of the GLUT1 and p63 status of thyroid carcinomas in revealing tumorigenesis. We studied GLUT1, p53, and p63 immunoexpression in a total of 86 cases of various thyroid carcinoma types to determine the biological significance of GLUT1 and p63 expression in thyroid carcinomas. GLUT1 was detected in six cases of anaplastic carcinoma and in one case of poorly differentiated carcinoma with membranous staining. p63 was detected in five cases of anaplastic carcinoma, in one case of poorly differentiated carcinoma, and in five cases of papillary carcinoma with nuclear positivity. p53 was detected in six cases of anaplastic carcinoma, in one case of poorly differentiated carcinoma, and in one case of follicular carcinoma with nuclear positivity. Five of seven cases of anaplastic carcinoma expressed all three of these markers. The results suggest that GLUT1, p63, and p53 are not expressed in well-differentiated thyroid carcinomas, and that they are usually expressed late in the course of thyroid tumor progression. These data strongly suggest that in anaplastic carcinomas, impairment of p53-mediated repression results in increased GLUT1 and p63 expression, and that this probably reflects the differential regulation of hypoxia-responsive pathways and basal/stem cell regulatory pathways.     

Telomerase activity, MIB-1, PCNA, HPV 16 and p53 as diagnostic markers for cervical intraepithelial neoplasia

The present investigation evaluated the relationship between dysplasia of the uterine cervix and telomerase activity, expression of p53, MIB-1 and PCNA. Telomerase activity was measured on cervical cytobrush material from 126 women suspected of having dysplasia and 61 controls using the telomeric repeat amplification protocol. Immunohistochemistry was used to detect the tumor suppressor protein p53 and cell proliferation, the latter by MIB-1 and PCNA expression. Infection with human papillomavirus 16 was detected by PCR amplification and Southern blot hybridization of DNA extracted from the same brush material. Positive telomerase activity was found in 5 of 43 (11.6%) normal samples, 12 of 57 (21.1%) samples with inflammation or koilocytosis, 7 of 17 (41.2%) CIN 1 (cervical intraepithelial neoplasia, grade 1), 8 of 20 (40.0%) CIN 2, and 25 of 42 (59.5%) CIN 3/ CIS. Telomerase activity was significantly related to the level of dysplasia (p<0.001) and proliferation measured by MIB-1 (p=0.019), but not to the level of PCNA (p=0.445), HPV 16 status (p=0.098) or staining for p53 (p=0.271). Dysplasia was also related to PCNA, MIB1, p53, and presence of HPV 16. A sequential increase in the examined parameters, paralleling the progression of abnormality, was observed. PCNA and telomerase showed an increase in CIN 1, MIB-1 and HPV16 in CIN 2, and finally p53 in CIN 3/CIS.     

p63 overexpression associates with poor prognosis in head and neck squamous cell carcinoma

p63 belongs to a protein family that includes 2 structurally related proteins, p53 and p73. The aim of this study was to investigate the biologic role of p63 in oral tumorigenesis and its possible role as prognostic marker in oral cancer. Ninety-four cases of oral squamous cell carcinoma and 10 cases of normal mucosa were analyzed for p63 expression by immunohistochemistry. Normal oral mucosa showed a basal and parabasal expression of p63. Five (5.3%) cases of oral cancer showed less than 10% of positive tumor cells; in 33 (35.1%) cases the positive tumor cells comprised between 10% and less than 30%, in 36 (38.3%) cases the positive tumor cells comprised between 30% and less than 50%, and in 20 (21.3%) cases the positive tumor cells were more than 50%. There was also a statistically significant correlation between p63 expression and tumor differentiation: p63 expression was amplified in poorly differentiated tumors (P < .05). When analyzed for prognostic significance, patients with perineural infiltration had poorer survival rates than the group with no perineural infiltration (P < .05) and patients with increased p63 expression had poorer survival rates than the group with reduced p63 expression (P < .05). The statistical analysis showed no significant correlation between p63 expression, sex, age, tumor size, staging, recurrence, and metastasis. Cases with diffuse p63 expression were more aggressive and poorly differentiated and related to a poorer prognosis. These data suggest that p63 expression may be useful to identify cases of oral squamous cell carcinoma with more aggressive and invasive phenotype providing novel diagnostic and prognostic information on individual patient survival with oral cancers.     

Immunohistochemical staining with MIB1, bcl2 and p16 assists in the distinction of cervical glandular intraepithelial neoplasia from tubo-endometrial metaplasia,endometriosis and microglandular hyperplasia

AIMS: Preinvasive endocervical glandular lesions, termed cervical glandular intraepithelial neoplasia, are increasing in incidence. The distinction of cervical glandular intraepithelial neoplasia from benign mimics, especially tubo-endometrial metaplasia, endometriosis and microglandular hyperplasia, can be difficult. This study investigates the value of immunohistochemical staining with MIB1, bcl2 and p16 in the distinction of cervical glandular intraepithelial neoplasia from these benign mimics. METHODS AND RESULTS: Immunohistochemical staining using the monoclonal antibodies MIB1, bcl2 and p16 was performed on cases of cervical glandular intraepithelial neoplasia (n = 21), tubo-endometrial metaplasia (n = 13), endometriosis (n = 7) and microglandular hyperplasia (n = 14). With tubo-endometrial metaplasia and microglandular hyperplasia staining with MIB1 was either negative or involved <10% of cells, while with cervical glandular intraepithelial neoplasia the majority of cases (86%) exhibited >10% positive cells. Two cases of endometriosis exhibited a MIB1 index of 10-30% while in the other cases <10% cells stained. With bcl2 the cells of microglandular hyperplasia were negative although there was staining of associated reserve cells in 43% of cases. All cases of tubo-endometrial metaplasia except one and all cases of endometriosis stained diffusely positive with bcl2. Cases of cervical glandular intraepithelial neoplasia were negative or exhibited focal staining. With p16 all cases of cervical glandular intraepithelial neoplasia exhibited diffuse strong positivity, generally involving 100% of cells, while all cases of microglandular hyperplasia were negative. Sixty-two percent of cases of tubo-endometrial metaplasia showed focal positivity, the remainder being negative. Cases of tubo-endometrial metaplasia were never diffusely positive with p16. In three cases of endometriosis there was staining of >50% of cells while the other cases were either focally positive or negative. CONCLUSIONS: A panel of antibodies, comprising MIB1, bcl2 and p16, is a useful adjunct to histology in distinguishing cervical glandular intraepithelial neoplasia from tubo-endometrial metaplasia, endometriosis and microglandular hyperplasia. Cases of cervical glandular intraepithelial neoplasia are diffusely positive for p16 and generally exhibit a high proliferation index with MIB1, while bcl2 is negative or, at most, focally positive. Tubo-endometrial metaplasia and endometriosis are characterized by strong diffuse positivity with bcl2 and a low proliferation index with MIB1 (although occasional cases of endometriosis show moderate proliferative activity). p16 is negative or exhibits focal positivity in tubo-endometrial metaplasia but in endometriosis there may be quite widespread positivity. Microglandular hyperplasia shows a low proliferation index with MIB1 and is negative for bcl2 and p16.     

Expression of p53 in oesophageal squamous epithelium from surgical specimens resected for squamous cell carcinoma of the oesophagus,with special reference to uninvolved mucosa

Accumulation of p53 protein has been considered an intermediate biomarker in multistage oesophageal carcinogenesis. The aim of the present study was to investigate p53 expression by immunohistochemistry in 13 thoroughly sampled oesophagectomy specimens from a geographical area with a high oesophageal cancer incidence (Basse Normandie, France). Expression of p53 was looked for in tissue samples of cancer, intraepithelial neoplasia, and uninvolved mucosa. The streptavidin biotin peroxidase complex method was used for p53 immunostaining. p53 expression was found in invasive squamous cell carcinoma in 8 out of 11 cases and in intraepithelial neoplasia in 10 out of 11 cases. In all 13 cases, in uninvolved oesophageal mucosa, expression of p53 was focally present in areas of chronic oesophagitis. Chronic oesophagitis has been regarded by epidemiologists as a precursor lesion for squamous cell carcinoma of the oesophagus. Since oesophageal carcinogenesis is a multistage process, the study of precursor lesions could provide information on the timing of p53 gene abnormalities during oesophageal carcinogenesis. These preliminary data require to be confirmed by molecular analysis of the p53 gene. © 1997 John Wiley & Sons, Ltd.     

Expression of cell cycle-regulatory proteins, MIB-1, p16, p53, and p63, in squamous cell carcinoma of conjunctiva: not associated with human papillomavirus infection

Squamous cell carcinoma (SCC), the most common primary malignant tumor of the conjunctiva, has a variable clinical presentation and immunohistochemical profile. Abundant cell cycles exist, including MIB-1 (Ki67 antigen), p16, p53, and p63, within the conjunctiva SCC. This investigation first reports the expressions of cell cycle markers in SCC. A retrospective study was conducted between December 1976 and June 2004, comprising 13 consecutive patients with conjunctiva SCC who were treated with surgical excision. Detailed clinical parameters were also reviewed. Overexpression of MIB-1, p16, p53, and p63 genes were studied by immunohistochemistry. Genechip containing 39 subtypes was used to elucidate human papillomavirus (HPV). The study group contained 13 (100%) men, with a mean age of 68±18 years and follow-up period of 20±17 months. The sample included four (33%) SCC located in the left eye and two (17%) recurrent SCC. Overexpression of the p53 and p63 was considerably higher than that of the p16 (P<0.01). HPV DNA was not detected in any of the 13 cases. This work first examined the immunohistochemical overexpression of cell cycle (MIB-1, p16, p53, and p63) in SCC. This investigation then showed that the expression of cell cycles in SCC was associated with key tumor clinicopathological features. This approach can help distinguish the potential roles of cell cycle in the development of SCC.     

细胞周期素D1和抗原Ki-67在宫颈上皮内瘤样病变和宫颈鳞癌的表达及与人乳头瘤病毒感染转归的关系

目的 研究细胞周期素D1(cyclin D1)、抗原Ki-67在宫颈上皮内瘤样病变(CIN)和宫颈鳞癌发生发展中作用及其与人乳头瘤病毒(HPV)感染转归的关系.方法 2002年1月至2006年12月广州医学院第一附属医院HPV阳性患者104例,分2组:(1)研究组:82例,即病理确诊CIN Ⅰ组17例、CINⅡ组19例、CINⅢ组23例、宫颈鳞癌组23例.(2)对照组:柱状上皮异位22例.应用EnVision法检测宫颈病变组织中cyclin D1、Ki-67蛋白的表达,杂交捕获试验检测宫颈分泌物或阴道残端中HPV感染情况,随访各组患者术后1年内的HPV变化.结果 (1)cyclin D1在各组宫颈组织细胞核内均有表达.其阳性率CINⅢ组[82.61%(19/23)]、宫颈鳞癌组[86.96%(20/23)]与对照组[27.27%(6/22)]、CINⅠ组[58.82%(10/17)]比较,差异有统计学意义(P<0.05),在宫颈鳞癌组与CINⅡ组[68.42%(13/19)]阳性率比较差异有统计学意义(P<0.05).(2)Ki-67在各组宫颈组织细胞核内均有表达,其对照组阳性率[31.82%(7/22)]与CINⅢ组[86.96%(20/23)]、宫颈鳞癌组[91.30%(21/23)]比较差异有统计学意义(P<0.05),而在宫颈鳞癌组与CINⅠ组[58.82%(10/17)]、CIN Ⅱ组[63.16%(12/19)]比较差异有统计学意义(P<0.05).(3)术后1年内各组HPV的转阴率分别与cyclin D1、Ki-67的表达强度呈负相关(rs=-0.299,rs=-0.367,P<0.05).结论 cyclinD1和Ki-67在CIN和宫颈鳞癌的细胞增殖活动中起作用,且两者可能与HPV感染转阴率有关.     

Immunohistochemical expression of p63, p53 and MIB-1 in urinary bladder carcinoma. A tissue microarray study of 158 cases

P63 is a member of the p53 family, which plays a role in the differentiation of urothelium and is supposed to play a role in urothelial carcinogenesis. P53 and MIB-1 are recognised in many studies as predictive markers of progression, but few studies in the literature have examined p63. The aims of our study were to explore the expression of p63 in bladder carcinomas and to compare this expression to p53 and MIB-1, as well as to stage and grade. Tissue microarrays were performed on 158 urothelial carcinomas (56 pTa, 45 pT1 and 57≥pT2). Immunohistochemical studies were performed with p63, p53 and MIB-1 antibodies. In our study we observed that p63 immunostaining is present in all cell layers in papillary urothelial neoplasm of low malignant potential (PUNLMP), but partially lost in non-invasive papillary urothelial carcinoma low grade (NILGC) and in pT1/≥pT2 bladder cancers. P53 and MIB-1 displayed lower expression in PUNLMP/NILGC vs non-invasive papillary urothelial carcinoma high grade (NIHGC)/pT1, but there was no correlation between the expression of p63, p53 and MIB-1. Our study demonstrates that p63 expression distinguishes between PUNLMP/NILGC and NIHGC/pT1 (p=4.10⁵). A statistical difference disserving pTa and pT1/≥pT2 with a statistical significance (p<10⁻⁶) could also be observed. P63 should be considered as an additional biomarker that might help pathologists to classify their patients.     

Immunohistochemical study of DNA topoisomerase I, DNA topoisomerase II alpha, p53, and Ki-67 in oral preneoplastic lesions and oral squamous cell carcinomas

Human DNA topoisomerase I (topo I) is the molecular target of the camptothecin group of anticancer drugs. Laboratory studies have shown that the cellular response to topo I-targeted drugs depends on the topo I expression and DNA replication rate and the apoptotic pathway activity. In this study, we tested potential indicators of the sensitivity of topo I-targeted drugs in 36 cases of oral squamous cell carcinoma (OSCC). Formalin-fixed, paraffin-embedded tissue sections were immunostained with monoclonal antibodies against Ki-67, p53, and topo I, and with polyclonal antibodies against DNA topoisomerase II-alpha (topo II-alpha). These markers were also tested in 18 epithelial hyperplastic lesions and 18 mild dysplasias. Immunostaining was quantified by the percentage of stained nuclei in each sample (the labeling index); 200 immunoreactive epithelial nuclei were counted per case for each antibody. The results support the possibility of using topo II-alpha staining for assessing the proliferative activity. High expression of topo II-alpha and topo I in OSCCs suggests that they may serve as potential indicators of sensitivity to topo I inhibitors. However, the apoptotic pathway assessed by p53 immunostaining was found to be uninformative. Analysis of the relationship between immunohistochemical results and clinical and pathologic parameters (the T and N stages and differentiation) showed that only the differentiation parameter correlated with the topo I expression rate. Thus, significant increase in the topo I expression in the poorly differentiated OSCCs suggests their higher sensitivity to drug treatment.     

Genetic alterations in head and neck squamous cell carcinoma: The next-gen sequencing era

Head and neck squamous cell carcinoma is the sixth most common cancer in the world with approximately 650000 new cases diagnosed annually. Next-generation molecular techniques and results from phase 2 of the Cancer Genome Atlas becoming available have drastically improved our current knowledge on the genetics basis of head and neck squamous cell carcinoma. New insights and new perspectives on the mutational landscape implicated in head and neck squamous cell carcinoma provide improved tools for prognostication. More importantly, depend on the patient’s tumor subtypes and prognosis, deescalated or more aggressive therapy maybe chosen to achieve greater potency while minimizing the toxicity of therapy. This paper aims to review our current knowledge on the genetic mutations and altered molecular pathways in head and neck squamous cell carcinoma. Some of the most common mutations in head and neck squamous cell carcinoma reported by the cancer genome atlas including TP53, NOTCH1, Rb, CDKN2A, Ras, PIK3CA and EGFR are described here. Additionally, the emerging role of epigenetics and the role of human papilloma virus in head and neck squamous cell carcinoma are also discussed in this review. The molecular pathways, clinical applications, actionable molecular targets and potential therapeutic strategies are highlighted and discussed in details.     

Expression of CD34, alpha-smooth muscle actin, and transforming growth factor beta1 in squamous intraepithelial lesions and squamous cell carcinoma of the larynx and hypopharynx

AIM OF THE STUDY: There is increasing evidence that stromal reaction in cancer has an important diagnostic and prognostic significance. Recent studies have shown that CD34-positive stromal cells and myofibroblasts may play an important role in host response to invasive cancer. The aim of our study was to analyze the expression of CD34, alpha-smooth muscle actin (SMA), and transforming growth factor beta1 (TGFbeta1) in squamous intraepithelial lesions (SILs) and squamous cell carcinoma (SCC) of the larynx and hypopharynx, to establish their significance, and to elucidate the mechanism of myofibroblast formation. METHODS: Immunohistochemistry was performed on samples of 42 resected larynges and 12 laryngeal biopsies of SILs and SCC using antibodies against SMA, CD34, CD31, TGFbeta1, and TGFbeta1 receptors. The expression of TGFbeta1 mRNA was detected with RNA in situ hybridization using specific oligonucleotides for TGFbeta1. RESULTS: The stroma in normal laryngeal mucosa and SILs contained scattered CD34-positive cells, but there were no SMA-positive myofibroblasts. In contrast, the stroma of SCC contained SMA-positive myofibroblasts, but there were no CD34-positive stromal cells. This pattern of stromal reaction was also observed in the peritumoral zone. In adjacent normal tissue, there were CD34-positive stromal cells and no myofibroblasts. We found more intense TGFbeta1 expression in carcinoma cells than in the normal laryngeal epithelium and positive staining for both TGFbeta1 receptors on stromal cells of the normal mucosa. In SCC, many myofibroblasts expressed TGFbeta1 and both receptors for TGFbeta1. Expression of TGFbeta1 mRNA was similar to expression of TGFbeta1 protein. CONCLUSION: Our study shows that disappearance of CD34-positive stromal cells and appearance of SMA-positive stromal myofibroblasts are associated with transformation of laryngeal SILs to SCC. This pattern of stromal reaction was found not only in the tumor but also in the peritumoral zone, defined as a band of host tissue between the invasive tumor front and adjacent normal tissue. Our findings also support the suggestion that overproduced TGFbeta1 in carcinoma cells mediates one of the mechanisms of transformation of stromal cells to myofibroblasts in laryngeal carcinogenesis.     

Immunohistochemical analysis of p53, APE1, hMSH2 and ERCC1 proteins in actinic cheilitis and lip squamous cell carcinoma

Souza L R, Fonseca‐Silva T, Pereira C S, Santos E P, Lima L C, Carvalho H A, Gomez R S, Guimarães A L S & De Paula A M B
(2011) Histopathology 58 , 352–360
Immunohistochemical analysis of p53, APE1, hMSH2 and ERCC1 proteins in actinic cheilitis and lip squamous cell carcinoma Aims: This study has compared the tissue expression of the p53 tumour suppressor protein and DNA repair proteins APE1, hMSH2 and ERCC1 in normal, dysplastic and malignant lip epithelium. Methods and results:  Morphological analysis and immunohistochemistry were performed on archived specimens of normal lip mucosa (n = 15), actinic cheilitis (AC) (n = 30), and lip squamous cell carcinoma (LSCC) (n = 27). AC samples were classified morphologically according to the severity of epithelial dysplasia and risk of malignant transformation. LSCC samples were morphologically staged according to WHO and invasive front grading (IFG) criteria. Differences between groups and morphological stages were determined by bivariate statistical analysis. Progressive increases in the percentage of epithelial cells expressing p53 and APE1 were associated with increases in morphological malignancy from normal lip mucosa to LSCC. There was also a significant reduction in epithelial cells expressing hMSH2 and ERCC1 proteins in the AC and LSCC groups. A higher percentage of malignant cells expressing APE1 was found in samples with an aggressive morphological IFG grade. Conclusions:  Our data showed that epithelial cells from premalignant to malignant lip disease exhibited changes in the expression of p53, APE1, hMSH2 and ERCC1 proteins; these molecular change might contribute to lip carcinogenesis.     

Immunohistochemical detection of p53 homolog p63 in solid cell nests, papillary thyroid carcinoma, and hashimoto's thyroiditis: A stem cell hypothesis of papillary carcinoma oncogenesis

Most models suggest that the cell of origin of papillary carcinoma is the mature thyroid follicular epithelial cell. In a recent study, p63 was detected in papillary carcinoma, Hashimoto's thyroiditis, and in squamoid aggregates and solid cell nests (SCNs), embryonic remnants found sporadically in the fully developed thyroid. In the present study, the relationship between solid cell nests and papillary carcinoma was investigated further. Four-micrometer sections from 88 routinely fixed and processed archival thyroidectomy specimens were pretreated with citric acid pH 6.0 for antigen retrieval, then incubated overnight with anti-p63 monoclonal antibody 4A4. Slides were stained with a streptavidin-biotin kit and diaminobenzidine as chromogen and were counterstained with hematoxylin. Squamoid aggregates or SCNs were noted in 21 specimens. Several morphologic variants of SCNs were found, all of which displayed p63 positivity. These included undifferentiated SCNs and those displaying commitment toward squamoid and ciliated glandular differentiation. Small, morphologically inconspicuous aggregates of p63-positive cells were commonly found in Hashimoto's thyroiditis. Commitment of p63-positive undifferentiated cells toward thyroid follicular epithelial differentiation was occasionally noted. One SCN variant, also associated with Hashimoto's thyroiditis, was a floretlike arrangement of p63-positive cells with fusiform nuclei. p63 staining was strong and uniform in some SCNs, but in other SCNs it was compartmentalized and homologous to stem cell-staining patterns in normal squamous or bronchial epithelia. Stem cell-like staining, associated with compartmentalized p63 staining or p63-positive undifferentiated cells, was noted in 7 of 27 papillary carcinomas. p63 immunostaining is a highly sensitive means of detecting SCNs. p63 expression patterns in SCNs and a subset of papillary carcinomas are closely homologous to stem cell-associated p63 staining patterns that have been described elsewhere in squamous and bronchial epithelia. We propose a stem-cell-associated model of papillary carcinoma oncogenesis that suggests that (1) p63-positive embryonal remnants rather than mature follicular cells are the cells of origin of a subset of papillary carcinomas; (2) these p63-positive cells are pluripotent and may stay undifferentiated or undergo benign squamoid or glandular maturation, may undergo thyroid follicular epithelial differentiation, may undergo oncogenic change leading to papillary carcinoma, or may trigger an immune reaction, resulting in lymphoid infiltration and Hashimoto's thyroiditis; and (3) Hashimoto's thyroiditis and papillary carcinoma may therefore be linked etiologically, because both disorders may be initiated by the same population of pluripotent p63-positive embryonal stem cell remnants.     

Utility of WT-1, p63, MOC31, mesothelin, and cytokeratin (K903 and CK5/6) immunostains in differentiating adenocarcinoma, squamous cell carcinoma, and malignant mesothelioma in effusions

To distinguish carcinoma, either adenocarcinoma (ADC) or squamous cell carcinoma (SCC), and malignant mesothelioma (MM) in effusion can be a diagnostic challenge based on morphology alone. This study evaluates the utility of WT-1, p63, MOC31, mesothelin, and cytokeratin (K903 and CK5/6) immunostains in effusions when ADC and SCC of the lung are in the differential diagnosis with MM. A cohort of 43 effusions consisting of lung ADC (N = 10), SCC (N = 15), and MM (N = 18, mostly (16) pleural based), was subjected to immunostains using the above mentioned antibodies. WT-1 was positive in 100% MM, 0% ADC, and 0% SCC cases while p63 was positive in 0% MM, 30% ADC, and 80% SCC cases. Stain for MOC31 was positive in 100% ADC, 67% SCC, and 35% MM cases. Similarly, mesothelin antibody stained 100% ADC, 60% SCC, and 47% MM cases. Antibodies for K903 and CK5/6 stained 100% SCC cases but fewer ADC cases (40 and 10%, respectively). In conclusion, in this cohort of mostly pleural malignant effusion, MM can be identified with positive staining for WT-1 and negative staining for p63. Conversely, negative staining with WT-1 and positive staining for p63 exclude MM. Used as part of an immunostain panel, cytokeratin markers (CK5/6 and K903) are useful in differentiating SCC from ADC when MM is already excluded, and MOC31 might have limited value in differentiating ADC from MM. A negative stain with MOC31 can exclude lung ADC. Mesothelin, on the other hand, is not useful in the differential diagnosis of ADC, SCC, and MM.